Sam Rhine - Genetic Update Conferences - www.samrhine.com
- GWAS and Stem Cells Revisited Genetics of Common Human Traits and Diseases…..
their Multifactorial / Polygenic Origins
A. NEW ERA in HUMAN GENETICS…..the OLD ERAS…..the TWO BIG SECRETS
BOX 1 - Cytogenetics
and
BOX 2 - Monogenic
‘Genetic Medicine’ - use our knowledge about chromosomal syndromes (>6100)
and single gene disorders (>23,000) to improve the diagnosis
and treatment of our patients
B. CLASSIC GENETIC CLASSROOM: Teaching Genetic Medicine…..SECRET #1
1. BOX 1 - Cytogenetics: Chromosome Syndromes…..Padlocked for centuries - 1956!
a.
b.
c.
d.
e.
Down Syndrome
Trisomy 21
(1/800)
Patau Syndrome
Trisomy 13
(1/10,000)
Edwards Syndrome
Trisomy 18
(1/6000)
Klinefelter Syndrome
XXY
(1/500 males)
Turner Syndrome
Monosomy X
(1/5000 females)
EASY to Understand - via Karyotype - but most syndromes are very RARE
2. BOX 2 - Mendelian: Monogenic Traits…..Padlocked for Centuries - 1865 / 1903!
a.
b.
c.
d.
AR - Sickle Cell Anemia / Cystic Fibrosis / Tay-Sachs Disease
AD - Huntington Disease / Neurofibromatosis / Marfan Syndrome
XL - Fragile X Syndrome / Hemophilia / Duchene Muscular Dystrophy
AR, AD, XLR or MITO - Retinitis Pigmentosa
EASY to Understand - via Punnett Square - but most conditions are very RARE
3. Why don't you genetics people study the genetics of…..Common Conditions:
Adult Type 2 Diabetes / Macular Degeneration / Hypertension
High Cholesterol / Heart Attack & Stroke / Tumors & Cancer
Bi-Polar Disorder (Manic/Depressive) / ADHD - Attention Deficit / Alzheimer
Alcohol Dependency / Autism Spectrum - ASD / Epilepsy / Asthma / etc.
…..COMMON DISEASES…..but COMPLEX GENETIC TRAITS
4. Why don't we study the genetics of …..Autoimmune Diseases:
Type 1 Diabetes (insulin dependent) / Rheumatoid Arthritis / Multiple Sclerosis
Lupus Erythematosis / Scleroderma / Crohn's Dx / Grave's Dx / Psoriasis
…..COMMON DISEASES…..but COMPLEX GENETIC TRAITS
C. SECRET #2: There has always been a THIRD BOX…..that was ignored for many decades!
Padlocked for centuries…..April 25, 2003 - Human Genome Project Completed
1. MULTIFACTORIAL - Box 3 - Complex Genetic Traits - COMMON DISEASES
the KEY to the New Era of Human Genetics…..the HUMAN GENOME
the Most Important Box! - the ‘New Era’ in human Genetics
2. GENOME: The sum total of all the genetic material for any biologic organism,
may be DNA or RNA, expressed as the total number of nucleotides.
Large genome: Human (DNA) > 3,000,000,000 nts / ~35,000 genes
Small genome: HIV virus (RNA)
9,749 nts /
9 genes
Loblolly Pine Tree - >23,000,000,000 nucleotides
3. NEW ERA in HUMAN GENETICS: ‘Genomic Medicine’
Use our knowledge of the entire GENOME plus ENVIRONMENTAL factors
in POPULATIONS of people…..to understand COMMON MEDICAL conditions
a. ‘Genetics of Common Diseases’
b. NO 'Normal'…..no normal human genome
c. 'Variants' - millions of differences among individuals in a population
SNPs - small variants and
CNPs - large variants
d. 'Mutation' - DNA variant that is pathologic - causes genetic disease or cancer
"We are all Mutants" - ~60 new mutations per generation
Person to Person Genomic Sequence - 99.6% identical DNA Sequences
0.4% different DNA Sequences
0.4% different = ~24,000,000 nucleotides variants in the genome
~24,000,000 variants in DNA sequence between any two people.
4. 'COMMON DISEASES / COMMON VARIANTS HYPOTHESIS'
a. Mid 1990s - Hypothesis to try to explain Common Diseases
b. Persons with the same COMMON DISEASE…..
would have a unique set of DNA VARIANTS in COMMON
c. Variant = DNA change found in 1% to 5% of the population
d. Quantitative Human Traits: Traits that are easily measured - easily quantified
Continuous Distribution in a Population
Most Common Diseases are Part of a Quantitative Trait
5. MULTIFACTORIAL HUMAN CONDITIONS - Box 3
a. BIOLOGIC BASIS of COMMON DISEASES
b. QUANTATATIVE HUMAN TRAITS –
easily measurable traits - easy to quantify:
continuous distribution in a POPULATION
Height, Weight, Finger Print Ridge Count,
Blood Pressure, Blood Glucose Level,
IQ,
etc.
Human Quantitative Traits in a general population…..
normal distribution - bell shaped curve
Low
Average
High
*COMMON DISEASES are part of a QUANTATIVE TRAIT*
c. VARIABLE EXPRESSION in DISEASES - from very mild to very severe
d. COMMON DISEASES - known to have a GENETIC COMPONENT
run through families
twin studies - compare MZ (identical) v. DZ (fraternal)
no Mendelian inheritance pattern…..AR, AD, XLR - not part of Box 2
chromosomes are fine - not part of Box 1
e. POLYGENES - POLYGENIC - many pairs of gene involved
ie: human height…..predicted to be ~700 polygenes involved
f. ENVIRONMENTAL FACTORS also play a significant role
HERITABILITY: H - the proportion of Phenotype variation for a
particular trait that is strictly due to Genetic
differences in a certain population at a certain time
Height
Cystic Fibrosis
Adult II Diabetes
HIV / AIDS
H
H
H
H
= ~80%
= ~90%
= ~55%
= ~5%
E
E
E
E
=
=
=
=
~20%
~10%
~45%
~95%
6. HUMAN QUANTATIVE TRAITS…..Locating Polygenes on a Chromosome
IF we could find one of the polygenes for a quantitative trait
a. locate it's QTL - Quantitative Trait Locus - e.g. 6p24.2
(chromosome #6, short arm (p), band region 2, subregion 4.2)
b. then check the genes at 6p24.2 from human genome map database
c. relate that polygene to the gene function - what does that gene do?
d. how could that gene contribute to that quantitative trait?
7. Flipping Pennies Model…..within a Normal Curve in a Population
a. each penny represents one polygene for a quantitative trait - height
ten polygenes (actually ~700) …..five from Mom and five from Dad
b. Height: Head = Tall polygene / Tail
= Short polygene
c. Polygenes - Additive or Cumulative - NO Dominant & Recessive
d. Reassortment - the mixing of genes and genetic material from the parents
into new combinations of genes in their offspring
d. This Means: Two parents of average height…..
WILL USUALLY have an average height children - most likely - near the Mean
BUT…..two average height parents can have a very tall child
OR……two average height parents can have a very short child
WHY?…..Genetic Reassortment:
that is how polygenes work…..all the polygenes reassort every generation
yielding potentially thousands of new and unique possibilities
e. Polygenes and Medical Conditions - Add THRESHOLD effect to the curve:
Blood Pressure:
Most people have average blood pressure
High Blood Pressure - cross threshold - Cardiac Risk
Blood Glucose Level:
Most people have average blood glucose levels
High Blood Glucose - cross the threshold - Diabetes
Neuronal Synaptic Pruning in Early Development:
Most people have the average number of synaptic connections
Less Pruning = Excessive Connections - cross threshold - Autism (ASD)
Extra Pruning = Fewer Connections - cross opposite threshold
f. TWO perfectly NORMAL parents…..from the middle of the normal curve
Can have an AFFECTED child at one extreme or the other!!
Because of the polygenes Reassortment every generation
D. QUANTATIVE TRAITS and POLYGENES in COMMON DISEASES:
What we do NOT know…..
a. How many polygenes for a particular common disease or trait?
b. Where are the polygenes located - their QTLs = chromosomal address
c. What do those genes do - what is the function of that one gene?
d. One day in the future - PREVENT that COMMON DISEASE!
…..Modify the polygenes in such a way so that we could
…..Nobody ever again Crosses the Threshold!
E. What would it take to find the Polygenes for Common Diseases?
1. HGP - Human Genome Project - Director: Dr. Francis Collins
Largest ever Scientific Endeavor!
15 Year International Cooperative effort - 20 Countries - mainly US + UK
Began: October 1, 1990 - projected completion date: Sept. 30, 2005
Actual Completion Date: April 25, 2003 - Watson / Crick Anniversary 1953
How many human genes? 100,000 predicted > 23,000 coding genes found
Plus the addresses of all the gene loci
>3,000,000,000 nucleotide pairs
~1.5% Coding DNA / ~98.5% Non-Coding DNA
'Rough Draft' - June 26, 2000 - US / British joint announcement
1600 Pennsylvania Avenue in DC / 10 Downing Street in London
Human DNA - we are all 99.6% Identical DNA Sequence…..0.4% different
Same among and between all international people groups
No DNA basis for the term 'Race'
Rough Draft Published: Nature - February 15, 2001 / Science - February 16, 2001
"To determine our DNA Sequence is to achieve an historic step
forward in Human Knowledge"
Genome Magazine - FREE subscription - http://genomemag.com
2. SNPs and CNPs
a. SNPs - Single Nucleotide Polymorphisms = SNVs - Single Nucleotide Variant
"snip" - found in >1% of the population = common
how many SNPs in the average person? ~3,750,000
b. CNPs - Copy Number Polymorphisms = CNVs - Copy Number Variants
"cnip" - found in >1% of the population = common
c. We are all 99.6% identical in our DNA sequences
d. Differences that cause common diseases - must be in 0.4% where we are different
SNPs = SNVs - Small Scale variants - ~80% of the 0.4%
Change one nucleotide…..A > T; G > C
CNPs = CNVs - Large Scale variants - ~20% of the 0.4%
Add or Lose 1 kilobase up to 5 megabases - add or lose a gene
2 copies of a gene > 1 copy or 2 copies of a gene becomes > 3
SNPs & CNPs
- Common - Normal Variants in the population
e. How do we find the SNPs? Search for groups of SNPs - Haplotypes
Haplotype - Sets of nearby SNPs - located close together on a chromosome
HAPMAP - Haplotype Map - find the addresses of the SNP loci
Hapmap I
2005 - 1,000,000 most common SNPs
Hapmap II
2006 - 10,000,000 SNPs
Hapmap III 2010 - 1,440,616 SNPs for detailed studies
Makes Possible: Routine, Affordable SNP & CNP testing
Average Person: 1 SNP every ~800 bp = ~3,750,000
f. HYPOTHESIS: COMMON DISEASES / COMMON VARIANTS…..OR
COMMON DISEASES / COMMON SNPs
F. GWAS - GenomeWide Association Studies
1. Evaluate the entire genome of thousands of people, in a population, all at one time
2. See if any unique set of SNPs might be Associated with a particular Common Disease
3. GWAS Example: T2D = Type 2 Diabetes - usually adult onset
a.
b.
c.
d.
Experimental Group: 2500 people - medically confirmed DO have T2D
Control Group: 2500 persons - medically confirmed - DO NOT have T2D
Scan their 5,000 genomes and check to see which SNPs variants are present
Question? Do the 2500 persons with T2D have a unique set of SNPs
in Common that persons without T2D DO NOT HAVE? YES
e. AND…..we know the QTLs of those unique SNPS…..
one unique T2D SNP is located at 10p12.3
one of the polygenes for T2D must be located at 10p12.3 - Candidate Gene
f. go to Human Genome Map - to 10p12.3 - see which genes are there
g. attempt to link that SNP to a known gene Function
4. We have TESTED the CD/CV HYPOTHESIS and it is VALID
COMMON DISEASE / COMMON VARIANTS HYPOTHESIS = VALID
G. GENOMICS of COMMMON DISEASES - MIT & Harvard - Sept 6 - 9, 2008
'The Beginning of the New Genetics'
GWAS STUDIES:
1. T2D - Adult Onset Type 2 Diabetes
a. 18 SNPs = 18 Polygenes - 18 QTLs / today = 53
b. QTLs connected to glucose metabolism pathway genes…..300 total
2. T1D - Juvenile Onset Type 1 Diabetes - autoimmune disease
a. 18 previous SNPs + 14 new SNPs = 32 Polygenes / QTLs / today = 45
b. QTLs connected to HLA - immune system genes
3. Inflammatory Bowel Disease - Crohn's and Ulcerative Colitis
a. Crohn's Disease - 32 SNPs - 32 QTLs / today = 71
b. Ulcerative Colitis - 10 SNPs - 10 QTLs / today = 47
4. GIANT Studies - Genomic Investigation of ANthropormorphic Traits
GWAS for BMI, Obesity, Height, Weight, Adiposity
a. Obesity - 6 SNPs = 6QTLs - BMI and Risk of Obesity Genes / today = 34
9 SNPs = 15 QTLs - Hypothalmic Weight Control Axis in brain
Obesity is a polarizing polygenic trait with thin habitus
b. Height - H = 80% - 180 SNPs = 180 QTLs - only 20% of H - 700 total?
Single Gene Mutation - override 700 polygenes effect - achondroplasia
5. Blood Lipids - major indicator of heart disease
100,000 persons tested - 95 distinct SNP and CNP variants
59 new - never found before
6. SNPs and I.Q. - g = general cognitive ability - Scientific American - Oct 2008
7. SNPs and Human Personality Traits:
Neuroticism / Extraversion / Openness /
Agreeableness / Conscientiousness / Music Ability / Leadership Skills
Leadership Skills……….All Human Behaviors polygenic/multifactorial??
8. SNPs and Human Facial Appearance - VisiGen - International Visible Trait Genetics
a. 5 polygenes found that affect human facial shape
b. 24 polygenes for eye and hair color - Forensic - facial ID from blood SNPs
c. Hope to identify victims / perps face from DNA in blood samples at crime scenes
9. Five major psychiatric disorders with shared risk effects; 60,000 person GWAS
a. ASD - Autism Spectrum Disorders; b. Schizophrenia;
c. Bipolar Disorder; d. Major Depression; e. ADHD
These five conditions have a group shared SNPs…..
2 SNPs were for genes affecting calcium channels – might lead to therapy?
10. GWAS - Big Questions…..
a. ONLY ~12% of GWAS SNPs are located in coding DNA gene regions
b. ~40% of GWAS SNPs are in non-coding introns
c. ~49% of GWAS SNPs are in intergenic regions
d. may indicate intronic or intergenic gene control elements
e. "Missing Heritability"
H. CNPs = CNVs - Large Scale variants - ~20% of the 0.4%
Add a gene or Lose a Gene: Add or Lose from 1 kilobase up to 5megabases of DNA
1. Normal - 2 copies of a gene / CNP = 1 gene copy or 3 gene copies, etc.
2. Large Scale Variation - add or lose a whole gene or genes
3. INDELS: INsertions / DELetions or Rearrangements
a. MAY be INHERITED…..passed from the parents , or
b. MAY arise DE NOVO…..mistake in meiosis - not passed from parents
‘Genetic but not Inherited’
c. Copy Number Variant - three instead of two, or one instead of two
d. Most Common Example - Down Syndrome - all genes on chromosome #21 - x3
4. Epilepsy / HIV / Heart / Schizophrenia / Tourette / Obesity / Autism
I. AUTISM EXAMPLE - Autism Spectrum Disorders
1 in 68 children.....1 in 42 males.....Males : Females = 4 : 1
1. Spectrum - from very mild to very severe effects
2. may be Autism only - Simplex Family - one child with autism
Multiplex Family - two or more children w/ autism
3. maybe a Syndrome: Fragile X, Rett, Tuberous Sclerosis, Angelman
4. Many forms of Autism = Many different causes
Neurodevelopmental disorder characterized by:
Social Impairments, Cognitive Impairments
Communications Difficulties, Repetitive Behaviors
Can occur in any ethnic, socioeconomic and age groups.
some children with autism appear normal before age 1 or 2 then suddenly 'regress' and
lose language skills they had previously gained - this is the ‘Regressive’ type of autism.
1
2
3
GWAS RESULTS: GWAS Trios - normal mother and normal father with autistic child
1.
2.
3.
4.
Children with autism carry a higher load of rare CNVs
Some inherited from parents as polygenes - others having arisen de novo
de novo CNVs account for 5-8% of simplex cases - paternal age effect
Recent Information - perhaps the main cause of ASD are “Ultra-Rare” mutations of
genes that are classified as “Vulnerable” genes which harbor an LGD - “Likely Gene
Disruption” - and the LSD mutations can occur de novo – genetic but not inherited.
PNAS - on line, September 23, 2015
5. Similar results Mental Retardation - numerous de novo mutations with Exome Trios
6. Examples of CNVs and Reciprocal Variants, +1 gene v -1 gene - Phenotypic Opposites
Obesity < 1
- Underweight >1
Macrocephaly < 1 - Microcephaly > 1
Autism < 1
- Schizophrenia > 1
MUTATION and HUMAN DISEASE - Science (special issue) - September 25, 2015
J. SNPs (SNVs)and CNPs (CNVs) associated with:
Migraines / Fatty Liver Disease / Glaucoma / Narcolepsy / Alzheimer Disease
Scleroderma / Intracranial Aneurysms / Essential Tremor / Epilepsy
Osteoarthritis / Osteoporosis / Restless Leg Syndrome / Tobacco Dependency
Cerebral Palsy / Attention Deficit Hyperactivity Disorder / Psoriasis
Asthma / Voter Turnout / Criminal Behavior?.....one Indy family…..50 convictions!
K. All GWAS results available at One Web Site: www.genome.gov/gwastudies
L. Early Genetics…..1800's - 19th Century
Beginning in 1900's
Box 3
- Inheritance = BLENDING
Blending of Characters of Mother & Father
- 20th Century - Inheritance = PARTICULATE
Particles called genes - Mendelian Inheritance
- 21st Century - BLENDING of PARTICLES called POLYGENES
from Mother and Father reassort every generation
M. DEPENDENCY STUDIED
1. SAGE
- Study of Addiction Genetics and Environment
2. COGA
- Collaborative Study of Genetics of Alcoholism
3. FSCD
- Family Study of Cocaine Dependence
4. COGEND - Collaborative Genetic Study of Nicotine Dependence
N. TCGA
1.
2.
3.
4.
- THE CANCER GENOME ATLAS - discover all human predisposing SNPs
Prostate Cancer
- 45 predisposition SNPs
Breast Cancer
- 29 predisposition SNPs…..Hereditary Breast & Ovarian Cancer
Colorectal Cancer - 20 predisposition SNPs
Lung Cancer - Tumor cells contain up to 50,000 SNPs…..Cancer Causing Mutations!
O. HUMAN MICROBIOME PROJECT - sum total of all the microbes of human body
“the Other Genome” - Major Environmental Factors
1. 800,000,000,000,000 - Cells in Human Body…..American Academy Microbiology
80,000,000,000,000 - Human Cells…..~220 different types
720,000,000,000,000 - Human Microbiome - Bacteria, Fungi, Protozoa - Live in Us and On US
>2,000 different species - weighs ~2 ½ pounds
2. You think you are ONLY human…..You better think again!
Human Microbiome holds the key to many future medical treatments
3. “Instead of declaring war on microbes, we need to think in the context of microbial
ecosystems within our bodies. Figuring out how to encourage good microbes to thrive,
while eliminating the bad ones will be of increasing future importance.”
“We are all surrounded by our own personal microbial cloud…..24/7”
4. “We are the Sum of Our Parts”
http://www.the-scientist.com//?articles.view/articleNo/43379/title/The-Sum-of-Our-Parts/
5. Personal Microbiome Evaluation - $99:
* http://humanfoodproject.com/americangut *
6. MULTICACTORIAL = GENOME + ENVIRONMENTAL EFFECTS
P. OPEN TREE of LIFE: the first comprehensive genomic tree of life
You can check how closely related you are to any other biologic organism
http://tree.opentreeoflife.org/
Q. THE '1000 GENOME PROJECT' - began in January 2008 - http://www.1000genomes.org/
1.
2.
3.
4.
Collaboration among US, UK, China, Germany and nine other countries
Produce an extensive international catalogue of human genetic variation - SNPs & CNPs
Goal was to sequence at least 1000 persons to provide a resource for almost all human variants
Evaluated 2,504 total genomes from 26 people groups from six continents - form the basis for
ethnic group comparisons and tracking people groups
5. Project Completed - Nature (cover article) October 1, 2015
R. YOUR PERSONAL TOTAL GENOME EVALUATION
1. 'The Language of Life' - book by Dr. Francis Collins - Director of NIH
2. Construct Family History / Pedigree Online via HHS - www.familyhistory.hhs.gov
3. Faster and Cheaper in the Future…..Whole Genome: 2015 - $1000 / 2020 - $100
4. www.23andMe.com - can check anyone today for ~35 medical conditions for $199!
S. YOUR PERSONAL DNA CHIP
1. 'Genomic Era of Medicine' - 'Personalized Medicine' or ‘Precision Medicine’
2. Pharmacogenetics - Personalized / Precision Prescriptions and Cancer Therapy
3. GPP - your personal Genetic Predispositions Profile - your Medical Future
a. Do you want to know? b. Who else should know?
c. Who pays? Insurance? d. Who else should have access?
e. Where will you keep the information? f. Who will explain it?
4. GINA - Genetic Information Nondiscrimination Act passed in 2008
5. Massive Parallel Sequencing - Next Generation Sequencing
Chromosome Evaluation without Karyotyping
6. Prenatal Diagnosis and SNP / CNP evaluation of the unborn?
c f DNA = cell free DNA in mother’s blood at 8 weeks of pregnancy
Your unborn baby is:
a. XY - boy
b. height SNPS - he will be about 6'1"
c. T2D SNPs - ~4.1% chance for adult onset Type 2 Diabetes
d. ~1.9% chance of developing adult rheumatoid arthritis
e. ~2.6% chance of developing adult osteoporosis
f. ~8.1% chance of childhood attention deficit hyperactivity disorder
g. ~4.9% chance of developing hypertension
h. ~0.6% chance of becoming alcohol dependent
i. ~16.2% chance of developing prostate cancer as an adult
j. ~57.7% chance of being gifted musically
k. ~5.9% chance of having an agreeable personality
l. ~39.4% chance of being very conscientious
What if there was ~75% chance of migraines?
What if there was ~80% chance of being autistic?
First Non-Invasive Human Prenatal Genome Evaluation - July 2012
Scientific American - December 2012
Time Magazine - Cover - Dec. 24, 2012
Now - Prenatal Diagnosis / Stem Cell Therapy for Osteogenesis Imperfecta
T. SNP / CNP WHOLE GENOME EVALUATIONS - will soon be routine and cheap!
Over 2,000,000,000 evaluations in the next ten years
The amount of Data Storage will be greater than Twitter and YouTube together
Human Cloning and Stem Cells…..
Current Applications and the Future of Medicine
A. Scientific Literacy…..Become Well Informed - GTFS - Informed Decisions!
B. Cloning: Two Procedures…..
1. Reproductive Cloning
2. Therapeutic Cloning
How do those procedures work?
How are they different?
How are they similar?
What is the end point?
C. Stem Cells: Four Main Types
1.
2.
3
4.
Embryonic Stem Cells (ESCs): 10 - natural existence 10 days
Adult Stem Cells (ASCs): 4 types; first recognized in BM 1961
Cancer Stem Cells (CSCs): mutate to cancer
iPSCs (induced Pluripotent Stem Cells) - 2006 / 2007
- natural
- natural
- natural
- man-made
Nobel Prize in Physiology or Medicine - 2012
What are those cells? When are they available for study and to use?
Where would we find them? Why would we want them? How would we use them?
D. Somatic Cells: ~220 Types
1. Normal Body Cells - Specialized for Normal Body Functions
2. Examples: Skin, Nerve, Gland, Muscle, Liver etc.
E. Why are stem cells called ‘stem’
1.
2.
3.
4.
Word is ‘borrowed’ from the Plants
The Stem of a plant gives rise to all the branches
ESCs give rise to all the branches…..220 Somatic Cells
When and Where? …..initially in the developing human embryo
F. Stem Cell Jargon - Potency
1. Pluripotent - a stem cell with the potential to become
ANY of the 220 specialized cells: 1. ESCs / 4. iPSCs
2. Multipotent - a stem cell with the potential to become
MANY of the 220 specialized human cells: 2. Adult Stem Cells
3. Unique characteristics of Stem Cells
Self Renewal - the unique ability to self renew with unique stem cell mitosis
Mitosis - two different mechanisms
Symmetric Mitosis - the two resulting cells are identical
mitosis results in cell expansion
make identical copies of existing cells
Asymmetric Mitosis - Stem Cells only - two resulting cells are different
produces one cell
- copy of original cell - Self Renewal
brings into existence - one new somatic cell - Differentiation
Asymmetric mitosis a million times…..
> 1 stem / > 1,000,000 somatic cells
G. Stem Cell Categories…..
1. Embryonic Stem Cells - gone in 10 days
DO NOT disappear in 10 days - they transform into…..
2. Adult Stem Cells - four types:
HSCs - Hematopoietic Stem Cells
MSCs - Mesenchymal Stem Cells
ESCs - Endodermal Stem Cells
NSCs - Neural Stem Cells
Adult Stem Cells - in the Embryo and Fetus…..
Produce all the ORIGINAL somatic cells
Today…..in all adults
Produce all the REPLACEMENT somatic cells
Examples:
Adult Stem Cells in bone marrow - HSCs
make all the replacement blood cells: RBCs, WBCs, etc.
Adult Stem Cells in skin
- skin is replaced every 7 days:
Epidermal Stem Cells - make new replacement epidermis
Melanocyte Stem Cells - make new replacement melanocytes
Dermal Stem Cells
- make new replacements dermis
“STEMNESS” - the unique characteristics of Stem Cells
1. Self Renewal - Asymmetric Mitosis
2. Differentiate - produce any oy 220 Somatic Cells
3. Biologic Immortality - Telomerase enzyme maintains
chromosome telomere tips
II. BIOLOGIC BASIS of CLONING and STEM CELL TECHNOLOGY
A. Human Embryology - Applied Embryology
Fetus at 7 weeks - 14 grams, the size of a quarter
Fetus at 8 weeks - size of a silver dollar with 220 specialized cells
B. DIFFERENTIATION - cells become Different 220 ways
Cells become DIFFERENT one from another.
Cells become SPECIALIZED 220 different ways.
Adult Stem Cells Differentiate into 200 Somatic Cells
O
N
E
S
E
T
S
W
A
Y
of
S
S
T
R
E
E
T
S
S
I
G
N
A
L
S
Differentiation:
Amazing process - 220 fetal somatic cells become 'Specialized'…..220 Ways
220 One Way Streets = 220 Sets of Signals
Terminal Differentiation - end point of 220 Streets - 220 Specialized Cells
All Embryology - One Way Street - Basic Tenet of Human Embryology
C. What are the signals? Transcription Factors / Epigenetic Chromatin Regulators
They can Activate or Repress dozens of genes at one time
Transcription Factors - turn transcription ON and OFF
a. attaches to the Promoter - Promoter acts as 'Docking Site'
b. TF Activators - bind to promoter and turns genes ON
c. TF Repressors - bind to promoter and turns gene OFF
d. Enhancers - can also bind to DNA and enhance protein output
microRNA = miRNA - negative regulators - turn genes OFF
a. microRNA, ~22 nts, complementary to and binds to 3' UTR of transcript
b. helps usher RISC (RNA Induced Silencing Complex) to the 3' UTR
c. RISC blocks the ribosome > blocks translation > turns gene off
d. referred to as miRNA 'Gene Silencing'
Epigenetics - 'Epi' means…..upon / on top of / above and beyond
a. Chromatin - epigenetic control mechanism acts on chromatin
1. DNA (Genome) interacting with
2. Histone Proteins…..DNA / Histone Complex
b. Histone Proteins:
Nucleosome - DNA wrapped around the NCP
Nucleosome Fiber - strand of multiple nucleosomes
DNA…..'Never Acts Alone'
c. Gene Control: the Degree of Chromatin Compaction
i. Methylation of Cytosine in the DNA of Chromatin at CpG Islands
add methyl groups - compact = 'Closed' chromatin = OFF
lose methyl groups - loose
= 'Open' chromatin = ON
ii. Acetylation of Histone Proteins = 'Open' chromatin = ON
d. Methylation and Acetylation
Epigenetic Marks on the Chromatin - Remodels the Chromatin
e. Enzyme Controlled Mechanism:
Methylation Enzymes: DNMT - DNA Methyl Transferase
de novo
DNMT-3
maintenance
DNMT-1
Acetylation: HAT - Histone Acetylase / HDAC - Histone DeAcetylase
EPIGENETIC MARKS: Open Chromatin - genes ON
Closed Chromatin - genes OFF
EPIGENOME: Sum total of all the epigenetic marks
E. CELLS, GENES and CHROMATIN - EPIGENETIC MARKS
Liver:
OPEN Liver Genes
CLOSED Kidney, Nerve, and Spleen Genes
Kidney: OPEN Kidney Genes CLOSED Liver, Nerve, and Spleen Genes
Nerve:
OPEN Nerve Genes CLOSED Liver, Kidney and Spleen Genes
Spleen: OPEN Spleen Genes CLOSED Liver, Kidney, and Nerve Genes
220 Cellular Phenotypes
Example - Differentiation Pathway for Pancreatic Beta cells
F. Human Embryome Project - International effort
find all 220 sets of signals - know the signals to make any human cell
then…..any pluripotent stem cell and the appropriate signals…..
would will be able to make any human cell…..in a laboratory
G. Fertilization to Embryonic Stem Cells…..
Fertilization or Conception - egg and sperm unite
Zygote = fertilized egg - size = point of a straight pin
2 cell embryo at ~24 hours - 4 cell embryo at ~48 hours - 32 genes working
8 cell embryo at ~72 hours - only 129 genes working
16 cells > 32 cells > 64 cells
Tohonen, et. al. Nature Communications, September, 2015
Blastocyst at ~ 6 days
'basketball' - size of the point of a pin = implantation stage
inside blastocyst - fluid filled - one small cluster of cells = ICM
ICM - Embryonic Stem Cells - ESCs - becomes fetus / amnion
'Ten Day Rule' - ESCs only exist 10 days under natural conditions
H. Embryonic Stem Cells…..immediately morph into four major branches:
ADULT STEM CELLS:
HSCs - Hematopoetic SCs > forms all blood cells - RBCs, WBCs etc.
MSCs - Mesenchymal SCs > forms muscle, bone, cartilage, etc.
ESCs - Endodermal SCs > forms liver, stomach, lungs, pancreas
NSCs - Neural SCs
> brain, spinal cord, epidermis
I. Adult Stem Cells:
INITIAL function - differentiate the ORIGINAL 220 somatic cells in the fetus
ADULT function - differentiate REPLACEMENT 220 cells the rest of your life
Where are they found today?
HSCs - in bone marrow (and in umbilical cord)
produce replacement blood cells…..~15,000,000 RBC per second
MSCs - in bone marrow, adipose, amniotic fluid
produce replacement muscle, bone, cartilage, tendon, heart,
blood vessels, etc. plus produce paracrine signals for tissue repair
ESCs - in lining of stomach and lining of intestine
produce replacement cells
NSCs - in hippocampus and sub-ventricular zones of the brain
produce replacement neural cells
HSCs and MSCs are easily obtained from bone marrow or adipose
used therapeutically for ~ 60 years - 1st BM transplant was 1959
Classic Terms: Ectoderm = NSCs, Endoderm = ESCs, Mesoderm = MSC & HSCs
III. REPRODUCTIVE CLONING
A. Clone - definition: a genetically identically copy of…..anything you choose
B. Robert Briggs - ‘Father of Cloning’ (December 10, 1911 - March 4, 1983)
first to clone an animal - frogs in 1952 - IU
C. Reproductive Cloning - Definition
reproduce an individual…..create a genetically identical copy
of a particular individual…..clone will be born as a new baby
First Mammalian Clone: Animal - Sheep
Names - Megan & Morag - cloned from donated embryo nucleus
Second Mammalian Clone: DOLLY
First clone from donated adult somatic donor nucleus
D. Reproductive Cloning Procedure: S C N T - Somatic Cell Nuclear Transfer
1. Oocyte Retrieval - obtain egg from the sheep ovary
2. Enucleation - remove the nucleus from egg - via cell surgery
3. SCNT - transfer donor somatic nucleus into the egg
new set of instructions - make a copy of the donor
“the DONOR will be CLONED”
4. Artificial Activation of the egg > 2cell, > 4 cell, > 8 cell embryo…..
5. ET - Embryo Transfer from the petri dish into the uterus of the
surrogate mother - who carries the clone embryo to term
E. Dolly: born July 5, 1996 / died February 14, 2003 - death from virus infection - not due to cloning
1. Ian Wilmut's lab in Roslin, Scotland
2. Dolly today: special exhibit - Royal Museum in Edinburgh
F. END POINT for Reproductive Cloning…..
BABY born to surrogate mother - too risky to try in humans
G. Other Mammalian clones:
Sheep, Mouse, Cattle, Pig, Goat, Gaur & Mouflon (endangered species), Rabbit, Cat,
Mule, Rat, African Wildcat, Dog, Water Buffalo, Horse, Ferret, Wolf, Banteng, Camel,
Wooly Mammoth? - 1. SCNT with Asian elephant as surrogate mother
2. Build a Mammoth / Elephant hybrid with ‘Genome Engineering’
Prometea - first cloned horse - donor was also the surrogate
1/841 = .12% success rate - surrogate delivered her own identical twin
Dolly
- 1/277 = .36% success rate - very inefficient process - not epigenetically identical
H. Most important lesson from Dolly:
1. A highly specialized, terminally differentiated, somatic donor
nucleus…..when placed into an enucleated egg, is reprogrammed
from a highly specialized nucleus - reverts back to a pluripotent nucleus
2. What in the oocyte reprograms the nucleus back to pluripotency?
3. Must be a cytoplasmic factor - because the egg nucleus in gone
4. Educated guess…..it must be SIGNALS in the cytoplasm - how many?
5. WHAT are the SIGNALS - “Stemness Signals”
III. THERAPEUTIC CLONING and STEM CELLS
A. Therapeutic Cloning Procedure: via Somatic Cell Nuclear Transfer
1. Human Oocyte Retrieval - very difficult procedure
2. Enucleation of the egg – surgical removal of nucleus from human oocyte
3. SCNT - transfer donor somatic nucleus to the egg
4. Artificial Activation of the egg > 2 cell, 4 cell, 8 cell…..
5. Embryo continues to grow in lab for 6 days - clone in the petri dish
Clone of my friend’s embryo, remove ECSs from embryo, grow in the lab
Establish a cell line from ESCs, grow until you have ~10,000,000,000 ESCs
Directed Differentiation:
a. give those ESCs the signals to differentiate into Spinal Motor Neurons
b. result: > ~10,000,000,000 Spinal Motor Neurons
genetically identical to my friend
c. the 'DREAM' - those cells to repair his spinal cord - one day he walks again!
B. Therapeutic Cloning = NT (Nuclear Transfer) Cloning
Create a genetic clone (copy) of an individual's embryo;
grow ECSs from the embryo; then use the appropriate signals;
produce any of 220 human somatic cells use for medical therapy
END POINT: a CELL that can be used for medical therapy
REPRODUCTIVE CLONING - END POINT:
BABY born to surrogate mother
THERAPEUTIC (N.T.) CLONING - END POINT:
CELL used for medical therapy
IV. ALTERNATIVE WAYS TO PRODUCE PLURIPOTENT STEMS
“Safe” and "Ethical" Pluripotent Stems: Develop new lab procedures that would yield
pluripotent stem cells but, DOES NOT involve destruction of an embryo
1. Biopsy 8-cell embryo,
2. ANT - Altered Nuclear Transfer,
3. Living cells from 'Dead Embryo', 4. Parthenogenesis
5. REPROGRAMMING
V. REPROGRAMMING = DeDifferentiation
1.
Take one of the 220 Differentiated, Specialized, Somatic Cells
BACKWARDS to a pluripotent state
Differentiation:
Stem Cell becomes Somatic Cell - Differentiation Signals
DeDifferentiation: Somatic Cell becomes Stem Cell - ‘Stemness’ Signals!
2.
How? The lesson learned from Dolly!!
Signals in oocyte cytoplasm - 4 signals - ‘Stemness’ Signals
3. Who? Shinya Yamanaka - Kyoto University in Japan / UCSF
Mouse iPSCs - July 7, 2006
Human iPSCs - November 20, 2007
iPSCs……induced Pluripotent Stem Cells
4 signals: Japan - OCT 3/4, SOX2, c-MYC, KLF4
'Yamanaka Factors' - OSMK Signals
Yamanaka & Gurdon - Nobel Prize – 2012
iPSCs ARE NOT: Adult Stem Cells (HSCs / MSCs / ESCs/ NSCs)
iPSCs ARE:
Adult Somatic Cells converted into Stem Cells
iPSCs and Embryonic Stem Cells are Functionally Equivalent
Nature Biotechnology, On Line, October 26, 2015
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CRISPR - Clustered Regularly Interspaced Short Palindromic Repeats
Bacterial immunity enzyme system that removes a harmful gene and inserts a good
gene in its place, may lead to therapy for R.P., AIDS, Sickle Cell, Hemophilia
CRISPR/Cas9 - TARGETED GENE THERAPY: New Era in Molecular Biology
CRISPR - localizes the target DNA / Cas9 cleaves the DNA
EDIT the HUMAN GENOME…..remove a bad gene and replace it with a good gene!!
Great YouTube Video: https://youtu.be/2pp17E4E-O8
Therapy for CF and SSA:
http://cysticfibrosisnewstoday.com/2015/10/27/vertex-crispr-use-gene-editing-search-new-cystic-fibrosistreatments/
Human Gene Editing Summit.....experts continue to discuss the logistics and ethical
considerations of editing human genomes at a historic meeting in Washington, DC.
http://www.the-scientist.com//?articles.view/articleNo/44686/title/Let-s-TalkHuman-Engineering/
CRISPR Therapy in a Dish - Muscular Dystrophy
CRISPR is best known for its use in gene editing - but scientists have used it boosted levels
of a protein in cells for a genetic disease called Duchenne muscular dystrophy (DMD).
http://www.the-scientist.com//?articles.view/articleNo/44727/title/CRISPR-Therapy-in-a-Dish/
V. POTENTIAL APPLICATIONS of PLURIPOTENT STEM CELLSs:
1. Cell Replacement Therapy: autologous = no rejection
Spinal Motor Neurons - spinal cord injury
Pancreatic Beta Cells to produce insulin - Type I Diabetes
Dopamine producing neurons - Parkinson Disease
Red Blood Cells - Sickle Cell Anemia…..~216 other possibilities
2. Human Disease Modeling: ‘Disease in a Dish’
Follow the cellular development of any genetic disease in vitro…..
Lou Gerhig's Disease (ALS) donor > motor neurons
Huntington Disease donor > medium spiny neurons
Type I Diabetes donor
> pancreatic beta cells
3. Drug Therapy Screening:
Test thousands of small molecules at one time in a petri plate
Find one of 1000s that has a beneficial therapeutic effect
4. Regenerative Medicine / Tissue Engineering:
Produce cells in the laboratory for use for production
human tissues and organs in vitro
5. Combination: #2 - Disease Modeling & #4 - Regenerative Medicine
Cell Replacement Examples:
Cell Stem Cell - July2, 2015
iPSCs from patients with type 1 diabetes - DiPS Diabetic iPS
August 31, 2009
Alginate Capsules protect the iPS Beta Cells from Immune System T-Cells!
Viacyte - human trials underway
http://viacyte.com/
VIDEO: https://www.youtube.com/watch?v=Q6U5kf5ByNE
Acinar > Beta Differentiation - Direct Cellular Reprogramming in vivo
Nature Biotechnology - Vol 32, p. 1223, December 2014
Direct Reprogramming: Somatic Cell > Desired Somatic Cell
No iPSC stage = No Teratoma Risk!
Fibroblasts > Functional Neurons
August 5, 2011
Fibroblasts > Functional Dopaminergic Neurons - August 11, 2011
Fibroblasts > Functional Lung Tissue
Blindness - 25,000,000 World Wide - visually impaired or blind
>190 Genes Mutated to cause blindness
AMD - Age Related Macular Degeneration
iPSCs > RPE - Retina Pigment Epithelium - 99% pure
First Successful Human Trials
Lancet - January 23, 2012
iPSCs > RPE ‘Patches’ - transplanted to back of retina / or RPE cell injections
Stem Cells Portal - October 5, 2015
RP - Retinitis Pigmentosa - Most Common Inherited Vision Loss
AD, AR, XLR, Mito - Rod Cell Loss
Achondroplasia - most common cause of short stature
Neural Applications
Human Brain: brain cells…..100 billion neurons
100 trillion synapses
100 billion astrocytes
human astrocytes - Special LTP (Long-Term Potentiation) Function
glia cells - billions of non-electrical cells in the brain
astrocytes: active at all synaptic junctions
microglia
oligodendrocytes: myelin producing cells
myelin - insulates axons in a protective sheath
Neural SC Engraftment and Re-Myelination in the Human Brain
Pelizaeus-Merzbacher Dx - PMD - hypomyelination of axons
Oligodendrocyte Deficiency - no myelination
huCNS-SCs - human Central Nervous System Stem Cells
 huGPCs - human Glial Progenitor Cells
injected into the PMD brains > form new oligodendrocytes
myelinated the brain axons > functional improvement
Therapy for Demyelinating Diseases…..Multiple Sclerosis?
Science Translational Medicine October 2012
Human Astrocytes - special adaptations for LTP
huNSCs > huGPCs > human astrocytes – injected in newborn mice
‘Smarter’ mice with human astrocytes in their brains
Future huiPSCs > huNSCs > therapy for ASD, Schizophrenia ?
Cell Stem Cell - March 7, 2013
Huntington Disease - AD - CAG Triplet Repeat Mutation – 4p16.3
Gene elongates every generation - Classic ‘Late Onset’ Condition
Huntingtin elongated gene > Huntingtin elongated protein
Huntingtin Protein > Loss of Medium Spiny Neurons in Striatum
Huntington Stem Cell Therapy:
HD Patient Fibroblasts > HD iPSCs
HD iPSCs Genetically Corrected CAG72 > CAG21
Corrected HD iPSCs > Medium Spiny Striatal Neurons
Patient specific, genetically corrected, HD patient iPSCs
Cell Stem Cell - August 3, 2013
ALS - Amyotrophic Lateral Sclerosis = Lou Gerhig’s - Slow loss of motor neurons
‘Neural Progenitors Derived from Human iPSCs Survive and
Differentiate upon Transplantation in a Rat Model of ALS
Stem Cell Translational Medicine - March 2013
Parkinson Disease - Loss of Dopaminergic Neurons in Substantia Nigra of midbrain
‘Rapid Generation of Functional Dopaminergic Neurons from
Human iPSCs through a Single-Step Procedure using Cell
Lineage Transcription Factors: ASCL1 / NURR1 / LMX1A
Stem Cell Translational Medicine - Vol 4, pp. 31-43, December 2014
Reproductive Applications:
Creating Spermatids from Skin - male infertility
Creating Oocytes from Skin
- female infertility
‘Egg Engineers’ - Nature, p.392, August 22, 2013
Oocyte Stem Cells - Women can procreate for ever!
Nature Medicine - March 2012
iPOD Link: http://www.nature.com/news/stem-cells-created-in-living-mice-1.13725#/ref-link-1
Disease Modeling Examples:
“Organoids” - primitive organ-like 3-D masses of differentiated cells grown in matrigel
Rise of the Organoids – Nature, July 30, 2015
Inner Ear Organoid: sensory epithelia via 3-D Culture - IUMC
Nature, August 8, 2013
Retinal Rod Cell Organoids: from iPSCs for Retinitis Pigmentosa Therapy
Nature Biotechnology August, 2013
Eye Organoids: Optic Cup Retina
Nature - April 7, 2011
Liver Organoids: Vascularized and Functional Human Liver from
iPSCs-liver bud transplant into mesentery
Nature - July 23, 2013
Heart Organoids: Cardiac Repair after MI (Myocardial Infarction)
Cardiac Fibroblasts > Cardiomyocytes - 3 signals
Cell - August 6, 2010
Large Intestine Organoid: Study the development stages of colon cancer
Sequential Cancer Mutations, Nature, May 7, 2015
Living Colon Cancer Patient Organoid Biobank - Cell, 161, pp. 933-945, May 7, 2015
Kidney in a Dish…..First Man-Made Mini-Kidney
Human kidney organoid with a full set of renal cell types!
Cover Article - Nature, vol 526, pp. 512 and 564, October 22, 2015
Vocal Cords in a Dish…..First Cells for Potential Vocal Cord Replacement
Science Translational Medicine
The Brain Maker: mouse ESCs > mouse NSCs
Primitive Optic Cup Retina / Cerebral Cortex / Primitive Cerebellum
Primitive Hypothalamus with hormone production
Nature - August 23, 2012
Human Brain Organoids - Minibrains - size of an apple seed
Resembles the Fetal Brain at ~9 Weeks / Microcephaly Brain Organoid - smaller
‘Cerebral organoids model human brain development and microcephaly’
Nature - August 28, 2013
Model for study of ASD - autism using the skin from a child with ASD diagnosis
Model for developing drugs for various brain related conditions
Human 5 Week Fetal Brain in a petri plate made from iPSCs - size of a pencil eraser
99% of normal genes working - contains part of spinal cord and retina
Takes ~12 weeks in vitro to reach the 5 week in vivo stage
May be a model to study experimental drugs and causes of brain disease
May be as a model to study: ASD, Parkinsons, Alzheimers - using their skin
https://news.osu.edu/news/2015/08/18/human-brain-model/
“Organs on a Chip” - Multichannel, 3-D, microfluid, cell culture on a plastic, chip…..
with living vascular cells to mimic the body’s circulatory system
Simulates the basic anatomy and function of entire organs.
can be used to test chemical toxicity, normal cells v. disease
Lung on a Chip
Heart on a Chip
Nephron on a Chip
Artery on a Chip
Bone Marrow on a Chip
Person on a chip
Microfluidic organs-on-chips - Nature Biotechnology, Vol 32, August 2014, pp. 760-770
“IDEAL STEM CELL":
1. Easy Access - from blood or bone marrow
2. Timely Access - obtain cells anytime
3. No Tumor Risks - no teratomas
4. Multipotent or Pluripotent
5. Homing Mechanism - migrates to site of injury or tumor
6. Non-Immunogenic - ImmunoModulatory
7. Paracrine effect - secrete factors to benefit neighboring cells
IMPOSSIBLE! - Adult Stem Cells - Mesenchymal Stem Cells = MSCs
MSCs - Leukemia - Decrease GVHD in BM / UC Transplants
Amniocytes to produce new heart valve before birth
Parental adult MSC stems correct O.I. = osteogenesis imperfecta
Adipose Derived Stem Cells (ADSC) for heart therapy
MCSs from bone marrow and cord blood - therapy for E.B.
E.B. = Epidermolysis Ballosa
http://www.ahc.umn.edu/eb/home.html
How do they work? ‘Home’ to area of injury of inflammation
‘Home’ to tumor and carry drug - interferon beta - to kill tumor
Heart Vein made in vitro - MSCs on man-made tubular scaffold
Science, August 26, 2011
Tracheas made from scratch - MSCs plus man-made scaffold - new scaffolds from 3-D printer
3D Bioprinting of tissues and organs - Nature Biotechnology, August 2014
Cartilage made from scratch - MSCs in seeded on polymer scaffold
Finger Regeneration - cellular matrix powder from a pig bladder and finger nail stem cells
Meat Lab - make burger and sausage from muscle stem cells
First Test-Tube burger made from 3,000 tiny strips of meat grown from muscle
muscle stem cells taken from a cow’s muscle tissue…..cost…..$383,875
#4 Regenerative Medicine Applications
- produce human tissues / organs in the lab
Scaffold = framework - basic shape of that tissue or organ
Make a Heart, Lungs, Liver or Kidney in the Lab? Can it be done?…..heart scaffold is too complex!
Video: http://www.nature.com/news/tissue-engineering-how-to-build-a-heart-1.13327
BUT…..there is a normal, natural heart scaffold - inside every heart
Decellularization - remove all heart cells and endothelial cells with detergent, under
pressure, left with a ‘deflated’ acellular human heart scaffold
Recellularization – in a Bioreactor: add new heart cells and endothelial cells made
ahead of time in the lab, from iPSCs of the recipient
Biomaterials - vol. 52, pp. 103-112, June, 2015
Lungs - Decelluarization and Recellularization
Pig lung formed in a Bioreactor in lab / successfully transplanted back
http://tmcnews.tendenciapp.com/articles/team-at-utmb-seeks-to-rebuild-donated-human-lungs/
Liver
- Decelluarization and Recellularization
Bioengineered transplantable porcine livers – Biomaterials, February, 2015
Kidneys - Decelluarization and Recellularization
Place human replacement cells in pig kidney scaffold
Regeneration of Bioengineered Kidney - Nature Medicine, OL, April 14, 2013
Limbs
- Decelluarization and Recellularization
Detergent removal of all cells, leaving cell free scaffold of blood vessels, tendons and
muscle from a rat…..took several weeks…..then in a bioreactor, supplied the progenitor
cells for replacement cells, even outer skin covering - and the new limbs responded
normally to electric stimuli and the fingers contracted.
http://www.nydailynews.com/life-style/health/watch-mass-researchers-grow-bioengineered
-rat-limb-article-1.2249891
Severed hand grafted to patient’s leg…..then later transferred back to the arm
http://www.cnn.com/2015/07/24/health/severed-hand/index.html?sr=fb072415severedhand1pStoryGalLink
Chimeric Mammalian Embryos - for neo-organ production
Chimera - an organism that is made up of a mixture of cells from two or more organisms
Make a rat pancreas in a mouse embryo…..mouse/rat chimera
Inject ‘blue’ rat iPSCs into mutant mouse embryo - can’t make a pancreas
At birth, the newborn mouse survives because it has a ‘blue’ rat pancreas
In vivo organogenesis - make an organ of one animal in the embryo of another
Make a human pancreas in the embryo of a pig…..pig/human chimera
Inject human iPSCs into mutant pig embryo - can’t make a pancreas
At birth, new newborn pig survives because it has a human pancreas
When pig reaches maturity - transplant pancreas from pig to human iPSC donor
Animal Embryo Chimeras used to produce human organs!
Cell Stem Cell - Vol 15, pp. 406-409, October 2, 2014
What about using Pig Organs - transplanting pig organs into humans??
Two Problems: 1. PERVs - Porcine Endogenous RetroViruses
2. Pig antigen genes that would cause rejection
Both eliminated in pig embryos by CRISPR genome editing - PNAS Conference - October 5, 2015
Future
- Yamanaka developing an iPS Cell Bank in Japan
~75 HLA compatible iPS donors would provide iPS for ~80% of the Japanese population
“When it gets here, when it arrives,,,,,Science Fact is always more amazing than Science Fiction”
Sam Rhine, Oct. 13, 2015
Sam Rhine - College and Career suggestions:
Check Out: http://www.kumc.edu/gec/prof/career.html
1. Go to you favorite Undergraduate college and obtain your 4-year Bachelor's degree.
Major in biology, biochemistry, molecular biology, bio-engineering etc.
Make sure you satisfy the Pre-Med requirements so you can apply to
medical school if you decide that is the best route for you.
2. Medical School is four years and the curriculum is very similar at all
medical schools in the US. The reason for that is that everyone must pass
the same national exam after finishing medical school - therefore the
schools must cover the basic subject matter. If you pass that exam the summer
after finishing medical school, you can then put M.D. behind your name.
3. Residency is then 4 - 8 years of ‘Specialty Training’ to become a pediatrician,
obstetrician, orthopedic surgeon, oncologist, neurosurgeon or whichever
specialty you choose. If you want to pursue a career in Tissue Engineering
then you might want to get a residency with Dr. Anthony Atalla at Wake
Forest University. If you want to use CD-47 antibodies to functionally disable
cancer stem cells you might want to do your residency in oncology at
Stanford University with Dr. Irv Weissman or Dr. Michael Clarke.
Keep your ‘antennae out’ during your four years of medical school – to determine
who is doing the research you want to pursue for a career - and go do your residency
with that person - he or she.
_________________________________________________________________________
2. For those who are not interested in medical school - they might want to
pursue a career in research and they will go on after their undergrad work
and get their Masters and Ph.D. which may be 4 to 6 more years.
3. The Ph.D. is usually followed by Post Doctoral studies, Post Doc, for 2 - 4 years to
gain more special expertise for the research career you want to follow. Then
you will be ready to join the faculty at a university to do research and
teach. Others will opt to get a job doing research in industry for biotech companies.
Also, some of these people are getting their Ph.D. in biostatistics or computer
sciences where they will help with the planning and evaluation of research data.
4. M.D. / Ph.D. Most major Medical Schools offer a combined M.D. / Ph.D. for a person who
may one day be the chairperson of the Department of Molecular Medicine at
some medical college - check that out for each individual medical school.
5. Masters Degree in Genetic Counseling - is another option for some.
There are almost 30 places in the US where those programs are available.
For more information - check out this web site:
http://www.nsgc.org/iframepages/GeneticCounselingTrainingPrograms/tabid/336/Default.aspx
6. Teaching - Also Remember…..many people who will make a major contribution to all
These careers in the future will do so by majoring in Education in college and
will be preparing young people in the future…..as your Teachers have been
preparing you!!
"Teachers Make All Other Professions Happen!"
Also Consider: Physician Assistant (PA): http://www.aapa.org/
Student Academy: http://www.aapa.org/saaapa/
MD/MS Genomic Medicine:
http://admissions.med.miami.edu/md-programs/md-ms-in-genomic- medicine
UPDATED: January 9, 2016