MDS DISSERTATION PROPOSAL 2012
TITLE
THE
COMPARITIVE EFFECT OF IN-SITU APPLICATION OF
SIMVASTATIN GEL WITH PRF AND PRF ALONE INTHE SURGICAL
MANAGEMENT OF OSSEOUS DEFECTS IN CHRONIC PERIODONTITIS
–A Quasi experimental study.
INVESTIGATOR
DR.ANU.R
MDS Student (2012)
GUIDE:
DR. SEBA A BRAHAM
PROFESSOR
DEPARTMENT OF PERIODONTICS
PMS COLLEGE OF DENTAL SCIENCE & RESEARCH ,TRIVANDRUM
1
CONTENTS
Introduction ................................................................................................................................ 3
Review of Literature: ................................................................................................................. 3
Rationale: ................................................................................................................................... 5
Aims & Objectives ..................................................................................................................... 6
Methodology .............................................................................................................................. 6
Study question…………………………………………………………………………………………………………………………………..6
Study hypothysis………………………………………………………………………………………………………………................6
Study Period ........................................................................................................................... 7
Study Design .......................................................................................................................... 7
Sample Size ............................................................................................................................ 7
Study Subject: ........................................................................................................................ 8
Criteria for Selection of Study Subjects ............................................................................. 8
Procedure .............................................................................................................................. 10
Periodontal status evaluation ............................................................................................ 10
Radiographicevaluation…………………………………………………………………………………………………………………..11
surgical procedure…………………………………………………………………………………………………………………………..11
follow up…………………………………………………………………………………………………………………………………………12
Outcome Measurements........................................................................................................... 12
Statistical Analysis: .................................................................................................................. 13
Ethical Considerations: ............................................................................................................ 13
References ................................................................................................................................ 14
2
INTRODUCTION
Periodontitis, an inflammatory disease occurring adjacent to alveolar
bone, leads to bone resorption, creating bony defects that may terminally cause tooth loss.
Topical applications of biological growth factors may augment bone growth, including the
use of bone morphogenic protein. However protein growth factors are expensive, may
degrade rapidly at the treatment site and could potentially elicit immune responses.
Statins are widely used cholesterol lowering drugs which inhibit cholesterol
biosynthesis by inhibiting Hydroxymethylglutaryl coenzyme A (HMG-COA) reductase, an
important enzyme for mevelonate pathway. These agents are widely used to lower cholesterol
and treat hyperlipidemia and arteriosclerosis. Statins seem to modulate bone formation by
increasing the expression of bone morphogenic protein-2, decreasing inflammation and
increasing angiogenesis.1 Animal studies showed that Simvastatin assists in bone
regeneration and has an anti-inflammatory effect when delivered or applied locally.2 In this
study local application of simvastatin has been chosen to minimize the adverse effects of
systemic uptake like stomach upset, diarrhea, stomach pain etc.
Choukroun platelet-rich fibrin (PRF), a secondgeneration platelet concentrate3,
is defined as an autologous leukocyte and PRF biomaterial. PRF consists of an intimate
assembly of cytokines, glycanic chains, and structural glycoproteins enmeshed within a
slowly polymerized fibrin network. These biochemical components have well known
synergetic effects on healing processes. PRF has been shown to act as suitable scaffold for
breeding human periosteal cells in vitro, which may be suitable for bone tissue engineering
applications.4
The purpose of present study is to evaluate the clinical and radiographic effect of
in-situ application of simvastatin gel along with PRF in surgical management of intrabony
defects in patients with chronic periodontitis.
REVIEW OF LITERATURE:
Animal Studies
Mundy et al.(1999) first reported that statins stimulate in vivo
bone formation in rodents and increase new bone volume in mouse calvaria cell cultures. He
identified that simvastatin may help in periodontal regeneration by inducing BMP-2 and TGF
3
beta in osteoblasts. The findings of their study were comparable to those seen in similar
conditions after direct application of BMP-2 and Fibroblast Growth Factor-1 (FGF-1).5
Goes et.al.(2010) found that Atrovastatin (ATV) reduced alveolar bone
loss by over 47% (p<0.05), when compared to the group of untreated rats and concluded that
ATV was able to prevent alveolar bone loss seen on a ligature-induced periodontitis model.6
Human Studies-Systemic Administration Of Statin
A study examined the association of statin use and clinical markers of chronic
periodontitis and concluded that patients on statin medication exhibit fewer signs of
periodontal inflammatory injury than subjects without the statin regimen.7
A study repoted that statin medication appears to have an effect on the
periodontium that is dependent on the inflammatory condition of the periodontium. The study
was based on a subpopulation of the Health 2000 Survey, which included dentate nondiabetic, non-rheumatic subjects who did not smoke, aged 40-69 years (n=2032).8
In a study reported the effect of Atrovastatin (ATV) treatment on bone loss
prevention in subjects with chronic periodontitis and reported that ATV have beneficial
effects on alveolar bone loss and tooth mobility in subjects with periodontal disease.9
A study reported that relative to the general population,
hyperlipidemic subjects are more prone to periodontal disease and also stated that statins
have a positive impact on periodontal health.10
Animal Studies- on Locally Delivered Statins
In this study devices for sustained or intermittent release of simvastatin
hydroxyacid were formed using a blend of cellulose acetate phthalate and a poly(ethylene
oxide) and poly(propylene oxide) block copolymer, and they were implanted directly over the
calvarium of young male rats. Drug-free devices were used as controls.
After 9, 18, or 28 days, specimens were histologically evaluated for new bone formation.
Intermittent delivery of simvastatin hydroxyacid in rats calvarium resulted in localized
osteogenesis.( Ju Hyeong Jeon et al 2008)11
Local application of statins in healing sites or defects has been shown
effective in new bone formation. Statin/collagen matrix grafts applied to the rabbit’s calvaria
caused expression of BMP-2, vascular endothelial growth factor and core binding factor 1 in
healing bone within 5 days, and 308% more bone than collagen matrix controls.12
4
Human Studies- on Locally Delivered Statins
Currently human studies using locally delivered simovastatin gel in periodontal
defects have been reported.
A study investigated the effectiveness of Simvastatin (SMV), 1.2 mg, in an
Locally delivered SMV provides a comfortable and flexible method not only to improve
clinical parameters but also enhances bone formation. indigenously prepared biodegradable
controlled-release gel as an adjunct to scaling and root planing (SRP) in the treatment of
chronic periodontitis and reported that there was a greater decrease in gingival index and
probing depth and more CAL gain with significant bone fill at sites treated with SRP plus
locally delivered SMV in patients with chronic periodontitis.13
A study showed that combination of allograft with a solution of simvastatin
leads to significantly greater reduction in probing depth, gain in clinical attachment level, and
linear defect fill than when the graft is used alone in the treatment of human periodontal
infrabony defects.(2010)14
A study showed the effectiveness of simvastatin (SMV),1.2% on
indigenouly prepared biodegradable controlled release gel as an adjunct to scaling and root
planing(SRP) in the treatment of chronic periodontitis in type II diabetes patients and
reported that there was more clinical attachment gain with significant intrabony defect fill at
sites treated with SRP and locally delivered simvastatin.15
RATIONALE Inhibition of the enzyme HMG-CoA reductase and the subsequent blockade
of the mevalonate pathway is probably the most important mechanism of inhibition of bone
resorption by statins. The reduction in mevalonate pathway intermediates by statins also
prevent the synthesis of isoprenoid intermediates, farnesyl pyrophosphate(FPP) and geranyl
geranyl pyrophosphate(GGPP). Isoprenoids are lipids attached by post translational
modification
to
some
small
G-proteins
including
Ras
and
Ras
like
proteins
(Rho,Rap,Rab,Ral).These proteins play important roles in cellular proliferation and
differentiation, and, therefore, any perturbation of their activity influences cellular activity.
Thus interference with the generation of isoprenoids leads to disruption of vesicular fusion
and ruffled border formation of osteoclasts, which are essential for their bone resorbing
activity. As a result, osteoclast inactivation occurs and bone resorption is inhibited. The role
of inhibition of mevalonate pathway is further elucidated by the finding that the effects of
statins on bone are inhibited or even reversed by products of this pathway. Local stimulation
of Bone Morphogenic Protein (BMP-2), a major bone growth regulatory factor, can lead to
5
new bone formation. Mundy et al. (1999) identified that lovastatin, and simvastatin,
mevastatin, and fluvastatin increased gene expression for BMP-2 in osteoblasts. The findings
of their study were comparable to those seen in similar conditions after direct application of
BMP-2 and Fibroblast Growth Factor-1 (FGF- 1). There was also a striking increase in
osteoblast cell numbers after statin application. Additionally, it has been observed that statins
like simvastatin, atorvastatin, and cerivastatin markedly enhance gene expression for vascular
endothelial growth factor (VEGF) in MC3T3-E1 cells (preosteoblastic murine cells).VEGF, a
bone anabolic factor, in osteoblasts regulate osteoblast function by increasing the expression
of bone sialoprotein (BSP), osteocalcin (OCN), and type I collagen, as well as suppressing
the gene expression of collagenases such as MMP- 1and MMP-13.
OBJECTIVES
1) Evaluate the periodontal tissue response to in-situ application of simvastatin gel
along with PRF and PRF alone in the surgically debrided intrabony osseous
defects in chronic periodontitis cases by recording clinical parameters (Probing
depth, Clinical attachment level, recession).
2) Assess the effect of in-situ application of simvastatin gel along with PRF and
PRF alone in the surgically debrided intrabony osseous defects in chronic
periodontitis cases by recording radiographic parameters over a period of 9
months.
3) Compare the periodontal tissue response and radiographic parameters to in-situ
application of simvastatin gel along with PRF and PRF alone in the surgically
debrided intrabony osseous defects in chronic periodontitis cases.
METHODOLOGY
STUDY QUESTION : Is the efficacy
of PRF with simvastatin gel more than PRF alone in the
surgical management of osseous defects in chronic periodontitis
HYPOTHESIS: (null hypothesis): There is no significant difference in the efficacy of PRF
with simvastatin gel compared to PRF alone in the surgical management of osseous defects
in chronic periodontitis.
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ALTERNATE HYPOTHYSIS: the efficacy of prf with simvastatin is more than prf alone in the
surgical management of osseous defects in chronic periodontitis.
STUDY SETTING:
Tertiary care setting
THE STUDY POPULATION: Will consist of subjects diagnosed with generalized
or
localized chronic periodontitis belonging to both the sexes. All the subjects will be selected
from patients referred to the department of periodontics in
PMS
dental
college
,Vattapara,thiruvanathapuram, India, during a period of 1year, after obtaining a written
informed consent from the subjects selected for the study
STUDY PERIOD:One and half year.
STUDY DESIGN
: Quasi experimental study.
SAMPLE SIZE: Sample size is calculated using the formula
n = 2σ2
(Zα +Zβ)2
δ2
Where ‘n’ is the minimum sample size required
Zα = type 1 error
Zβ = type 2 error
σ2 = standard deviation of difference
δ= difference of mean of difference
As per the formula the sample size calculated as 22.as we expect some dropouts we are
setting the sample size as 30.
Sample size – 30 subjects contributing 60 sites
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Sampling Method –Quasi design
STUDY SUBJECT:
Subjects selected are both male & female patients who come to Department of
periodontics aged between 20 and 60 years.
CRITERIA FOR SELECTION OF STUDY SUBJECTS
INCLUSION CRITERIA:

Systemically healthy patient diagnosed with generalised or localized (more
than two sites) chronic periodontitis with bilateral intrabony defects

Clinical probing pocket depth ≥ 5 mm in 2 sites .

Radiographic evidence of bilateral intrabony defects in any 2 region.
EXCLUSION CRITERIA:

History of antibiotics in last 2 months

History of periodontal therapy in last 6 months

Suspected allergy to statin group

Patients on systemic statin therapy

Aggressive periodontitis

Tooth with grade 3 mobility

Current smokers

Immunocompromised patient

Pregnant and lactating females

Blood disorders

Any other contraindication for periodontal surgery

Patients indicated for multiple extractions/undergone multiple extractions

Diabetes

Any systemic disorders.

Patients with dental infections like chronic periapical lesions,Apthous
stomatitis,Oral lichenplanus
PROCEDURE

Detailed case history of the patients will be recorded.

Periodontal status will be assessed using
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 plaque index (Sillness & Loe, 1963).
 Gingival index- Loe H and Silness J in 1963,
 Mobility,
 Furcation involvement,
 Pocket depth,
 Loss of attachment ,
 Gingival recession.

All
clinical parameters were standardized by measuring with Williams graduated
periodontal probe.

Informed consent of the patient will be taken.
A total of 60 sites will be selected from 30 patients with chronic periodontitis in the age
group of 20 to 60 years and probing pocket depth of ≥ 5mm with radiographic evidence of
intrabony defects in more than two sites.
All patients recruited for the study will receive oral hygiene instructions and full
mouth scaling and root planing prior to surgical treatment and Re-evaluation is done .
Clinical and radiographic parameters will be recorded at baseline, three, six, and nine months
post surgery. Clinical periodontal probing will be done by Williams graduated periodontal
probe using customized acrylic stents with guiding grooves for reproducible probing sites and
direction

Occlusal stents for positioning measuring probes will be fabricated with cold cure
acrylic resin on a cast/model obtained from alginate impression. The stent will be
made to cover the occlusal surfaces of the tooth being treated and the occlusal
surfaces of at least one tooth in the mesial and distal directions. Stents will also extend
apically on the buccal and lingual surfaces to cover the coronal 1/3rd of the tooth
involved. Vertical grooves will be placed so that measurements made post surgically
could be at the same position and angulations as those made prior to surgery. The
lower margin of the occlusal stent will be taken as the fixed reference point.
Radiographic Evaluation of Intrabony Defects :
Bone fill and alveolar crest height will be evaluated at baseline, three, six
,and nine months post surgically using Intraoral periapical radiographs. Radiographic film
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with grid will be used in this study . The following landmarks will be marked on the image
of the radiograph
Cemento-enamel junction

Alveolar crest

Base of defect
Surgical procedure:Selected sites will be randomly assigned as either control or experimental. After adequate
anesthesia of surgical site, a full thickness mucoperiosteal flap will be reflected and the
osseous defect will be exposed. After thorough surgical debridement, 0.1 ml of simvastatin
gel (1.2mg) with PRF will be placed at the osseous defect on the experimental site and on the
control site only PRF will be placed. Following this, the flap will be repositioned and
secured in place by using 3-0 black silk sutures.
PERIODONTAL STATUS EVALUATION
Periodontal status will be examined before surgery . 3 months after first clinical
examination then follow up in 6th and 9th
. It includes plaque index (Silness and
Loe1963)and gingival index (Loe and Sillness H 1963).Full mouth periodontal examination
will be done which comprises measurement of bleeding on probing,pocket depth, loss of
attachment, gingival recession, furcation involvement, mobility.
Plaque index
The plaque in the gingival margin was evaluated using the plaque index (PI) reported
by Silness and Löe. The PIs were recorded on four tooth surfaces (mesial, distal, buccal, and
lingual). The scores for the PI are:
SCORE
CRITERIA
0
No plaque in the gingival area
1
A film of plaque adhering to the free gingival
margin and adjacent area of the tooth. The
plaque may be recognized
by running a
probe across the tooth surface
2
Moderate accumulation of soft deposits
within the gingival pocket and on the
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gingival
margin
and/or
adjacent
tooth
surface that can be seen by the naked eye
3
Abundance of soft matter within the gingival
pocket and/or on the gingival margin and
adjacent tooth surface
Gingival index- Loe H and Silness J in 1963
SCORE
CRITERIA
0
Absence of inflammation /normal gingival
Mild inflammation slight change in
1
colour, slight edema,no bleeding on
probing
Moderate inflammation, moderate glazing
2
redness edema and hypertrophy, bleeding
on probing
Severe inflammation, marked redness and
3
hypertrophy
ulceration.
Tendency
to
spontaneous bleeding
Total score=Total scores around each tooth /number of surface examined x 4
Gingival index score is interpreted as:
0.0-normal
0.1-1.0-mild gingivitis
1.1-2.0-moderategingivitis
2.1-3.0-severe gingivitis
Pocket depth:is measured from the crest of the marginal gingiva to the probable depth of the
pocket.
Furcation involvement classified according to Glickman in 1953 as:
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Grade 1- pocket formation into the flute of the furcation but interradicular bone is intact
Grade 2- loss of interradicular bone and pocket formation of various depths into the furcation
but not completely probable to the opposite side of the tooth
Grade 3 –complete loss of interradicular bone with pocket formation that is completely
probable to the opposite side of the tooth
Grade 4- loss of attachment and recession that has made the entire furcation clinically visible.
Mobility classified as:
Degree 1: mobility of crown of the tooth 0.2-1 mm in horizontal direction
Degree 2: mobility of crown of the tooth exceeding 1mm in horizontal direction
Degree 3: mobility of crown of the tooth in vertical direction as well.
Gingival Recession is measured in millimetres as the distance from thecemento-enamel
junction (CEJ) to gingival margin.
Loss of attachment:is defined as the distance from the cemento-enamel junction (CEJ) to the
base of the pocket.
.
Formulation of 1.2% Simvastatin gel:Carbopol 934 P containing 1.2 mg/0.1ml Simvastatin gel will be prepared by using
1.2 gm (1200 mg) simvastatin in 100 ml of distilled water. 3% w/v of Carbopol will be used
in formulation of gel.
FOLLOW UP :
Follow up will be done at 3, 6,and 9, months respectively for measuring the
clinical and radiographic parameters.
OUTCOME MEASUREMENTS
Primary outcome: Plaque index , probing pocket depth, clinical attachment level (CAL)` ,
Recession,.(in mms)
Secondary outcome: radiographic evaluation of bone fill.
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STASTICAL ANALYSIS
(intention to treate analysis will be done)
All the parameters will be evaluated at 3, 6, and 9, months period and compared
with base line data using paired t- test.
Efficacy will be assed using paired t test as the comparison is between two sites of the same
patient.
SIGNIFICANCE Level will be fixed at 5%
ETHICAL CONSIDERATION:
Patient’s informed consent will be taken prior to the study. Patient will be given
written information either in English or Malayalam regarding the procedures. Patient will
have the freedom to withdraw at any point during study. Reasons for withdrawal will be
explored.
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REFERENCES
1) Mundy G, Garrett R, Harris S, Chan j, Chan D, Rossini G, et al. Stimulation of bone
formation in vitro and in rodents by statins. Science 1999;286:1946-1949
2) Stein D, Lee Y, Schmid M J, Killpack B, Genrich M A, Narayana N,et al. Local
simvastatin effects on mandibular bone growth and inflammation. J periodontol
2005;76:1861-1870.
3) Dohan DM, Choukroun J, Diss A, et al. Platelet-rich fibrin (PRF): A second-generation
platelet concentrate. Part I: Technological concepts and evolution. Oral Surg Oral Med Oral
Pathol Oral Radiol Endod 2006;101:e37-e44.
4) Three-Dimensional Architecture and Cell Composition of a Choukroun’s Platelet-Rich
Fibrin Clot and Membrane David M. Dohan Ehrenfest, Marco Del Corso, Antoine Diss,
Jaafar Mouhyi, and Jean-Baptiste CharrierJ Periodontol • April 2010;546-554
5) Mundy G, Garrett R, Harris S, Chan J, Chen D,Rossini G, Boyce B, Zhao M, Gutierrez G.
Stimulation of bone formation in vitro and in rodents by statins. Science, 1999; 286: 19461949.
6) Goes P, Lima AP, Melo IM, Rêgo RO, Lima V. Effect of Atorvastatin in radiographic
density on alveolar bone loss in wistar rats. Braz Dent J. 2010;21(3):193-8.
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periodontal lesions: A retrospective study. BMC Oral Health. May 2008 ; 15;8:16.
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chronic periodontitis: a randomized pilot study. J Clin Periodontol. Nov 2010; 37(11):101622.
10) Sangwan A, Tewari S, Singh H, Sharma RK, Narula SC.Periodontal status and
hyperlipidemia-J Periodontol 2011 April 2.
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11) Ju Hyeong Jeon, Ward T. Piepgrass, Yi-Ling Lin, Mark V. Thomas. Localized
Intermittent Delivery of Simvastatin Hydroxyacid Stimulates Bone Formation in Rats. J
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Maxillofac Surg. 2005; 43:46–50.
13) Pradeep AR, Thorat MS.Clinical effect of subgingivallydelivered simvastatin in the
treatment of patients with chronic periodontitis: a randomized clinical trial.JPeriodontol. Feb
2010; 81(2):214-222.
14)Kinra P, Gupta H, Khan S. Evaluation of the Relative Efficacy of an Allograft used alone
and that in Combination with Simvastatin in the Treatment of Human Periodontal Infrabony
Defects – A Clinical and Radiological Study.Journal of Taibah University Medical Sciences
2010; 5(2): 75 - 88.
15) Pradeep AR, Rao NS, Bajaj P, Kumari M.Efficacy of Subgingivally Delivered
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