University Hospitals
Case Medical Center
MetroHealth
Medical Center
Louis stokes va
Medical Center volume 5 issue 1 | 2008
volume 4 issue 1 | 2007
Case Western reserve University

© 2008 University Hospitals ORT-00038
ORT-00038 journal ad_r3.indd 1
Iris S. and Bert L. Wolstein
Research Building –
Case Western Reserve University
School of Medicine
University Hospitals
Alfred and Norma
Lerner Tower
University Hospitals
Samuel Mather Pavilion
University Hospitals
New Cancer Hospital
Facility to open in 2010
Case Western Reserve University
School of Medicine
University Hospitals
Lakeside Hospital
University Hospitals
Rainbow Babies &
Children’s Hospital
University Hospitals
Adult Center for Emergency and Marcy R. Horvitz
Pediatric Emergency Center
New facility to open in 2010
9/8/08 11:43:09 AM

© 2008 University Hospitals ORT-00038
ORT-00038 journal ad_r3.indd 1
Iris S. and Bert L. Wolstein
Research Building –
Case Western Reserve University
School of Medicine
University Hospitals
Alfred and Norma
Lerner Tower
University Hospitals
Samuel Mather Pavilion
University Hospitals
New Cancer Hospital
Facility to open in 2010
Case Western Reserve University
School of Medicine
University Hospitals
Lakeside Hospital
University Hospitals
Rainbow Babies &
Children’s Hospital
University Hospitals
Adult Center for Emergency and Marcy R. Horvitz
Pediatric Emergency Center
New facility to open in 2010
9/8/08 11:43:09 AM

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5

Resident Editors
Matthew Smith, M.D.
Editor-in-chief
Andrew Islam, M.D.
Editor-in-chief Elect
Parke Oldenburg, M.D.
Anthony Skalak, M.D.
Senior Editors
Michael Chen, M.D.
Steven Fitzgerald, M.D.
Junior Editors, Advertising
Daniel Masters, M.D.
Ryan Garcia, M.D.
Patrick Messerschmitt, M.D.
James Murphy, M.D.
Junior Editors
Faculty Editors
Randall Marcus, M.D.
Edward Greenfield, Ph.D.
J. Robert Anderson, M.D.
Shana Miskovsky, M.D.
Ellen Greenberger
Secretary
Phyllis Lie
Accounting
Letter from the Editor-in-chief .............................................................................................................................7
Dedication to Dwight T. Davy , Ph.D. ..................................................................................................................8
Year in Review
Chairman’s Report – Dr. Randall Marcus ..................................................................................................10
Louis Stokes Cleveland Department of Veterans Affairs Medical Center Orthopaedic Surgery Service
Annual Report – Dr. Patrick Getty .............................................................................................................13
Research Section of the Department of Orthopaedics at Case Western Reserve University School of Medicine – Dr. Edward Greenfield ........................................................................................................14
Photos from Throughout the Year .....................................................................................................................15
University Hospitals Attendings ........................................................................................................................20
MetroHealth Attendings ....................................................................................................................................21
Basic Science Faculty .......................................................................................................................................22
VAMC Attendings ..............................................................................................................................................23
Awards Section
Nicolas Andry Lifetime Achievement Award .............................................................................................24
Orthopaedic Research Society - OREF Distinguished Investigator Award ..............................................25
Arthur H. Heune Memorial Award .............................................................................................................25
Kingsbury G. Heiple, M.D., and Fred A. Lenon Profesor of Orthopaedics Endowed Chair .....................26
American Orthopaedic Association Resident Leadership ........................................................................27
Manuscripts
Developmental, Topographical, and Radiographic Anatomy of the Femoral Intercondylar Notch ..........28
Metastatic Osteosarcoma of the Spine in an Adolescent ........................................................................33
Paralyzed Nerve Transfer for Denervation: A Case Report .......................................................................38
The Cre-loxP System in Bone and Cartilage Research ............................................................................40
ERK1 and ERK2 Negatively Regulate the Size of Cartilaginous Skeletal Elements .................................47
Surgical Management of Piriformis Syndrome .........................................................................................53
Minimally Invasive Aproach for Drilling of the Posterior Cruciate Ligament Femoral Tunnel ...................57
Paralyzed Nerve Transfer for Denervation: Histological Results from an Animal Model .........................59
Expansion of the Coordinator Role in Orthopaedic Residency Program Management ...........................66
SOX9 and RUNX2: the Two Master Regulators of Skeletogenesis ..........................................................71
How Can Bone Turnover Modify Bone Strength Independent of Bone Mass? ........................................77
Reherniation and Failure Following Lumbar Discectomy .........................................................................84
Chief Resident Research Symposium ...............................................................................................................88
Visiting Professors
Bohlman Visiting Professor – Dr. Lawrence Lenke ...................................................................................89
RBC Visiting Professor – Dr. Peter Waters ................................................................................................89
Allen Fellow Professor .............................................................................................................................90
Obituaries
Dr. Lawrence Samuel Cohen ....................................................................................................................91
Dr. Curtis W. Smith ....................................................................................................................................91
Exiting Residents’ Future Plans .........................................................................................................................92
Incoming Interns - Class of 2013 ......................................................................................................................93
Instructions for Authors .....................................................................................................................................94
6

Matthew Smith, M.D.
W e are pleased to bring you the fifth edition of the Case
Orthopaedic Journal . We strive to offer the highest quality publication that highlights the outstanding academic achievements in the Department of
Orthopaedics at Case Western Reserve
University. This journal is made possible because of our institution’s commitment to research and the hard work of those conducting the research. It is an honor to share these accomplishments with you.
This edition of the Case Orthopaedic
Journal is dedicated to Dr. Dwight
Davy, one of our distinguished professors in the Department of
Mechanical and Aerospace Engineering.
Dr. Davy has made significant contributions to orthopaedics through his research endeavors. He has also served as a committed teacher, a reliable mentor and a trusted friend to many in our department. Dr. Davy is retiring this year. We will miss both his presence in the laboratory and his enthusiasm for education. We wish him the very best.
The Department of Orthopaedics at Case Western Reserve University continues a proud tradition of producing exceptional research while maintaining a firm commitment to outstanding patient care. It has been a privilege to be a part of this tradition. It has also been an honor to work with an outstanding editorial staff that offered their valuable time to put this journal together. I would like to thank them for their hard work and dedication.
I would also like to thank the many others not on the editorial staff for their support and commitment to make this journal a success. I hope that the Case
Orthopaedic Journal reflects the pride we all have in our research program and our department.
OrtHOPaeDiC JOUrnaL | Case Western reserve University | vOL.5, nO.1 | 2008 | 7

8
O n July 1, 2008, Dwight Davy,
Ph.D., became an emeritus faculty member of the Department of
Mechanical and Aerospace Engineering and the Case School of Engineering at
Case Western Reserve University. At this important career transition, it is an opportune time to honor Dwight and his many accomplishments.
Dwight began his career at the
University of Nebraska where he received his B.S.M.E. in Mechanical
Engineering in 1962, followed by an
M.S. in Engineering Mechanics in
1964. California called to Dwight and his wife Kathlene, and he worked as a
Project and Research Engineer on the mechanics of materials and structures for radiation and blast resistance at
Lawrence Livermore Laboratories from
1964 to 1969. However, his love for learning and academics brought him back to the Midwest where he pursued a Ph.D. at the University of Iowa in
Mechanics and Hydraulics. The grad student lifestyle was stimulating but also a bit of a “scramble” for Dwight and Kathlene, as they were raising three young children (Sheri, Randy and Lane) during this time. Following receipt of his Ph.D. in 1972 (on a topic totally unrelated to orthopaedics – a theoretical study of nonlinear wave propagation), he then became an
Assistant Professor at the University of Iowa. A turning-point in his career occurred as he was completing his
Ph.D. dissertation; he began to feel a pull towards orthopaedics and he extended his stay in Iowa City for a year to work on a project on hip mechanics with Dick Johnston,
M.D. Dwight then landed a faculty position as an Assistant and then
Associate Professor in Engineering
Mechanics Mechanic and Hydraulics at the University of
Nebraska-Lincoln.
His academic career kept him busy teaching three to four core engineering courses per semester (e.g.,
Statics, Dynamics,
Numerical Methods).
However, he was motivated to pursue his research interest in orthopaedic biomechanics and he established a relationship as an
Adjunct Professor in Orthopaedics at the University of
Nebraska Medical
Center. Through this appointment, he conducted research on fracture repair and fracture fixation with John Connolly, M.D., then Chair of the Department of Orthopaedics.
Then, in 1978, another pivotal point in his career occurred when Dwight had the opportunity to spend the summer as a Visiting Scientist with Albert
Burstein, Ph.D, in the Department of Biomechanics at The Hospital for
Special Surgery. It was during this time that he formulated his strong desire to join a University where he could actively pursue his research goals in addition to his academic goals. So, he,
Kathlene and the children were “on the road again”, this time for Case Western
Reserve University.
Dwight came to CWRU in 1979 as an
Associate Professor in the Department of Mechanical and Aerospace
Engineering with a secondary appointment in Orthopaedics. Dwight took up the baton and became the steward of the collaboration between
Orthopaedics and Mechanical and
Aerospace Engineering that was initiated by Al Burstein, Victor Frankel,
M.D., and Kingsbury Heiple, M.D. in 1967. He co-directed and then directed the Orthopaedic Engineering
Labs from 1982 to 2000. He was promoted to Full Professor in 1987.
Over the years, Dwight developed an outstanding ability to bridge fundamental biomechanics and clinical research, as evidenced by his appointment as Director of Research in the Department of Orthopaedics from
1990 to 1999 and by his position as
Chair of the NIH Orthopaedics and
Musculoskeletal Study Section from
1991-1993. To date, Dwight is still

one of only two engineers to serve in that latter role. Dwight’s leadership excellence also led to his election as
Program Chair (1996) and President
(1999) of the Orthopaedic Research
Society. Sadly, in 2000, Dwight lost his dear wife and partner, Kathlene, following a prolonged illness.
Throughout his career, Dwight has been funded nearly continuously by the NIH to conduct research on joint mechanics and bone damage mechanics. Notably, along with King
Heiple, M.D., and Victor Goldberg,
M.D., he conducted ground-break research on in vivo hip force analysis using telemetrized implants in the late
1980s. Their work was translational and interdisciplinary long before anyone had thought of such labels.
Among many honors, Dwight was a co-recipient of an AAOS Kappa Delta
Young Investigator Award for studies on the biology and mechanics of bone allografts. He has published numerous articles, book chapters, and technical reports. He is a co-author (along with
Don Bartel, Ph.D. and Tony Keaveny,
Ph.D.) of a textbook for engineering students on musculoskeletal mechanics:
“Orthopaedic Biomechanics
: Mechanics and Design in
Musculoskeletal Systems” (Prentice-
Hall, 2006). Over the years, Dwight contributed significantly to the scholarly and academic mission of the University: he advised more than
30 M.S. and Ph.D. students and more than 20 post-doctoral fellows
(including Harry Figgie III, M.D. and
Matthew Kraay, M.D.). All of this was accomplished while teaching core courses in mechanical engineering to hundreds of undergraduate and graduate students, and also advising them and guiding them in their career development.
In closing, Dwight is a highly accomplished scholar and researcher.
However, he is also a man whose integrity, values, and faith have guided his life and career. All who know
Dwight know that his heart is as big as the “big sky” in the state of Kansas in which he was born. He has been and continues to be a close friend and wise mentor to me and to many others.
As he now embarks on his emeritus
“career”, we send our best wishes to
Dwight and his soon-to-be new wife,
Judy.
Clare Rimnac, Ph.D.
Chair, Mechanical and Aerospace
Engineering
September, 2008
OrtHOPaeDiC JOUrnaL | Case Western reserve University | vOL.5, nO.1 | 2008 | 9

10
I am delighted to introduce this year’s volume of the Case Orthopaedic
Journal, which highlights the outstanding achievements of the
Department of Orthopaedics at
Case Western Reserve University
School of Medicine. The Department continues its ranking as one of the top orthopaedic departments in the United
States, which makes me extremely proud of the outstanding achievements and excellent work carried out during the past year by our clinicians, scientists, residents and staff.
The Department of Orthopaedics at Case Western Reserve University consists of our three medical centers, our research laboratories and, most importantly, the outstanding people who have earned our reputation for excellence. Our medical centers include:
• University Hospitals Case Medical
Center, which includes Rainbow Babies and Children’s Hospital,
• MetroHealth Medical Center, our
Level I trauma hospital, and
• Louis Stokes Veterans Administration
Medical Center here on our Case campus.
Our basic science laboratories are located:
• in the School of Medicine, with our
Molecular Biology division in the
Biomedical Research Building,
• in the Case School of Engineering, in the Musculoskeletal Mechanics and
Materials Laboratory, and
• at MetroHealth Medical Center and the Veterans Administration Medical
Center, where our Functional Electrical
Stimulation Laboratories are located.
Additionally, our Anatomic Research
Laboratory resides at the Cleveland
Museum of Natural History, the home of the Hamann-Todd bone collection.
Departmental Achievements
The Department’s excellence in clinical activities was once again recognized by
U.S. News & World Report (17th). Our national leadership in musculoskeletal research is confirmed by our continued high ranking (4th) as one of the topfunded orthopaedic departments in the
United States by the National Institutes of Health (NIH). Our residency program received 501 applications for our 6 residency positions, and the
Department matched 6 of our top selections. We welcome to the program
Dr. Shane Hanzlik from the University of Nevada, Dr. Scott Kling from the University of Pennsylvania, Dr.
Ethan Lea from Case Western Reserve
University, Dr. James Learned from the
University of Southern California, Dr.
Jonathan Macknin from the University of Pennsylvania and Dr. Lorraine Stern randall e. Marcus, M.D.
from George Washington University.
In addition, we welcome our two new
Spine Fellows, Dr. Anil Kesani from the
University of Medicine and Dentistry of
New Jersey and Dr. Jason Tinley from the Fort Worth Orthopaedic Residency
Program, and our Pediatric Orthopaedic
Fellow, Dr. Amir Abdelgawad from the
University of Toledo Medical Center.
This year’s Trauma Fellow is Dr. Andrew
Steiner from the State University of
New York. The two Allen Research
Fellowships were awarded this year to
Dr. Troy Mounts , who will be working in our tissue engineering laboratory, and
Dr. Erik Schnaser , who will be working in our bone biology laboratory.

YEAR IN REVIEW
Congratulations to Faculty Members and Residents
On June 9, 2008, Dr. Heather A.
Vallier was appointed the inaugural holder of the Clyde L. Nash, Jr,
MD, Professorship in Orthopaedic
Surgery at Case Western Reserve
University. The Nash Professorship was established by generous donations from
Dr. Nash’s friends and grateful patients.
It honors Clyde L. Nash, Jr, MD,
Professor Emeritus of Orthopaedics, who directed the Department of
Orthopaedics at MetroHealth Medical
Center from 1992 to 1998. Dr. Nash, an internationally renowned expert in scoliosis, is the Past President of the
Scoliosis Research Society (1982) and an honorary member of the British
Scoliosis Research Society. He is the author of numerous peer-reviewed articles and book chapters in spinal deformity surgery. His seminal work in spinal cord monitoring during operative spinal procedures has markedly improved the safety of spinal-deformity surgery throughout the world. Dr. Heather Vallier received her B.A. degree in biochemistry from Northwestern University and her medical degree from Stanford
University. Following her orthopaedic residency at the University of
Wisconsin and her Trauma Fellowship at the University of Washington, she joined our Department at MetroHealth
Medical Center in 2001. Dr. Vallier has received national recognition for her expertise in orthopaedic traumatology and was honored by the American
Orthopaedic Association when she was selected as a North American Traveling
Fellow in 2005.
Dr. Richard Grant received a
$916,000 grant from the St. Luke’s
Foundation as part of the establishment of the Timothy L. Stephens, Jr, MD,
Orthopaedic Fellowship at University
Hospitals Case Medical Center. Dr.
Stephens, an Emeritus member of the Department of Orthopaedics at
Case Western Reserve University, was the first African-American member of our Department. Dr. Stephens was also Chairman of Orthopaedics at St. Luke’s Hospital for many years. This Fellowship will support underrepresented minority medical students as they perform research in the
Department of Orthopaedic Surgery at Case Western Reserve University.
Additionally, Dr. Grant was selected by
Black Enterprise Magazine as one of the top African-American physicians in the United States and one of four in the
State of Ohio.
Dr. Victor M. Goldberg received the
1st annual Distinguished Investigator
Award given jointly by the Orthopaedic
Research Society and the Orthopaedic
Research and Education Foundation at the annual AAOS meeting in San
Francisco this year.
Dr. Brendan Patterson , Chief of
Orthopaedic Surgery at MetroHealth
Medical Center, was elected as a member in the American Orthopaedic
Association and was elected as a
Director of the Orthopaedic Trauma
Association.
Dr. Henry H. Bohlman , Director of the Spine Center at University
Hospitals Case Medical Center, received the Nicolas Andry Award sponsored by the Association of
Bone and Joint Surgeons. This award recognizes Dr. Bohlman for a lifetime of achievements and advances in the treatment of spinal disorders. Dr.
Bohlman was also the honored guest at the 15th Annual Advanced Techniques in Cervical Spine Seminar in St. Louis,
Missouri.
Dr. George H. Thompson , Director of Pediatric Orthopaedic Surgery, was awarded the Huene Award by the
Pediatric Orthopaedic Society of North
America. The statement regarding the award observes:
“The Arthur H. Huene Memorial
Award for Excellence and Promise in
Pediatric Orthopaedics . . . is made to an individual who has made an outstanding contribution in the field of Pediatric Orthopaedics and who offers the potential of continuing that contribution in the future.”
This award, to a senior-level pediatric orthopaedic surgeon, is the highest award given by the Pediatric
Orthopaedic Society of North America.
Dr. Jack Wilber presided this year as President of AO North America and as a Trustee of the International
AO Foundation. He was the invited guest lecturer at the Trauma Center
Symposium organized by the National
College of Surgeons of France held in Paris last spring. He was also the international guest lecturer at the AO
Nordic Course held in Stockholm,
Sweden, this year.
Dr. Edward M. Greenfield , Director of Orthopaedic Research, Dr. Patrick
J. Getty , Director of the Division of
Orthopaedic Oncology, Dr. Patrick
J. Messerschmitt , orthopaedic resident, Dr. Ryan M. Garcia , orthopaedic resident, and Ashley
Rettew and Robert Brookover , both research associates in the Department of Orthopaedics, were invited to participate in the Clinical Orthopaedics and Related Research Symposium on
Molecular Genetics and Sarcoma, for their research involving the regulation of osteosarcoma cells.
Dr. Ronald Triolo was invited to be a
Visiting Professor at Rutgers University, where he presented his work on neuroprostheses.
In Shun Murakami, MD, PhD ’s laboratory, Dr. Takehiko Matsushita
OrtHOPaeDiC JOUrnaL | Case Western reserve University | vOL.5, nO.1 | 2008 | 11

CHAIRMAN’S REPORT
12 received the New Investigator
Recognition Award at the 54th Annual
Meeting of the Orthopaedic Research
Society in San Francisco.
Dr. Eben Alsberg was selected as editor of a special edition of Tissue
Engineering , “Technologies for
Enhancing Tissue Engineering.”
Dr. Fadi Abdul-Karim published the book chapter, “Fine needle aspiration biopsy of soft tissue tumors,” in the recently republished text Soft Tissue
Tumors.
Further honors received by the
Department included awards to our orthopaedic residents. Dr. Christopher
McAndrew was honored with the
American Orthopaedic Association/
Orthopaedic Research and Education
Foundation Resident Leadership
Forum Award and he represented the Department at the 121st Annual
Meeting of the American Orthopaedic
Association in Quebec City.
Case Orthopaedic residents swept the
Annual Barry Friedman Orthopaedic
Research Awards for Northeast Ohio, with Dr. Robert J. Gillespie ’s study,
“Biomechanical Evaluation of Threepart Proximal Humeral Fractures: A
Cadaveric Study,” winning 1st place and Dr. Raymond W. Liu ’s study,
“Prospective Comparison of Supine
Bending, Push Prone, and Traction
Under General Anesthesia Radiographs in Predicting Curve Flexibility and
Postoperative Correction in Adolescent
Idiopathic Scoliosis,” receiving 2nd place. Dr. Liu’s research work was also selected for presentation at the Annual
Meeting of the American Academy of Pediatrics and at the International
Pediatric Orthopaedic Symposium.
Dr. Daniel Master received 2nd place in the Ohio Orthopaedic Society’s
Annual Resident Research papers, with his study on “Paralyzed Nerve
Transfer for Denervation in the Setting of Spinal Cord Injury: An Animal
Model.” Dr. Patrick J. Messerschmitt was a 3rd place winner in the
Orthopaedic Research and Education
Foundation Ohio State Society
Resident Competition for his work,
“3,4-Methylenedioxyβ -nitrostyrene
(MNS) Reduces the Motility and
Colony Formation of Osteosarcoma
Cells.”
This year’s chief residents, who graduated in June, were another outstanding class. They are all advancing on to Fellowships in their subspecialty areas of choice, and we welcome them into the Case/Herndon
Alumni Association and wish them all the best in their future careers:
• Clayton Dean, MD , Spine
Fellowship, Emory University
• Jason Eubanks, MD , Spine
Fellowship, University of Pittsburgh
• Brian Hardy, MD , Hand and Upper
Extremity Fellowship, Jacksonville,
Florida
• Parke Oldenburg , MD, Trauma
Fellowship, University of Washington,
Seattle
• Matt Smith, MD , Sports Medicine
Fellowship, University of Michigan,
Ann Arbor
• Glenn Wera, MD , Adult
Reconstruction and Joint Replacement
Fellowship, Rush Medical Center,
Chicago
Once again, it has been a privilege to lead this fabulous Orthopaedic
Department in its 101st year. I hope you will enjoy this volume of the Case
Orthopaedic Journal , which highlights the outstanding work that typifies the faculty, residents and staff of this
Department.

YEAR IN REVIEW
T he Orthopaedic section at the
Cleveland Veterans Affairs Medical
Center (VAMC) continues to be a hub of education and service. During the 2007-2008 academic year, we performed 479 operations (with eleven different attending surgeons) and 6,165 outpatient evaluations. The VAMC section provides this tremendous clinical experience for the four residents and spine fellows that are here for their four month rotations.
The section is divided into the Sports/
Spine team and the Arthroplasty/
Upper Extremity team. Each team consists of one chief resident, one
PGY-3 resident and various attendings with a spine fellow on the spine team. Attendings actively involved at the VA include Nicholas Ahn, MD
(spine), J. Robert Anderson, MD
(upper extremity), Patrick Getty, MD
(orthopaedic oncology and general orthopaedics), Richard Grant, MD
(arthroplasty), Randall Marcus, MD
(foot/ankle and amputation), Thomas
McLaughlin, MD (sports medicine and general orthopaedics) and William
Petersilge, MD (arthroplasty). We performed 121 joint replacements, 118 upper extremity operations, 64 spine operations, 59 sports operations, 28 foot and ankle operations and 89 other
(mostly trauma) operations.
The tradition of funded research continues with the ongoing functional electrical stimulation program for spinal cord injury patients under the direction of Ron Triolo, PhD.
Our current clinical research project, headed by Richard Grant, MD, is studying Joint Replacement Utilization
Disparities in the VA patient population and is nearing completion.
Patrick Getty, M.D.
OrtHOPaeDiC JOUrnaL | Case Western reserve University | vOL.5, nO.1 | 2008 | 13

14
I am pleased that this issue of the COJ is dedicated to Dwight Davy, who recently became an Emeritus Professor in the Engineering School. Dwight’s collaborations with Victor Goldberg and other members of our department have been the foundation for the long-standing interactions between the
Department of Mechanical Engineering and the Department of Orthopaedics.
Dwight was one of the first people
I met at CWRU and he has always been enthusiastically supportive. He’s looking forward to devoting more time to research now that his teaching and administrative responsibilities are less demanding. We look forward to his continued long-term successes.
Each year, two of our residents are selected as Allen Fellows, who join a research lab for a full-time, year-long, experience. The 2007-2008 Allen
Fellows were Dan Master, who worked with Harry Hoyen and Bob Kirsch
(Biomedical Engineering) on nerve repair, and James Murphy, who worked with Matt Kraay and Clare Rimnac on polyethylene damage. Dan and Jim have already made numerous presentations and submitted multiple publications based on their Fellowship research. The two Allen Fellows for the 2008-2009 year are Troy Mounts and Erik Schnaser.
Troy’s primary project involves cartilage tissue engineering in Jim Dennis’ lab and
Erik’s primary project involves regulation of PTH signaling in my lab. We look forward to both of them following in the footsteps of the previous Allen Fellows by being extremely productive.
The 2007 Allen Fellows Society Visiting
Professor was Randy Rosier, MD, from the Department of Orthopaedic
Surgery at the University of Rochester.
Randy presented a Grand Rounds
Talk and a Research Seminar on his long-time research effort to understand chondrogenesis and his more recent efforts to translate this understanding into clinical trials. In addition to the talks, Randy met with the Allen Fellows to provide them with perspective on the roles that research can play in the careers of orthopaedic surgeons. We look forward to the 2008 Allen Fellows
Society Visiting Professor, who will be
Bang Hoang, MD, from the Department of Orthopaedic Surgery at the University of California at Irvine. As a former
Allen Fellow himself in the not that distant past, we anticipate that Bang’s perspective will be especially interesting to the current Allen Fellows.
Last year in this column, I listed the six trainees supported at that time by the CWRU/NIH Musculoskeletal
Training Grant. Since that time, we’ve been fortunate to welcome the following additional trainees:
Katie Polasek, PhD is a post-doctoral fellow with Ron Triolo and Dustin Tyler in the FES Center
Jevan Furmanski, PhD is a post-doctoral fellow with Clare Rimnac and Chris
Hernandez in Mechanical Engineering
I am pleased to report that our department edward Greenfield, Ph.D.
has returned to the top ranks of NIH funding. In the most recently released information, which covers the 2007 fiscal year, we ranked fourth among all
Orthopaedics Departments nationally with $3.2 million in funding comprising
12 funded projects. This makes us the only
Orthopaedics Department in Ohio that is ranked in the top 30. I am hopeful that we will continue to move up in the rankings, especially since we trail third-place
Washington University by only $9,000.
Finally, I would like to congratulate
Randy Marcus on becoming the
President of both the American Board of
Orthopaedic Surgery and the Association of Bone and Joint Surgeons. I’m also looking forward to continuing to work with Randy in his roles as Chairman of the Department of Orthopaedics and as Co-Director of the CWRU/NIH
Musculoskeletal Training Grant.

YEAR IN REVIEW
As a result of his endless contributions to spine surgery–in clinical practice, research and teaching–
Henry H. Bohlman, MD, Director, University
Hospitals Spine Institute, was named the first
Henry H. Bohlman, MD, Chair in Spine Surgery, at University Hospitals Case Medical Center
(UHCMC).
The Henry H. Bohlman, MD, Chair in Spine
Surgery is a $2.2 million endowed chair that will allow UHCMC to remain at the forefront of clinical care, research and education. As a testament to the respect Dr. Bohlman has garnered, his former spine fellows and residents worldwide contributed
$1.2 million to the endowment. Additionally a $1 million contribution came from The Spine Research and Education Foundation.
Dr. Bohlman, a world-renowned spine surgeon, joined University Hospitals in 1972 and became
Director of the University Reconstructive and
Traumatic Spine Surgery Center in 1985 before assuming his current responsibilities in 1996.
Also Professor of Orthopaedic Surgery at Case
Western Reserve University School of Medicine and Director of the UHCMC Spine Fellowship
Program, Dr. Bohlman earned his medical degree from the University of Maryland School of Medicine. He completed his orthopaedic residency at The Johns Hopkins Hospital where he participated in National Institutes of Healthsponsored spine research. In the 1970’s shortly after joining the faculty at UHCMC, he was among the first surgeons in the country to investigate a new surgical approach from the front for decompression and fusion after spinal cord injuries that ultimately lead to reduced neurological risks and improved recovery from paralysis.
Over the course of his 33-year career, he has trained over 75 spine fellows who have gone on to positions at major medical centers throughout the world and who have published more than 2,000 articles in peer-reviewed journals.
OrtHOPaeDiC JOUrnaL | Case Western reserve University | vOL.5, nO.1 | 2008 | 15

PHOTOS fROM THROugHOuT THE YEAR
Len Greenburger, Laurel Blakemore, and Dan Cooperman
John feighan, sandy emery, and ellen Greenberger
Mike Chen focused on his golf swing.
Mike Paczas, ray Liu, Clay Dean, steve fitzgerald and Brian Hardy watch the other golfers.
16
Dr. McLaughlin looking for his ball.
Drs. Marcus, Bohlman and Goodfellow enjoy the presentations after dinner.

John feighan, sandy emery, and ellen Greenberger
YEAR IN REVIEW
OrtHOPaeDiC JOUrnaL | Case Western reserve University | vOL.5, nO.1 | 2008 | 17

PHOTOS fROM THROugHOuT THE YEAR
Cynthia Gordon, alisa Mounts and katie schnaser Patrick Messerschmitt, robert Coale and kasra ahmadinia tara Hardy and erin Wera Chris Mcandrew, Julie Chen and Mike Paczas
18 ryan Garcia and his wife, andrea Jarchow roger Wilber, Doug armstrong and rob Gillespie

YEAR IN REVIEW
erik schnaser and troy Mounts man the grills at the picnic.
Chris and Bridget Mcandrew and Beth Gillespie enjoy lunch.
two of the new interns, shane Hanzlik and Lorraine stern.
Dr. anderson serves the ball as his teammates look on.
kasra ahmadinia, Cynthia and Zack Gordon.
OrtHOPaeDiC JOUrnaL | Case Western reserve University | vOL.5, nO.1 | 2008 | 19

nicholas ahn robert anderson Douglas armstrong Henry Bohlman susannah Briskin Daniel Cooperman
Christopher furey Patrick Getty allison Gilmore reuben Gobezie victor Goldberg richard Grant
Donald Goodfellow amanda Weiss kelly Matthew kraay stephen Lacey randall Marcus shana Miskovsky
William Petersilge John shaffer Joe son-Hing George thompson Brian victoroff John Wilber
20 roger Wilber

YEAR IN REVIEW
Daniel Cooperman robert de swart Michael eppig Harry Hoyen Michael keith stephen Lacey kevin Malone tim Moore Clyde nash Brendan Patterson
John sontich Heather vallier John Wilber roger Wilber
OrtHOPaeDiC JOUrnaL | Case Western reserve University | vOL.5, nO.1 | 2008 | 21

eben alsberg Dwight Davy Jim Dennis edward Greenfield thomas Hering Christopher Hernandez Joseph Mansour* shunichi Murakami
22
P Hunter Peckham Clare rimnac
* Modified copy of image [Source] property of Case Western Reserve University Archives.
ronald triolo Guang Zhou

YEAR IN REVIEW
nicholas ahn robert anderson Patrick Getty victor Goldberg richard Grant randall Marcus thomas McLaughlin William Petersilge
OrtHOPaeDiC JOUrnaL | Case Western reserve University | vOL.5, nO.1 | 2008 | 23

NICOlAS ANdRY lIfETIME ACHIEVEMENT AWARd
Dr. Henry H. Bohlman, M.D., was awarded the nicolas andry
Lifetime achievement award from the association of Bone and Joint surgeons. this award was given at the associations annual meeting in Jackson, Wyoming.
James Carville and randall Marcus
24 randall Marcus and Bob Derkash

AWARdS
ORTHOPAEdIC RESEARCH SOCIETY - OREf dISTINguISHEd INVESTIgATOR AWARd
Dr. Victor M. Goldberg, M.D. was awarded with the ORS-OREF
Distinguished Investigator Award at the 2008 Annual Meeting of the American Academy of
Orthopaedic Surgeons. This award recognizes the vast contribution
Dr. Goldberg has made to the field of orthopaedics during his career through research, leadership and advocacy.
Courtesy of OREF
Dr. Goldberg first became interested immunology and rheumatoid arthritis during his post-residency training in Norwich, England, at the Clinical Research Center. When Dr. Goldberg returned to the United States he became the director of a comprehensive arthritis center at Case Western Reserve University. Dr. Goldberg began building parallel clinical and research programs which led to many important discoveries in the fields of rheumatology and immunology and eventually in joint implant design and loosening.
Dr. Goldberg has a long history of involvement with the OREF serving as a trustee for 9 years including 2 years as chair. He and his wife, Harriet, are longtime supporters of the OREF, supplying funds that will hopefully allow for the development of the next generation of clinician-scientists.
thomas a. einhorn, M.D., presents Dr. Goldberg with the Ors-
Oref Distinguished investigator award. Courtesy of OREF
ARTHuR H. HEuNE MEMORIAl AWARd
The Arthur H. Huene Memorial Award for excellence and promise in Pediatric
Orthopaedics was initiated in 1991. The award consists of a commemorative certificate and a monetary award of $25,000, generously provided by the St. Giles Foundation, intended to be applied by the recipient to continuing work in Pediatric Orthopaedics.
The award is made to an individual who has made an outstanding contribution to the field of Pediatric Orthopaedics and who offers the potential of continuing that contribution in the future.
The 2008 Arthur H. Heune Memorial Award was presented to Dr. George Thompson at the
2008 POSNA meeting.
Dr. thompson delivers his address at the 2008 POsna meeting. Courtesy of POSNA.
Dr. thompson receiving the Heune Memorial award at the 2008 POsna meeting in albuquerque, new Mexico. Courtesy of POSNA.
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KINgSbuRY g. HEIPlE, M.d., ANd fREd A. lENNON
PROfESSOR Of ORTHOPAEdICS ENdOWEd CHAIR
Matthew Kraay, M.D., was appointed the first holder of the Kingsbury G. Heiple, M.D., and
Fred A. Lennon Professor of Orthopaedics during a ceremony at University Hospitals Case
Medical Center. Dr. Kraay is the Director of
Joint Reconstruction and Arthritis Surgery at the facility and is a national leader in research evaluating the long-term effectiveness of joint replacement.
Prior to joining University Hospitals Case
Medical Center where he completed his orthopaedic residency, Dr. Kraay earned his
Master’s Degree in Science-Bioengineering from the University of Michigan. In 1983, he received his medical degree from Wayne State
University. Dr. Kraay completed his fellowship in the Comprehensive Arthritis Program at The
Hospital For Special Surgery, Cornell Medical
Center in New York.
The Fred A. Lennon Charitable Trust created the Heiple-Lennon Chair to support research and technology advancements in orthopaedics at UHCMC and Case, while honoring Dr.
Kingsbury Heiple, former chairman of the
UHCMC Department of Orthopaedic Surgery, for his numerous contributions to the hospital.
Fred A. Lennon was the founder of the Swagelok
Company, a privately held company that designs, manufactures and delivers an expanding range of the highest-quality, fluid system products and solutions.
Dr. Kingsbury G. Heiple served as chairman of the Department of Orthopaedic Surgery from
1982 to 1988. He served as president of the
American Board of Orthopaedic Surgery and an editor of the Journal of Bone and Joint Surgery.
Matthew kraay (seated), (standing left to right), randall Marcus, kingsbury
Heiple and fred rothstein
26

AWARdS
AMERICAN ORTHOPAEdIC ASSOCIATION RESIdENT lEAdERSHIP
The AOA-OREF Resident Leadership Forum identifies PGY4 residents as young orthopaedic leaders by recognizing their accomplishments to-date and by giving them an opportunity to further develop their leadership skills.
Attendees are asked to think critically about leadership and how they can apply it to the specialty.
Dr. Christopher McAndrew was selected from our department to attend this forum in Quebec City,
Quebec.
Dr. Christopher Mcandrew and ellen Greenberger
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28
IntroductIon
A nterior cruciate ligament (ACL) injuries are very common.
As a result, over 100,000 ACL reconstructions are performed each year in the United States. Good to excellent long term results can be expected in the vast majority of patients.1-3 The rate of failure following ACL reconstruction approaches 10 percent. Technical error remains the most common cause of graft failure, with anterior malposition of the femoral tunnel being most common.4-5 Multiple biomechanical studies have demonstrated how small errors in tunnel placement can have serious implications with respect to knee kinematics and graft tension.6-9
Malposition of the ACL femoral tunnel has also been implicated as a cause of clinical failure.10
Proper placement of both the ACL and posterior cruciate ligament (PCL) femoral tunnels require a thorough knowledge of the intercondylar notch surface anatomy. In the coronal plane, the ACL femoral tunnel is traditionally placed 1 -2 mm anterior to the outlet of the intercondylar notch, the socalled over-the-top position.5,11 In the axial plane, the femoral tunnel has historically been placed at the
1 to 2 o’clock position in left knees
(10 to 11 o’clock in right knees).5,11
PCL injuries are much less common.
As such, our clinical knowledge concerning reconstruction of the
PCL lags far behind that for the ACL.
Traditionally, the PCL femoral tunnel is placed at the 11 o’clock position in left knees (1 o’clock in right knees), and just a few millimeters posterior to the articular cartilage border at the inlet of the notch.
Recently there has been much interest in anatomic reconstruction of the ACL and PCL. While current clinical evidence is lacking to support the routine use of double bundle reconstruction in all patients, the anatomic principles emphasized in these techniques has led to a better understanding of, and more anatomic graft placement during single bundle reconstruction. Our institution has recently published four papers on the developmental, topographical, and radiographic anatomy of the intercondylar notch.12-15 It is our hope that a better understanding of femoral intercondylar notch surface anatomy will help avoid technical errors in femoral tunnel placement during both ACL and PCL reconstruction.
topographIc anatomy
The intercondylar notch anatomy is very consistent. The lateral intercondylar ridge is a bony prominence which represents the anterior border of the ACL femoral attachment. William G. Clancy previously termed this landmark the
“resident’s ridge,” as it is commonly mistaken as the over-the-top position by inexperienced surgeons in training.16 Errant placement of the
ACL femoral tunnel anterior to this landmark results in significant anterior malposition of the femoral tunnel, the most common cause of graft failure following ACL reconstruction.4-5
We have shown that the lateral intercondylar ridge is present in the majority of skeletal specimens.12

manuscripts
Figure 1: simulated arthroscopic view (a) and profile (B) of the lateral wall of the intercondylar notch. Lateral intercondylar ridge (solid arrow). Posterolateral rim
(empty arrow).
!
measures aid with proper identification of the lateral intercondylar ridge. Viewing the lateral intercondylar wall from the medial portal allows for better visualization compared to the standard lateral viewing portal. Debridement of the residual ACL femoral stump with electrocautery versus a motorized shaver prevents inadvertent removal of the ridge.
Additionally, we no longer routinely perform notchplasty in all patients as more anatomic graft placement has led to a lower incidence of roof impingement.
The medial intercondylar ridge represents the posterior border of the posterior cruciate ligament femoral attachment. To our knowledge, our institution is the first to describe this osseous landmark.12 Although not as common as the lateral intercondylar
!
ridge, the PCL ridge is present in the majority Figure 2:
Posterior view of intercondylar notch. Probe on medial intercondylar ridge
Additionally, there is little variance in the location and morphology of this osseous landmark (Figure 1).12 As such, the lateral intercondylar ridge is a reliable landmark for placement of the ACL femoral tunnel. During ACL reconstruction, this landmark should be identified and the ACL femoral tunnel placed just posterior to it, into the native ACL footprint. Multiple of skeletal specimens.12
This ridge is more vertically oriented in the notch compared to the lateral
Figure 3:
Posterolateral rim morphology. Probe on type 1 (a), type 2 (B), and type 3 (C) posterolateral rims.
intercondylar ridge (Figure 2).12 The utility of this osseous landmark for femoral tunnel placement during PCL reconstruction is unknown as the PCL footprint is more easily visualized than the ACL footprint due to its location at the inlet of the intercondylar notch.
The posterolateral rim represents the posterior outlet of the intercondylar notch. Specifically, it is the region where the roof of the intercondylar notch transitions to the popliteal surface of the distal femur. This landmark has also been termed the over-the-top position or the linea intercondylaris.17 The posterolateral rim is a common reference for placement of the ACL femoral tunnel.
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ANATOMY Of THE fEMORAl INTERCONdYlAR NOTCH
30
Traditionally, the posterior edge of the ACL femoral tunnel should be 1
– 2 mm anterior to the posterolateral rim.18-19 We identified 3 types of posterolateral rim morphology.12
Type I rims had a sharp, straight border (Figure 3A).12 Type II rims had a sharp, V-shaped border (Figure
3B).12 Type III rims have an indistinct border (Figure 3C).12 For surgeons who reference the posterolateral rim during ACL reconstruction, Type
III rims could potentially lead to errant femoral tunnel placement.20
Specifically, commercial femoral tunnel aiming guides which reference the posterolateral rim for guide pin placement require that the aimer tongue be placed accurately behind the posterolateral rim.20 Intuitively, type ridge.14 This method is based on the ridge’s radiographic relationship to Blumensaat’s line. Blumensaat’s line is a radiographic landmark that is easily identified and is familiar to most orthopaedic surgeons.
The location of the lateral intercondylar ridge can be estimated by multiplying the length of Blumensaat’s line by
0.79, the Blumensaat’s-ridge ratio.14 The Blumensaat’sridge ratio identifies the point where the lateral intercondylar ridge intersects Blumensaat’s line.14 From this point, the lateral intercondylar ridge runs at a 75.5 degree angle with respect to Blumensaat’s line, the Blumensaat’s-ridge angle (Figure 4).14 These calculations can be used to estimate the anterior
!
border of the native ACL femoral attachment, the lateral intercondylar ridge.
A properly placed femoral tunnel should lie in the native ACL footprint, just posterior to the lateral intercondylar ridge. For
Figure 4:
Blumensaat’s-ridge ratio is determined by dividing the inlet-to-ridge distance (a-B) by the length of
Blumensaat’s line (a-C). Blumensaat’s-ridge angle is indicated by double arrow.
III rims might prohibit accurate guide pin placement. Intraoperatively, all soft tissues should be carefully cleared from the outlet of the notch to correctly identify the posterolateral rim prior to guide pin placement. Additionally, the lateral intercondylar ridge can be identified and used to confirm proper guide pin placement.
radIographIc anatomy
Recently we described the radiographic location of the lateral intercondylar the young arthroscopist or the arthroscopist who only performs a few ACL reconstructions per year, the described method can aid with femoral tunnel positioning during primary ACL reconstruction. In the postoperative period our method may be used for quality assurance, allowing the surgeon to alter technique and femoral tunnel drilling position as necessary in subsequent patients.
In the patient with a poor outcome following previous ACL reconstruction, femoral tunnel position can be accurately evaluated in the outpatient setting with a lateral plain film image.
This allows for quick and inexpensive evaluation of femoral tunnel position
Figure 5: radiographic classification of posterolateral rim.
(a) radiographic type 1 rims have a well-defined transition (arrowhead) from Blumensaat’s line
(double arrowhead) to the posterior femoral cortex
(arrow). (B) radiographic type 2 rims have an ill-defined transition (arrowhead) from Blumensaat’s line (double arrowhead) to the posterior femoral cortex (arrow) prior to obtaining more advanced imaging which may take days or weeks to perform. Finally, the described method may be used intraoperatively to aid with femoral tunnel placement in revision cases where arthroscopic anatomy is obscured or when anatomic guide pin placement is in doubt.
Posterolateral rim morphology can also be delineated on lateral plain film images.13 The clinical implication of the posterolateral rim morphology has been discussed above. Well-defined rims have a characteristic appearance on lateral plain film images, radiographic
Type 1 rims (Figure 5A).13
Likewise, ill-defined rims also have a characteristic appearance on lateral
!
!

MANuSCRIPTS plain film images, radiographic Type
2 rims (Figure 5B).13 For surgeons who reference the posterolateral rim for ACL femoral tunnel placement, this classification system can be utilized in the preoperative period to identify patients with indistinct posterolateral rim morphology. When patients with indistinct posterolateral rims are identified preoperatively, the surgeon should be prepared to confirm proper guide pin placement with alternative landmarks such as the lateral intercondylar ridge or the residual ACL femoral stump.
Alternatively, fluoroscopy may be utilized intraoperatively to confirm anatomic guide pin placement prior to tunnel drilling.
developmental anatomy
ACL ligament tears are becoming more commonplace in pediatric patients, with a corresponding increase in ACL reconstructions in skeletally immature patients.21 Kocher et al. performed a survey which revealed 15 cases of leg length discrepancy after ACL reconstruction.21 Twelve of these occurred in the distal femur, with 11 of the 12 attributed to technical errors.21
Given the fact that ACL reconstructions in skeletally mature patients are infrequent surgeries even for specialists,21 a better understanding of the anatomy of the developing intercondylar notch may help decrease the incidence of technical errors.
Our study found that the lateral intercondylar ridge was a common landmark in specimens ages 13 years and older, while it was present in only about half of younger specimens.15
Thus, in most surgical cases the lateral intercondylar ridge remains a useful landmark that can guide the location of the femoral tunnel. The distance between the middle of the lateral intercondylar ridge and the posterolateral rim averaged above on tunnel size.
Figure 6:
Measurements of the angle between the femoral surface at the aCL footprint and the postero-lateral aspect of the distal femoral physis. (a) the left side is lateral in this figure. this posterior view demonstrates the obliquity of the femoral tunnel in the coronal plane if it was drilled perpendicular to the femoral surface. a drillhole directed more medially would decrease the obliquity. (B) this lateral view demonstrates the obliquity of the femoral tunnel in the sagittal plane if it was drilled perpendicular to the femoral surface. a drillhole directed more posteriorly would decrease the obliquity. Photo adapted from Liu rW et al., J
Pediatr Orthop 2008;28:177-183
8mm in children ages 13-15 years, and above 7mm in children ages 10-
12 years.15 However, this distance averaged approximately 1mm less in females versus males. Thus, the surgeon can expect to drill through the lateral intercondylar ridge when making femoral tunnels in females, while in males there may be adequate space between the lateral intercondylar ridge and the posterolateral rim depending
!
!
To our knowledge, the orientation of the femoral surface with respect to the distal femoral physis had not been well described prior to our study. A drill hole that is oriented more perpendicular with respect to the physis will result in a smaller hole in the physis, which would theoretically decrease the risk of growth arrest.22 Figure 6 demonstrates that the angle between the femoral surface at the ACL footprint and the postero-lateral aspect of the physis is dramatic in both the coronal and lateral planes. Based on these observations, a femoral tunnel drilled perpendicular to the femoral surface will penetrate the physis obliquely, while aiming the drill more medial and posterior will decrease the obliquity through the physis. While there were statistical differences in these angles between different age groups, there was no clear clinically significant trend, and were not any statistical difference between males and females.
Summary
This article summarizes four studies which investigated the bony anatomy of the femoral intercondylar notch and its relevance to cruciate ligament reconstruction.12-15
Ultimately, our aim is to provide clinically relevant anatomical information to help minimize the incidence of technical errors, which in ACL reconstruction are the most common cause of graft failure in adults,4-5 and postoperative growth disturbance in children.21
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32
ANATOMY Of THE fEMORAl INTERCONdYlAR NOTCH acknowledgementS
The authors would like to acknowledge
Michael R. Chen, Robert J. Gillespie,
Patrick J. Messerschmitt and Mark S.
Robbin for their contributions to the previously published studies.
REfERENCES
1. Howe JG, Johnson RJ, Kaplan MJ, Fleming
B, Jarvinen M.
Anterior cruciate ligament reconstruction using quadriceps patellar tendon graft. Part I. Long-term follow-up. Am J Sports
Med . 1991;19:458-462.
2. Kaplan MJ, Howe JG, Fleming B, Johnson
RJ, Jarvinen M. Anterior cruciate ligament reconstruction using quadriceps patellar tendon graft. Part II. A specific sport review. Am J
Sports Med.
1991;19:458-462.
3. Kornblatt I, Warren RF, Wickiewicz TL.
Long-term follow-up of anteriorcruciate ligament reconstruction using the quadriceps tendon substitution for chronic anterior cruciate ligament insufficiency. Am J Sports
Med. 1988;16:444-457.
4.
Allum, R. Aspects of current managment.
Complications of arthroscopic reconstruction of the anterior cruciate ligament.
J Bone Joint
Surg Br.
2003;85B:12-16.
5. Fu FH, Bennett CH, Ma CB, MenetreyJ,
Latterman C.
Current trends in anterior cruciate ligament reconstruction. Part II.
Operative procedures and clinical correlations.
Am J Sports Med. 2000;28:124-130.
6. Loh JC, Fukuda Y, Tsuda E, Steadman RJ,
Fu FH, Woo SL. Knee stability and graft function following anterior cruciate ligament reconstruction: Comparison between the
11 o’clock and 10 o’clock femoral tunnel placement. Arthroscopy. 2003;19:297-304.
7. Markolf KL, Hame S, Hunter DM, Oakes
DA, Zoric B, Gause P, Finerman GA. Effects of femoral tunnel placment on knee laxity and forces in an anterior cruciae ligament graft. J
Orthop Res. 2002;20:1016-1024.
8. Norwood LA, Cross MJ. The intercondylar shelf and the anterior cruciate ligament. Am J
Sports Med. 1977;5:171-176.
9. Odenstein M, Gillquiest J. Functional anatomy of the anterior cruciate ligament and a rationale for reconstruction. J Bone Joint Surg
Am. 1985;67:257-262.
10. Lee MC, Seong SC, Lee S, Chang CB,
Park YK, Jo H, et al. Vertical femoral tunnel placement results in rotational laxity after anterior cruciate ligament reconstruction.
Arthroscopy. 2007;23:771-778.
11. Bylski-Austrow DI, Grood E, Hefzy E,
Holden JP, Butler DL. Anterior cruciate ligament replacements: a mechanical study of femoral attachment location, flexion angle at tensioning and initial tension. J Orthop Res.
1990;8:522-531.
12. Farrow LD, Chen MC, Cooperman DC,
Victoroff BN, Goodfellow DB.
Morphology of the femoral intercondylar notch. J Bone Joint
Surg Am. 2007;89:2150-2155.
13. Farrow LD, Chen MR, Goodfellow DB,
Cooperman DC., Robbin MS. Radiographic classification of the intercondylar notch posterolateral rim. Arthroscopy, In Press.
14. Farrow LD, Gillespie RJ, Victoroff BN,
Cooperman DR. Radiographic location of the lateral intercondylar ridge. Its relationship to
Blumensaat’s line. Am J Sports Med, In Press.
15. Liu RW, Farrow LD, Messerschmitt PJ,
Gilmore A, Goodfellow DB, Cooperman
DR. An anatomical study of the pediatric intercondylar notch. J Pediatr Orthop,
2008;28:177-183.
16. Hutchinson MR, Ash SA. Resident’s ridge: assessing the cortical thickness of the lateral wall and roof of the intercondylar notch.
Arthroscopy. 2003;19:931-935.
17. Petersen W, Zantop T. Anatomy of the anterior cruciate ligament with respect to its two functional bundles. Clin Orthop Relat Res.
2006;454:35-47.
18. Ahn JH, Lee SH. Anterior cruciate ligament reconstruction with with hamstring tendon autografts. Arthroscopy . 2007;23:109.e1-109.e4.
19. Giron F, Cuomo P, Edwards A, Bull
AM, Amis AA, Aglietti P. Double bundle
“anatomic” anterior cruciate ligament reconstruction: A cadaveric study of tunnel positioning wih a transtibial technique.
Arthroscopy . 2007;23:7-13.
20. McGuire DA, Hendricks SD, Grinstead
GL. Use of an endoscopic aimer for femoral tunnel placement in anterior cruciate ligament reconstruction. Arthroscopy . 1996;12:26-31.
21. Kocher MS, Saxon HS, Hovis WD, Hawkins
RJ.
Management and complications of anterior cruciate ligament injuries in skeletally immature patients: survey of the Herodicus
Society and The ACL Study Group. J Pediatr
Orthop.
2002;22(4):452-457.
22.
Makela EA, Vainionpaa S, Vihtonen K, et al. The effect of trauma to the lower femoral epiphyseal plate. J Bone Joint Surg Br.
1988;70:187-191.

MANuSCRIPTS
IntroductIon
A lthough osteosarcoma is the most common bone sarcoma in children and adolescents, overall incidence of osteosarcoma is low with approximately 400 new diagnoses each year in the United States.
10
Primary osteosarcoma of the spine is rare, comprising less than 3% of all new osteosarcoma cases, as the most common primary site is the metaphyseal region of the distal femur, proximal tibia, or proximal humerus.
2,10,18 The overall prognosis for patients with osteosarcoma arising in the axial skeleton is worse compared to the prognosis of those patients with osteosarcoma arising in the appendicular skeleton.
2,4,18 On initial presentation, 20% of patients with osteosarcoma will have clinically detectable metastases.
3,11 Despite surgical resection and chemotherapy regimens, the 10-year survival rate for patients with clinically detectable metastases on presentation is 20-
30%, 3,7,10 with the most common cause of death remaining to be respiratory failure secondary to pulmonary metastatic disease.
11,18 caSe report
An otherwise healthy, fifteen-year-old male presented with a 4 week history of progressive low back pain and right leg pain, without an identifiable traumatic event. The low back pain began insidiously and was diffuse and achy in nature. The right-sided radicular pain originated in the right buttocks and proceeded down the posterolateral aspect of the leg into the calf. The patient was beginning to experience difficulty walking secondary to pain and had a concomitant feeling of right leg weakness.
Past medical history was noncontributory,
Figure 1. anteroposterior (aP) and lateral roentgenograms of the lumbar spine revealing overall good alignment. a. aP roentgenogram demonstrating subtle changes at the L4 level, including asymmetry of the pedicles and mild loss of vertebral body height. Contrast material in the colon obstructs the view of the L5 vertebral body. B. Lateral roentgenogram revealing sclerotic changes of the L4 vertebral body with overlying bowel contrast material. and the patient denied any fevers, night sweats, and weight loss.
The patient was a non-smoker.
function and absent dorsiflexion/ plantarflexion. Examination of the left lower extremity was unremarkable. No pathologic reflexes were present and normal bladder/bowel function existed.
Physical examination revealed a healthy appearing teenager with an antalgic gait. There was mild, diffuse tenderness to palpation over the lumbar spine, but no skin abnormalities.
Lasegue’s sign was positive in the right leg at 30 degrees of elevation.
Strength examination of the right lower extremity revealed 3/5 quadricep
Plain radiographs demonstrated nonspecific changes involving the
L4 vertebral level with mild pedicle asymmetry and loss of vertebral body height (Figure 1). Magnetic resonance
(MR) imaging revealed heterogenous signal change of the L4 vertebral body with a large, right-sided, paraspinal mass involving the psoas muscle (Figure
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OSTEOSARCOMA Of THE SPINE
34
2). There was noted compression of the inferior vena cava (IVC) and evidence of thrombus within the vessel. Computed tomography
(CT) of the chest showed multiple, peripheral pulmonary nodules and several pulmonary emboli involving the segmental pulmonary arteries in the right upper lobe and right lower lobe (Figure 3). No obvious infarction of the lung parenchyma existed. CT of the abdomen and pelvis showed the right psoas mass at the L4 vertebral body level with soft tissue calcifications. Bone scan and positron emission tomography (PET) scan failed to reveal any skip lesions or distant metastases to bone.
CT-guided biopsy of the right-sided psoas muscle mass revealed atypical spindle cells, osteoid, and focal areas with anaplastic features (Figure 4).
The histopathologic findings on tissue biopsy confirmed the impression
Figure 2. Magnetic resonance (Mr) with contrast of the lumbar spine. a. sagital t-2 weighted Mr demonstrating abnormal signal within the L4 vertebral body representing edema and cysts without obvious cortical disruption. B. axial t-2 weighted Mr revealing a large and complex, heterogeneous right-sided paraspinal mass, measuring
4.5cm x 5.2cm, centered within the psoas muscle. the mass extends through the L4-5 neural foramen into the epidural space, ventral to the thecal sac, with compression of the neural elements.
(preoperative) chemotherapy consisting of adriamycin and cisplatin. Additionally, the patient was started on low-molecular-weight heparin anticoagulation and underwent IVC filter placement for prophylaxis against further pulmonary emboli secondary to the
IVC thrombus. During the preoperative period, the patient experienced several episodes of exacerbated low back and radicular right leg pain.
There was a concern for further neurolobical deterioration, and therefore, the patient underwent four emergent radiation treatments targeting the
L4 vertrebral body and paraspinal mass.
Figure 3. axial computed tomography (Ct) of the chest showing a 8.0mm peripheral lung nodule in the right lower lobe of primary osteosarcoma of the
L4 vertebral body with soft tissue extension into the paraspinal region and metastatic disease to the lungs.
Following placement of a central venous catheter, the patient underwent a 6-week course of neoadjuvant
Upon completion of neoadjuvant chemotherapy, the patient underwent restaging with repeat CT of the chest, abdomen, and pelvis. The number of pulmonary nodules remained stable, but fortunately all pulmonary nodules had decreased in size. The largest pulmonary nodule on initial CT of the chest measured 8.0mm and subsequently measured
4.2mm following the preoperative course of chemotherapy. Repeat PET scan revealed a dramatic decrease in uptake signal of the L4 vertebral body and right-sided paraspinal mass, both consistent with an excellent

MANuSCRIPTS stability to the spinal column. The wound was closed over drains.
Figure 4. mass.
axial Ct of the abdomen during Ct-guided core needle biopsy of the paraspinal response to neoadjuvant chemotherapy.
The patient’s pain was dramatically improved following the radiation treatments and his neurological status remained stable.
Approximately three months following initial presentation, the patient was taken to the operating room for surgical resection. The patient was placed in the left lateral decubitus position and a retroperitoneal approach was undertaken. The IVC and iliac vessels were dissected free from the spinal column with the assistance of a pediatric surgeon. The paraspinal mass involving the psoas muscle was then carefully dissected away from the vertebral column. Next, the paraspinal tumor was resected out of the psoas muscle along with a healthy cuff of muscle on gross appearance. The exiting L3 and L4 nerve roots on the right side were penetrating through the tumor mass and therefore required ligation. Frozen surgical specimen sent for intraoperative evaluation revealed an absence of tumor in the healthy cuff
Three days later, the patient was returned to the operating suite for the second stage of the procedure. He was positioned prone and the lumbosacral region was exposed through a midline incision. It appeared that a portion of the right hemilamina of L4 was replaced by tumor. A complete L4 laminectomy was then performed.
Next, the inferior facet of L3 and the superior facet of L4 as well as the transverse process of L4 were removed completely as one unit on the right side. It was then assured that adequate decompression of the neural elements existed and all visible tumor had been removed. Posterolateral fusion of L2-S1 was performed with instrumentation, autogenous iliac crest bone graft, allograft bone, and bone morphogenic protein soaked sponges (Figure 5). The wound was closed over drains. of psoas muscle.
The next step of the operation involved performing complete diskectomies at the L3-4 and L4-5 levels. A subtotal en bloc L4 corpectomy, including the rightsided L4 pedicle, was then completed with removal of the posterior cortical wall, revealing the underlying thecal sac. There did not appear to be any significant amount of gross tumor or bone destruction in the L4 vertebral body during resection.
An appropriate sized Pyramesh cage
(Metronics, Memphis,
TN) filled with demineralized bone matrix and bone morphogenic protein soaked sponges was secured to restore anterior
Figure 5. Postoperative lateral roentgenogram of the lumbar spine demonstrating L2-s1 instrumentation and
Pyramesh cage.
OrtHOPaeDiC JOUrnaL | Case Western reserve University | vOL.5, nO.1 | 2008 | 35

OSTEOSARCOMA Of THE SPINE
36
Pathology reports of the paraspinal mass excision revealed skeletal muscle with a central focus of nonviable osteosarcoma reflecting near 100% tumor necrosis. The vertebral body specimens revealed portions of cancellous bone with interspersing areas of nonviable osteosarcoma reflecting near 100% tumor necrosis.
The patient went on to complete an additional 6 weeks of adjuvant
(postoperative) chemotherapy. This regimen was similar to the preoperative regimen due to the excellent tumor necrosis noted on pathologic examination.
At two years following initial presentation, the patient had made a dramatic recovery in his lower extremity function. Despite ligation of the right-sided L3 and L4 nerve roots, full motor strength eventually returned.
Examination revealed 5/5 strength in both lower extremities with only a mild decrease in light touch sensation along the L4-L5 dermatome in the right leg.
Unfortunately at 30 months following initial presentation, a solitary pulmonary nodule was found on screening CT of the chest, which was not present at the two year followup. The patient underwent successful surgical resection of the pulmonary nodule and is currently underway with repeat chemotherapy. Advanced imaging of the spine has not revealed any local recurrence. dIScuSSIon
Primary osteosarcoma of the spine is an exceedingly rare occurrence. While osteosarcoma of the extremities most commonly occurs in the 2 nd or 3 rd decade of life, osteosarcoma arising in the spine generally occurs in patients in their 30’s or 40’s.
6,10,18 Patients with osteosarcoma of the spine almost uniformly present with back pain at the affected level and 50-70% have neurological symptoms when first evaluated.
5,9,13,18
The time of symptom onset to diagnosis commonly exceeds 6 weeks, 15 but may be related to patient and/or family delay in obtaining a medical evaluation or a delay in referral by the primary care physician to a spine specialist. In our case, four weeks lapsed before initial evaluation by the Orthopaedic spine surgeon, but tissue diagnosis was established within the subsequent 72 hours with the cooperation of a multidisciplinary team.
Prior to updated chemotherapy protocols and innovative surgical procedures, survival rates for patients with osteosarcoma of the spine was reported to be 6-10 months in studies conducted during the 1970’s and
1980’s.
reviewed in 13 More recent reports indicate survival may be higher with aggressive treatment, including both preoperative and postoperative chemotherapy and attempts at widemargin surgical resection. Ozaki reported 19 of 22 (86%) patients alive >1 year following treatment and three patients (without metastases on presentation) surviving >6 years.
13 In another study, Delamarter reported one patient who presented without metastases and was disease-free at five years following treatement.
5
Poorer prognosis has been associated with metastases, large tumors, sacral tumors, or resection not achieving marginal or wide margins.
13 However, if an intralesional resection is the only feasible operation, overall survival has still been found to be improved over those patients without surgery.
13,18
The percentage of tumor necrosis within a primary tumor quantifies the response to neoadjuvant chemotherapy.
Tumor necrosis has been shown to be the most sensitive treatment-related predictor of survival in patients with osteosarcoma.
12,17 Improved survival rates correlate with greater than 90% tumor necrosis following neoadjuvant chemotherapy.
11 Fortunately our patient experienced close to 100% tumor necrosis, and therefore, his postoperative chemotherapeutic drugs were similar to those used preoperatively.
Surgical resection of vertebral osteosarcoma is challenging, as typically the tumor location is in close proximity to neural elements and major vascular structures. Newer surgical techniques, such as total ( en bloc ) spondylectomy, have been proposed in an attempt to obtain a wide or marginal margin instead of the traditional curettage and intralesional resection. This strategy may provide protection from contaminating surrounding tissue with tumor cells leading to lower recurrence rates.
8,19 In our patient, a wide excision of the paraspinal lesion and subtotal en bloc corpectomy was accomplished anteriorly and coupled with a posteriorbased en bloc resection of the involved lamina, facet, and transverse process.
Children and adolescents with sarcoma, including osteosarcoma, are at an increased risk for thromboembolism.
1,14
Tumor cells stimulate cytokine production and increase tissue factor levels which predispose patients to a prothrombotic state.
16 It was recently reported that 14-16% of children and adolescents with sarcoma develop thromboembolism, 1,14 with approximately 22% of these cases involving pulmonary embolism.
14 Half of the clots were associated with tumor compression on vasculature.
14 Also shown, patients with metastatic disease on presentation were 2.5 times more likely to develop thromboembolism.
14
As with the patient in our case report, tumor extension into the paraspinal

MANuSCRIPTS region led to a mass effect on the inferior vena cava most likely creating a flow disturbance which predisposed to the thrombosis and resultant pulmonary emboli.
In summary, osteosarcoma of the spine is a rare occurrence in the pediatric and adolescent population, but early diagnosis and initiation of aggressive treatment is critical to survival. Patients with osteosarcoma of the spine are at an increased risk for thromboembolism and may require anticoagulant medications and/or intravenous filters to prevent more devastating embolic events.
REfERENCES
1. Athale, U.; Cox, S.; Siciliano, S.; and Chan,
A. K.: Thromboembolism in children with sarcoma. Pediatr Blood Cancer, 49(2): 171-6,
2007.
2. Barwick, K. W.; Huvos, A. G.; and Smith,
J.: Primary osteogenic sarcoma of the vertebral column: a clinicopathologic correlation of ten patients. Cancer, 46(3): 595-604, 1980.
3. Bielack, S. et al.: Prognostic factors in high-grade osteosarcoma of the extremities or trunk: an analysis of 1,702 patients treated on neoadjuvant cooperative osteosarcoma study group protocols. J. Clin. Oncol., 20: 776-790,
2002.
4. Bielack, S. S.; Wulff, B.; Delling, G.;
Gobel, U.; Kotz, R.; Ritter, J.; and Winkler,
K.: Osteosarcoma of the trunk treated by multimodal therapy: experience of the
Cooperative Osteosarcoma study group
(COSS). Med Pediatr Oncol, 24(1): 6-12, 1995.
5. Delamarter, R. B.; Sachs, B. L.; Thompson,
G. H.; Bohlman, H. H.; Makley, J. T.; and Carter, J. R.: Primary neoplasms of the thoracic and lumbar spine. An analysis of 29 consecutive cases. Clin Orthop Relat Res, (256):
87-100, 1990.
6. Green, R.; Saifuddin, A.; and Cannon,
S.: Pictorial review: Imaging of primary osteosarcoma of the spine. Clin Radiol, 51(5):
325-9, 1996.
7. Kager, L. et al.: Primary metastatic osteosarcoma: presentation and outcome of patients treated on neoadjuvant Cooperative
Osteosarcoma Study Group protocols. J Clin
Oncol, 21(10): 2011-8, 2003.
8. Kawahara, N.; Tomita, K.; Fujita, T.;
Maruo, S.; Otsuka, S.; and Kinoshita, G.:
Osteosarcoma of the thoracolumbar spine: total en bloc spondylectomy. A case report. J Bone
Joint Surg Am, 79(3): 453-8, 1997.
9. Kelley, S. P.; Ashford, R. U.; Rao, A. S.; and
Dickson, R. A.: Primary bone tumours of the spine: a 42-year survey from the Leeds Regional
Bone Tumour Registry. Eur Spine J, 16(3): 405-
9, 2007.
10. Longhi, A.; Errani, C.; DePaolis, M.;
Mercuri, M.; and Bacci, G.: Primary bone osteosarcoma in the pediatric age: state of the art. Cancer Treat. Rev., 32: 423-436, 2006.
11. Meyers, P.; Heller, G.; Healey, J.; Huvos, A.;
Applewhite, A.; Sun, M.; and LaQuaglia, M.:
Osteogenic sarcoma with clinically detectable metastasis at initial presentation. J. Clin.
Oncol., 11: 449-453, 1993.
12. Meyers, P. A.; Heller, G.; Healey, J.; Huvos,
A.; Lane, J.; Marcove, R.; Applewhite, A.;
Vlamis, V.; and Rosen, G.: Chemotherapy for nonmetastatic osteogenic sarcoma: the
Memorial Sloan-Kettering experience. J Clin
Oncol, 10(1): 5-15, 1992.
13. Ozaki, T. et al.: Osteosarcoma of the spine: experience of the Cooperative Osteosarcoma
Study Group. Cancer, 94(4): 1069-77, 2002.
14. Paz-Priel, I.; Long, L.; Helman, L.
J.; Mackall, C. L.; and Wayne, A. S.:
Thromboembolic events in children and young adults with pediatric sarcoma. J Clin Oncol,
25(12): 1519-24, 2007.
15. Pollock, B. H.; Krischer, J. P.; and Vietti,
T. J.: Interval between symptom onset and diagnosis of pediatric solid tumors. J Pediatr,
119(5): 725-32, 1991.
16. Prandoni, P.; Falanga, A.; and Piccioli, A.:
Cancer and venous thromboembolism. Lancet
Oncol, 6(6): 401-10, 2005.
17. Provisor, A. J. et al.: Treatment of nonmetastatic osteosarcoma of the extremity with preoperative and postoperative chemotherapy: a report from the Children’s
Cancer Group. J Clin Oncol, 15(1): 76-84,
1997.
18. Shives, T. C.; Dahlin, D. C.; Sim, F.
H.; Pritchard, D. J.; and Earle, J. D.:
Osteosarcoma of the spine. J Bone Joint Surg
Am, 68(5): 660-8, 1986.
19. Tomita, K.; Kawahara, N.; Baba, H.;
Tsuchiya, H.; Fujita, T.; and Toribatake, Y.:
Total en bloc spondylectomy. A new surgical technique for primary malignant vertebral tumors. Spine, 22(3): 324-33, 1997.
OrtHOPaeDiC JOUrnaL | Case Western reserve University | vOL.5, nO.1 | 2008 | 37

38
abStract
S pinal cord injury and brachial plexus injury often result in denervation. Current surgical techniques for re-innervation in this setting are severely limited. Paralyzed nerve transfer is a new technique which allows for re-innervation without the sacrifice of voluntary function. In this technique, a paralyzed but nondegenerated nerve is transferred onto a degenerated nerve. We have performed the first case with this technique for upper extremity re-innervation and report our clinical results.
IntroductIon
Denervation can result from damage to lower motor neuron (LMN) cell bodies in the spinal cord and/or discontinuity between LMN cell bodies and a given muscle.
(1,2) Damage to LMN cell bodies occurs with spinal cord injury (SCI) at the level of the lesion.
Discontinuity between LMN cell bodies and a given muscle can occur with root avulsion or peripheral nerve injury.
Denervation due to injured LMN cell bodies is significantly more difficult to address, in comparison to discontinuity, due to concomitant SCI. In this case, the denervated muscle is surrounded by involuntary nerves and muscles making conventional tendon and nerve transfers unfeasible. One potential reconstructive option is tendon transfer of an involuntary muscle onto the denervated muscle in conjunction with the application of a functional electrical stimulation (FES) system. (3,4)
However, functional returns after such a procedure may be limited due to poor donor muscle mass and loss of strength from the transfer.
(5) Furthermore, paralyzed tendon transfer and FES system application may not be possible in high cervical SCI due to a lack of appropriate donor muscles.
As such, the senior author developed a new technique for re-innervation in the setting of SCI. Paralyzed nerve transfer consists of transferring a paralyzed but electrically excitable (LMN cell bodies intact) nerve onto a paralyzed but no longer electrically excitable nerve
(LMN cell bodies injured). By using this technique in conjunction with an
FES system, it is possible to re-animate denervated muscle without sacrificing voluntary function (from conventional tendon transfer or nerve transfer) and without paralyzed tendon transfer.
caSe report
The patient presented with an incomplete cervical spinal cord injury
(SCI), central cord syndrome, and a left sided C5-C6 brachial plexus injury one week post-injury. On initial physical examination, the patient demonstrated grade zero deltoid, rotator cuff, elbow flexor, wrist dorsiflexor, finger flexor, and interosseous muscle function on manual motor testing. However, grade five trapezius function was preserved on the injured side. Serial electromyography revealed denervation of the deltoid, corcaobrachialis, brachialis, and biceps brachii muscles and the patient did not recover any motor function following his injury.
Surgical intervention was initiated nine months post-injury and included one conventional nerve transfer (spinal accessory to suprascapular nerve) and two paralyzed nerve transfers
(paralyzed radial nerve branch to long head of triceps to degenerated axillary nerve and paralyzed ulnar fascicular to degenerated musculocutaneous nerve).
Intra-operatively, the spinal accessory, radial, and ulnar nerves were electrically excitable confirming the presence of intact LMN cell bodies. In contrast, the axillary and musculocutaneous nerves were not electrically excitable confirming the presence of Wallerian degeneration. A conventional spinal accessory to suprascapular nerve transfer was performed first to allow for restoration of rotator cuff function.
Following this, the paralyzed radial nerve to the long head of the triceps was transferred onto the axillary nerve for deltoid re-innervation. Finally, a paralyzed ulnar fascicle was transferred onto the musculocutaneous nerve for re-innervation of the elbow flexors.
At the time of this report, the patient is 44 months post surgery and serial electromyography reveals ongoing re-innervation of the deltoid,

MANuSCRIPTS brachialis, and biceps brachii muscles.
Consequently, the patient is now eligible for FES system implantation to restore active shoulder abduction and elbow flexion. dIScuSSIon
Denervation poses a significant clinical dilemma with only limited techniques available for successful re-animation.
Furthermore, all current techniques rely upon the restoration of continuity between intact and “voluntary” LMN cell bodies (i.e. LMN cell bodies still in continuity with the cortex) and a given target muscle. Current strategies for re-innervation include primary repair (6) , nerve transfer
(4,7,8,9) , motor nerve transplantation
(11) , direct muscle neurotization
(12,13,14,15,16) , muscle-nervemuscle neurotization (17) , and free microneurovascular transfer (18,19) .
However, in the setting of denervation with concomitant SCI, many of these strategies are no longer feasible due to a lack of voluntary donor nerves.
Unlike the aforementioned techniques, paralyzed nerve transfer only requires an intact LMN cell body in continuity with target muscle for successful re-innervation. Reliance on intact but “involuntary” LMN cell bodies for re-innervation has yielded excellent short-term results in a rat animal model performed at this institution. In addition, this technique has previously been employed for diaphragm re-innervation with equally promising results. Six patients with
C3-C5 tetraplegia (and confirmed diaphragm denervation) underwent a T4 intercostal (paralyzed) transfer onto a degenerated phrenic nerve. A diaphragmatic pacemaker lead was then placed one centimeter distal to the neurorrhaphy site and stimulation was applied on a monthly basis until diaphragmatic contractions could be detected. At the conclusion of this study, all patients were capable of using diaphragmatic pacing alone (in place of mechanical ventilation) to support ventilatory function. (20)
In conclusion, paralyzed nerve transfer holds great promise as a new reconstructive technique for reinnervation in the setting of spinal cord injury. It has been used successfully for diaphragmatic re-innervation and with excellent long-term results for upper extremity re-innervation in this report. Furthermore, paralyzed nerve transfer can be paired with FES in the upper extremity for active re-animation of denervated muscle. Alternatively, paralyzed nerve transfer may potentially prove useful as a stand-alone procedure for the prevention or reversal of contractures associated with severe denervation. (21)
REfERENCES
1.
Berman SA, Young RR, Sarkarati M, Shefner
JM. Injury zone denervation in traumatic quadriplegia in humans. Muscle Nerve.
1996
Jun;19(6):701-6. PMID: 8609919
2. Gorman PH, Kikta DG, Peckham PH.
Neurophysiologic evaluation of lower motor neuron damage in tetraplegia. Muscle Nerve.
1998 Oct;21(10):1321-3. PMID: 9736062
3. Keith MW, Kilgore KL, Peckham PH,
Wuolle KS, Creasey G, Lemay M.
Tendon transfers and functional electrical stimulation for restoration of hand function in spinal cord injury. J Hand Surg [Am]. 1996 Jan;21(1):89-
99.PMID: 8775202
4. Wood MB, Murray PM. Heterotopic nerve transfers: recent trends with expanding indication. J Hand Surg [Am]. 2007
Mar;32(3):397-408. Review. PMID: 17336851
5. Hoyen H, Gonzalez E, Williams P, Keith
M. Management of the paralyzed elbow in tetraplegia. Hand Clin.
2002 Feb;18(1):113-33.
Review. PMID: 12143409
6. Gamo K, Kiryu-Seo S, Yoshikawa H, Kiyama
H. Suture of transected nerve suppresses expression of BH3-only protein Noxa in nervetransected motor neurons of C57BL/6J mouse.
J Neurotrauma. 2007 May;24(5):876-84.
PMID: 17518541
7. Merrell GA, Barrie KA, Katz DL, Wolfe
SW. Results of nerve transfer techniques for restoration of shoulder and elbow function in the context of a meta-analysis of the
English literature. J Hand Surg [Am]. 2001
Mar;26(2):303-14. PMID: 11279578
8. Terzis JK, Kostopoulos VK. The surgical treatment of brachial plexus injuries in adults.
Plast Reconstr Surg.
2007 Apr 1;119(4):73e-92e.
Review. PMID: 17496583
9. Battiston B, Lanzetta M. Reconstruction of high ulnar nerve lesions by distal double median to ulnar nerve transfer.
J Hand Surg
[Am]. 1999 Nov;24(6):1185-91. PMID:
10584939
10. Ustün ME, Oğün TC, Büyükmumcu M,
Salbacak A.
Selective restoration of motor function in the ulnar nerve by transfer of the anterior interosseous nerve. An anatomical feasibility study. J Bone Joint Surg Am. 2001
Apr;83-A(4):549-52. PMID: 11315783
11. Gray WP, Keohane C, Kirwan WO.
Motor nerve transplantation.
J Neurosurg.
1997
Oct;87(4):615-24. PMID: 9322851
12. Brunelli G.
Direct neurotization of severely damaged muscles.
J Hand Surg [Am]. 1982
Nov;7(6):572-9. PMID: 7175128
13. Brunelli G, Monini L.
Direct muscular neurotization. J Hand Surg [Am]. 1985
Nov;10(6 Pt 2):993-7. PMID: 4078294
14. Brunelli G, Brunelli LM. Direct neurotization of severely damaged denervated muscles. Int
Surg.
1980 Nov-Dec;65(6):529-31. PMID:
7203873
15. Brunelli GA, Brunelli GR.
Direct muscle neurotization. J Reconstr Microsurg.
1993
Mar;9(2):81-90; discussion 89-90. PMID:
8468705
16. Becker M, Lassner F, Fansa H, Mawrin
C, Pallua N.
Refinements in nerve to muscle neurotization. Muscle Nerve.
2002
Sep;26(3):362-6. PMID: 12210365
17. Urbanchek MG, Ganz DE, Aydin MA, van der Meulen JH, Kuzon WM Jr. Muscle-nervemuscle neurotization for the reinnervation of denervated somatic muscle. Neurol Res.
2004
Jun;26(4):388-94. PMID: 15198864
18. Chuang DC.
Functioning free-muscle transplantation for the upper extremity. Hand
Clin. 1997 May;13(2):279-89. Review. PMID:
9136041
19. Canale ST (ed.) Single-stage tissue transfer
(free flaps). Campbell’s Operative Orthopaedics.
10 th ed. St Louis, MO: Mosby Inc. 2003.
20. Krieger LM, Krieger AJ. The intercostal to phrenic nerve transfer: an effective means of reanimating the diaphragm in patients with high cervical spine injury. Plast Reconstr Surg.
2000 Apr;105(4):1255-61. PMID: 10744213
21. Bryden AM, Kilgore KL, Lind BB, Yu DT.
Triceps denervation as a predictor of elbow flexion contractures in C5 and C6 tetraplegia.
A rch Phys Med Rehabil.
2004 Nov;85(11):1880-
5. PMID: 15520985
OrtHOPaeDiC JOUrnaL | Case Western reserve University | vOL.5, nO.1 | 2008 | 39

lOxP
40 abStract
G enetically engineered mouse models are powerful tools for studying molecular mechanisms under physiologic and pathologic conditions. The CreloxP system, which in living mice allows the tissue-specific recombination of a “gene of interest”, has been widely used in bone and cartilage research. Recently, this system has been successfully used in a number of fracture studies. In this review, we describe the technology and some of the most widely used mouse lines for the CreloxP system in bone and cartilage.
IntroductIon
Bone and cartilage formation and maintenance involve many regulatory molecules. A previous advance in mouse genetics allowed the breeding of knockout mice, in which a “gene of interest” is specifically disrupted and inactivated. Those genetically engineered mouse models helped to identify many important regulatory mechanisms. That systemic inactivation of genes, however, often led to early embryonic death, precluding the analysis at later stages of development.
To circumvent this embryonic lethality, the CreloxP system has been developed. This DNA recombinasebased technology allows the tissuespecific recombination of a “gene of interest” in mice that are living. This technology has been used in numerous studies addressing the mechanisms of bone and cartilage development and maintenance. In fracture studies, for example, researchers have used the CreloxP system to show that
Bmp2 is essential for fracture callus formation 1 , that Bmp4 is dispensable for fracture callus formation 2 , and that beta-catenin enhances fracture healing 3 . Studies using the Cre-loxP system have also helped to elucidate the molecular mechanisms for osteoblasts, chondrocytes, periosteum, and longbone ossification centers (Table 1, 2).
I. the creloxP system
Cre recombinase
Cre recombinase is an enzyme that catalyzes the site-specific recombination of DNA between loxP sites. The loxP recognition site is a 34 base pair (bp) sequence. Cre recombinase efficiently excises the DNA fragment that is flanked—and thus identified—by two loxP sites in the same orientation.
By creating genetically engineered mice that harbor loxP sites in a gene of interest, and by expressing Cre recombinase in a tissue of interest, it is possible to recombine the target gene in a tissue-specific manner. Depending on the design of the DNA construct and the mutations introduced, the activation or inactivation of the gene of interest can be achieved in vivo.
The CreloxP mediated recombination takes place when cells start to express
Cre recombinase, and all the progeny cells derived from these cells harbor the recombined allele. Therefore, it is possible to genetically mark the cells and trace the fate of the progeny cells.
CreER
To control the timing of CreloxP mediated recombination, CreER—a fusion molecule between Cre recombinase and the ligand-binding domain (LBD) of estrogen receptor (ER) —was created
4 . Upon binding with the ligand,
CreER translocates into the nucleus and recombines genes at the loxP sites. To avoid activation by endogenous estradiol, modified CreER molecules have been generated in which Cre has been fused to a mutated LBD that is insensitive to endogenous estradiol. CreER T is a fusion molecule between Cre recombinase and
5 a mutated LBD G521R of human ER
. CreER T2 has G400V/M543A/L544A mutations, making the fusion protein up to 10 fold more sensitive to 4-hydroxytamoxifen than CreER T 6,7 . CreER TM is a fusion molecule between Cre recombinase and the mutated LBD G525R of mouse
ER 8 . These mutated CreER molecules are activated by tamoxifen, but not by estradiol.
II. regulatory elements and transgenic mouse lines for cre expression in bone and cartilage
A number of transgenic mouse lines have been generated to express Cre and CreER in bone and cartilage. The expression of Cre and CreER is driven by regulatory sequences that direct transgene expression in a tissue-specific manner. Here, we will focus on some of the most widely used regulatory elements for bone and cartilage expression and the Cre- and CreERexpressing transgenic mouse lines created by using these sequences.

MANuSCRIPTS table 1.
Timing and domains of transgene expression driven by the Col1a1, Col2a1 , and Prx1 promoters.
Promoter
3.6kb Col1a1
3.2kb Col1a1
2.3kb Col1a1
Col2a1
Prx1
Expressed gene
CAT
GFP
Cre
LacZ
CAT
LacZ
GFP
GFP, CAT
Cre
Cre
CreERT2
LacZ
Cre
Cre
Cre
Cre
CreERT
Cre TM
CreERT2
LacZ
Cre
CreERT2
Author
Pavlin
Kalajzic
Liu
Rossert
Bogdanovic
Rossert
Kalajzic
Marijanović
Dacquin
Liu
Kim
Zhou
Ovchinnickow
Haigh
Sakai
Long
Nakamura
Hilton
Chen
Martin
Logan
Hasson
Year
1992
2002
Timing Expression domain bone, tooth, tendon bone, tail tendon
2004 E18
1995 E13-14
1994
2000 E11.5
osteoblasts, osteocytes, perichondrium, periosteum, articular cartilage, skin, tendon, knee ligament, some of the growth plate chondrocytes osteoblasts, odontoblasts, skin(faint), ligaments, tendons osteoblasts, odontoblasts
1995 E13-14
2002 osteoblasts, odontoblasts, skin(faint) bone, tail tendon, nonosseous tissue, periosteal fibroblasts
2003 E11, E14 long bones, tendon, calvaria, teeth
2002 E14.5
skull, long bone ossification center (osteoblasts), skin of the digits and face
2004 E18
2004 E16.5
1995 E10.5
osteoblasts, osteocytes, skin, some of the growth plate chondrocytes bones, teeth (osteoblasts, odontoblasts) chondrocyte, pre-chondrocytes, notochord
2000 E9.5
cartilaginous primordia, submandibular glands, notochord cartilage, eye, heart, neuroepithelium, epidermis, cranial mesenchymal cells
2001 E10.5
2001 E12.5
2006 E10
2007 E9.5
2007 E15.5
2000 E10.5
2002 E9.5
2007 E8.5
cartilaginous lineage including sclerotome and mesenchymal condensations, notochord, developing brain, perioptic region, ligament, tendons, osteoblasts in primary spongiosa, periosteum, perichondrium cartilaginous structures(ectoderm and mesenchyme), facial mesenchyme, CNS chondrogenic lineage, perichondrium
Chondrocytes, perichondrium, primary spongiosa, joint capsule chondrocytes limb bud mesenchyme, tendons, periosteum, lateral craniofacial mesenchyme limb bud mesenchyme, cranial mesenchyme, interlimb flank mesoderm, germline limb mesenchyme
1. Col1a1 promoter
Type I collagen is a major constituent of bone matrix. Type I collagen consists of two alpha 1 chains and one alpha 2 chain that form a triple helix. Type I collagen is synthesized by osteoblasts and various other cell types, including periosteal cells, tendon cells, odontoblasts, and dermal fibroblasts.
Osteoblast-specific enhancer element has been identified in the mouse and rat Col1a1 gene, the gene for alpha
1 chain. Both mouse and rat 2.3 kb promoter sequences direct transgene expression in osteoblasts 9 (Table
1). The promoter is also active in odontoblasts in the teeth. A longer mouse 3.2 kb promoter and rat 3.6 kb promoter drive expression of the transgene also in the tendon, skin and periosteum 10,11 . Both the 2.3 kb and
3.6 kb promoter sequences become active in long bones from E13-14 9 .
Col1a1-Cre mice
A number of research groups have generated Cre-expressing transgenic mouse lines using promoter sequences of Col1a1 . At least two groups have generated mice using the mouse 2.3
OrtHOPaeDiC JOUrnaL | Case Western reserve University | vOL.5, nO.1 | 2008 | 41

THE CRElOxP SYSTEM
42 kb Col1a1 promoter 12,13 . At E14.5,
CreloxP mediated recombination is detected in all long bone ossification centers and the skin of the digits and face 12 . These mice have been successfully used to inactivate a number of genes in osteoblasts (Table 2).
Rat 2.3 kb and 3.6 kb Col1a1 promoter sequences have been also used to drive
Cre recombinase expression in the bone
11 . Interestingly, when these transgenic mice were used to recombine the Igf1 gene, the Igf1 locus was rearranged in some of the progeny in the absence of inheritance of the Cre transgene.
The incidence was 50 and 28% with
Col2.3-Cre and Col3.6-Cre females, respectively, and 15 and 18% with
Col2.3-Cre and Col3.6-Cre males, respectively 14 . This may be caused by the early activation of Col1a1 promoters in gamete progenitor cells such that cytoplasmic Cre mRNA/ protein is distributed to haploid cells during meiotic division. Therefore, careful confirmation of tissue-specific gene recombination would be necessary when using these mice.
Col1a1-CreER mice
Transgenic mice that express CreER in osteoblasts under the control of a
2.3 kb and 3.2 kb Col1a1 promoter have been generated 15,16 . These mice allow the timed activation of Cre recombinase in osteoblasts. These mice may be useful for studying postnatal bone repair and bone maintenance.
However, the utility of these mice for such studies remains to be examined.
The 3.2 kb Col1a1-CreER transgenic mice have been used to genetically mark and trace perichondrial osteoblasts, which migrate to and populate the primary ossification center
16 .
2. Col2a1 promoter/enhancer
Type II collagen is the most abundant collagen in cartilage. Col2a1 , the gene encoding the pro-alpha1(II) collagen chain, is a principal marker of chondrocyte differentiation. There are two splice isoforms, type IIA and type IIB 17 . Type IIA is expressed in the periosteum and perichondrium and other non-cartilaginous tissues, while type IIB is expressed in differentiated chondrocytes. During embryonic development, the earliest
Col2a1 expressions are seen in nonchondrogenic tissue at E9.5 18 . Col2a1 is also expressed in non-cartilaginous tissues, such as the notochord, heart, epidermis and discrete areas of the brain. Col2a1 expression in the developing limb starts as early as
E10.5-11.5
.
Cis-acting regulatory elements of Col2a1 that direct transgene expression in chondrocytes have been identified. Two widely used constructs are a combination of 3.0 kb
Col2a1 promoter and 3.0 kb intron 1 sequences 19 and a combination of 1.0 kb promoter and 650-800bp intron 1 sequences 20-22 .
Col2a1-Cre, CreER mice
Several research groups have generated mice that express Cre and CreER in chondrocytes using these sequences
23-27 . When using the Col2a1-Cre mice, recombination in cartilage starts around E12.5. These transgenic mouse lines have been successfully used to recombine numerous genes in chondrocytes (Table 2). Col2a1-
CreER T2 mice have been also shown to efficiently recombine genes in articular chondrocytes 28 .
Recombination outside of cartilage
Off-target recombination outside of cartilage has been reported for some of the transgenic mouse lines.
Notably, there have been conflicting reports about recombination in the perichondrium and periosteum.
Because a splice isoform type IIA is expressed in the periosteum and perichondrium 17 , it is possible that
Col2a1 regulatory sequences also direct transgene expression in these tissues. In addition, if the transgene is expressed in the osteo-chondroprogenitor cells that give rise to both chondrocytes and osteoblasts, CreloxP mediated recombination may be observed in both chondrocytes and osteoblasts.
Ovchinnikov et al. 23 created a transgenic mouse line using the 3.0 kb Col2a1 promoter and 3.0 kb intron 1 sequences and showed no
CreloxP mediated recombination in the perichondrium of neonatal tibia. Baffi et al. 29 also analyzed the same transgenic mouse line by in situ hybridization and detected no Cre expression in the perichondrium of phalanges at E15.5 29 . In contrast, Day et al. 30 showed recombination in the perichondrium of tibia at E15.5 using the same transgenic mouse line 30 .
Another Col2a1-Cre transgenic mouse line created by using a similar promoter and intron 1 sequences also showed recombination in the perichondrium and osteoblasts in the primary spongiosa 25 .
CreloxP mediated recombination outside of cartilage has been also reported in transgenic mice expressing
CreER. In a Col2a1-CreER T mouse line,
CreloxP mediated recombination was detectable in the perichondrium at 24 h and 36 h after tamoxifen injection at
E13.5 31 . However, no recombination was detected in the perichondrium and periosteum of tibiae at age 7 and
14 days when tamoxifen was injected at birth using the same Col2a1-
CreER T mouse line 32 . Hilton et al. also expressed CreER TM under the control of Col2a1 regulatory sequences 27 . They administered tamoxifen between E11.5 and E14.5 and found that tamoxifen administration at or after E12.5

MANuSCRIPTS restricted recombination largely to chondrocytes and the primary spongiosa in long bones.
Our own laboratory has also examined
CreloxP mediated recombination in the perichondrium using the
Col2a1-Cre transgenic mice generated by
Ovchinnikov et al. These mice were crossed with
ROSA26 reporter mice, and ROSA26; Col2a1-
Cre offspring were stained with X-gal at age 3 days. We observed intense X-gal staining in the perichondrium and periosteum of the tibia in addition to underlying epiphyseal cartilage. We also found X-gal staining in the periosteum of ribs, while we did not observe X-gal staining in the periosteum and perichondrium of the sternum of the same mouse. These observations indicate that even in the same mouse, the extent of
CreloxP recombination in the periosteum and perichondrium varies considerably depending on the bone. Therefore,
CreloxP recombination of the target gene in the periosteum and perichondrium should be examined for each skeletal element at each developmental stage.
3. Prx1 promoter
The paired -related homeobox gene-1 (Prx1, table 2.
Col1a1-Cre, Col2a1-Cre, Prx1-Cre
Mouse line Targeted gene Author Journal
Col1a1-Cre Cx43
Mmp13
G(s) alpha
PTHrP
Nf1
Mdm2
Stat3
Cx43
AR
Igf 1 beta-catenin
Presenilin-1/2
Notch1
Castro
Stickens
Sakamoto
Miao
Elefteriou
Lengner
Itoh
Chung
Notini
Cochrane
Chen
Engin
Zanotti
Cell Commun Adhes
Development
J Biol Chem
J Clin Invest
Cell Metab
J.Cell Biol
Bone
J Cell Sci
J Bone Miner Res
Genesis
PLoS Med.
Nat Med
Endocrinology
Col2a1-Cre
Col2a1-CreER Ihh
Smo
Prx1-Cre Sox9
Fgfr3
Bmp4
Vegfa
Presenilin-1/2
Dicer
Notch1
Tbx4, 5
Sox9
Bmpr1a
Bmp2
Tgfbr2
HIF 1alpha
Tgfbr2
Tbx4
Kif3a, Ift88
Bmp4
Psen1,2, Notch1,2
Fgfr1, Fgfr2
Vegfa
Fgfr3 K644E
HIF-1alpha
Smo
Sox9
CYP27B1
ILK
Jun
Haigh
Iwata
Schipani
Long
Akiyama
St-Arnaud
Terpstra
Behrens
Smo
Vegfa
Vhlh
Fosl2
Long
Zelzer
Pfander
Karreth
Tgfbr2 Baffi
X-linked puDeltatk Chen
G(s) alpha
Smad4
Ihh beta-catenin
Hoxa2
Kif3a
Pten
Wnt4
Igf1
Bmp4 cRNAi
Development
Hum Mol Genet
Genes Dev
Development
Genes Dev
J Cell Biochem
J Cell Biol
Development
Development
Development
Development
Development
Dev Biol
Nucleic Acids Res
J Bone Miner Res
Dev Biol
Sakamoto
Zhang
Razzaque
Rodda
Massip
Song
Ford-Hutchinson J Bone Miner Res
Lee
Govoni
Li
J Pathol
Development
Differentiation
Dev Biol
PLoS ONE
Physiol Genomics
Yi Chuan
Maeda
Hilton
Akiyama
Murakami
Selever
Zelzer
Pan
Harfe
Francis
Minguillon
Akiyama
Ovchinnikov
Tsuji
Seo
Provot
Spagnoli
Naiche
Haycraft
Tsuji
Hilton
Yu
Proc Natl Acad Sci USA
Dev Biol
Genes Dev
Genes Dev
Dev Biol
Development
Dev Biol
Proc Natl Acad Sci USA
Dev Dyn
Dev Cell
Proc Natl Acad Sci USA
Dev Biol
Nat Genet
Dev Biol
J Cell Biol
J Cell Biol
Development
Development
J Bone Joint Surg Am
Nat Med
Development
Prx1-CreER Tbx5 Hasson Development
104
308
102
295
38
310
177
177
134
134
286
102
234
8
16
18
276
131
90(Suppl 1)
14
135
134
Volume Page Year
131
131
131
131
276
32
20
284
127
9
15
128
16
88
162
130
22
2
30
30
207
133
75
305
10
131
280
115
4
172
39
119
3
45
4
14
Apr 17
445-50
5883-95
2402-11
441-51
909-21
505-12
2003
2004
21369-75 2005
2005
2006
2006
2006
4187-98 2006
347-56
17-20
2007
2007 e249 2007
299-305 2008
2008
1445-53 2000
1603-13 2000
2865-76 2001
5099-108 2001
2813-28 2002
245-51 2003
2003 139-48
103-9 2003
1309-18 2004
2161-71 2004
2497-508 2004
5717-25 2004
124-42 e161
663-71
311-22
453-61
2004
2004
2005
2005
2005
3231-44 2006
256-67
202-16
1245-59 2007 e450
354-62
341-6
2007
2007
2007
2007
2008
6382-7
93-105
2007
2007
2813-28 2002
290-305 2004
268-79 2004
2161-71 2004
472-82 2005
10898-903 2005
1006-15 2005
75-84 2005
14665-70 2005
103-15
1242-9
2006
2006
2007
2007
304-16
451-64
1105-17 2007
93-103
307-16
14-8
306-14
483-91
2007
2007
2008
2008
2008
85-92 2006
OrtHOPaeDiC JOUrnaL | Case Western reserve University | vOL.5, nO.1 | 2008 | 43

THE CRElOxP SYSTEM
44 formerly called MHox1) belongs to the family of vertebrate homeobox genes which play essential roles in embryonic development. Endogenous
Prx1 expression starts to be expressed at E7.5 in the ectoderm, mesoderm, and part of the allantois 33 . Cserjesi et al. reported that Prx1 expression starts in the lateral mesoderm at around
E9.5 34 . Prx1 is expressed in the limb and cranium mesenchyme, somites, branchial arches, heart and developing brains. The endogenous Prx1 is not expressed in differentiated chondrocytes and bone cells, while its expression is maintained in the perichondrium
33,35,36 .
2.4 kb Prx1 promoter
A 2.4 kb Prx1 promoter has been shown to direct transgene expression in undifferentiated mesenchyme in the developing limb buds 37 . The transgene expression is detectable as early as E10.5. The 2.4 kb Prx1 promoter directs transgene expression also in the maxillary process and lateral cranial mesenchyme 37 . The transgene expression is extinguished in the condensing mesenchyme, and the expression is confined to the periosteum of the long bones and tendons of the limbs at E15.5.
Prx1-Cre transgenic mice
Prx1-Cre transgenic mice that express
Cre recombinase under the control of the 2.4 kb Prx1 promoter have been generated 38 . These mice have been successfully used to recombine genes in the limbs, cranium, and sternum (Table
2). i) limbs
Cre recombinase activity is detectable in the developing limb bud as early as
E9.5. While the transgene is active in the undifferentiated mesenchyme, it is not expressed in the surface ectoderm, including AER (apical ectodermal ridge). The Prx1-Cre transgene has been successfully used to inactivate
Tbx4 and Tbx5 , showing their importance in hindlimb and forelimb development, respectively 39,40 . When a master chondrogenic factor, Sox9 , was inactivated using the Prx1-Cre transgene, both chondrocyte and osteoblast lineages failed to develop
41 . These observations clearly indicate that the Prx1-Cre transgene targets osteochondral progenitor cells that give rise to chondrocytes and osteoblasts.
The Prx1-Cre transgene has been also used for postnatal analyses and fracture studies of long bones 1,2 . ii) cranium
The Prx1-Cre transgene is also expressed in the cranial mesenchyme around the lambdoid suture. It has been useful for examining the roles of target genes in calvaria development. The inactivation of beta-catenin and TGF-beta type II receptor (Tgfbr2) resulted in a loss of parietal and interparietal bones 42,43 .
The inactivation of Tgfbr2 caused intracranial hemorrhage resulting in perinatal lethality. We have also inactivated ERK2 in the ERK1 -null background using the Prx1-Cre transgene. The lambdoid suture failed to close at least up to 12 days of age, indicating the essential roles of ERK1 and ERK2 in cranial bone development
44 . iii) sternum
The Prx1-Cre transgene is also active in the interlimb flank mesoderm that gives rise to the sternum 38 . The inactivation of Sox9 resulted in a lack of the sternum, which apparently resulted in perinatal lethality of Sox9 mutant mice 41 . We have also used the
Prx1-Cre transgene to recombine the
Fgfr3 allele so that an achondroplasia mutant of Fgfr3 is expressed. These mice showed premature closure of the sternal synchondroses, indicating that the Prx1-Cre transgene is useful for analyzing the mechanisms of sternum development. iv) germline
An important aspect of the Prx1-
Cre mice is germline recombination.
A partially penetrant germline recombination was observed in the offspring of female Prx1-Cre mice harboring an allele with loxP sites 38 .
In contrast, germline recombination was not observed in the offspring of male Prx1-Cre mice. Because germline recombination results in the systemic recombination of the target allele, the
Prx1-Cre transgene should be inherited from male mice for tissue-specific recombination.
Prx1-CreER transgenic mice
For timed activation of Cre recombinase, mice expressing CreER T2 have been also generated 45 . Single oral gavage of tamoxifen at E8.5 induces uniform Cre recombination throughout the limb bud by E10.5.
These mice were used to define the time frame during which Tbx5 is required for limb development.
4. other transgenic mouse lines
Various other mouse lines have been generated for expressing Cre recombinase in a subset of cell populations in bone and cartilage.
These include GDF5-Cre mice for articular chondrocytes 46 , Col10a1-Cre mice for hypertrophic chondrocytes
47 , Sox9-Cre and Dermo1-Cre mice for osteo-chondroprogenitor cells 48,49 , and Osteocalcin-Cre and Osterix-Cre mice for osteoblasts 50,51 . These mouse lines would be useful for studying the functions of target cell populations.
The CreloxP system in orthopaedic research
Recently, the CreloxP system has been used in a number of fracture studies 1-3 .
Prx1-Cre transgenic mice were used to inactivate Bmp2 and Bmp4 in fracture

MANuSCRIPTS healing. When Bmp2 was inactivated using the Prx1-Cre transgene, no callus formation was observed, indicating that Bmp2 is essential for fracture callus formation 1 . In contrast, Bmp4 was dispensable for fracture callus formation 2 . Furthermore, the Col1a1-
Cre transgene was used to activate or inactivate beta-catenin signaling in osteoblasts during fracture healing
3 . While the inactivation of betacatenin significantly inhibited bone regeneration, the activation of betacatenin enhanced fracture healing.
Genetic manipulation using the CreloxP system, therefore, is a powerful approach to evaluate the outcome of gain-of-function or loss-of-function of regulatory molecules in fracture healing. The CreloxP system would be also useful for studies such as bone and cartilage regeneration and tissue engineering. Whether or not the commonly used Cre-expressing mouse lines are suitable for these studies remains to be examined. In addition, the disruption of developmentally important genes often leads to preexisting phenotypes that can preclude the postnatal experiment. In such cases, the use of CreER-expressing mouse lines should circumvent the problem. However, the promoter activity and the expression levels of the transgene in adult mice can be much lower than in embryos. The identification of regulatory elements that drive high levels of transgene expression in adult mice—and the establishment of new Cre and CreERexpressing mouse lines—may be necessary for future orthopaedic studies using the CreloxP system.
ACKNOWLEDGMENTS
We thank Valerie Schmedlen for editorial assistance.
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51. Rodda SJ, McMahon AP.
Distinct roles for
Hedgehog and canonical Wnt signaling in specification, differentiation and maintenance of osteoblast progenitors. Development.
2006;
(133)3231-44.

MANuSCRIPTS
abStract
E
RK1 and ERK2 Mitogen-activated protein kinases (MAPK) are major downstream effectors in FGFR3 signaling. Activating mutations in
FGFR3 and molecules in the MAPK pathway cause achondroplasia and other skeletal developmental disorders.
To study the role of ERK1 and ERK2 in skeletal development, the CreloxP system was used to inactivate
ERK1 and ERK2 . The inactivation of ERK1 and ERK2 in chondrocytes resulted in a larger vertebral body with greater anterior-posterior length and cross-sectional area. In addition, these embryos showed longer long bones with wider epiphyses. These findings indicate that ERK1 and
ERK2 negatively regulate the size of cartilaginous skeletal elements.
Histologically, the vertebral synchondroses had an expansion of the zone of hypertrophy with disorganization of the growth platelike architecture, indicating ERK1 and
ERK2 play an important role in the organization of the synchondroses.
Interestingly, the inactivation of ERK1 and ERK2 also resulted in a smaller vertebral foramen. This finding might be a concern for future therapies for achondroplasia directed at inhibition of the MAPK pathway.
IntroductIon
The Mitogen activated protein kinase (MAPK) pathway consists of a series of phosphorylating kinases that play important roles in skeletal development. The signaling cascade begins with a MAPKKK (Raf) which phosphorylates a MAPKK (MEK 1/2).
MAPKK phosphorylates MAPK (ERK
1/2) which can then activate other kinases such as RSK 2. Mutations in the MAPK pathway, its upstream activators, and downstream effectors have been shown to cause a number of human skeletal disorders, including
Noonan, Costello, Coffin-Lowry, and cardio-facio-cutaneous syndromes.
One of the key activators of the MAPK pathway is the FGF receptor family.
A ctivating mutations in FGF receptor
3 (FGFR3) cause achondroplasia, thanatophoric dysplasia, and hypochondroplasia 1 . Achondroplasia is the most common form of shortlimb dwarfism and affects about 1 in
15,000 live births 2 .
A number of achondroplasia mouse models have been generated by expressing Fgfr3 with gain-of-function mutations 3,4 . These mouse models have shown that increased Fgfr3 signaling leads to an inhibition of chondrocyte proliferation and maturation as well as abnormal growth plate vascularization resulting in a dwarf phenotype. In contrast,
Fgfr3 -deficient mice show an increase in hypertrophic chondrocytes and an expansion of the zone of hypertrophy in the growth plate 5,6 . These animals show a thicker growth plate cartilage as well as skeletal overgrowth with larger long bones.
Recent studies on the MAPK pathway have indicated that the MAPK pathway is a critical downstream effector of Fgfr3. The growth plates of mice expressing the human achondroplasia mutation have increased levels of phosphorylated
MEK1, a MAPKK that phosphorylates ERK1/2 7 . In addition, mice expressing a constitutively active MEK1 in chondrocytes show an achondroplasia-like phenotype 8 .
These animals had incomplete hypertrophic differentiation of chondrocytes and reduced bone growth. The expression of a constitutively active MEK1 mutant in chondrocytes rescued skeletal overgrowth in Fgfr3 -deficient mice, suggesting that much of the negative regulation of bone growth by Fgfr3 is mediated by the MAPK pathway.
Experiments examining the effects of C-type natriuretic peptide (CNP) on skeletal development have also suggested that the MAPK pathway plays important roles in the regulation of endochondral bone growth. CNP is a peptide consisting of 22 amino acids that is expressed in growth plate chondrocytes. CNP has been shown to block activation of the
MAPK cascade in response to FGF in vitro 9, 10 . Targeted overexpression of CNP in chondrocytes causes skeletal overgrowth and rescues the dwarf phenotype in a mouse model of achondroplasia 11 .
The rescue of the dwarf phenotype was associated with
OrtHOPaeDiC JOUrnaL | Case Western reserve University | vOL.5, nO.1 | 2008 | 47

ERK1 ANd ERK2
48 reduced phosphorylation of ERK1/2, supporting the notion that CNP promotes bone growth by inhibiting
ERK1/2. However, the exact mechanism by which CNP promotes bone growth requires further investigation, since CNP signaling is also mediated by other signaling molecules such as cGMP-dependent kinase II and GSK3bata 12 .
Recent work in our laboratory has also indicated that Fgfr3 and the MAPK pathway regulate synchondrosis closure.
A synchondrosis consists of two opposed growth plates with a common zone of resting chondrocytes 13 . We observed premature synchondrosis closure in the cranial base and vertebrae of mice expressing the Fgfr3 G374R achondroplasia mutation and mice expressing a constitutively active
. MEK1 mutant in chondrocytes 14
These observations suggest that premature synchondrosis closure accounts for the foramen magnum and spinal canal stenosis in achondroplasia. A majority of individuals with achondroplasia suffer from spinal canal stenosis 15 .
Foramen magnum and cervical spinal stenosis associated with compression of the brainstem and spinal cord can lead to sudden death during early childhood 16 . In addition, spinal canal stenosis can cause other neurologic complications such as radiculopathy, claudication, and paraparesis.
While increased MAPK signaling inhibits endochondral bone growth and accelerates the closure of synchondrosis, whether or not reduced activity of MAPK promotes the growth of cartilage and increases the spinal canal dimensions remain to be investigated. In this study, we examined the effects of inactivation of ERK1 and ERK2 on the growth of vertebrae and epiphyses in the long bones. We show here that inactivation of ERK1 and ERK2 in chondrocytes resulted in increased dimensions of the epiphysis of the long bones and vertebral bodies, while ERK1 and
ERK2 inactivation caused reduced dimensions of the spinal canal. materIalS and methodS
Mice
ERK1 -null mice and mice with the floxed ERK2 allele were provided by Dr. Gary Landreth (Dept. of
Neurosciences, Case Western Reserve
University). While ERK1 -null mice develop normally, systemic inactivation of ERK2 results in lethality at embryonic day 6.5 (E
6.5) 1 . Therefore, the CreloxP system was used to inactivate ERK2 in chondrocytes of ERK1 -null mice.
Col2a1-Cre transgenic mice were used to express Cre recombinase specifically in chondrocytes 17 . To inactivate ERK1 and ERK2 in chondrocytes, we crossed
ERK1 -/+ ; ERK2 flox/flox ; Col2a1-Cre mice with ERK1 -/; ERK2 flox/flox mice.
By using this mating strategy, we successfully generated mice in which one allele (1-allele KO: ERK1 -/+ ; ERK2 flox/flox
-/+
), three alleles (3-allele KO: ERK1
; ERK2 flox/flox ; Col2a1-Cre ), and all four alleles (4-allele KO: ERK1 -/;
ERK2 flox/flox ; Col2a1-Cre ) of ERK1 and
ERK2 were inactivated. While mice with at least one functional allele of either ERK1 or ERK2 are viable and fertile, 4-allele KO mice die perinatally due to chondrodysplasia.
The genotype of each embryo was determined by
PCR using specific primers for ERK1 and Cre recombinase.
Skeletal Preparations and
Measurements
Embryos were skinned, eviscerated, and fixed in 95% ethanol for several days.
Samples were cleared using 1% KOH for 1-3 hours and then stained using a solution of Alizarin red for four hours.
Samples were then cleared in 0.1%
KOH for two days before being placed into a 1:1 solution of 95% ethanol and glycerol. Photographs of isolated bones were taken along with a ruler using a dissection microscope, a digital camera, and Leica Application software. Scion
Image software (Scion Corporation) was used to measure bone lengths along the longitudinal axis as well as epiphyses widths at their widest point.
Histological examination
A section of thoraco-lumbar vertebrae were dissected out of the embryos for histological analysis. The tissues were dimineralized in 0.5 M EDTA before embedding. Tissues were fixed in 10% formalin and embedded in paraffin. 7 µm thick sections were stained with hematoxylin, eosin, and alcian blue. Photographs were taken of the stained sections utilizing Leica
Application software and measurements of the vertebral body and foramen were made using Scion Image software.
Anterior-posterior measurements were made at the midline, and lateral diameter was measured by taking the lateral dimension at the midpoint of the anterior-posterior midline (Figure
1-D).
Statistical Analysis of Data
Analysis was performed using Analyse-it software for Microsoft Excel (Analyseit Software, Ltd.) . A 1-way ANOVA analysis was performed with a post-hoc pairwise Scheffe test. Results with 95% or greater confidence were considered significant. reSultS
Vertebral body measurements were made according to the scheme in Figure
1-D. There were significant differences in the vertebral body dimensions and architecture of synchondroses between embryos in which all four alleles of

MANuSCRIPTS
ERK1/2 were inactivated (4-allele
KO: ERK1 -/; ERK2 flox/flox ; Col2a1-Cre ) and embryos in which only one allele flox/flox ) (1-allele KO: ERK1 -/+ ; ERK2 or three alleles (3-allele KO: ERK1 -/+ ;
ERK2 flox/flox ; Col2a1-Cre ) of ERK1/2 were inactivated. The anterior-posterior length of the vertebral body was much longer in 4-allele KO embryos than in the other groups (Figure 1-A). Similar tendency was also observed in the lateral diameter of the vertebral body
(Figure 1-B). The cross-sectional area of the vertebral body was also significantly larger in 4-allele KO embryos (Figure
1-C).
Histological analysis of the vertebrae showed profound differences between
4-allele KO embryos and 1-allele
KO embryos (Figures 2). We did not observe obvious differences between
1-allele KO and 3-allele KO embryos
(data now shown). While the vertebral body of 1-allele KO embryos (Figure
2-E) was mostly ossified, and the synchondroses consisted of growth plate-like zones, the vertebral body of 4-allele KO embryos remained entirely cartilaginous and consisted of hypertrophic chondrocytes. The increased number of hypertrophic chondrocytes and its persistent presence may account for the larger crosssectional area and anterior-posterior length of the vertebral body. 4-allele
KO embryos also showed persistent presence of hypertrophic chondrocytes in the vertebral arch (Figures 2-B and
2-D).
Figure 1: vertebral body measurements. a: anterior – posterior length B: Lateral diameter C: Cross-sectional area. D: scheme for measurements. values are the mean ± sD (error bars indicate the sD)
*p<0.05, **p<0.01, ***p<0.001. (1 allele, n=6) ERK1 -/+ ; ERK2 n=4) ERK1 -/+ ; ERK2 flox/flox alleles, n=3) ERK1 -/-
Cre.
; Col2a1-Cre
; ERK2 flox/flox flox/flox
; (3 alleles,
; (4
; Col2a1-
Figure 2: Cross section of lower thoracic vertebrae. a, C: ERK1 -/+ ; ERK2 flox/flox embryos at e 18.5. B, D: ERK1 -/the vertebral body of ERK1 embryo (f) at e 18.5.
; ERK2
-/+ flox/flox ; Col2a1-Cre embryos at e18.5. e, f: Magnified views of
; ERK2 flox/flox embryo (e) and ERK1 -/; ERK2 flox/flox ; Col2a1-Cre
OrtHOPaeDiC JOUrnaL | Case Western reserve University | vOL.5, nO.1 | 2008 | 49

ERK1 ANd ERK2
Vertebral foramen measurements were also made according to the scheme in Figure 1-D. In contrast to the measurements of the vertebral body, the cross-sectional area of the vertebral foramen was significantly larger in
1-allele KO embryos in comparison to
3-allele KO and 4-allele KO embryos
(Figure 3-A). In addition, 1-allele KO embryos had a larger lateral diameter in comparison to the other genotypes
(Figure 3-C). The anterior-posterior lengths were not significantly different between 1-allele KO and 4-allele KO; however, 3-allele KO embryos had significantly smaller lateral diameters and anterior-posterior lengths in comparison to the other genotypes
(Figures 3-B,C). These differences in vertebral foramen measurements could be related to the increased size of the vertebral body in 4-allele KO embryos (Figure 1-A,C). Also, there was a marked difference in the shape of the vertebral foramen between
1-allele KO and 4-allele KO embryos
(Figure 2-A,C) (Figure 2-B,D). The vertebral foramen of 4-allele KO embryos was less circular, and the spinal cord appeared to be compressed.
The irregularity in the shape of the vertebral foramen probably reflects the abnormalities in the ossification process and persistent presence of hypertrophic chondrocytes in the vertebral body and vertebral arch.
The skeletal preparations of the embryos were analyzed for long bone length in the humerus, radius, ulna, femur and tibia. The lengths were measured along the longitudinal axis as shown in Figure 4-E. The most
50
Figure 3: vertebral foramen measurements. a: Cross-sectional area.
B: anterior-Posterior length. C. Lateral diameter. values are the mean ± sD (error bars indicate sD). *p<0.05, **p<0.01,
***p<0.001. (1 allele, n=6) ERK1 -/+ ;
ERK2 flox/flox ; (3 alleles, n=4) ERK1 -/+ ; ERK2 flox/flox ; Col2a1-Cre ; (4 alleles, n=3) ERK1
-/; ERK2 flox/flox ; Col2a1-Cre.
Figure 4: Long bone measurements. a: Length of humerus, ulna, radius, femur, and tibia in ERK1 -/+ ; ERK2
, and ERK1 -/flox/flox (1 allele, n=6), ERK1
; ERK2 flox/flox
-/+ ; ERK2 flox/flox ; Col2a1-Cre (3 alleles, n=4)
; Col2a1-Cre (4 alleles, n=3) embryos at e18.5
.
. B: epiphysis width of proximal and distal humerus. C: epiphysis width of proximal and distal femur. D. epiphysis width of proximal and distal tibia. e: scheme for measurements. values are the mean ± sD (error bars indicate sD). *p<0.05, **p<0.01, ***p<0.001.

MANuSCRIPTS proximal long bones (humerus and femur) were significantly longer in
4-allele KO embryos in comparison to 1-allele KO and 3-allele KO embryos (Figure 4-A). The epiphyses widths were also measured as shown in Figure 4-E. 4-allele KO embryos had significantly wider epiphyses in the proximal and distal humerus and femur. 4-allele KO embryos also showed wider epiphyses in the distal tibia. dIScuSSIon
Our histological analyses and skeletal measurements clearly indicated that
ERK1/2 plays an important role in the regulation of endochondral bone growth. The inactivation of all four alleles of ERK1 and ERK2 resulted in longer and wider epiphyses of the long bones. The ERK1/2 inactivation also resulted in a larger vertebral body with a greater anterior-posterior length and cross-sectional area. These observations are consistent with the notion that MAPK signaling negatively regulates endochondral bone growth and the MAPK pathway mediates
Fgfr3 signaling in endochondral ossification. The most proximal long bones (humerus and femur) were significantly longer in 4-allele KO embryos similar to Fgfr3 -deficient mice 5 . This phenotype is opposite to human achondroplasia, which is characterized by rhizomelic dwarfism, a disproportionate dwarfing of the proximal bones 5 . Similar to vertebral bodies, the long bones of 4-allele
KO embryos were characterized by remarkable expansion of the zone of hypertrophic chondrocytes 18 .
Therefore, the increased length of long bones of 4-allele KO embryos is consistent with previous data showing that hypertrophic chondrocytes are a major determinant of bone length 19 .
In addition, it has been estimated that chondrocyte hypertrophy contributes about 44-59% of bone growth depending on the speed of ossification in the growth plate 20 . The epiphyses of 4-allele KO embryos were also wider. The decreased activity of
ERK1 and ERK2 may account for the larger femoral head and tibial condyles observed in Fgfr3 -deficient mutants 6 .
Histological analysis of the vertebrae of 4-allele KO embryos indicated a remarkable delay in ossification. There are an abundance of hypertrophic chondrocytes in the vertebral body, developing neurocentral synchondrosis, and vertebral arch.
Normal synchondroses consist of two opposed growth plates with distinct zones of reserve, proliferative, and hypertrophic chondrocytes. Chondrocytes stack up like coins in the proliferative zone to form organized columns of chondrocytes. In 4-allele KO embryos , no distinct zones were observed in the developing neurocentral synchondrosis, and there is a disorganized mass of hypertrophic chondrocytes in the vertebral body. These observations indicate that ERK1 and ERK2 are essential for the proper organization of the synchondrosis. We have observed similar disorganization of the epiphyseal growth plate in the long bones of 4-allele KO embryos embryos 18 . While both 4-allele KO embryos and Fgfr3 -deficient mice show an expansion of the zone of hypertrophic chondrocytes, the growth plate architecture is well maintained in Fgfr3 -deficeint mice.
Because various cytokines and growth factors signal through the MAPK pathway, it is possible that ERK1 and
ERK2 are activated by other signals in the absence of Fgfr3.
Due to the perinatal lethality of
4-allele KO mice, it is not possible to determine the effects of the loss of
ERK1 and ERK2 on the closure of the neurocentral synchondrosis, which normally takes place at age 12-14 days.
However, the pronounced delay in the ossification of the vertebral body and the abundance of hypertrophic chondrocytes in the developing neurocentral synchondrosis of 4-allele
KO embryos suggest that the overall endochondral ossification process is remarkably delayed in the absence of ERK1 and ERK2. Because loss of ERK1 and ERK2 also results in increased dimensions of cartilaginous skeletal elements, the inactivation of
ERK1 and ERK2 may delay the timing of neurocentral synchondrosis closure and expand the dimensions of the vertebral canal. However, 4-allele KO embryos showed smaller vertebral canal with respect to cross-sectional area and lateral dimensions at E18.5. The increased growth of the vertebral body toward the spinal canal may account for the smaller vertebral canal in 4-allele
KO embryos. In addition, the delayed ossification of the vertebral arch and continued presence of hypertrophic chondrocytes apparently resulted in the alteration of the shape of the vertebral arch contributing to the narrower spinal canal. While embryos with at least one functional allele of ERK1 and ERK2 showed a circular vertebral canal and round spinal cord, the vertebral canal of 4-allele KO embryos was irregular and the spinal cord was apparently compressed. Alteration of the spinal canal dimensions and overgrowth of the vertebral body might be a concern in future therapies for achondroplasia targeting the ERK
1/2 MAPK pathway.
In conclusion, the inactivation of
ERK1 and ERK2 resulted in an increased size of the epiphyses in the long bones and vertebral bodies, indicating ERK1/2 negatively
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THE EffECTS Of AgE ANd gENdER
ERK1 ANd ERK2
52 regulates the size of cartilaginous skeletal elements. These observations further support the notion that
ERK1/2 mediates Fgfr3 regulation of endochondral bone growth.
While inhibiting ERK1/2 may rescue the long bone dwarfism in achondroplasia, it remains to be determined how such inhibition might affect development of the vertebrae and spinal canal. Our histological analysis of the vertebrae of 4-allele
KO embryos indicate that overgrowth of the vertebrae and alteration of the shape of the spinal canal are possible consequences of therapies targeted at inhibiting ERK1/2. Such complications may be avoided, if
ERK1/2 is inhibited after the vertebral body and arch ossify. Future studies that inactivate ERK1 and ERK2 postnatally in chondrocytes should determine how the inhibition of
ERK1 and ERK2 affects the postnatal development of the vertebrae and long bones.
REfERENCES
1. Matsushita T, Murakami S.
The mitogenactivated protein kinase pathway in skeletal development. Case Orthop J. 2007; (4) 34-40.
2. Rousseau F, Bonaventure J, Legeai-Mallet L,
Pelet A, Rozet JM, Maroteaux P, Le Merrer
M, Munnich A.
Mutations in the gene encoding fibroblast growth factor receptor-3 in achondroplasia. Nature. 1994; (371) 252-4.
3. Wang Y, Spatz MK, Kannan K, Hayk H, Avivi
A, Gorivodsky M, Pines M, Yayon A, Lonai
P, Givol D. A mouse model for achondroplasia produced by targeting fibroblast growth factor receptor 3. Proc Natl Acad Sci U S A. 1999;
(96) 4455-60.
4. Segev O, Chumakov I, Nevo Z, Givol D,
Madar-Shapiro L, Sheinin Y, Weinreb M,
Yayon A. Restrained chondrocyte proliferation and maturation with abnormal growth plate vascularization and ossification in human
FGFR-3 (G380R) transgenic mice. Hum Mol
Genet. 2000; (9) 249-58.
5. Colvin JS, Bohne BA, Harding GW, McEwen
DG, Ornitz DM. Skeletal overgrowth and deafness in mice lacking fibroblast growth factor receptor 3. Nat Genet. 1996; (12) 390-97.
6. Deng C, Wynshaw-Boris A, Zhou F, Kuo A,
Leder P. Fibroblast growth factor receptor 3 is a negative regulator of bone growth. Cell. 1996;
(84) 911-21.
7. Matsushita T, Murakami S. FGFR3 signaling in endochondral bone formation: A review. Case
Orthop J. 2005; (2) 52-58.
8. Murakami S, Balmes G, McKinney S, Zhang
Z, Givol D, Crombrugghe B. Constitutive activation of MEK1 in chondrocytes causes
Stat1-independent achondroplasia-like dwarfism and rescues the Fgfr3-deficient mouse phenotype. Genes Dev. 2004; (18) 290-305.
9. Chrisman TD, Garbers DL. Reciprocal antagonism coordinates C-type natriuretic peptide and mitogen-signaling pathways in fibroblasts. J. Biol. Chem. 1999; (274) 4293-99.
10. Suganami T, Mukoyama M, Sugawara A, Mori
K, Nagae T, Kasahara M, Yahata K, Makino
H, Fujinaga Y, Ogawa Y, Tanaka I, Nakao K.
Overexpression of brain natriuretic peptide in mice ameliorates immune-mediated renal injury.
J. Am. Soc. Nephrol. 2001; (12) 2652-63.
11. Yasoda A, Komatsu Y, Chusho H, Miyazawa
T, Ozasa A, Miura M, Kurihara T, Rogi
T, Tanaka S, Suda M, Tamura N, Ogawa
Y, Nakao K. Overexpression of CNP in chondrocytes rescues achondroplasia through a MAPK-dependent pathway. Nat Med. 2004;
(10) 80-86.
12. Kawasaki Y, Kugimiya F, Chikuda H,
Kamekura S, Ikeda T, Kawamura N, Saito
T, Shinoda Y, Higashikawa A, Yano F,
Ogasawara T, Ogata N, Hoshi K, Hofmann
F, Woodgett J R, Nakamura K, Chung U,
Kawaguchi H. Phosphorylation of GSK-3 β by cGMP-dependent protein kinase II promotes hypertrophic differentiation of murine chondrocytes. J Clin Invest. 2008; 2506-2515.
13. Matsushita T, Murakami S. Regulatory mechanisms of growth plate closure. Case
Orthop J. 2006; (3) 75-82.
14. Matsushita T, Givol D, Murakami S. FGFR3
Regulates growth plate closure through the
MAPK pathway. Trans Orthop Res Soc. 2006;
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Annegers FJ. Mortality in Achondroplasia. Am
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Leavitt LA, Ver Hoeve J, Hall JG, Partington
MW, Jones KL, Sommer A, Feldman W,
Langer LO, Rimoin DL, Hecht JT, Lebowitz
R. Apnea and Sudden Unexpected Death in
Infants with Achondroplasia. J Pediatr. 1984;
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19. Breur GJ, VanEnkevort BA, Farnum CE,
Wilsman NJ. Linear relationship between the volume of hypertrophic chondrocytes and rate of longitudinal bone growth in growth plates. J.
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(14) 927-936.

MANuSCRIPTS
abStract
P iriformis syndrome is an uncommon condition in which the sciatic nerve is compressed by the piriformis muscle or other closely approximated structures within the gluteal region. Patients experience buttock pain, with or without lower extremity radiculopathy similar to that caused by nerve compression within the lumbar spine. Limited descriptions within the literature have led to controversy and skepticism of the existence or frequency of piriformis syndrome as a true clinical entity.
The diagnosis is made with a high degree of suspicion and with exclusion of the more common etiologies of lumbar disc herniation and spinal stenosis. In appropriately diagnosed patients, surgical management with piriformis resection and sciatic nerve decompression is effective in relieving symptoms and improving function. Currently we present our long-term experience with the surgical management of patients with piriformis syndrome.
materIalS
95 patients over a 9 year period
(1996-2004) with piriformis syndrome underwent surgical treatment in the form of piriformis muscle resection and sciatic nerve decompression. There were 60 women and 35 men. Average age was 52 years (range 18-78 years).
Average follow-up was 4.2 years (range
2.0-8.0 years).
The chief complaint on presentation was isolated buttock pain in 62 patients
(65%) and both buttock pain and radiculopathy in 33 patients (35%).
11 patients (12%) recalled a specific inciting event to their symptoms, most commonly a direct fall on the buttocks or back. 76 patients (80%) described sitting as the most provocative position for the production of symptoms. 11 patients (12%) identified limitation of athletic activity (running, biking) as the most provocative activity. 8 patients
(8%) described no specific provocative activity. The average duration of symptoms prior to presentation was 14 months (range: 6 months – 14 years).
20 patients (21%) had undergone prior lumbar spine procedures
(12 microdiscectomies, 6 lumbar laminectomies, 2 lumbar fusions).
16 of 20 patients having lumbar surgery (80%) reported no lasting improvement following their surgery.
Of these 16 failed lumbar spine procedures, 12 had undergone microdiscectomy and 4 had undergone laminectomy.
Localized sciatic notch tenderness was the most common positive finding on physical exam, present in 76 patients (80%). A positive Lasegue sign was noted in 66 patients (69%).
The Lasegue sign was defined as reproduction of pain or tenderness to palpation of the sciatic notch within the sciatic notch with the knee extended and the hip flexed at
90 degrees. A positive Freiberg’s sign
(pain with passive internal rotation of the hip) was noted in only 20 patients
(21%). Focal neurologic deficits were uncommon; 4 patients had mild weakness in the L5 or S1 distribution and 2 patients had decreased sensation noted to light touch.
Each patient had a trial of some type of conservative management of at least
6 weeks duration, typically involving physical or aquatic therapy, antiinflammatory medications or localized injections.
80 patients (84%) underwent MRI imaging and diagnostic injections performed by a single musculoskeletal radiologist with extensive experience with this clinical entity. An obvious anatomic abnormality was documented by MRI in 57 patients (60%). An abnormally enlarged piriformis muscle was diagnosed in 25 patients, anomalous fibrous bands in 22 patients, and anomalous vascular bundle in 13 patients. No documented abnormality was found on MRI in 38 patients (40%). At the completion of the imaging, localized injection of the piriformis region was performed with a combination of short and long-acting local anesthetic and a corticosteroid.
Relief of pain with the diagnostic injection was used as a diagnostic tool and considered as further confirmation of piriformis syndrome.
Surgery was performed by one of the two authors. The senior author
OrtHOPaeDiC JOUrnaL | Case Western reserve University | vOL.5, nO.1 | 2008 | 53

SuRgICAl MANAgEMENT Of PIRIfORMIS SYNdROME
54
(H.H.B.) performed 89 surgeries and the junior author (C.G.F.) performed 6 surgeries.
SurgIcal technIQue
All procedures were performed under a general anesthetic. Patients were positioned in the lateral decubitus position employing a bean bag. The approach was a limited form of a standard posterior approach to the hip with an incision starting at the tip of the greater trochanter and extended proximally for 6-8 cm. The deep fascia was incised in line with the skin incision and the gluteal fibers bluntly dissected. The sciatic nerve was identified and a vessel loop or penrose drain placed around it for protection and gentle retraction. Special attention was made to identify any anatomic anomalies of either the sciatic nerve or the piriformis muscle. The piriformis was incised at its tendinous insertion to femur and lifted to expose the sciatic nerve. The muscle belly was resected at its base and removed. The entire length of the sciatic nerve was examined for any additional compression by vascular bundles or fibrous bands. If an anomalous bipennate piriformis muscle was noted, both heads of the muscle were resected. The wound was closed in layers with absorbable sutures. Small penrose drains were placed for the first
35 consecutive cases, but subsequently no drains were placed. The majority of patients remained hospitalized for a single overnight stay. Activity as tolerated was allowed for all patients.
reSultS
Surgical Findings
indentation of the sciatic nerve by either the piriformis muscle or an adjacent structure was definitively seen in 85 patients (89%), while 10 patients
(11%) had no obvious abnormality of the nerve upon resection of the piriformis muscle. Visible compression solely by the piriformis muscle was found in 46 patients (48%), solely by anomalous fibrous bands in 14 patients (15%), by both an enlarged muscle and fibrous bands in 10 patients
(11%), solely by abnormal vessels in
10 patients (11%), and by an enlarged muscle and abnormal vessels in 5 patients (5%). In the remaining 10 patients, no obvious nerve indentation, nor other anomalous structure compressing the sciatic nerve was found; in each of these patients, the piriformis muscle was resected in an identical fashion as all other cases.
A split sciatic nerve was noted in 3 patients (3%) while split piriformis muscle was present in 2 patients (2%).
Clinical Results
The visual analogue pain score (VAS) decreased from a pre-operative average of 8.7 to a post-operative average of 2.0 at most recent follow-up. 81 patients
(85%) reported complete relief of pre-operative symptoms, 11 patients
(12%) partial relief and 3 patients
(3%) no relief or worsened symptoms.
85% of patients reported excellent satisfaction with the procedure,
12% good satisfaction, and 3 % no satisfaction. 91% of patients were able to return to all desired activities, 6% partial activities, and 3% were unable to return to any level of activity. 94% of patients were willing to repeat the procedure in retrospect.
Operative findings concurred with preoperative MRI findings in 62 patients
(65%). Compressive fibrous bands were most accurately identified, noted in
91% of the 22 patients with evidence on pre-operative MRI.
No neurological injuries occurred and no patient developed a post-operative neurologic deficit. 3 patients (3%) required further surgery. One patient required urgent re-exploration on post-op day two for severe recurrence of buttock and radicular leg pain; a hematoma was found and evacuated with subsequent resolution of symptoms. 2 patients underwent reexploration at an average of 6 months for ongoing symptoms, with definitive evidence of nerve compression from fibrous bands being noted in one of these patients.
dIScuSSIon
Yeoman was the first to describe a relationship between the piriformis muscle and sciatica, concluding that inflammation within the sacroiliac joint caused a “periarthritis” within the piriformis muscle and sciatic nerve resulting localized buttock and radicular pain (21). In 1937, Freiberg described effective relief of sciatica in
12 patients treated with surgical release of the gluteals and the piriformis (8).
Beaton and Anson identified four distinct anatomical variations in the course of the sciatic nerve about the piriformis muscle and postulated these anomalies as potential causes of sciatic nerve compression and inflammation
(2). In 1947, Robinson was the first to formally describe piriformis syndrome as an entrapment neuropathy of the sciatic nerve (16). He identified six classic features of the condition : (1) history of trauma to the sacroiliac and gluteal region; (2) pain in the sacroiliac joint, greater sciatic notch, and piriformis, extending down the leg and causing difficulty walking; (3) acute exacerbation of pain brought on by stooping or lifting, with moderate relief by traction on the affected extremity with the patient supine; (4) presence of a tender, palpable mass within the piriformis region; (5) a positive Lasegue maneuver; and (6) gluteal atrophy.
He reportedtwo patients treated with operative release of the piriformis who experienced immediate pain relief and no recurrence on long-term follow-up
(16).

MANuSCRIPTS
Over time, descriptions in the literature have been limited to case reports and small series of patients with piriformis syndrome (9,12,17,19-20).
Specific pathologic lesions, including hematoma, myositis ossificans, and polymyositis have been implicated as causes for piriformis syndrome, due to direct mass effect upon the sciatic nerve within the gluteal region
(3-5,11,14-15). Benson reported 14 patients over a 5 year period with posttraumatic piriformis syndrome who were effectively treated with surgical release (4). All patients had history of blunt trauma to the buttock, localized buttock pain, sciatic notch tenderness, and reproduction of pain with hip flexion, adduction and internal rotation. Consistent surgical findings were adhesions between the piriformis muscle, sciatic nerve, and roof of the greater sciatic notch.
More common is the description of an entrapment neuropathy (as initially described by Robinson) in which the sciatic nerve is compressed by the overlying piriformis muscle or an immediately adjacent fibrous band or vascular structures. Adams reported successful surgical treatment of 4 patients, three of whom were noted to have vascular anomalies compressing the sciatic nerve and two with anomalous passage of the sciatic nerve anterior to the muscle (1).
Foster reported effective pain relief in
7 patients treated with surgical release of a taught piriformis muscles and surrounding fibrous bands (7).
Most reports emphasize that piriformis syndrome is a clinical entity that is appropriately diagnosed based on a high index of suspicion from a patient’s history and physical exam (13,17-18).
The hallmark symptom is localized buttock pain that is aggravated by sitting. In particular, prolonged sitting or driving frequently identified as the patient’s chief complaint. In younger, athletic individuals, biking, rowing, or running may cause exacerbation of symptoms. Radicular symptoms of pain or parathesias will frequently accompany the buttock pain (35% of patients in our study).
Pertinent findings on physical exam may assist in arriving at the diagnosis of piriformis syndrome.
Localized tenderness with the sciatic notch is generally present in most patients. A positive straight leg raise is an inconsistent and non-specific finding. Provocative maneuvers are accomplished with active contraction of the muscle against resistance or with passive stretch of the muscle. The
Lasegue maneuver is reproduction of pain or tenderness to palpation of the sciatic notch with the hip flexed at 90 degrees and the knee extended.
The Pace test assesses pain, weakness, or loss of function with resistance to abduction with the flexed hip in the sitting position.
There is limited evidence for the use of imaging modalities in diagnosing suspected piriformis syndrome.
Jankiewicz used CT and MRI scans to define an enlarged piriformis muscle in a patient with classic symptoms and subsequent injection o local anesthetic and corticosteroid brought complete pain relief (10). Beauchesne reported positive finding of uptake on bone scan within the hemipelvis of an individual with piriformis syndrome due to myositis ossificans following remote trauma (3). However, in Benson’s study of post-traumatic cases, ten of
14 patients had a bone scan, none of which provided a positive finding
(4). There is also limited evidence to support the use of neurodiagnostics to accurately localize the piriformis region as a source for nerve compression (4,6).
In Benson’s study, 6 of 8 patients had abnormal EMG findings indicative of extrapelvic compression of the sciatic nerve (4).
In our current series, which is believed to be the largest to date, effective and predictable results were achieved in patients with piriformis syndrome with surgical release of the piriformis release and resection of associated compressive structures, most commonly fibrous bands or vascular bundles compressing he sciatic nerve. The chief complaint of each patient included localized buttock pain and was associated with radicular symptoms in 35%. Aggravation with prolonged sitting and driving were very common associated presenting complaints. A history of localized trauma was occasionally noted, but not present in a majority. History of unsuccessful lumbar spine surgery was a common occurrence in our series, with
21% of all patients giving a history of a prior lumbar surgery. The majority with prior lumbar surgery had incomplete relief of pain, including all 12 patients who had undergone a microdiscectomy.
This finding underscores the importance of considering all causes of sciatica in all patients who present to a spine care provider. While the vast majority of patients with sciatica will have pathology in the lumbar spine (disc herniation, spinal stenosis), special consideration should be given to the patient who presents with radicular pain but without associated low back pain and those whose clinical symptoms do not correlate with lumbar spine neuroimaging. The diagnosis was made on a clinical basis and supported by MRI images and a diagnostic injection performed by an experienced musculoskeletal radiologist.
Localized sciatic notch tenderness and reproduction of pain with the ipsilateral hip flexed and knee extended
(a positive Lasegue sign) was the most common finding on physical exam in our series. A positive straight leg raise
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56
SuRgICAl MANAgEMENT Of PIRIfORMIS SYNdROME however was less commonly noted and focal neurologic deficits were very uncommon, found in less than 5% of our patients.
MRI scans were obtained on the majority of patients in our series and while helpful in confirming the diagnosis in some patients, it was not a critical modality to establish the diagnosis. Identifiable abnormalities were found on pre-operative MRI in 60% of our patients, though in only 62% of these patients was the
MRI finding corroborated by surgical findings. These results underscore the fact that piriformis syndrome is a clinical diagnosis and that an accurate history and physical exam with a high index of suspicion are the gold standards to achieving the correct diagnosis. concluSIon
Piriformis syndrome is an uncommon entity when compared to other causes of lower extremity radiculopathy.
However, with a high index of suspicion in patients with buttock and leg pain and absence of lumbar spine pathology, piriformis syndrome can be accurately diagnosed and successfully managed surgically in selected cases.
The majority of patients in our series had predictable pain relief and reported satisfaction with the surgical procedure and ability to return to their desired level of function. Complication rates were low and re-operation rate uncommon (3%).
REfERENCES
1. Adams JA. The pyriformis syndrome-Report of four cases and review of the literature. S African
J. Surg.
1980;18:13-18.
2. Beaton LE and Anson BJ.
The sciatic nerve and the piriformis muscle: their interrelation as a possible cause of coccygodynia. J. Bone Joint
Surg.
1938;20:686-688.
3. Beauchesne RP and Schutzer SF. Myositis ossificans of the piriformis muscle. An unusual cause of piriformis syndrome. A case report. J.
Bone Joint Surg.
1997;79-A:906-910.
4. Benson ER and Schutzer SF.
Posttraumatic piriformis syndrome: diagnosis and results of operative treatment. J. Bone Joint Surg.
1999;74-A:941-949.
5. Chen W-S.
Sciatica due to piriformis pyomyositis: Report of a case. J. Bone Joint
Surg. 1992;74-A;10:1546-1548.
6. Fishman LM and Zybert PA.
Electrophysiologic evidence of piriformis syndrome. Arch. Phys. Med. And Rehab.
1992;73:359-364.
7. Foster MR.
Piriformis syndrome. Orthopedics
2002;25:821-825.
8. Freiberg AH.
Sciatica pain and its relief by operations on the muscle and fascia. Arch Surg.
1937;34:337-350.
9. Hughes SS, Goldstein MN, Hicks DG and
Pellegrini VD.
Extrapelvic compression of the sciatic nerve. An unusual cause of pain about the hip: report of five cases. J. Bone Joint Surg.
1992;74-A:1553-1559.
10. Jankiewicz JJ, Hennrikus WL, Houkom
JA. The appearance of the piriformis muscle syndrome in computed tomography and magnetic resonance imaging: A case report and review of the literature. Clin. Ortho. Rel. Res.
1991;262:205-209.
11. Ku A, Kern H, Lachman E, Nagler W. Sciatic nerve impingement from piriformis hematoma due to prolonged labor. Muscle Nerve .
1995;18:789-790.
12. Mizuguchi T.
Division of the piriformis muscle for the treatment of sciatica. Postlaminectomy syndrome and osteoarthritis of the spine. Arch.
Surg.
1976;111:719-722.
13. Pace JB and Nagle D.
Piriformis syndrome.
West. J. Med.
1976;124:435.
14.
Papadopoulos SM, McGillicuddy JE, Albers
JW. Unusual cause of ‘piriformis muscle syndrome’. Arch. Neurol.
1990;47:1144-1146.
15.
Peh WC and Reinus WR.
Piriformis bursitis causing sciatic neuropathy. Skeletal. Radiol.
1995;24:474-476.
16. Robinson DR. Piriformis syndrome in relation to sciatic pain. Am J. Surg.
1947;73:355-358.
17. Solheim L, Siewers P and Paus B.
The piriformis muscle syndrome. Sciatic nerve entrapment treated with section of the piriformis muscle. Acta Orthop. Scandinavica
1981;52:73-75.
18.
Steiner C, Staubs C, Ganon M, Buhlinger C.
Piriformis syndrome: pathogenesis, diagnosis, and treatment. J. Am. Osteopath. Assoc.
1987;87:318-324.
19.
Synek VM. The pyriformis syndrome: review and case presentation. Clin.and Exper. Neurol.
1987;23:31-37.
20. Vandertop WP and Bosma WJ. The piriformis syndrome. A case report. J. Bone Joint Surg.
1991;73-A:1095-1097.
21. Yeoman W. The relation of arthritis of the sacro-iliac joint to sciatica, with analysis of 100 cases. Lancet 1928;ii:1119-1122.

MANuSCRIPTS
P osterior cruciate ligament (PCL) injuries occur much less frequently than injuries to the anterior cruciate ligament (ACL). As such, the medical literature and experience with management of PCL injuries have lagged far behind that for the ACL.
Traditionally PCL injuries have been treated nonoperatively. Our current understanding of the long-term sequelae of chronic PCL injuries has led to a lower threshold for surgical management.
On the tibial side, both tibial inlay and endoscopic transtibial techniques are commonly utilized for reconstruction of the PCL. Multiple options also exist for preparation of the PCL femoral tunnel. In the inside-out technique, the femoral tunnel is drilled endoscopically through the lateral arthroscopic portal.
The inside-out technique puts the articular cartilage of the lateral femoral condyle at risk during tunnel drilling.
The inside-out method also creates a sharply angled femoral tunnel that theoretically may put the graft at risk.
This “critical corner” is synonymous to the so-called “killer turn” created at the tibial tunnel aperture during the endoscopic transtibial technique
(Figure 1). The killer turn has been shown to cause graft thinning and subsequent graft failure in cadaveric specimens. Considering these issues with the inside-out technique, some surgeons prefer the outside-in technique for femoral tunnel drilling.
This technique typically requires a medial incision overlying the distal femur with subsequent splitting of the vastus medialis fibers or elevation of the vastus medialis from the distal femur.
While the outside-in technique allows for more favorable positioning of the femoral tunnel (figure 1), the medial incision and insult to the quadriceps mechanism may result in increased postoperative pain and quadriceps inhibition. Quadriceps inhibition is a concern as the quadriceps mechanism is an important dynamic stabilizer to posterior tibial translation. Less invasive techniques may avoid quadriceps inhibition and protect the new graft by maintaining the dynamic stabilization effects of the quadriceps mechanism.
Here we describe a new method for minimally invasive drilling of the PCL femoral tunnel that avoids major insult to the quadriceps mechanism.
We utilize the Arthrex RetroDrill system to drill the femoral tunnel from inside-out (Arthrex Inc., Naples,
Florida). After diagnostic arthroscopy, the graft is sized and the PCL femoral footprint is identified. For singlebundle reconstruction the anatomic position for the anterolateral (AL) bundle is identified. The medial intercondylar ridge, as previously described by us 2 , can be used to identify the AL bundle footprint. As the medial intercondylar ridge represents the posterior border of the PCL femoral attachment, the AL bundle footprint lies just anterior to this ridge at the 1 o’clock position in the right knee (11 o’clock in left knees).
Once the AL bundle footprint is identified the appropriately sized
RetroCutter, correlating to the size of the prepared graft, is passed through the medial portal and positioned in the center of its footprint (Figure 2).
The RetroDrill guide sleeve is then positioned over the medial aspect of the distal femur (Figure 3). A small poke hole is then made over the medial aspect of the knee and the RetroDrill guide sleeve is advanced to the outer metaphyseal cortex (Figure 3). At this time, the length of the femoral osseous tunnel can then be estimated directly from the calibrated drill sleeve. The
3mm guide pin is then passed from outside-in to capture the waiting
RetroCutter. The RetroCutter is reverse threaded. As the pin is advanced on forward it captures the RetroCutter which is then spun free from the
RetroCutter holder. While on forward, a blind-ended femoral tunnel is drilled to the appropriate length, at least 25 –
30 mm. At this time the RetroCutter is advanced back into joint and recaptured by the holder (Figure 4).
The pin is then placed in reverse and disengaged from RetroCutter.
A wire loop or Hewson suture passer may be passed from outsidein to facilitate graft passage into the femoral tunnel. The graft is fixed with a standard interference screw passed through the lateral portal or a
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POSTERIOR CRuCIATE lIgAMENT fEMORAl TuNNEl
RetroScrew (Arthrex). If a RetroScrew is utilized, the femoral tunnel must be completed through the outer cortex to facilitate passage of the screw driver.
The described technique provides a minimally invasive approach for preparation of the PCL femoral tunnel.
The minimally invasive approach may result in less pain and avoid inhibition of the quadriceps mechanism in the early postoperative period.
REfERENCE:
Farrow LD, Chen MC, Cooperman DC,
Victoroff BN, Goodfellow DB. Morphology of the femoral intercondylar notch. J Bone Joint
Surg Am . 2007;89:2150-2155.
Figure 1: radiograph demonstrating difference in femoral tunnel angles with outside-in and inside-out technique (Courtesy John A.
Bergfeld, MD).
Figure 3: retroDrill guide in position.
58
Figure 2: retroCutter positioned in aL bundle footprint.
Figure 4: retroDrill being recaptured on holder.

MANuSCRIPTS
1
2
3
1
2
3
purpoSe
Develop an animal model of denervation in the setting of spinal cord injury. Evaluate the efficacy of paralyzed nerve transfer for axonal regeneration within this model. We hypothesized that paralyzed nerve transfer would allow for a mean of 50% axonal regeneration which would be less than primary neurorrhaphy. methodS
Fischer 344 rats underwent lower thoracic spinal cord injury (SCI) followed by unilateral tibial nerve transsection and delayed peroneal
(paralyzed) to tibial nerve transfer
(Group A) or primary neurorrhaphy
(Group B). Control groups underwent
SCI and a unilateral hindlimb approach only (Group C) or a unilateral hindlimb approach and transsection of both the tibial and peroneal nerves
(Group D). Three months following surgery, tibial and peroneal nerves were harvest from the bilateral hindlimbs.
Transverse sections were stained with anti-Neurofilament protein (160 kD) to determine total axon counts. Operative side axonal regeneration was expressed as a percent ratio of total axon count on the operative side over total axon count on the non-operative side. reSultS
Mean axonal regeneration was 51%
(46, 56) for Group A (surviving N=9),
73% (71, 75) for Group B (surviving
N=4), 99% (97, 100) for Group C
(surviving N=4), and 2% (1, 2) for
Group D (surviving N=4). One-way
ANOVA for independent samples and the Tukey HSD test revealed significant differences between all groups (p <
.01). concluSIonS
Paralyzed nerve transfer allows for a mean of 50% axonal regeneration which is less than primary neurorrhaphy. In future research, this model will provide a foundation for defining axonal regeneration physiology via retrograde neuronal tracing. In addition, the model will be used to test novel stem cell therapies for motor axon regeneration in the peripheral nervous system.
Introduction
Denervation results from damage to lower motor neuron (LMN) cell bodies in the spinal cord and/or discontinuity between LMN cell bodies and a given target muscle.
(1) Damage to LMN cell bodies occurs during spinal cord injury (SCI) at the level of the lesion.
Discontinuity between LMN cell bodies and a given target muscle can be caused by damage to nerve roots (root avulsion), damage to peripheral nerves
(crush injuries, penetrating trauma), or damage to the neuromuscular junction and motor endplate zone (avulsion of nerve from muscle).
Damage to LMN cell bodies is unique from discontinuity between LMN cell bodies and target muscle because it is typically associated with SCI.
In this case, the denervated muscle is surrounded by paralyzed nerves and muscles which severely limits reconstructive options for re-animation.
Furthermore, previous research addressing denervation has primarily focused on isolated denervation
(i.e.without concomitant paralysis) where donor nerves and muscles are readily available. Experimental and clinical models have included primary repair (2) , embryonic spinal cord grafting (3,4,5,6,7), peripheral nerve grafting (2,3), nerve transfer
(8,9) , direct muscle neurotization
(10,11,12,13,14,15,16,17,18) , motor nerve transplantation (19) , and even muscle-nerve-muscle neurotization (20) .
However, all of these techniques rely on the presence of nearby voluntary nerves with LMN cell bodies that are still in continuity with the motor cortex. We hypothesized that LMN cell body continuity with the motor cortex via intact upper motor neurons is not necessary for the regeneration of axons into denervated muscle.
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PARAlYzEd NERVE TRANSfER fOR dENERVATION
60
Instead, the only requirement for successful regeneration is an intact
LMN cell body in continuity with the target muscle. In this case, involuntary
(i.e.“paralyzed”) nerves with intact
LMN cell bodies can be transferred onto degenerated nerves to allow for re-innervation in the setting of concomitant SCI and denervation.
Materials and Methods
This protocol was approved by the
Institutional Animal Care and Use
Committee and all animal care complied with the guidelines set forth by the National Institutes of Health.
Based upon pilot study data, a mean axonal regeneration rate of 50% was assumed for group A rats, 75% for
Group B rats, 100% for Group C rats, and 0% for Group D rats. Based upon these estimates, F-test calculations projected a minimum sample size of four animals per group needed for
ANOVA comparison between mean percent axonal regeneration among the groups (alpha < 0.05 and power >
0.8). However, a 70 to 80% morbidity and mortality rate was also estimated from pilot study data necessitating a minimum of 20 animals per group to ensure adequate power.
Spinal Cord Injury
A total of one hundred Fischer 344 rats were utilized for this study. All rats underwent a lower thoracic dorsal spinal approach followed by a T10 to
T11 laminectomy. A modified forceps was used to compress the cord for a minimum of 40 continuous seconds.
After this initial injury, meticulous hemostasis was obtained, the cord was re-visualized, and compressed a second time for a minimum of 40 continuous seconds. Double compression was used to ensure a complete spinal cord injury as previous single compression experiments yielded incomplete injury patterns.
Hindlimb Surgery
Following SCI, rats underwent a unilateral hindlimb approach exposing the distal sciatic, tibial, and peroneal nerves. During this approach, all soft tissues and vessels were handled with extreme care to avoid any devascularization of the hindlimb.
Group A rats underwent tibial nerve transsection 4 to 5 millimeters proximal to the neuromuscular junction and then were allowed to recover from spinal shock for a minimum of fourteen days. During the third post-operative week, the hindlimb incision was re-opened and a peroneal
(paralyzed but innervated) to distal tibial (paralyzed and denervated) nerve transfer was performed using 10-0 polypropylene suture.
Group B rats underwent tibial nerve transsection at the same level as
Group A rats followed by primary neurorrhaphy. Group C rats underwent the same hindlimb approach with only visualization of the distal sciatic, tibial, and peroneal nerves. Group D rats underwent the hindlimb approach followed by transsection of both the tibial and peroneal nerves without primary or delayed repair.
Terminal Surgery
At the conclusion of a three month recovery period, surviving rats underwent a terminal operative procedure consisting of premortem muscle force testing of the gastrocnemius muscles bilaterally, spinal cord lesion testing, and peripheral nerve lesion pathway testing.
These biomechanical testing results are published in a separate report. At the completion of biomechanical testing, animals were euthanized and the tibial and peroneal nerves were harvested from the bilateral hindlimbs for histological analysis.
For groups A and B rats, one centimeter nerve sections were harvested immediately proximal and immediately distal to the neurorrhaphy site. For group C rats, one centimeter sections of tibial nerve and peroneal nerve were harvested 4-5 mm proximal to the neuromuscular junction. For group D rats, one centimeter sections of the distal tibial nerve stump and the distal peroneal nerve stump were harvested immediately proximal to the neuromuscular junction.
Tissue Preparation and Analysis
After harvest, all specimens were fixed in 10% formalin for 4 hours and then transferred to 0.01 molar phosphate buffered saline. All matched specimens from each animal (i.e. experimental side tibial nerve section adjacent to control side tibial nerve section and experimental side peroneal nerve section adjacent to control side peroneal nerve section, etc.) were then embedded together in a single paraffin block. Embedding was performed in this manner to minimize sample-tosample and slide-to-slide variability.
Paraffin blocks were serially sectioned at 10 micrometers of thickness.
Thirty sections were cut from each block with every third section being stained with hematoxylin and eosin.
The remaining twenty sections were prepared for immunohistochemical staining with anti-Neurofilament protein. These sections were air dried for 60 minutes and fixed in Acetone for 10 minutes. Slides were washed in 50 millimolar Tris Buffered Saline
(TBS, pH 7.6) for 5 minutes, treated with a blocking reagent for 10 minutes, and then anti-Neurofilament antibody diluted in blocking reagent for 60 minutes. Sections were then incubated at 4 degrees Celsius overnight. The following day, sections were treated with biotinylated secondary antibody diluted in blocking reagent for 30 minutes at 25 degrees Celsius. Next,

MANuSCRIPTS sections were treated with ABC complex-HRP antibody for 30 minutes at 25 degrees Celsius. Slides were developed in 3,3’ diaminobenzidene tetrahydrochloride (DAB) for 5 minutes at 25 degrees Celsius. Finally, the sections were dehydrated, cleared, and mounted in DPX mountant. (21)
An inverted light micrsocrope (40X magnification) with a digital camera was then used to take multiple photographs covering the entire transverse section of a given nerve.
Each digital image was then stitched back together to create a single merged image for a given transverse section.
Color-sensitive Meta Morph software
(MDS Analytical Technologies,
Toronto, Canada) was then used to identify and count all axons stained with the anti-Neurofilament antibody which appeared dark brown (see
Figure 1 ). In addition, each image was individually inspected and immunhistochemical artifacts were excluded from the total axon count values.
Statistical Analysis
Percent axonal regeneration was calculated by dividing the total axon count from the experimental side tibial nerve by the total axon count from the control side tibial nerve.
Mean percent axonal regeneration and
95% confidence intervals were then calculated for each group of animals.
The mean value from each group was then compared with the mean value of all other groups utilizing a one-way
ANOVA.
Results
Mean axonal regeneration was
51% (46, 56) for Group A animals
(surviving N=9), 73% (71, 75) for
Group B animals (surviving N=4),
99% (97, 100) for Group C animals
(surviving N=4), and 2% (1, 2) for
Group D animals (surviving N=4).
Raw total axon counts and mean operative side axonal regeneration for animals in all groups can be found in
Table 1.
One-way ANOVA for independent samples and the Tukey HSD test revealed significant differences between all groups (p < .01). In addition, axonal regeneration closely paralleled functional recovery of gastrocnemius force as measured by biomechanical testing. Biomechanical testing results are published in a separate report.
Discussion
Denervation poses a significant clinical dilemma with only limited techniques available for successful re-animation.
Furthermore, all current techniques rely upon the restoration of continuity a b
Figure 1. Stained Experimental and Control Side Tibial Nerves.
Distal tibial nerve sections were stained with antineurofilament protein which appears dark brown under light microscopy. a) experimental side tibial nerve section from animal 1a2. B)
Control side tibial nerve section from animal 1a2.
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PARAlYzEd NERVE TRANSfER fOR dENERVATION table 1. total axon count data
Id Experimental side count
3D
4D
1D
2D
2C
3C
4C
1C
2a5
1B3
1B6
1B10
1B13
3a2
10a1
3a5
1a6
19a
18a
2a4
9a3
1768
2031
1460
3661
1865
1950
2137
2397
2985
3589
3229
3386
3584
4437
5190
3999
5989
103
67
87
58
Control side count % Regeneration
Mean
95% Ci
Mean
95% Ci
Mean
95% Ci
Mean
95% Ci
4279
5051
3235
7615
3641
3603
3671
4040
4750
5154
4416
4562
4769
4506
5127
4108
6057
5234
4067
5604
5876
51
54
58
59
41
40
45
48
63
51
(46, 56)
98
101
97
99
99
(97, 100)
2
1
2
2
2
(1, 2)
70
73
74
75
73
(71, 75) table 1.
Total Axon Count Data. total axon counts from experimental and control side tibial nerve sections are listed for Groups a, B, C, and D. Percent regeneration was calculated for each animal by dividing the experimental side axon count over the control side axon count. Mean percent regeneration and the 95% confidence interval was calculated for each group. One-way anOva for mean percent regeneration revealed statistically significant differences between all groups (p < 0.01).
62 between intact and “voluntary” LMN cell bodies (i.e. LMN cell bodies still in continuity with the cortex) and a given target muscle. This principle has been applied from the spinal cord all the way down to the muscle in a variety of different experimental models.
At the cord level, research has shown that transplantation of fetal spinal cord cells allows for peripheral nerve-like structure formation. Sieradzan et al. implanted embryonic spinal cord cells into adult rat spinal cords and found that these motor neurons eventually formed axons which extended into the adjacent musculature.
(5) Similarly,
Nogradi et al. found that concomitant fetal spinal cord transplantation and nerve root avulsion allowed for targeted axonal regeneration into specific muscles. In this experiment, L4 ventral roots were avulsed and embryonic motor neurons were simultaneously grafted into the adjacent spinal cord.
Following a recovery period, animals demonstrated targeted re-innervation of the extensor digitorum longus and tibialis anterior.
(6)
Duchossoy et al. performed fetal spinal cord transplantation in conjunction with peripheral nerve grafting. Fetal spinal cord cells (embryonic day 14) were implanted into cervical spinal cord lesions in rats and a peripheral nerve was grafted from the implanted spinal cord cells to the distal end of a transsected musculocutaneous nerve still in continuity with the biceps brachii. In a control group, the distal end of the peripheral nerve graft was not connected to the musculocutaneous nerve stump. Transplanted embryonic spinal cord cells and native spinal cord cells extended from the spinal cord through the peripheral graft and into the biceps brachii in the experimental group only.
(2,3) These results suggest that the target muscle most likely has a trophic effect on nerve regeneration.
At the level of the peripheral nerve, simple primary repair of transected nerves allows for regeneration. Gamo et al. transected the hypoglossal nerves in mice and then either left the nerve transsected or performed primary epineural repair. Primary repair allowed for regeneration while unrepaired nerves did not regenerate.
(7) Alternatively, spinal cord cell transplantation at the site of peripheral nerve injury may also allow for

MANuSCRIPTS regeneration. Erb et al. transplanted dissociated ventral spinal cord cells
(embryonic days 14 and 15) into the distal stump of axotomized tibial nerves in adult rats. These cells survived for up to 18 weeks and developed into large multi-polar cells resembling alpha motoneurons with axons entering the tibial nerve stump and the gastrocnemius with the formation of neuromuscular junctions. (4)
At the level of the neuromuscular junction, re-innervation can be achieved by transplantation of a donor motor end-plate zone or by regeneration of the native motor end-plate zone. In motor nerve transplantation (MNT), a motor nerve and a portion of its native motor end-plate are transplanted into the denervated muscle. Gray et al. performed MNT in New Zealand white rabbits and observed successful motor axon regeneration. (19)
In direct muscle neurotization (DMN) peripheral nerve fascicles are directly inserted into denervated muscle without donor motor end-plate zone transplantation. Therefore, the success of DMN relies upon regeneration of the native motor end-plate zone which increases the time needed for reinnervation. (16) During this process of native motor end-plate regeneration, the acetylcholine receptor inducing activity (ARIA) glycoprotein most likely plays a crucial role. In embryonic development, ARIA glycoproteins are released from motor neurons to stimulate the synthesis of acetylcholine receptors on post-synaptic muscle fibers. Furthermore, animal models suggest that specific ARIA isoforms are needed for re-innervation after nerve injury. Ng et al. identified three active isoforms of the ARIA glycoprotein in chick muscle which were variably expressed during development. ARIA beta 1 was the major form expressed in adults and all three active isoforms
(ARIA beta 1, ARIA alpha 2 and ARIA beta 2) were expressed in embryonic and young chick muscle. However,
ARIA alpha 2 and ARIA beta 2 were temporarily expressed in adult chick muscles after nerve injury. (22) of specific ARIA glycoprotein active isoforms is most likely crucial for reinnervation.
A need for continuity between motor nerve and target muscle was clearly illustrated in a sciatic nerve injury model in rats. In this experiment, sciatic nerves were injured in one of three ways: transsection with proximal stump ligation, crush, or transsection followed by implantation into the gastrocnemius muscle (i.e. DMN).
Subsequently, the injured sciatic nerves were harvested for histological analysis. Rats that underwent transsection followed by sciatic nerve implantation into muscle (i.e. DMN) demonstrated a significantly higher percentage of motor nerve survival.
(18) In accordance with these findings,
Payne et al also demonstrated both native and ectopic motor end-plate zone regeneration with DMN. In this case, the tibial nerve was used for DMN of the denervated soleus muscle. Furthermore, tibial nerves were implanted in one of two different locations with respect to the native motor endplate zone of the soleus. In the “near” group, the tibial nerve was implanted close to the original motor endplate zone and, in the “far” group, the nerve was implanted far from the original motor endplate zone. Animals in the near group produced a greater number of motor endplates at the native zone than animals in the far group. However, animals in the far group produced a greater number of motor endplates total. (17)
Therefore close motor nerve continuity with target muscle and the expression
Muscle-nerve-muscle (MNM) neurotization also restores continuity between LMN cell bodies and target muscle but with one additional link in the chain. In MNM neurotization, a peripheral nerve graft is used to connect an innervated donor muscle to a denervated recipient muscle.
Urbanchek et al. performed MNM neurotization between an innervated extensor digitorum longus and a denervated peroneus digiti quinti.
This technique allowed for 62% re-innervation of the peroneus digiti quinti with no detectable force deficit of the extensor digitorum longus. (20)
In addition to the chosen technique, the timing of reconstruction has a significant effect on outcome. In almost all cases, earlier reconstructions provide better functional outcomes.
Keilhoff et al. performed DMN for the gracilis with the sciatic nerve either immediately after denervation or 2, 4,
6, or 8 weeks after denervation. The best functional recovery was achieved with immediate or early (< 2 weeks after denervation) reconstruction.
Rats that underwent DMN at 4 or 6 weeks after denervation demonstrated moderate regeneration and rats that underwent DMN at 8 weeks after denervation demonstrated negligible regeneration.
(23) Furthrermore, temporary electrical stimulation of denervated muscles does not prolong the ideal window for reconstruction.
Dow et al. applied electrical stimulation to rat muscle immediately after denervation. This stimulation was associated with higher muscle mass and greater force production. However, it did not allow for faster recovery of function after reconstruction. (24)
In terms of functional recovery, only partial recovery can be achieved for re-innervation of denervated muscles even under ideal circumstances.
Yoshimura et al. transected the peroneal
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PARAlYzEd NERVE TRANSfER fOR dENERVATION
64 nerves of rats and then performed immediate repair. Rats recovered for four months and then underwent force testing of the extensor digitorum longus. The extensor digitorum longus demonstrated decreased force production and power output in comparison to the control hindlimb.
(25)
In summary, all of the aforementioned reconstructive techniques attempt to restore continuity between intact and “voluntary” LMN cell bodies (i.e.
LMN cell bodies still in continuity with the cortex) and denervated muscle. However, in the setting of concomitant SCI and denervation, sole reliance on “voluntary” LMN cell bodies severely limits the available reconstructive options. In an attempt to overcome this limitation, we developed paralyzed nerve transfer for re-innervation in the setting of spinal cord injury. In this technique, a paralyzed but non-degenerated nerve is transferred onto a degenerated nerve.
As such, paralyzed nerve transfer only requires intact and “involuntary” LMN cell bodies (i.e. LMN cell bodies no longer in continuity with the cortex) for re-innervation.
This technique has previously been applied in one clinical report of diaphragm rei-nnervation. Six patients with C3-C5 tetraplegia (and confirmed diaphragm denervation) underwent a T4 intercostal (paralyzed) transfer onto the degenerated phrenic nerve.
A diaphragmatic pacemaker lead was then placed one centimeter distal to the neurorrhaphy site and stimulation was applied on a monthly basis until diaphragmatic contractions could be detected. At the conclusion of the study, all patients were capable of using diaphragmatic pacing alone (in place of mechanical ventilation) to support ventilatory function. (26)
We have achieved similar results with paralyzed peroneal to degenerated tibial nerve transfer in this animal model.
After a minimum of three months, rats demonstrated a mean of 51% axonal regeneration which closely paralleled functional return of gastrocnemius strength. This data strongly suggests that “voluntary” LMN cell bodies are not necessary for axonal regeneration.
As such, paralyzed nerve transfer holds great promise as a new reconstructive option for extremity re-innervation in the setting of spinal cord injury when combined with functional electrical stimulation. In addition, paralyzed nerve transfer may also be performed as a stand-alone procedure to prevent or reverse contractures associated with denervation. (9,27)
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2001 Jun 1;64(5):476-86. PMID: 11391702
4. Duchossoy Y, Kassar-Duchossoy L, Orsal
D, Stettler O, Horvat JC. Reinnervation of the biceps brachii muscle following cotransplantation of fetal spinal cord and autologous peripheral nerve into the injured cervical spinal cord of the adult rat. E xp Neurol.
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5. Erb DE, Mora RJ, Bunge RP.
Reinnervation of adult rat gastrocnemius muscle by embryonic motoneurons transplanted into the axotomized tibial nerve. Exp Neurol. 1993 Dec;124(2):372-
6. PMID: 8287933
6. Sieradzan K, Vrbová G. Replacement of missing motoneurons by embryonic grafts in the rat spinal cord. Neuroscience.
1989;31(1):115-30. PMID: 2771053
7. Nógrádi A, Vrbová G. Improved motor function of denervated rat hindlimb muscles induced by embryonic spinal cord grafts. Eur
J Neurosci. 1996 Oct;8(10):2198-203. PMID:
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8. Battiston B, Lanzetta M.
Reconstruction of high ulnar nerve lesions by distal double median to ulnar nerve transfer. J Hand Surg
[Am]. 1999 Nov;24(6):1185-91. PMID:
10584939
9. Ustün ME, Oğün TC, Büyükmumcu M,
Salbacak A. Selective restoration of motor function in the ulnar nerve by transfer of the anterior interosseous nerve. An anatomical feasibility study. J Bone Joint Surg Am. 2001
Apr;83-A(4):549-52. PMID: 11315783
10. Brunelli G.
Direct neurotization of severely damaged muscles. J Hand Surg [Am].
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11. Brunelli G, Monini L. Direct muscular neurotization. J Hand Surg [Am]. 1985
Nov;10(6 Pt 2):993-7. PMID: 4078294
12. Brunelli G, Brunelli LM.
Direct neurotization of severely damaged denervated muscles. Int
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13. Brunelli GA, Brunelli GR. Direct muscle neurotization. J Reconstr Microsurg. 1993
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14. Papalia I, Lacroix C, Brunelli F, d’Alcontres
FS. Direct muscle neurotization after end-toside neurorrhaphy. J Reconstr Microsurg.
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15. Becker M, Lassner F, Fansa H, Mawrin
C, Pallua N. Refinements in nerve to muscle neurotization. Muscle Nerve. 2002
Sep;26(3):362-6. PMID: 12210365
16. Bielecki M, Skowroński R, Skowroński
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A comparative morphological study of direct nerve implantation and neuromuscular pedicle methods in cross reinnervation of the rat skeletal muscle. Rocz Akad Med Bialymst.
2004;49:10-7. PMID: 15631308
17. Payne SH Jr, Brushart TM. Neurotization of the rat soleus muscle: a quantitative analysis of reinnervation. J Hand Surg [Am]. 1997
Jul;22(4):640-3. PMID: 9260619
18. Zhao D, Wang W, Kang K, Jing T, Wang
T, Yu X, Yang L, Cui X. Protection effect of nerve implantation after peripheral nerve injury to rats. Zhonghua Wai Ke Za Zhi.
2002
Nov;40(11):862-4. PMID: 12487867
19. Gray WP, Keohane C, Kirwan WO. Motor nerve transplantation. J Neurosurg. 1997
Oct;87(4):615-24. PMID: 9322851
20. Urbanchek MG, Ganz DE, Aydin MA, van

MANuSCRIPTS der Meulen JH, Kuzon WM Jr. Muscle-nervemuscle neurotization for the reinnervation of denervated somatic muscle. Neurol Res. 2004
Jun;26(4):388-94. PMID: 15198864
21. Kawasaki Y, Yoshimura K, Harii K, Park S.
Identification of myelinated motor and sensory axons in a regenerating mixed nerve.
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1997;9(2):132-43. PMID: 9245497
23. Keilhoff G, Fansa H. Successful intramuscular neurotization is dependent on the denervation period. A histomorphological study of the gracilis muscle in rats. Muscle Nerve.
2005
Feb;31(2):221-8. PMID: 15736301
24. Dow DE, Carlson BM, Hassett CA, Dennis
RG, Faulkner JA.
Electrical stimulation of denervated muscles of rats maintains mass and force, but not recovery following grafting.
Restor Neurol Neurosci. 2006;24(1):41-54.
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25. Yoshimura K, Asato H, Cederna PS,
Urbanchek MG, Kuzon WM.
The effect of reinnervation on force production and power output in skeletal muscle. J Surg Res. 1999
Feb;81(2):201-8. PMID: 9927541
26. Krieger LM, Krieger AJ. The intercostal to phrenic nerve transfer: an effective means of reanimating the diaphragm in patients with high cervical spine injury. Plast Reconstr Surg.
2000 Apr;105(4):1255-61. PMID: 10744213
27. Bryden AM, Kilgore KL, Lind BB, Yu DT.
Triceps denervation as a predictor of elbow flexion contractures in C5 and C6 tetraplegia.
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5. PMID: 15520985
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1
1
2
1
2
reprinted with kind permission of springer science and Business Media.
Original Citation: Grant re, Murphy La, Murphy Je. expansion of the coordinator role in orthopaedic residency program management.
Clin Orthop Relat Res.
2008. 466(3):737-42.
66 abStract
The Accreditation Council of Graduate
Medical Education’s (ACGME) Data
Accreditation System indicates 124 of 152 orthopaedic surgery residency program directors have 5 or fewer years of tenure. The qualifications and responsibilities of the position based on the requirements of orthopaedic surgery residency programs, the institutions that support them, and the ACGME Outcome Project have evolved the role of the program coordinator from clerical to managerial.
To fill the void of information on the coordinators’ expanding roles and responsibilities, the 2006 Association of Residency Coordinators in
Orthopaedic Surgery (ARCOS) Career survey was designed and distributed to 152 program coordinators in the United States. We had a 39.5% response rate for the survey, which indicated a high level of day-to-day managerial oversight of all aspects of the residency program; additional responsibilities for other department or division functions for fellows, rotating medical students, continuing medical education of the faculty; and miscellaneous business functions.
Although there has been expansion of the role of the program coordinator, challenges exist in job congruence and position reclassification. We believe use of professional groups such as
ARCOS and certification of program coordinators should be supported and encouraged.
IntroductIon
The Accreditation Council of Graduate
Medical Education (ACGME)
Outcome Project [17], initiated in 1999, heralded an intentional shift in emphasis for granting
Graduate Medical Education (GME) accreditation from a program’s potential to educate to a program’s accomplishments, as revealed by assessment of program outcomes.
Expectations for increased emphasis on outcome assessment are reflected in changes to program and institutional requirements that require programs to
(1) identify learning objectives related to ACGME’s general competencies, (2) use increasingly more dependable and objective methods of assessing residents’ attainment of these competency-based objectives, and (3) use outcome data to facilitate continuous improvement of resident and residency program performance. The ACGME Outcome
Project has a four- phase 10-year time line for creation of models of excellence for the core competencies of GME.
The first two phases conducted from
July 2001 through June 2006 focused on the development of an initial response to changes in requirements and sharpening of the focus and definition of the competencies and assessment tools. The third phase of the project that began in July 2006 and runs through June 2011 calls for full integration of the competencies and their assessment with learning and clinical care. The project will initiate its final phase in July 2011 with expansion of the competencies and their assessment to develop models of excellence.
This shift has led to redefinition of the professional responsibilities and requirements of the residency coordinator as an essential member of the management team of an integrated program. This agenda requires strengthening of administrative systems and day-to-day management oversight necessary to maintain accountability and relevancy for orthopaedic surgery residency programs. The program director holds ultimate responsibility

MANuSCRIPTS for overseeing and organizing the activities for an educational program; however, administrative staff involvement is critical given an annual program director turnover rate of
16% per year. According to ACGME’s
Data Accreditation System, 124 of
152 or 82% of all orthopaedic surgical residency program directors have 5 or fewer years of tenure.
Although the current literature addresses the evolution of orthopaedic residency education [2, 3, 6, 8, 9], proposals to improve orthopaedic surgery GME [16], evaluative tools
[5, 7, 10, 19, 20], and the impact of requirements and regulations from a clinician’s perspective [4, 11, 15], there is virtually no discussion regarding management of the administrative burden associated with these expansions and the director-coordinator relationship. Specifically, there is a paucity of literature on the evolution of the Association of Residency
Coordinators in Orthopaedic Surgery
(ARCOS) and its effort to increase the level of professionalism in the field.
Likewise, the literature contains little information on the coordinators’ new and expanding roles and responsibilities in response to ACGME requirements for the orthopaedic surgery residency program, institutional requirements, and integration of the ACGME
Outcome Project.
The survey designers addressed the following questions: What is the profile of a typical residency coordinator? What do they really do? What perceptions do they hold regarding their expanding roles and responsibilities? Collectively, what can be done to ensure their positional viability and promote professional development? materIalS and methodS
The 2006 ARCOS Career Survey was designed to generate qualitative information necessary to illuminate how the role of the orthopaedic surgery residency coordinator had expanded since the initiation of the third phase of the ACGME Outcome Project in
July 2006. Described as the integration phase [17], Phase 3 runs for 5 years with the goal of full integration of the competencies and their assessment with learning and clinical care.
In August 2006, the draft survey instrument was drawn up and presented for review to a focus group of four well-experienced orthopaedic residency coordinators. After being revised to incorporate focus group members’ input and finalized, the survey was distributed by US mail to the 152 orthopaedic residency program coordinators in the United States in
November 2006. The three-page survey instrument consisted of 27 questions grouped around five topics: (a) the residency coordinator profile; (b) stakeholder interactions; (c) resident coordinator roles and responsibilities pertaining to resident/fellow selection and program accreditation; (d) job satisfaction; and (e) special topics that related to resident/faculty diversity and a detailed description of the responsibilities of the residency coordinator (see Table 1 for specific questions). Included among the 27 questions were 10 short-answer, two numerical rating scale, four multiplechoice, four yes/no, and seven openended questions. Once distributed in hard copy, coordinators were reminded and asked to complete survey via two separate e-mails from ARCOS during the investigation period.
Responses were received from 60 of
152 orthopaedic surgery program coordinators, yielding a 39.5% return rate. For virtually all data reported, there was at least an 85% response rate to survey questions; the exceptions were the survey items on diversity, job congruence, and the number of residents per year.
Quantitative data were analyzed with
SPSS software (SPSS Inc, Chicago, IL). reSultS
Sixty-one percent of the respondents had been in their current positions more than 5 years (Table 1). Thirtyone percent had attained a bachelor’s or advanced degrees. On average, there were 22.3 residents currently in their programs, with 62% of respondents identifying the program director as a faculty member other than the chair of their department or division. Annual compensation averaged $43,308.
Ninety-two percent of respondents replied to a question concerning their detailed job responsibilities. Their comments ranged from ‘‘too much/too many to list’’ to multipage attachments.
Of those who provided detail, the commonality of the oversight and management of the residency program were consistent themes. Additionally, most respondents provided similar levels of oversight and management for other elements of the orthopaedic department or division, which included the fellows program, medical student clinical rotations, continuing medical education for faculty, human resources, travel arrangements, and miscellaneous business functions of the department.
Ninety-six percent of respondents indicated some level of involvement in ensuring successful resident and fellowship matching. Sixty-nine percent had either total responsibility for or coordinated the application process.
Ninety-eight percent of respondents indicated they had a role in the completion of the Program
Information Form for their program.
Seventy-eight percent of respondents indicated complete responsibility or sharing of responsibility with the
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ExPANSION Of THE COORdINATOR ROlE program director. All of the respondents were involved in the preparation for the
ACGME-Residency Review Committee site inspection, with 76% indicating a total or high level of responsibility for overseeing the site visitors’ inspection of their residency or fellowship program.
The majority of the respondents reported a consistent level of contact with the Chicago office of the
ACGMEResidency Review Committee, with 68% indicating contact one to six times a year and 16% indicating multiple contacts monthly.
table 1.
Key 2006 ARCOS survey data
Question
How long have you been in your position? (n = 60)
Response
<1 year
5%
1–5 years
33%
Response to questions posed regarding professional development and recruitment varied based on topic.
Eighty-seven percent of respondents had never taken college or graduate level courses relevant to their current positions, although 50% thought
5–10 years
35%
>10 years
26%
What is your current educational level? (n = 60)
Have you taken any special college or graduate level courses relevant to your current position? (n = 60)
Would your institution support these endeavors?
(n = 56)
Does your institution underwrite the cost of obtaining continuing medical education meetings to enhance your job skill set relevant to your position? (n = 59)
High school
22%
Yes
13%
Yes
50%
Yes
80%
What is your annual salary? (n = 57) Mean
$43,308
What do you envision your ideal salary based upon Mean your current job description and responsibilities? (n = 52) $51,259
Estimate the congruency of your current job description and your actual job duties. (n = 60)
High
23%
Do you work on weekends? (n = 59)
Number of weekends per academic calendar?
(n = 56)
Have you been successful in getting your job upgraded? (n = 52)
Should your job be upgraded?
(n = 55)
Yes
75%
Mean
4.31
Yes
42%
Yes
73%
What is your current level of satisfaction with your job benefits? (n = 60)
What is your current level of familiarity with
ACGME core competency regulations? (n = 55)
Very satisfied
48%
High
29%
What suggestions do you have for increasing the participation of women and underrepresented minorities in orthopaedic residency programs? (n = 60)
Responsive
43%
What suggestions do you have for increasing the number Responsive of women and underrepresented minorities within the 28% faculty of your department? (n = 60)
Some college
47%
No
87
No
34%
No
15%
Low
$25,000
Low
$35,000
Medium
22%
No
25%
Low
0
No
58%
No
27%
Satisfied
42% 1
Medium
42%
Nonresponsive
57%
Nonresponsive
72%
Bachelor’s degree Advanced degree
23%
Not sure
16%
Don’t know
5%
High
$87,000
High
$110,000
Low
18%
High
40
Not very satisfied
0%
Low/ No
29%
8%
Nonresponsive
37%
68
ARCOS = Association of Residency Coordinators in Orthopaedic Surgery; ACGME = Accreditation Council of Graduate Medical Education.

MANuSCRIPTS they would be supported by their institution if they chose to enroll for such coursework. Eighty percent reported employer reimbursement for costs associated with participation in continuing medical education meetings to enhance their job skills. When asked to indicate their level of familiarity with
ACGME core competency regulations,
29% were highly familiar, 42% were moderately familiar, and 29% had low familiarity or were unfamiliar. Response rates to questions on strategies for increasing the participation of women and underrepresented minorities in orthopaedic fellowships or residency programs and as faculty were low (43% and 28%, respectively).
When comparing their written position description with actual roles and responsibilities, 45% of respondents confirmed a high to medium level of congruence. Although 73% of the respondents believed their positions should be upgraded from clerical to managerial, only 42% had been successful in achieving this objective.
Respondents envisioned an ideal annual average salary of $51,259. Only 10% of the respondents were dissatisfied with employee benefits such as retirement and healthcare plans (Table 1).
dIScuSSIon
The ACGME Outcome Project
[17], initiated in 1999, heralded an intentional shift in emphasis for granting GME accreditation from a program’s potential to educate to a program’s accomplishments, as revealed by assessment of program outcomes.
The roles of directors and coordinators have been altered considerably by the
ACGME’s phased introduction of revised outcome metrics. The survey designers wanted to answer such questions as: What is the profile of a typical residency coordinator? What do they really do? What perceptions do they hold regarding their expanding roles and responsibilities? Collectively, what can be done to ensure their positional viability and promote professional development?
There are limitations of this survey.
First, although the 39.5% response rate of this followup instrument represented an increase of 8% over the initial survey of 2004, one should not overgeneralize the results. Second, the 2006 instrument was designed based on the first instrument; however, direct comparisons of data between the two are limited because of differences between the two questionnaires. The second instrument was designed with more open-ended questions to solicit a broader range of responses, but this design also increased the opportunity for response bias, which made it difficult to form specific conclusions.
Third, the focus group that reviewed the 2006 draft was chosen because of availability and not randomly selected from ARCOS membership. Focus group members represented the varied types of facilities, and they all were experienced or seasoned program coordinators. Finally, the data from an inactive program were included in the responses and data analysis and affected the lower end of range values in some instances.
A major challenge faced by orthopaedic residency program directors, as presented in ACGME’s essentials of accredited residencies in GME, is the classic tension between the provision of patient care and resident education.
While maintaining the qualifications necessary to be a program director
(specialty expertise, documented educational and administrative abilities,
American Board of Orthopaedic
Surgery certification, appointments in good standing, and station at the primary teaching site), they are responsible for organization and oversight of educational programming in all participating institutions, preparation of statistical and descriptive narratives of the program, and annual updating of program and resident records through ACGME’s
Accreditation Data System. In response to the exponential increase [16] in the functions of the program director, the role of the residency program coordinator has evolved with a more critical managerial component.
Although sparse, the recent literature does cover to some extent such topics as evolution of the orthopaedic residency education process and the impact of progressive ACGME regulations on residency programs.
We found discussions of the program director’s role, responsibilities, and challenges in specialty journals relating to ambulatory medicine, radiology, family medicine, and internal medicine [4, 8, 12, 21]. Of particular interest was a 2004 article [16] in a leading orthopaedic journal that proposed strengthening the program director’s position by eliminating the requirement that the chairperson be the program director in orthopaedic surgery residency programs.
Virtually nothing has been published on the expanding administrative burden of ACGME program requirements and the program coordinator’s role in surgical subspecialty programs, orthopaedic surgery included. Articles relevant to the coordinator position [13, 14] confirm the survey data with respect to breadth of their actual roles and administrative duties that encompass responsibilities for programs and functions beyond the residency program.
In a 2005 article [13], Ruth H.
Nawotniak, a past president of the
Training Administration of Graduate
Medical Education [18], details
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background
ExPANSION Of THE COORdINATOR ROlE
70 the stakeholders that a residency coordinator serves from the local level to the national level, the types of services provided, and the politics of the position. She writes, ‘‘The push and pull between the needs of education and the needs of service is often a topic of conversation between the program director and coordinator and the hospital administrators…
In each of these areas, the program coordinator needs to understand the issues and recognize the impact on their particular training schedules’’
[13]. Data provided by respondents supporting the rationale for upgrading the position concur with her discussion of how the program coordinator should be positioned to maximize the success of the residency program. ‘‘To be successful, the program director and the faculty must view the position of program coordinator as that of a mid-level manager’’ [13]. She writes,
‘‘When these perceptions come together, the coordinator is given the opportunity to become an active, productive participant, involved in achieving all of the goals and objectives of the training program’’ [13].
We found no literature or survey instruments focused on program coordinators, their roles and responsibilities in response to
ACGME standard requirements for the orthopaedic surgery residency program, institutional requirements, or the ACGME Outcome Project.
This lack of attention is unfortunate because expansion of the skills, resourcefulness, responsiveness, and professionalism of the orthopaedic residency coordinator is essential to attainment of accreditation goals and to full compliance with the ACGME
Outcome Project.
Based on the challenges identified by respondents in the 2006 Career
Survey and its mission, ARCOS appears uniquely positioned to provide professional development support to its membership and provide a platform for networking and information sharing nationwide. Similar actions have taken place in internal medicine by the
Association of Program Directors in
Internal Medicine who have endorsed the ‘‘program administrator position as one of professional stature’’ and prepared a standard job description with suggested qualifications and duties [1].
The orthopaedic surgery residency coordinator role has expanded to keep pace with the functional requirements of the program director, the medical education requirements of orthopaedic departments or divisions, and the impact of the turnover rate of the program director position. Adding to the educational and training requirements of an orthopaedic surgery residency education inherently affects clinical orthopaedics and the training of physicians, whether this is a structural linkage with academic medicine or orthopaedic surgery staffing. To maintain accreditation, long-term program viability, and institutional memory, all stakeholders have a role to play in encouraging the necessary increases in program coordinator professionalism and perceptions regarding their expanded role in the management of the orthopaedic surgery residency program.
Acknowledgments
We thank Ellen Greenberger, Residency
Coordinator for the Department of
Orthopaedic Surgery of University
Hospitals Case Medical Center, for extending the invitation to provide the keynote address to ARCOS membership during the 2006 AAOS convention in San Diego, CA. We also thank the members of ARCOS for an engaging, constructive focus group to refine the survey instrument; the wider
ARCOS membership for responding to and completing the 2006 career survey; and Richard J. Haynes, MD, Peter
Stern, MD, and Steven Nestler, PhD, for reviewing the draft presentation, the manuscript, and providing invaluable suggestions.
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T he study of human skeletal dysplasias has greatly facilitated the dissection of developmental pathways governing skeletogenesis.
Mutations in transcription factors
SOX9 and RUNX2 lead to the inherited skeletal dysplasias of campomelic dysplasia and cleidocranial dysplasia, respectively. Both SOX9 and RUNX2 play essential roles in bone and cartilage formation and in postnatal bone remodeling. In this review, we summarize our current knowledge about the functions of
SOX9 and RUNX2--including the interaction between these two master regulators--during skeletogenesis.
IntroductIon
The vertebrate skeleton consists of a variety of individual elements, each characterized by a specific size and shape. There are two main modes by which the skeletal elements are formed (1). Intramembranous ossfication occursduring the development of the flat bones in the skull and the medial part of the clavicles. Here, in intramembranous ossification, mesenchymal cells directly differentiate into bone.
Endochondral ossification, by contrast, forms the bones of the vertebral column, the extremities and the shoulder and pelvic girdles. Here, the skeletal elements are derived from cartilage precursors and are later replaced by bone tissues through a complex, multiple-step process (2,3).
This process includes the patterning of appendicular and axial elements, the commitment of multipotent mesenchymal cells to chondrogenic and osteogenic lineages, and the terminal differentiation of precursor cells into three specialized cell types: the hypertrophic chondrocyte in cartilage, the osteoblast and the osteoclast in bones (4,5). Skeletal development is highly regulated by a hierarchy of genetic, endocrine, and mechanical regulatory programs.
Molecular and biomedical studies in cell culture, gene targeting
(“knockout”) experiments in mice, and molecular studies of human skeletal dysplasia have demonstrated the essential role of a number of regulatory factors and their receptors, including TGF-β, bone morphogenetic proteins (BMPs), FGFs, Sonic and
Indian Hedgehog (SHH and IHH), and parathyroid hormone-related peptide (PTHrP) (2,3,6). Downstream from these signaling molecules, several transcription factors play essential roles in endochondral ossification, including SOX9 and its downstream effectors L-SOX5 and SOX6 in chondrogenesis, and
RUNX2 and its downstream effector zinc finger protein Osterix in bone formation (3,5)
I. SOX9 , campomelic dysplasia, and chondrogenesis
SOX9 belongs to a family of genes that encode transcription factors with a 79-amino-acid high-mobility group (HMG)-type DNA binding domain homologous to that of SRY
(Sry-type HMG box). The functions of the Sox protein family range from roles in early embryogenesis to roles in lineage specification and terminal differentiation events (7,8).
SOX9 contains a proline/glutamine/ alanine rich (P/Q/A) domain and a transactivation (TA) domain carboxyterminal to its HMG domain (9)
(Figure 1). Sox9 binds to essential sequences in the Col2a1 , Col11a2 , aggrecan chondrocyte-specific enhancers and can activate these enhancers in non-chondrogenic cells (10). In 1994,
SOX9 became the first transcription factor whose mutations were identified in the inherited skeletal disorders.
Heterozygous mutations in and around
SOX9 in humans cause campomelic dysplasia (CD), a severe skeletal malformation syndrome characterized by anomalies in all skeletal elements derived from cartilage (11) (12).
Campomelic dysplasia is characterized by the bowing of the femur and tibia, with an incidence of approximately 1 in
111,000 to 1 in 2000,000 live births.
It can be a lethal condition, although a proportion of CD children can survive into adulthood (13,14). Interestingly, three-quarters of XY individuals are either intersex or exhibit male-tofemale sex reversal, underscoring the essential roles of SOX9 in testis development (11) (12,15,16) . Indeed,
SOX9 has recently been implicated in the development of numerous other tissues, including skin, intestine, and pancreas (8).
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INHIbITION Of THE PI3K-AKT SIgNAlINg PATHWAY
SOx9 ANd RuNx2
72
Genetic study of the mouse has contributed greatly to our understanding of the pathogenesis of
CD, revealing that SOX9 is essential for chondrocyte condensation and cartilage formation. During mouse embryonic development, Sox9 is highly expressed in all chondrogenic precursor cells and in differentiated chondrocytes.
However, the expression of Sox9 is not detected in hypertrophic chondrocytes
(9) (17). Murine knockout studies demonstrate that Sox9 is required for chondrogenesis. Its downstream transcriptional effectors L-Sox5 and Sox6, two other HMG family members, act redundantly to promote chondrogenesis (18,19). Furthermore, haploinsufficiency of Sox9 in mice results in defective cartilage primordial, premature skeletal mineralization, and an enlarged hypertrophic zone in growth plates, resembling the
CD phenotype (20) (21). In a Sox9 knock-in mouse model in which Sox9 was over-expressed in proliferating chondrocytes, osteoblast differentiation was also delayed (22). Interestingly, recent mouse genetic studies showed that Sox9 -expressing precursor cells could eventually differentiate into tendons and osteoblasts (23).
Besides activating the initiation of chondrogenesis, SOX9 also controls chondrocyte hypertrophy. In vitro and in vivo studies suggest that Sox9 might be a physiological target of parathyroid hormone (PTHrP) signaling (24,25).
The PTHrP-dependent increased transcriptional activity of Sox9 helps maintain the chondrocyte phenotype of cells in the prehypertrophic zone and inhibit their maturation to hypertrophic chondrocytes. However, the downstream transcription targets for Sox9 that prevent chondrocyte hypertrophy are still largely unknown.
Besides its well-established role in embryonic skeletogenesis, SOX9 also plays an essential role in fracture healing, a process that recapitulates the successive steps of endochondral ossification. In a murine fracture healing model, Sox9 expression was increased markedly during the early stages of callus formation, followed by the up-regulation of type II collagen production. Moreover, bone morphogenetic protein 2 (BMP2) gene transfer into the fracture site led to the accelerated up-regulation of Sox9 and type II collagen when compared with control fractures (26). Thus, SOX9 is also involved in the activation and maintenance of chondrogenesis during fracture healing, and the enhancement of chondrogenesis by BMP2 is mediated via a SOX9-dependent pathway. However, the regulation of
SOX9 activity during fracture healing is still poorly understood.
II. RUNX2, osteogenesis, and CCD
By the mid 1990’s, a series of converging studies pointed to the critical function of RUNX2
(also called AML3, PEBP2 α A,
Osf2, and CBFA1) during skeletal morphogenesis and especially during osteoblast differentiation (27,28).
RUNX2 is one of three mammalian orthologs ( RUNX1, RUNX2, RUNX3 ) of the Drosophila Runt gene. It contains a unique N-terminal stretch of consecutive polyglutamine and polyalanine repeats (Q/A domain), a runt domain, and a C-terminal proline/serine/threonine-rich (PST) activation domain (Figure 1). The runt domain is a 128 amino acid polypeptide motif that has the unique ability to independently mediate DNA binding and protein heterodimerization (27). RUNX2 has been shown to regulate all the major genes expressed by osteoblasts in tissue culture (29). Runx2 null mice display a complete lack of both intramembranous and endochondral ossification, with no overt osteoblast differentiation (30,31). Runx2 also regulates postnatal bone formation and bone remodeling (32-34).
Importantly, mutations in RUNX2 result in cleidocranial dysplasia
(CCD), a dominantly inherited skeletal dysplasia characterized by hypoplastic clavicles, large fontanels, dental anomalies, and delayed skeletal development (35) (36)
(37) (38). The significant effect on
Figure 1. Schematic representation of SoX9 and runX2 domain structures.
HMG, high-mobility-group Dna binding; P/Q/a, proline/glutamine/alanine rich domain; ta, transactivation domain. Q/a, polyglutamine/polyalanines repeat domain; rUnt, runt domain; Pst, proline/serine/threonine-rich activation domain.

INHIbITION Of THE PI3K-AKT SIgNAlINg PATHWAY manuscripts
Figure 2. Severe osteopenia in Col1a1-SOX9 transgenic mice. top, schematic representation of Col1a1-SOx9 transgenic construct containing full-length
SOx9 under the control of an osteoblast-specific 2.3-kb Col1a1 promoter in the coat color vector. tyr, tyrosinase minigene; WPre, woodchuck hepatitis virus posttranscriptional regulatory element; Hs4, chicken β -globin insulator. Bottom, radiographic analysis of skeletons from 5-week-old wild-type (WT) and Col1a1-SOx9 transgenic mouse.
calvicular development supports the presence of a generalized dysplasia since the clavicle is among the first bones to ossify and does so via both intramembranous and endochondral ossification (39). The prevalence of
CCD is estimated to be 1 in 1 million,
Figure 3. working model of the effect of SoX9 and runX2 interaction on skeletogenesis. in pluripotent osteochondro-progenitor cells, sOx9 is an essential transcriptional activator that promotes these cells’ commitment and differentiation to chondrocytes. in parallel, at least in part by directly inhibiting rUnx2 function, sOx9 also acts as a transcriptional repressor to prevent these cells from differentiating into osteoblasts or hypertrophic chondrocytes prematurely.
though this is likely an underestimate.
CCD patients are often diagnosed by wide, open sutures and fontanels, along with characteristic hypoplastic clavicles that enable the patients to appose their shoulders (28,40).
Since there is little initial morbidity associated with these symptoms, CCD patients may escape diagnosis until they are seen by pediatric dentists for dental complications such as delayed loss of primary dentition, delayed eruption of permanent dentition, supernumerary teeth, and malocclusion (28,40).
Besides controlling osteoblast differentiation, RUNX2 also acts as a positive regulator of chondrocyte hypertrophy.
Runx2 null mice lack hypertrophic chondrocytes in some skeletal elements, and the expression of Runx2 in nonhypertrophic chondrocytes can induce chondrocyte hypertrophy (41) (42) (43) (44) (45).
Additionally, the continuous expression of Runx2 in nonhypertrophic chondrocytes partially rescues Runx2 deficient mice by restoring chondrocyte hypertrophy and vascular invasion
(44). Defective terminal chondrocyte hypertrophy is also present in CCD patients and is due in part to decreased
RUNX2 transactivation of the hypertrophic chondrocyte-specific type
X collagen gene ( Col10a1 ) (46).
III. RUNX2, osteoporosis, and tumorigenesis.
Interestingly, besides being the causal gene for the rare disease CCD, increasing evidence suggests that subtle polymorphic variations in RUNX2 could also regulate BMD in the general population (47-49). Most of the identified polymorphisms lie within the polyalanine and polyglutamine repeats in exon 1, whose functional relevance remains unclear (47). Thus, RUNX2 itself has been proposed as one of the candidate genes for osteoporosis (50).
Osteosarcoma is the 5 th most frequent malignancy in adolescents, and a deeper understanding of the basic biology of this tumor can lead to better treatment (51,52). Interestingly, more than 80% of osteosarcomas were either poorly differentiated or undifferentiated with the later marker of osteogenic differentiation--osteocalcin-undetectable (53). Thus, differentiation is antithetical to tumor development in osteosarcoma. However, the molecular basis underlining the relationship between malignant transformation and loss of differentiation is still poorly understood (53). Interestingly,
RUNX2 expression is consistently disrupted in osteosarcoma cell lines
(53). The ectopic expression of
RUNX2 in human osteosarcoma cells inhibits cell growth through the induction of CDK inhibitor p27 KIP1 ,
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SOx9 ANd RuNx2
74 leading to the dephosphorylation of the retinoblastoma tumor suppressor protein (pRb) (53). Thus it is proposed that the coupling of osteoblast differentiation to cell-cycle withdrawal is mediated through RUNX2 and p27 KIP1 , and these processes are disrupted in osteosarcoma (53). But how RUNX2 activity is downregulated in osteosarcoma remains poorly understood. On the other hand,
RUNX2 can also contribute to the ability of metastatic breast cancer cell lines to form osteolytic bone lesions in mouse models (54,55). Thus, the precise role of RUNX2 in osteosarcoma and bone metastasis is very complex and probably cell context-dependent.
IV. SOX9 represses RUNX2 activity during skeletogenesis
As a master regulator of osteoblast differentiation, chondrocyte hypertrophy, and vascular invasion of developing bones, RUNX2’s context-dependent specific function has to be tightly regulated to ensure proper bone formation. Indeed,
RUNX2 cooperates with numerous tissue-specific transcription factors or cofactors to integrate a variety of signals and regulate osteoblast differentiation and chondrocyte maturation (27,56).
Several transcriptional co-repressors have been identified to interact with RUNX2 that prevent its DNA binding, alter chromatin structure, and possibly block co-activator complexes
(56). We have recently shown that
SOX9 can directly regulate RUNX2 function during chondrogenic cell fate commitment as well as later during chondrocyte hypertrophy (57).
SOX9 directly interacts with RUNX2 and represses its activity in vitro via their HMG and RUNT domains, respectively. In a novel Col1a1-SOX9 transgenic mouse model, ectopic expression of full-length SOX9 in mouse osteoblasts results in an osteodysplasia characterized by severe dwarfism, osteopenia, down regulation of osteoblast differentiation markers, and reduced mechanical strength
(Figure 2, data not shown) (57).
Finally, this dominant inhibitory function of SOX9 is physiologically relevant in human campomelic dysplasia. In CD, haploinsufficiency of SOX9 resulted in the upregulation of the RUNX2 transcriptional target
COL10A1 as well as all three members of the RUNX gene family (57). Because
SOX9 and RUNX2 co-express during mesenchymal condensation, our study supports the dominance of SOX9 function over RUNX2 early in the progenitor cell fate decision between osteoblastic vs. chondrogenic lineages and later in chondrocyte hypertrophy.
IV. Future directions
While tremendous progress has been made in understanding the function and regulation of the two master regulators of skeletogenesis, SOX9 and
RUNX2, many more questions remain.
Interestingly, in addition to severe dwarfism, Col1a1-SOX9 transgenic mice (Figure 2) also exhibited altered mesenchymal stem cell
(MSC) differentiation with increased adipogenisis and decreased osteogenesis capability (data not shown). Thus, it would be interesting to further investigate whether SOX9 can directly interact with other factors during
MSC differentiation. Understanding the transcriptional network governing
MSC property may ultimately lead to better therapeutic options for various bone diseases and fracture healing. It would be also interesting to delineate the direct effect of SOX9 on RUNX2 during chondrocyte hypertrophy. Indeed, in our latest transgenic model with targeted
SOX9 expression in hypertrophy chondrocytes, there is decreased mineralization and surprisingly severe spontaneous osteoarthritis in 11-month-old transgenic mice
(data not shown). In summary, the study of rare skeletal dysplasias not only identifies the causal molecular alterations for specific diseases, but it also helps us to elucidate the complex network of normal skeletogenesis and bone remodeling (58,59). The study of SOX9 and RUNX2 has served as an excellent example of integrating clinical and basic research to achieve a better understanding of not only the pathogenesis of skeletal dysplasias, but also of the underlying molecular mechanisms of normal bone and cartilage formation.
Acknowledgements
Work in Dr. Guang Zhou’s laboratory is supported by a NIH grant (K22
DE015139). We thank Valerie
Schmendlen for editorial assistance.
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abStract
T he amount of bone turnover in the skeleton is has been identified as a predictor of fracture risk independent of areal bone mineral density (aBMD) and is increasingly cited as an explanation for discrepancies between areal bone mineral density and fracture risk. A number of mechanisms have been proposed to explain how bone turnover influences bone biomechanics, including regulation of tissue degree of mineralization, the disconnection or fenestration of individual trabeculae by remodeling cavities, and the ability of cavities formed during the remodeling process to act as stress risers. While these mechanisms can influence bone biomechanics, they also modify bone mass. If bone turnover is to explain any of the observed discrepancies between fracture risk and areal bone mineral density, however, it must not only modify bone strength but modify bone strength in excess of what would be expected from the associated change in bone mass. This article summarizes biomechanical studies of how tissue mineralization, trabecular disconnection and the presence of remodeling cavities might have an effect on cancellous bone strength independent of bone mass. Existing data support the idea that all of these factors may have a disproportionate effect on bone stiffness and/or strength, with the exception of average tissue degree of mineralization, which may not have an effect on bone strength that is independent of aBMD.
Disproportionate effects of mineral content on bone biomechanics may instead come from variation in tissue degree of mineralization at the microstructural level. The biomechanical explanation for the relationship between bone turnover and fracture incidence remains to be determined but must be examined not in terms of bone biomechanics but in terms of bone biomechanics relative to bone mass.
IntroductIon
Bone turnover represents the total volume of bone that is both resorbed and formed over a period of time
[1]. In adults, bone turnover occurs primarily through bone remodeling, a focal process that involves the coupled activity of osteoclasts and osteoblasts
[2]. Changes in the amount of bone turnover cause local changes in bone volume and the average age of tissue in a bone, resulting in alterations in tissue degree of mineralization and trabecular microarchitecture.
Clinical findings that biochemical markers of bone turnover can predict fracture risk independent of areal bone mineral density (aBMD) have led to the suggestion that the amount of bone turnover in the skeleton can have a biomechanical effect independent of bone mass 1 [3-8] and that bone turnover may help to explain discrepancies between aBMD and fracture risk [5, 6]. The biomechanical effects of alterations in bone turnover are commonly attributed to modifications in tissue degree of mineralization, the fenestration or disconnection of individual trabeculae and/or by remodeling cavities acting as stress risers [4, 6-9]. While these mechanisms can influence bone strength, they also modify bone mass. If one of these mechanisms is to explain any of the discrepancies between aBMD and fracture risk, however, it must have an effect on bone strength that is much larger than would be expected from the change in bone mass alone. The purpose of this article is to review the biomechanical effects of changes in bone that can be caused by bone turnover. The current article concentrates on the biomechanics of human cancellous bone specimens 3-5mm in smallest dimension, as that is the scale at which the mechanisms mentioned above all have biomechanical significance.
Biomechanics of bone at this size scale is also important because a factor that does not have a disproportionate biomechanical effect at this scale could not have a disproportionate biomechanical effect at the scale of the whole bone [10]. The biomechanical effects of a 6% difference in bone mass caused by different aspects of bone turnover are summarized in Table 1
1 The term “bone mass” will be used here to describe the total mass of mineralized tissue in a bone specimen (grams) and should not be confused with aBMD, a measure of bone density performed using dual-energy x-ray absorptiometry that is expressed in the units g/cm 2 .
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78 table 1.
the reduction in human cancellous bone stiffness and strength associated with a 6% difference in bone volume or mass is shown. the 95% confidence interval is reported if available, otherwise the range across all cited studies is shown. a factor that causes a greater change in bone stiffness or strength than that caused by a 6% reduction in bone volume (the first row) has the potential to explain how bone turnover can influence fracture risk independent of bone quantity.
Process through which bone remodeling modifies bone mass
Reduction in bone volume
Difference in Bone
Mass (%)
-6% (Volume)
Expected
Reduction in
Stiffness (%)
12-16%
Expected
Reduction in
Strength (%)
9-14%
Source
Empirical power law models [14,
17-21]
Reduction in average tissue degree of mineralization from
65% to 62% ash by weight.
Intraspecimen variation in tissue degree of mineralization
Removal of Trabeculae
-6%(Bone Mineral
Content)
0% Reduction in bone mass;
Increase in
COV of tissue mineralization from 20% to 50%
-6% (Volume)
11-13%
14-24%
3-39%
11-13%
Not Yet
Evaluated
95% confidence interval from empirical power law models [41]
Micro-computed tomography based finite element models [45,
46]
Addition of Remodeling Cavities -6% (Volume) 12-47%
18-35%
13-61%
3D cellular solid finite element models [50, 51]
Micro-computed tomography based finite element models
[53, 55]
(a 6% decline in bone mass has been selected as that is the estimated size of the remodeling space in the spine [11]).
Bone Volume
Because the first step in the remodeling process is bone resorption, each remodeling event is associated with the formation of a temporary cavity. The total volume of bone occupied by all remodeling cavities and unmineralized bone tissue (osteoid) is known collectively as the remodeling space
[12]. Increases in bone turnover result in an increase in the volume occupied by the remodeling space and cause a corresponding reduction in mineralized bone volume.
Biomechanical testing of cancellous bone specimens has shown that bone stiffness and strength are related to apparent density ( ρ , g/cm 3 ) through power law relationships. Because apparent density is directly related to bone volume fraction (BV/TV) [13], the same power law relationships are valid for bone volume fraction as well.
These power law relationships can be expressed as follows:
E ∝ ρ A ∝ (BV/TV) A , (1)
σ
Ult
∝ ρ B ∝ (BV/TV) B , (2) where E is the stiffness of the specimen
(elastic modulus), σ
Ult
is the strength
(ultimate stress) in compression and
A and B are constants [14] (for a comprehensive review please see [15,
16]). Studies of human cancellous bone [14, 17-21] have reported the exponent A to be as small as 1.2 [20] or as large as 3.0 [14], suggesting that a bone specimen with 6% less bone mass due to having less bone volume is expected be 7-17% less stiffness.
The exponent B used to predict bone strength has been reported to be as small as 1.48 [22] or as large as 2.47
[19], suggesting that a specimen with
6% less bone mass is expected to be
9-14% less strong (Table 1).
Tissue Degree of Mineralization
After a new volume of bone is formed, it begins to accumulate mineral in a process that can continue for years afterwards [23, 24]. As a result, the degree of mineralization of older bone tissue is greater than that of newly formed tissue, so that the amount of bone turnover can influence the average tissue degree of mineralization [25, 26].
Examination of mineralized tissues across a wide range of species
(including deer and whales) have associated increased tissue degree of mineralization with increased bone stiffness and strength and, in some cases, increased brittleness (the term
‘brittle’ is used here in an engineering sense expressing a material property)
[27, 28]. While these studies demonstrate that tissue degree of

manuscripts mineralization can be biomechanically important, most do not include analyses of bone porosity and therefore cannot be used to examine the biomechanical effects of tissue degree of mineralization independent of bone volume . In addition, it is important to keep in mind that trends observed among species do not necessarily apply within a species. For example, consider the commonly held idea that “hypermineralized” bone tissue is more brittle, a concept frequently noted when discussing possible adverse effects of long-term inhibition of bone turnover. Although comparisons among animals suggest that more highly mineralized tissue is more brittle, only two studies of human bone specimens have shown increased tissue degree of mineralization to be associated with increased brittleness
(evaluated as impact energy in cortical bone specimens) [29, 30]. Indeed, other studies of human bone specimens have found increased tissue degree of mineralization to be associated with reduced brittleness (measured as compressive toughness in cancellous bone [31], or fracture toughness evaluated in cortical bone [32]).
Additionally, a number of studies of human bone specimens did not observe a relationship between tissue degree of mineralization and brittleness
(measured as toughness or energy to failure in cortical or cancellous bone
[33-36]). While comparisons among species suggest that highly mineralized bone specimens are more brittle, it is not yet clear that specimens of human bone can become brittle through an increase in average tissue degree of mineralization alone.
With regard to bone stiffness and strength, few studies have been designed to separate the biomechanical effects of tissue degree of mineralization from those of bone volume. Follet and colleagues found that average tissue degree of mineralization
(measured through quantitative contact radiography) was positively correlated with cancellous bone stiffness, strength and brittleness (brittleness evaluated as toughness) [31], and that tissue degree of mineralization had a biomechanical effect independent of bone volume.
Others have used power law models to predict the biomechanical effects of tissue degree of mineralization in cortical bone [18, 37-40], but only two of these studies accounted for variation in bone volume fraction (both using non-human tissue [38, 40]). Currey suggested that the separate effects of bone volume and tissue degree of mineralization could be expressed with a two-parameter power law model and applied the approach to non-human tissue [38]. Hernandez and colleagues applied this statistical approach to human bone and found the following relationships [41]:
E (GPa) ∝ (BV/TV) 2.58±0.02
α 2.74±0.13
, (3)
σ
Ult
(MPa) ∝ (BV/TV) 1.92±0.02
α 2.79±0.09
, (4) where E, σ
Ult
and BV/TV are as defined above, α is the degree of mineralization
(measured as ash mass/dry bone mass), and the exponents are expressed as mean ± standard error. This analysis is the only study of cancellous bone to detect and quantify the independent effects of bone volume and average tissue degree of mineralization on bone biomechanics. With regard to bone strength (equation 4), the exponent applied to tissue degree of mineralization (2.79) is much greater than the exponent applied to bone volume fraction (1.92) suggesting that cancellous bone strength is much more sensitive to differences in tissue degree of mineralization. Clinical measures of bone mass may capture some of this effect, however. Clinical measures of bone mass evaluate the total amount of mineral present (both mineralized volume and degree of mineralization).
Assuming clinical evaluation of bone mineral content (BMC) are directly related to the inorganic content in bone (the ash content), BMC can be expressed as:
BMC ∝ (BV/TV) ρ t
α , (5) where ρ t
is the density of the mineralized bone tissue (in grams), a parameter that is linearly related to tissue degree of mineralization [13, 41].
By combining equations (3), (4) and
(5) we can estimate the differences in bone biomechanics associated with a
6% difference in BMC caused entirely by reductions in average tissue degree of mineralization (this change in BMC corresponds to a reduction in tissue degree of mineralization from 65% to
62% ash by weight). This difference in BMC is associated with a difference in specimen stiffness of 11-13% and a difference in bone strength of 11-13%
(both of these ranges are expressed using the 95% confidence interval of the regression coefficients above).
Another way of comparing the biomechanical effects of bone volume and tissue degree of mineralization is to examine the relationship between bone mineral content and bone strength
[10]. Figure 1 shows the percent change in bone strength expected from a hypothetical reduction in bone mineral content under two different conditions: 1) when changes in bone mineral content are caused entirely by bone volume (lower region with dark blue shading); and 2) when changes in bone mineral content are caused entirely by tissue degree of mineralization (upper region with lighter orange shading). That these two confidence intervals overlap suggests that alterations in tissue degree of mineralization may not modify the relationship between bone strength
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80
Figure 1. the predicted reduction in strength caused by a reduction in bone mineral content or bone mineral density (as would be measured by dual-energy x-ray absorptiometry) is shown 1) a hypothetical case where changes in bone mineral content are caused entirely by changes in volume fraction
(dark blue region) and 2) a hypothetical case where changes in bone mineral content are caused entirely by reductions in tissue degree of mineralization (light orange region). the overlapping regions are based on the 95% confidence interval of the regression model reported by Hernandez and colleagues[41].
and clinical measures of BMC. Hence, alterations in average tissue degree of mineralization may have little effect on the ability of BMC to predict bone strength and are therefore not expected to be responsible for discrepancies between fracture incidence and aBMD. Additional studies are needed to confirm this analysis (equations
3, 4) and to determine if the average degree of mineralization can have disproportionate effects on other mechanical properties (brittleness for example) or under different loading conditions (impact, shear, etc.).
While most studies have concentrated on the biomechanical effects of average tissue degree of mineralization, variation of tissue degree of mineralization at the micro-scale has also been implicated as a factor that can influence bone biomechanics. By altering the number and/or size of new remodeling events, bone turnover will not only modify the average tissue degree of mineralization, but also the variability of tissue degree of mineralization among osteons/hemiosteons. Changes in variation of tissue degree of mineralization have been associated with alterations in bone turnover during bisphosphonate therapy and in metabolic bone disease [42, 43].
Variability of tissue degree of mineralization can also differ among regions of the skeleton (iliac crest v. calcaneous) [31].
Micro-computed tomography based finite element models have been useful for studying the biomechanical consequences of variation in tissue degree of mineralization because they make it possible to consider the biomechanical effects of tissue degree of mineralization independent of bone volume or microarchitecture. Finite element studies suggest that an increase in intraspecimen variation in tissue degree of mineralization (modeled as local variation in tissue stiffness) can cause a reduction in cancellous bone stiffness, even when trabecular microarchitecture and average tissue degree of mineralization are maintained constant [44-46]. Jaasma and colleagues determined that an increase in the coefficient of variation (standard deviation/mean) of the tissue stiffness from 20% to 50% would result in a reduction in elastic modulus of cancellous bone by 19-24%, even when average tissue degree of mineralization was maintained constant. More recently, Bourne and van der Meulen measured variation in mineral content directly using calibrated microcomputed tomography and found that an increase in the coefficient of variation of tissue stiffness from 20% to 50% is expected to cause a 14% reduction in the elastic modulus.
Disconnection of Trabeculae
It is commonly stated that cavities formed during remodeling can disconnect or fenestrate trabeculae, modifying trabecular microarchitecture and potentially cause a disproportionate change in cancellous bone strength [7-9]. Quantifying the effect of trabecular disconnection experimentally is challenging because of technical difficulties in identifying and counting individual trabeculae
(only recently have techniques for directly counting individual trabeculae in micro-CT images of cancellous bone been presented [47, 48]).
Existing biomechanical analyses have therefore used cellular solid models with trabecular-like microarchitectures to mimic cancellous bone structure.
Two- and three-dimensional cellular solid models indicate that removal of individual trabeculae can result in reductions in cancellous bone stiffness and strength that are greater than would be expected from the associated change in bone volume [49-51]. Threedimensional models suggest that a
6% difference in bone volume caused by removal of trabeculae can reduce cancellous bone stiffness by 3% (only horizontal trabeculae removed) to
39% (only vertical/oblique trabeculae removed) and compressive strength by
18% (only horizontal trabeculae) to
35% (only vertical/oblique trabeculae)
[51]. As these simulations represent extreme cases where only horizontal or only vertical/oblique trabeculae are removed, the actual changes in bone biomechanics resulting from trabecular

manuscripts
Figure 2. three cylindrical trabeculae (150 microns in diameter) are shown with remodeling cavities wrapped around them circumferentially. the remodeling cavities in each image occupy the same volume (i.e. the same amount of bone turnover is depicted) but the number, surface size and depth of the cavities differs within the range observed histologically. the bending moment and critical load for euler buckling (calculated within the tapered region only) is presented relative to that in the first image and is shown to vary by as much as an order of magnitude among the three possibilities.
disconnection are expected to be somewhere in between these values.
Remodeling Cavities
It has been proposed that cavities formed during bone remodeling
(Howship’s lacunae) can act as stress risers, causing disproportionate reductions in the biomechanical performance of cancellous bone.
Experimental evaluation of the effects of remodeling cavities on cancellous bone has been limited because a repeatable technique for making threedimensional measures of remodeling cavities in cancellous bone has not yet been demonstrated. Existing data is therefore limited to predictions made from finite element models. A number of biomechanical analyses have illustrated how the presence of a cavity on the surface of a single trabecula may increase the stresses and strains in surrounding tissue [52-
54]. Additionally, two finite element analyses have suggested that remodeling cavities can have a disproportionate effect on cancellous bone stiffness and strength in 3-5mm specimens [53, 55].
A reduction in bone volume of 6% caused by the addition of remodeling cavities was predicted to reduce the elastic modulus by 12-47% and the compressive strength by 13-61%. The ranges for these predictions are large because the biomechanical effects of remodeling cavities can be influenced by a number of factors including the initial bone volume fraction (more porous bone can be more sensitive to remodeling cavities) and the placement of remodeling cavities in the cancellous bone structure. When placed in regions of high strain within the cancellous bone structure (where tissue microdamage and mechanical stresses are expected to be greatest) remodeling cavities can have a large, disproportionate effect on cancellous bone biomechanics [55]. As a result, the degree to which remodeling cavities are targeted to tissue damage or tissue strain (two factors believed to stimulate bone remodeling) will modulate the effect of bone remodeling on bone biomechanics.
Additionally, the number and size
(length, width, depth) of remodeling cavities may have biomechanical significance. Although an increase in bone turnover is commonly interpreted as an increase in the number of remodeling events, two-dimensional histomorphometry measurements cannot differentiate an increase in the number of remodeling events from an increase in the size of each individual event (width, length and depth) [56], a distinction that can result in very different stress distributions within cancellous bone. Simple mechanical analyses suggest that the number and size of remodeling cavities may influence the mechanical performance of a trabecula independent of bone volume or total amount of bone turnover
(Figure 2). The number and size of remodeling cavities may also influence intraspecimen variation in tissue degree of mineralization by determining the size of each osteon or hemi-osteon and may also influence the rate at which trabeculae are disconnected by remodeling events (deeper cavities are more likely to disconnect trabeculae [57]).
Unfortunately, little is known about the complete size and shape (length, width and depth) of remodeling cavities in human cancellous bone because two-dimensional techniques cannot obtain measure all three of these size dimensions at once [58,
59]. Micro-computed tomography is not as helpful as one would expect because few imaging systems can obtain the resolution needed to detect the scalloped surface of a remodeling cavity and those imaging systems with such high resolution can typically only observe one or two cavities per specimen, far too few to characterize the population of remodeling events in a region of the skeleton. Recently, serial block-face imaging using an automated microtome or milling machine has been used to image remodeling cavities in three dimensions, and may prove useful in determining the placement and size of remodeling cavities in human bone biopsies or cadaver tissue [54, 60].
concluSIonS
Table 1 provides a unique way of comparing the biomechanical effects of bone remodeling for a given difference in bone mass (in this case a 6% difference in bone mass). An aspect of bone that has the potential to explain discrepancies between aBMD
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82 and fracture incidence will have a biomechanical effect that is greater than would be expected from that caused by bone volume alone. For example, a 6% difference in bone mass caused by the average tissue degree of mineralization
(second row in Table 1) is associated with an 11-13% difference in bone strength, a range that is well within that expected for the same reduction in bone mass caused by bone volume
(9-14%, first row of Table 1). As a result, this analysis suggests that it is unlikely that differences in average tissue degree of mineralization can have a disproportionate effect on cancellous bone compressive strength and that average tissue degree of mineralization would be unlikely to contribute to a biomechanical explanation for discrepancies between aBMD and fracture incidence.
Two conclusions can be made from comparing the remaining factors in
Table 1 to the effect of bone volume.
First, existing experimental and computational data suggest that, while average tissue degree of mineralization can influence bone strength, it may not be able to explain discrepancies between bone biomechanics and clinical measures of bone mass.
Local variability of tissue degree of mineralization is a more likely explanation. Secondly, while existing biomechanical analyses support the idea that trabecular disconnection and remodeling cavities may have a disproportionate effect on bone biomechanics (i.e. the biomechanical effects can exceed those expected from difference in bone volume), there is considerable overlap between the biomechanical effects of these factors and the biomechanical effects of bone volume alone. As a result, further work is needed to determine if bone remodeling may have a disproportionate effect on bone biomechanics . Whether or not these aspects of bone remodeling have a biomechanically relevant effect independent of bone mass will depend on characteristics of bone remodeling that we currently know little about, such as the number and size of remodeling events and how well remodeling cavities are targeted to mechanical stress/strain and microscopic tissue damage.
Additionally there is growing evidence that aspects of collagen such as the concentration of naturally occurring non-enzymatic cross-links, can be modified by bone turnover and can influence the biomechanical performance of bone specimens [36,
61-65]. Unfortunately regression models accounting for these relationships in cancellous bone specimens are not available so the potential effect is not listed in Table
1. As it is unlikely that collagen cross-linking can be detected by aBMD, any biomechanical effect of collagen in bone biomechanics would be likely to have a disproportionate effect on bone biomechanics. Lastly, it is not clear whether all of these biomechanical effects are independent of one another or if they can interact to have synergistic effects. Further study of the biomechanical effects of bone remodeling and the degree to which those effects are explained by bone mass is necessary to truly understand the relationships between bone turnover and fracture risk.
ACKNOWLEDGEMENTS
This work was supported by NIH/
NIAMS R21 AR054448.
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52. Smit TH, Burger EH.
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reprinted with the permission of the Journal of Bone and Joint surgery.
Wera GD, Marcus re, Ghanayem aJ, Bohlman HH. failure within one year following subtotal lumbar discectomy. J Bone Joint Surg Am.
2008. 90(1):10-15.
84
This investigation was approved by the Institutional Review Board At
University Hospitals of Cleveland.
Structured abStract:
Study design: discectomies.
Retrospective review of 259 lumbar
Objective
To compare rates of reoperation after subtotal discectomy versus established rates after fragment excision.
Summary of Background Data
Herniated Nucleus Pulposes (HNP) and anular morphology influence rates of reherniation after discectomy. Certain patterns are linked to reherniation rates exceeding 20%.
Methods
We retrospectively reviewed 259 singlelevel lumbar discectomies performed between 1980 and 2005. Mean followup was 60.9 (range 24-230) months.
In each case, annulotomy and subtotal discectomy was performed in addition to excision of disc fragments. HNP morphology was classified according to the four-part system of Carragee
(Type 1: fragment/fissure; Type 2: fragment/defect; Type 3: fragment/ contained; Type 4: no fragment/ contained). Fisher’s exact test was used to compare our proportion of patients with reherniation and/or reoperation to Carragee’s series in which only fragment excision was performed.
Results
Of 259 patients, 12 (4.5%) reoperations were performed resulting in a significant difference in failure and reoperation rate noted in Type 2 herniations. There was a significantly lower rate of failure and reoperation for
Type 2 HNP after subtotal discectomy
(3.4%) when compared to fragment excision alone (21.2%), p<0.003. We also propose a 5 th disc morphology for future investigations: the calcified disc.
Conclusions
Subtotal discectomy is an acceptable technique that decreases reherniation after lumbar discectomy.
Key Words
Lumbar Disc Reherniation
Key Points
We classified Herniated Nucleus
Pulposes in 259 patients according to the Carragee system.
Our rates of reherniation and reoperation are lower than previously established rates.
Subtotal discectomy decreases reherniation and reoperation even in high risk disc morphology.
Mini Abstract/Précis
Anular morphology of lumbar discs is believed to influence rates of reherniation after discectomy. Certain patterns are linked to reherniation rates exceeding twenty percent. Therefore, consideration has been given to performing subtotal discectomy as opposed to fragment excision alone. In this study, we demonstrated significantly lower rates of failure after subtotal discectomy versus established norms for fragment excision alone.
Introduction:
Lumbar disc herniation is a prevalent spinal disease with variable re-operation rates reported in the literature. Lifetime occurrence rates of sciatica are high reaching as many as 40% of individuals.
Depending on severity, the rates of surgery for sciatica range from 2 to 10% of patients 1 . In the long term, large studies point to rates of repeat surgical intervention after lumbar discectomy

manuscripts to be on the order of 12 percent 2 .
Epidemiologic studies of lumbar surgery in general have also found high rates of re-operation (9.5%). Lumbar surgery itself is an independent risk factor for re-operation regardless of demographics, comorbidities, or anatomic type of index procedure 3 .
Recently, the morphology of lumbar disc herniation has proven a strong predictor of reherniation following limited lumbar discectomy for sciatica. Until 2003, the terms disc protrusion, noncontained, extrusion, and sequestration have been the main descriptors of the disc, annulus, and any fragments. Carragee et al classified lumbar disc morphology according to anular integrity and the presence of extruded fragments (Table
I) 4 . By this method, the highest rates of disc reherniation and reoperation occurred among patients with a large anular defect and a free fragment
(fragment-defect).
To our knowledge, no other investigation has classified a large number of lumbar disc herniations or reviewed outcomes after lumbar discectomy using the
Carragee system. Nor has this system been applied to a patient population undergoing discectomy and curettage of the nucleus rather than fragment excision alone. Of particular interest is the rate of reherniation and reoperation in these patients.
Materials and Methods
Institutional review board approval was obtained for this investigation.
We retrospectively reviewed a database of 1100 patients of lumbar surgery performed between 1980 and 2005.
This group was screened for primary cases of single level lumbar discectomy for herniated nucleus pulposes (HNP) performed by our group. 259 patients with a single level lumbar discectomy were identified that had adequate documentation for follow up. Mean follow-up was 97.6 months (range
2-350 months).
Inclusion criteria in the study were 1) history and physical consistent with sciatica, 2)radiographic confirmation of single level HNP on MRI or myelogram, and 3) index surgery performed by our group. Exclusion criteria included 1) infection, 2) instability requiring fusion, 3) malignancy, and 4) inadequate documentation.
Surgical Technique: Each patient underwent laminotomy or laminectomy and discectomy for lumbar disc herniation by our senior authors (HHB,
NUA, and UMA). All of our authors perform a subtotal discectomy with removal of extruded fragments and loose material within the disc space. A box shaped incision is created with a scalpel in the annulus and disc followed by debridement of the disc space with pituitary rongeurs and curettes.
In this review, all HNP and anular morphologies were classified by a chart review process. Operative notes, clinical documents, and radiology reports were evaluated by two independent reviewers
(GDW and NUA) in order to classify disc herniations. Differing opinions were settled in conference by a 3 rd reviewer (CLD). Cases with inadequate documentation were excluded. Each disc was classified as type I - IV based on the system of Carragee (Table I). Rates of reherniation in each class were calculated and then compared to established norms
(Table II). Fisher’s exact test was used to compare our proportion of patients with reherniation and/or reoperation to
Carragee’s series in which only fragment excision was performed (Table III).
A paired student t test was used to compare difference in mean age between groups.
Results
259 patients were identified that satisfied the study design (Table II).
We classified 61 (23.5%) into Carragee type I, 60 (23.1%) type II, 52 (20.0%) type III, and 86 (31.7%) type IV discs.
Of 259 cases, 12 (4.5%) reoperations were performed. The indications for reoperation were: lumbar disc reherniations at the same level in nine, postoperative instability in three, and symptomatic DDD (degenerative disc disease) in one. The average time to reherniation was 80.8 months with a range 7-350 months. No significant difference in reoperation rate was noted in any of the groups when compared to Carragee’s series except in Type 2 herniations. This series demonstrated a significantly lower rate of failure and reoperation for Type 2 HNP after subtotal discectomy (3.4%) when compared to fragment excision alone
(21.2%), p<.003. Among all groups, the patients in our series were 13.9 years older (p<.012) on average. Of the 3 cases of postoperative instability, fusions were required due to an unrecognized pars fracture in one, and two cases secondary to osteoarthritis.
Discussion
This investigation is the first retrospective study to apply the
Carragee classification system to a large group of lumbar disc herniations. Our purpose was to use this classification and identify whether disc and anular morphology influence rates of reoperation in the long term after subtotal discectomy. We found a significantly lower reoperation rate among the class II defects (p<.003).
This class (Fragment-Defect) is characterized by disc herniation with a large anular defect and extruded/ sequestrated fragments. We attribute the low reherniation rate in our series to the technique of discectomy and curettage of the disc space.
The Carragee classification succinctly
OrtHOPaeDiC JOUrnaL | Case Western reserve University | vOL.5, nO.1 | 2008 | 85

REHERNIATION ANd fAIluRE fOllOWINg luMbAR dISCECTOMY
86 stratifies disc herniation type based on the presence of extruded or subanular fragments in combination with anular integrity. Since it is based on operative and radiographic findings and has only four types; surgeons find it easy to apply. Moreover, class directs treatment strategy among other surgeons. Anular defects are explored and free fragments are removed. Disc space curettage was not performed. If necessary, large iatrogenic annulotomy was created to allow large disc herniations to be débrided back to the level of the vertebral wall. In our series however, we applied aggressive curettage of the disc space in all cases.
We adopted this 4 part disc classification system in order to evaluate whether anular competence and the presence of subanular fragments prognosticates outcomes in our centers.
Key differences between our use and the original classification relate to patient age, distribution of disc class, inclusion of intraforaminal herniations, and the presence of calcified discs.
With our technique of discectomy, we were able to include intraforaminal herniations in this study. We did not identify any contralateral reherniations so we did not view them as confounders. We did have 3 reoperations for instability (1.1%).
Only one of these 3 index cases included a foraminal herniation. We feel that including these cases reflects an accurate patient sample.
We found our patients to be 13.9 years older (p<.012) than Carragee’s series. This may reflect geographic or institutional selection bias. However, it may also reflect an evolution or transformation of anular competence in these patients as they age. Our distribution of disc class was roughly even (Table II). Whereas, 89 out of 180 of the (49.4%) were type I or fissurefragment in Carragee’s patients. If these patients were to present later, their class could change to reflect the even distribution of our population. The connection between age and disc class warrants further investigation.
We also identified a 5 th morphology not included in the Carragee system. The calcified or hard disc. We assigned these to class IV (no fragment-contained) as the intraoperative and radiographic descriptions were most similar to discs with a competent but hard annulus without free fragments. Contrary to popular belief, we did have 2 cases of calcified discs that reherniated.
We classified these as type IV for the purpose of this analysis. This amounted to half of our type IV reherniations.
During our preliminary analysis, we uncovered 8 cases of microdiscectomies that were referred to us from other centers. 2 of these were also calcified.
We also discovered 20 additional referrals for lumbar disc reherniations.
Five of these cases were also described as calcified or hard discs. Therefore, we would like to propose the hard disc as a
5 th class for future studies.
As we compare the results of this series with established norms, we attribute our low reherniation rates to the difference in technique of discectomy. In the Carragee series a modification of the Spengler technique was used 5 . Cases were performed with an operating microscope. In some patients, the discectomy was performed through the interlaminar space alone.
Otherwise small laminotomies were performed. Medial facetectomy was performed rarely and the midline interspinous ligaments were preserved in all cases. In our series, we employed a more aggressive technique under loupe magnification. The spinous processes and interspinous processes were taken down with bone cutters and
Leksell rongeurs. After developing a plane between the epidural space and ligamentum flavum a laminectomy was performed with Kerrison rongeurs.
We also frequently performed foramenotomies at the same level.
Discectomy was achieved by creating an annulotomy with a # 15 blade followed by removal of disc fragments with pituitary rongeurs and curettes.
These differences in surgical technique remain under investigation and debate. Balderston et al performed a retrospective study on reherniation and reoperation after fragment excision alone versus disc excision and curettage in 83 patients 6 . This study compared the results of two centers of excellence utilizing different techniques. This review found no difference in reherniation or reoperation rate between the two techniques.
However, there was a higher incidence of back pain in the curettage group.
The investigators felt the technique of disc space curettage was unwarranted and could increase surgical risk such as great vessel injury 6 . However, we had no cases of great vessel injury using the curettage technique and demonstrated a significant decrease in reoperation rates especially among type II discs.
Our investigation was not designed to capture the prevalence of back pain after subtotal discectomy. However, prospective data is beginning to emerge that suggests back pain is the trade off to reherniation risk in these patients. A study of 30 patients compared subtotal discectomy to 46 historical controls who underwent limited discectomy. That series’ reherniation rate was reduced from 18%-9% which is in line with established norms. However, back pain and Oswestry scores were worse in the subtotal discectomy group 7 . Rates of reoperation in the long term have been published on the order of 7.3 percent with a third of these stemming from reherniation 8 . In these studies,

manuscripts back pain was more prevalent after subtotal discectomy compared to fragment excision. Nevertheless, our series demonstrates lower reoperation rate (4.6%) across all classes of disc herniation in a large number of patients.
table I. Classification scheme and treatment protocol by Carragee EJ et al. Journal of Bone and Joint Surgery (American)
85:102-108 (2003).
CLASSIFICATION
Disc Fragment
Annulus Defect
Treatment
I. FRAGMENT-
FISSURE
Extruded or sequestrated
Minimal
Removal of fragments through slit-like anular defect
II. FRAGMENT-
DEFECT
Extruded or sequestrated
Large or massive
Removal of fragments through massive defect
III. FRAGMENT-
CONTAINED
Subanular detached fragments
Intact
Oblique Incision; removal of fragments
IV. NO FRAGMENT
CONATINED
No subanular detached fragment
Intact
Extensive annulotomy; piecemeal removal table II. Distribution of disc classification, and anular morphology.
CLASSIFICATION NUMBER OF
SUBJECTS
I. FRAGMENT
FISSURE
II. FRAGMENT
DEFECT
This Study
Carragee et al
259
180
23.5% (61)
49.4% (89)
23.1% (60)
18.3% (33)
III.FRAGMENT
CONTAINED
20.0% (52)
23.3% (42)
IV. NO
FRAGMENT
CONTAINED
31.7%(86)
8.8% (16) table III.
Rates of reoperation after lumbar discectomy based on classification.
CLASSIFICATION
This Study (n=259)
Carragee et al (n=180)
Significance
I. FRAGMENT
FISSURE
1.6% (1)
1.1% (1)
p<.61
II. FRAGMENT
DEFECT
3.3% (2)
21.2% (7)
p< .003
III. FRAGMENT
CONTAINED
9.6% (5)
4.8 % (2) p< .847
table Iv.
Age distribution and lumbar disc classification.
CLASSIFICATION I. FRAGMENT
FISSURE
This Study of 259 cases 53.1
Carragee et al study of
180 cases
37.8
II. FRAGMENT
DEFECT
52.6
37.0
III. FRAGMENT
CONTAINED
45.3
38.7
IV. NO FRAGMENT
CONTAINED
4.6% (4)
6.3% (1) p< .41
IV. NO FRAGMENT
CONTAINED
49.8
31.7
REfERENCES
1 JW Frymoyer, MH Pope, JH Clements, DG
Wilder, B MacPherson, and T Ashikaga
Risk factors in low-back pain. An epidemiological survey
J. Bone Joint Surg. Am., Feb 1983; 65: 213 -
218.
2 Keskimaki, Ilmo MD, DMSc *; Seppo
Seitsalo, MD, PhD,Heikki Osterman, MD, and Pekka Rissanen, PhD. Reoperations After
Lumbar Disc Surgery: A Population-Based
Study of Regional and Interspecialty Variations.
Spine. 25(12):1500-1508, June 15, 2000.
3 Hu RW, Jaglals, AxcellT, Anderson G.
A population-based study of reoperations after back surgery. Spine, 1997 Oct 1;22(19):2265-70.
4 Carragee EJ, Han MY, Suen PW, Kim D.
Clinical outcomes after lumbar discectomy for sciatica: the effects of fragment type and anular competence. J. Bone Joint Surg. Am., 2003
Jan;85-A(1):102-8.
5 Spengler DM, Ouellette EA, Battie M,
Zeh J.
Elective discectomy for herniation of a lumbar disc. Additional experience with an objective method.
J Bone Joint Surg Am,
1990;72: 230-7.
6 Balderston RA, Gilyard GG, Jones AA,
Wiesel SW, Spengler DM, Bigos SJ,
Rothman RH. The treatment of lumbar disc herniation: simple fragment excision versus disc space curettage. J Spinal Disord. 1991
Mar;4(1):22-5.
7 Carragee EJ, Spinnickie AO, Alamin TF,
Paragioudakis S.
A prospective controlled study of limited versus subtotal posterior discectomy: short-term outcomes in patients with herniated lumbar intervertebral discs and large posterior anular defect. Spine. 2006 Mar
15;31(6):653-7.
8 Loupasis GA, Stamos K, Katonis PG, Sapkas
G, Korres DS, Hartofilakidis G.
Seven- to
20-year outcome of lumbar discectomy. Spine.
1999 Nov 15;24(22):2313-7.
OrtHOPaeDiC JOUrnaL | Case Western reserve University | vOL.5, nO.1 | 2008 | 87

88
Tuesday May 27 th – Wednesday May 28 th , 2008
Visiting Professor:
Professor, Orthopaedic Surgery
Washington University School of Medicine
St. Louis, MO
Papers:
Subtrochanteric Femur Fracture Predicted by Implant Location with Respect to Lateral
Cortical Thickness
Frederick Parke Oldenburg, M.D.
Long-term Outcomes After Arthroscopic Lateral Release for Lateral Patellar Compression
Syndrome and Patellofemoral Chondromalacia
Matthew V. Smith, M.D.
richard H. Gelberman, M.D.
Ketorolac Use for Post-operative Pain Management Following Lumbar Decompression Surgery: A
Prospective Randomized Double-blinded Placebo Controlled Trial
Clayton Dean, M.D.
Revision Total Knee Arthroplasty in the Rheumatoid Arthritis Patient
Brian Hardy, M.D.
Prevalence of Spina Bifida Occulta and Its Relationship to Age, Sex, Race and the Sacral Table Angle: An
Anatomic Cadaveric Study of 3,100 Specimen s
Jason Eubanks, M.D.
Revision of Metal-on-metal Hip Resurfacing Arthroplasty
Glenn D. Wera, M.D.

rd
April 24 th – April 25 th , 2008
Visiting Professor:
Professor, Orthopaedic Surgery
Washington University
St. Louis, MO
The Annual Bohlman Lecture was given by Dr. Lawrence Lenke from
Washington University. The Seminar consisted of research presentations by the spine fellows and orthopaedic residents. After discussion of these presentations, Dr. Lenke gave two highly anticipated talks: “Extreme
Makeovers in Treatment of Spinal Deformity” and “The Lenke Classification of Adolescent Idiopathic Scoliosis.”
Lawrence Lenke, M.D.
June 10 th , 2008
Visiting Professor:
Professor, Orthopaedic Surgery
Harvard University
Boston, MA
Dr. Waters was the 2008 RBC Visiting Professor. The itinerary included case presentations including a live patient demonstration as well as multiple talks given by residents. Dr. Waters concluded the seminar with the talk “Neurovascular Injuries of the Upper Extremity.”
Peter Waters, M.D.
OrtHOPaeDiC JOUrnaL | Case Western reserve University | vOL.5, nO.1 | 2008 | 89

90 randall Marcus, James Murphy, Bang Hoang, Dan Master and ed Greenfield
T his years visitng professor was Bang Hoang, M.D. from UC Irivine. The program including presentations given by James Murphy, M.D. and Daniel Master, M.D. on research done during their Allen Fellowship the previous year.

L awrence Samuel Cohen passed away on February 15, 2008 surrounded by his family.
Born in Nashville, TN, he attended West High School and Vanderbilt University’s undergraduate and medical schools. After an orthopaedic residency at Case Western
Reserve University and two years service in the Air Force, he moved to Tampa, Florida.
He practiced there in private practice for 33 years and at the James Haley VA Hospital for six years.
The practice of medicine had been the focus of his life. He served as president of the
Hillsborough County Medical Association, a position of which he was particularly proud.
He is survivied by his wife, Betty, and their children.
Lawrence samuel Cohen, M.D.
P epper Pike orthopaedic surgeon Dr. Curtis W. Smith died July 19, 2008 after collapsing at the Ravenswood Golf Course. He was 60.
Smith was a member of Associates in Orthopaedics, Ohio’s first African-American orthopaedic group practice and one of the few minority orthopaedic surgery groups in the country.
Smith showed residents “how to live their lives professionally and personally,” said Dr. Audley
Mackel, a colleague of Smith’s.
Born in rural Mississippi into a family of educators, Smith entered Tougaloo College at age
14. He went on to earn a pair of master’s degrees, including one from Case Western Reserve
University, and later received his medical degree from CWRU in 1973.
Smith did his internship and residency at University Hospitals and in 1982 went into private practice where he has remained.
Dr. Curtis W. smith
OrtHOPaeDiC JOUrnaL | Case Western reserve University | vOL.5, nO.1 | 2008 | 91

92
Matthew smith, M.D.
sports Medicine fellowship
University of Michigan, ann arbor, Mi
Glenn Wera, M.D.
adult reconstruction fellowship rush University,
Chicago, iL
Brian Hardy, M.D.
Hand/Upper extremity fellowship
University of florida,
Gainesville, fL
Jason eubanks, M.D.
spine fellowship
University of Pittsburgh,
Pittsburgh, Pa
Parke Oldenburg, M.D.
trauma fellowship
Harborview Medical
Center, seattle, Wa not Pictured:
Clayton Dean, M.D.
spine fellowship emory University, atlanta, Ga

shane Hanzlik, M.D.
University of nevada school of Medicine scott kling, M.D.
University of Pennsylvania school of Medicine ethan Lea, M.D.
Case Western reserve University school of Medicine
James Learned, M.D.
University of southern California school of Medicine
Jonathan Macknin, M.D.
University of Pennsylvania school of Medicine
Lorraine stern, M.D.
George Washington University school of Medicine
OrtHOPaeDiC JOUrnaL | Case Western reserve University | vOL.5, nO.1 | 2008 | 93

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Effect of collagen denaturation on the toughness of bone. Clin Orthop. 2000;
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