IOC World Congress on Sport Science, Oct, 2003 Genes and Performance Wednesday, 1030am, Chair: Bengt Saltin
Genes and Health Greg collier, AGT Biosciences, LTD, Deakin Uni, International Diabetes Institute and South West foundation Texas
PARL a new gene involved in muscle function and type 2 diabetes
Literature is full of candidate gene studies – not worth the paper their written on
Unique DNA collection
AGT Biosceince cetre for Stat Genomics
Outbread Israeli Sand Rat Colony
Ezpress Technology Platform
AGT Biosceinces has access to number of human DNA sample collections
Access to unique DNA collections from worldwide populations
Major Collaborators
Dr. John Blangero, Texas - analysis of genetics of many complex diseases - devel opf new stat software - SOLAR
Native habitat is semi-arid regions of middle east
Israeli sand Rat develops diabetes and obesity in manner similar to humans
PARL – Preseneilins-associated rhomboid-like protein
7 transmembrane protein
predicted was located in mitochondria – regulating basic muscle function
Located at chromosome 3q27
Reduced gene expression in muscle of diabetic P. obesus
PARL gene expression was increased after exercise-training
Human gene expression data - Eric Ravussin, Anthony Civitarese (Pennington Biomedical Research Center)
Discovered by microarrary….
REF: McQuibban, Nature 423, 534 (2003) - in mitochondria, yeast equivalent…. - if knock gene out, mitochondria shrink and don’t function correctly
PARL cleaves human equivalent of OPA1… - in diabetic muscle, where decreased expression of PARL, this is a problem
Mitochondrial defects in genes occur before diabetes occurs
Dgene expression I skeletial musc asso with
REF: Kissebah, AH PNAS 97, 14478 (2000)
Genotyping 50 individuals, XXX
Association studies
Look at relationship of XXX
Responsible for about 5% of variation in insulin levels
Largest yet genetic variation causing insulin resistance
Discovered in muscle of animals
Leu26Val variant of parl asso with plasma insuli, with a strong genotype-by-age variant
Finding genes like this is not simple
Need to combine human linkage studies with expression
Not a worthwhile task to identify a gene and find an association
Brain health, and voluntary running Frank Booth
Epidemiological reports indicate that physically active elderly humans have less cognitive dystnfunction
Laurin Arch Neurol. 58:498, 2001 - women 65 yars or older, evaluated - highlevel of phys activ corresponsed to exercise
What is the mechanism? How phys activ tie into brain disfunction
Brain derived neuro..factor (BDNF)
Lu Learn, Mem. 2003, 10: 83-85
Wifdenfalk Neurosci Res 34 125: 1999 - animals running more had increase in XXX (i.e.
Located in hippocampus - Hippocampus is highly plastic structure normally asso with cog function, rather than motor
Malcangio Trends Pharmacol Sci 24:116, 2003-10-08
Increase in neurogenesis in hippocampus in animals that are running, compared to animals who are not running
Running primes t brain to enhance neuronal health
It is dogma in medicine that health indivs are the control group and t sick patients are treatment group
It is dogma in exercise that healthy indivs are t treatment group and t sicker population is t control group
Nby calling t phyaically active group t treatment, some others outside of exercise believe that being sedentary is health and thus see no reason to further supp exercise research
Carro, J. Neurosci. 21, 5678, 2001 Carro Mol Neurobi9ol 27, 153, 2003-10-08 - Increased sendetarism, contributes to an increasing incidence of neurological diseases - Sedentary life is a risk factor for neurodegenerative disease
What genese are changing in brain that will protect you from neural degeneration
(Saltin: Have identified candidate genes, now how activate?)
Kinetic Consideration of endurance training adaptations Name?
Approx 4 weeks before protein increase, through training
In 1980s, mRNA increase detected after 2 weeks
How do muscle cells adapt to exercise?
Metabolic genes: at transcription or mRNA (transient response)
Measure mRNA increases (transient) 48hrs after exercise – inc in protein concentration
Stress genes Priority Genes Metabolic/Mitochondrial Genes
Adaptations from training have to stem from acute stimulus (exercise bout)
Trying to indicate different categories of genes
Example: PDK4
protoocal after 4 weeks of one-legged knee extensor exercise train
24 hrs after exercise bout, mRNA back to basel levels
during exercise, why would muscle want to shut of a gene that produces carbohydrates? - hyp: as glucose ….muscle no longer want to use…
Endurance (PGC-1) David Hood, York University, Toronto
Endurance = high capacity of mitochondrial enzyme activity Not only applicable to endurane athletes, but also sedentary individuals – no exercise, low mitochondrial content and endurance capacity)
Ageing and low physical activity, bring mitochon down
Changes in energy status and calcium,. XXXX l
Leads to change of nucleus – transcription factors
Stimuli that affect transcription of nuclear genes
Mitochondria has its own genome - very limited, only codes for 13 genes (and 100s of genes reqd for mitochondrial function) - transcription factor for 13 genes is TFAM
Studied with animal model - chronic stimulation
typical marker for mitochondria (cytochrome C) - Freyseeenet, et al AJP 277, E26, 1999
Performance change - muscle force greater fatigue resistance (40% improvement)
PGC-1alpha - popular due to widespread effects in cell biology - mediates thyroid - influ on muscle fibre type - affects mitochondrial biogenesis
co-activator not a transcrioption factor not binding DNA, but binding transcritpopn factors - e.g. NRF-1, thyroid receptors (can enhance)
with chronic stim model, looked at PGC-1 level - nothing happened after 3 days, but by 5, 7, 10, increase in PGC-1 protein by 50% (Irrcher et al AJP 2003, 284)
used cells for better control induce to fuse together - immature muscle cell. Can make contract, or treat with drug - not physiological, but useful - is calcium important in mitochondrial biogenesis
number of transcriptors that could be involved in calcium response - 2.5x inc in PGC-1
effects of exercise on PGC-1 might be mediated by calcium
stimulation model - cells stimulated in disk - compared to non stimulated - looked at factors o PGC-1 coactives NRF-1 o NRF-1 transcriptionally activates Tfam and Cyto c) o Tfam imported to mito and inc mtDNA transcption and copy number o P38 MAP kinases phosphorylation stablizes PGC-1 protein o Irrcher et al AJP… (as abobe)
Effect of altering AMPKalpha activity with AICAR - inc in PGC-1 levels - which will ultimately affect mito levels
if understand PGC-1, can understand what affects mitochondrial levels
thyroid hormone is another potent stimulator of metabolism - high thyroid, high mito - hypo-thyroid is opposite - also acts by PGC-1
Conclusion
Contractilve activity-endurance signalling of mito bigenesis involves both calcium signals and changed I ATP turnove and is mediated by PGC-1alpha
Important role of PGC-1 in mataining normal levels…
How neuronal activity controls muscle fiber type and fiber size S. Schiuaffino, Padova
Dissecting signalling pathways involved in activity-dependant muscle gene regulation
Fiber types in skeletal muscle Genetics important, but other factors can modulate fibers - motorneuron activity is major factor
motorneuron modulate fibre size and fibre type
mechanical effects also important, and one of less explored areas - sports that generate tension in muiscle (e.g. weightlifting) can induce muscle hypertrophy metabolic changes - exercise can inc AMP and can activate…?
Try to identify important transaction pathways (activate and block) - How? o Pharma not helpful o Need a genetic approach • Somatic transgenesis • Transgenic mice: long procedure • Instead make transgenic muscles o Inject foreign DNA through plasmid o In few days, see effect in muscle phenotype e.g. Changes in MyHC gene expression inducaed by slow motor neuron…
injected mutant transducers to see if can block effect of nerve or to induce denervated muscle
Murgia et alNature Cell Biol, 2000 Serrano el al PNAS 2001 Pallafaccihna et al PNAS ,2002
Effects: two fibres (injected with foreign DNA) do not express slow myocin
More recently identify transcription factor - NFAT (transcip factor, protein binding DNA o Known to translocate o This translocation been able to….directly, linkd to GFP (fluorescent protein)
Same approach can be used to study muscle hypertrophy
(Saltin: people are on path to find out how performance is regulated)
An overwall enlargement of skeletal muscles is obtained by training and enhanced by anabolic steroids Name? (Female, Paris)
What’s important in skeletal muscle is, as muscle grows, inc proteins, mitochondria, and myonuclei and satellite cells are controlling this
What happens to nuclei in muscle that accompanies growth of muscle fibre As indiv trains (power train), is inc in myonuclear number, and anabolic steroids increase myonuclear number - amount of cytoplasm remains constant
if inc in no. of nuclei, means that cells have to be added in and must come from myonuclear component
skeletal muscle has cell, reserve cell
satellite cells can proliferate, to allow muscle to grow - also used to repair muscle
sat cells can be isolated from muscle and grow from biopsy
sat cells isol from human muscle fibres, have ltd capacity to cell
from birth can make 60-70 divisions - most of these lost during rapid growth
then, muscles stop growing - from 20-90yrs, maintain capacity to grow and repair skel muscle
Mitotic Clock - this is why there is a limited capacity, limited by Telemere (on each chromosome, is a piece of redundant DNA, each time cell divides, small part of this DNA will be lost, after while, signals to stop cells from further division, protects cells from cancer, but limits no. times can divide)
in athletes, no. of satellite cells increase on muscle fibre - to inc muscle mass, to repair muscle
same biopsies – look at telemere length, compared to sedentary - in elite pro athletes, is small decrease in Tele DNA, muscle is turning over much more than in sedentary population
Athletes with FAMS (Fatigued Athlete Myopathic Syndrome - Noakes) - suffering from chronic fatigue - connective tissue abundant - inc internal nuclei - abnormal mito nuclei
compared with control group of sports people
in this syndrome, where overtrain and genetic background, where cannot replenish sat cells, is pathologic – comparable to genetic disease
similar decrease in sat cells in weightlifters with large doping history
lose telemetric DNA each time cell divides
in cell culture, when isolated from elite athletes, compared to sedentary, doing small amount of regular activity is beneficial, because amateurXXXXX? - what level of activity is beneficial, compared to excessive exercise
Muscle satellite cells activation and skeletal muscle mass recognition Geoff Goldspink
Muscle mass muss be regulated locally and systemically - must be local growth factors
Alterantive Splicing of Human IGF-1 gene - now call mechano growth factor (only detected in ?) o derived from IGF-1 • IGF-1 genereal growth factor that makes cells inc in size
Normally think of IGF-1 as produced in liver
But MGF has different sequence and different action to systemic factor Activated by chemical signals quickly after exercise?
49 base insert - ie. Downstream shifts
put cDNA into different plasmids using same techniqe as earlier paper (inject to muscle of mouse)
made some constructs for in vitro when pt into mouse, found that a group of fibres that hve been tranZZ have inc in size within 2 weeks (25% larger) - potent growth factor)
how was it doing this? - looked at cells in culture - put interest into C2C12 cells o saw effects of systemic IGF-1, increased in mass, but …?
Subjected rat to mechanical damage, put in myotoxin agent - to see repair, etc
if look at marker for sat cell activation - sat cells inc in numbers quickly after injury - what sort is activating? o Discovered that shortly after injury MGF is expressed • Chief contender as major factor activiating sat cells Need to replenish pool of cells, but not too much
Muscles respond to mechanical signals – how? - not enough sat cells - seems that they are not activated - all muscles do not respond to exercise as well, since do not prod enough MGF
(Saltin: now health subject)
Muscle Contraction Febbraio?, Female, Brown hair
Plasma IL-6 - Ostrowksi et al J. Physiol 1998 - Ostrowski,
Is IL-6 produced during exercise? - yes, by working muscles, when it is released into circulation
role of muscle glycogen? - systemic vs. local effects - transcription rate inc with exercise, but further enhanced when muscle glycogen is XXX?
What does it mean to feed athletes carbohydrates during exercise? - IL-6 is inhibited
Febbraio et al 2003
If IL-6 was being produced to signal to liver to produce glucose? - not clear from findings
Febbraio hiscock, fischer, sachetti, Pederson - 2hrs cycling, at 40% VO2max….etc - IL-6 can influence glucose production, but co-factor is required
Does IL-6 induce lipolysis? - yes. Van hall, et al J clin endorcinol metab june 2003-10-08
Keller, FASEB, J, December, XXXX?
Nutrition – supplementation and sports performance
Ergogenic aids:food for performance or food for thought
Diet, training and ergogenic aids: t evidence from antiquity Louis Grivetti
Examples of
Doping Substances in Nutritional Supplements: Results of an IOC study Hans Geyer
Since 1996, Prohormones available over counter
Prohormones of Testosterone - DHEA
Prohormones of Nandrolone
Labelling of preparations does not reflect actual content
Parasrampuria M. et al 1998
Insufficient urveyence of prohormones
Now analyse non hormonal
Broad based study of international market
634 nutritional supplements purchased from Oct2000 – Nov 2001
in 13 countries
mainlybourght in shops (91.2%) and internet
289 supplements (49%) from prohorone selling companies
Results: 15% of nutritional supp,ements contained anaboligc androgenic not on label
most of positive nutritional included DHEA andendion
most positive products came from US companies (about 90%!)
do only prohormone companies have positive samples?
10% of products from non pro-hormone also positive
does application of such contamination of substances, lead to positive doping results - Yes, especially if prohormone of nandrolone
Many victims of contaminated substance? - Christie, ottie, etc
Conclusion
Problem of non-hormonal nutritional supplements containing prohibited anabolic-androgeneic is international problem
Consumption can lead to positive doping
Minimize risk, athletes should only buy nutritional supplements from companies which perform qualiy check for prohormones that guaranteee
In germany have companies that
www.osp-koeln.de - low risk suppplments (cannot excp
www.dopinginfo.de
in this study, cases were too low to have a physiological affect
example from Belgium where athlete has managed to gain compensation from
Muscle Glycogen: train low – compete high
Carbo ingestionduring exercise of longdurationin c performance
High muscle glycogen enhances time to exhaustion
What are infol of CHO ingestion and uiscle glycogen on training??
Training adaptation: what is influ of training at low versus high XXX
Training adaptation - what is that? - Molecular mechanisms? o Accumulation of proteins o How accumulate protein in muscle?
Prof. Choulis,
Framing it as a problem.
Number of durugs see
Questions
Randy Welberg, USOC presentations
Goldspink - china, factor 8 - haemophilia