Download Udvardy, A.1,Bosnyák, E.1, Trájer, E.1, Protzner, A.1, Komka, Zs.1

Complex DNA test method with a DNA chip among Hungarian athletes
Udvardy, A. 1,Bosnyák, E.1, Trájer, E.1, Protzner, A. 1, Komka, Zs. 1, Szendrei, B. 1,Györe, I.2, Tóth, M. 1 , Pucsok, J.2
SEMMELWEIS UNIVERSITY BUDAPEST1, NATIONAL INSTITUTE OF SPORT MEDICINE2
Hungary
Introduction:
We have been studying the genetic background of sport and exercise
physiology on several genes and their polimorphisms. We wanted to broaden
the number of measured genes and examine not only the specific sport genes
but the area of cardiovascular diseases, sudden cardiac death (SCD) and
metabolic syndrome(MetS) as well. Our research group established a complex
test to investigate 64 gene variants at one time (TaqMan OpenArray DNA
chip).
SCD can strike both professionals and amateurs, young and old. It is
particularly disturbing when it happens to an athlete and makes us stop and
re-evaluate exercising for health. The reason of the SCD in athletes is mostly
unclear, it can rarely be verified (trauma, embolism, aneuryzmarupture,
asthma etc.). Therefore it is essential to understand the genetic background
Genes, related to physical performance:
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beta(2)-adrenoceptor (ADRB2)
Arg16Gly rs1042713
Arg16
endurance
ACTN3
R577X (rs1815739)
R
speed, strength
PGC1A
Gly482Ser (rs8192673
Gly
endurance
C825T rs5443
guanine nucleotide-binding protein, beta polypeptide
3 (GNB3)
T
endurance
PPARA
rs4253778?
GG
endurance
Myostatin K153R (GDF8) 2q32.2
rs1805086
KK
strength (max. muscle contraction)
AMP deaminase-1 (AMPD1)
Gln12Ter rs17602729
C
endurance
beta(2)-adrenoceptor (ADRB2)
Gln27->Glu27 rs1042714
Gln27
endurance
Genes, related to MetS
and make it possible in the future to use it as a tool to filter out individuals who
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have risk factors.
The MetS is a cluster of risk factors including central obesity, hypertension,
hyperglycaemia and dyslipidaemia. There is research based evidence already
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APOE
rs405509 (-219G>T)
G
Higher total cholesterol level, lower response to statin therapy
beta(2)-adrenoceptor (ADRB2)
Gln27->Glu27 rs1042714
Gln27
MetS risk
alpha-adducin
Gly460Trp rs4961
Trp/Trp
high blood pressure risk
Gly460Trp exercise induced antihypertensive response is good
LIPG i24582 rs6507931
endothelial lipase (LIPG)
Peroxisome proliferator-activated receptorPro12Ala rs1801282
gamma (PPARgamma)
rs1800629
tumor necrosis factor ( TNF)
CC
exercise induced HDL increase is good
Ala
DM type 2 risk
GG
high blood pressure, hypercholesterinaemia risk
Plasminogen activator inhibitor-1 (PAI-1)
-844 A>G
G
MetS-ra veszélyeztetettebbek
response to physical training which is one of the basic therapies for MetS.
bradykinin B2 receptor gene (BDKRB2)
58C/T rs1799722
T
high blood pressure risk
HFE
H63D rs1799945
D
DM type 2 risk
Therefore it can be informative to study athlete population who live a healthy
preproneuropeptide Y
Leu7Pro rs16139
Pro
DM type 2 risk
calpain 10
SNP43-G allele rs3792267
GG
DM type 2 risk
lifestyle (nutrition, exercise, non-smoking) to associate the genetic variants
fatty acid binding protein 2 (FABP2
endothelial NO synthase gene (eNOS
gene)
rs1799883 ala 54 to Thr 54
Thr
MetS risk
Glu298Asp rs1799983
Asp
atherogenic lipid profile risk
of a complex interplay between genetic determinants and environmental
factors as it has been experienced that there are individual differences in the
with measurable phenotypes (blood pressure, lipid levels, etc.).
Genes, related to cardiovascular diseases
Studied population, genes:
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Our aim is to use a reliable, fast and complex research method to evaluate
the 64 set gene variants on elite athlete population.
muscle glycogen synthase gene (GYS1)
(XbaIC>T) rs8103451 XbalT
Angiotensin II type I receptor (AGTR1)
A1166C rs12721235 CC
The system has been validated with previously genotyped DNA samples.
We would like to introduce some of the chosen genes, based on international
angiotensinogen
M235T rs699
T
adiponectin receptor 2 (ADIPOR2)
rs767870
heterozygote
cardiovascular risk
adiponectin gene (ADIPOQ)
+45T >G
G
lower cardiovascular risk
CASP-1
G+7/in6A, rs501192
lack of A allele(in6) cardiovascular risk
AMP deaminase-1 (AMPD1)
Gln12Ter rs17602729 C
cardiovascular risk
il18
-137 G/C rs187238
Sudden cardiac death, high blood
pressure risk
studies related to the previously mentioned three areas, and our preliminary
results from the validation process (n=48,elite athletes in kayak-canoe,
rowing, triathlon and cycling).
GG
cardiovascular risk
better cardiovascular and
anthropometric parameters
exercise induced left ventricle
hypertrophy
Methods:
DNA is extracted from blood samples using a DNA isolating kit. 64 genes are
studied from each DNA sample. DNA detection is done with a DNA chip at Avidin
Ltd.
TaqMan OpenArray plate is divided into 48 subarrays (Pic. 1 and 2); each
subarray consists of 64 through-holes, which means that 3072 nanocapillary
quick reaction time (QRT) PCR can be run at one time. To measure the
polimorphisms fluorescently signaled Taqman probes are used. The isolated
DNA (150ng) from whole blood is mixed with OpenArray MasterMix and loaded
into the nanocapillary reaction chamber. The OpenArray plates are put into
Perkin Elmer 1000 Plate PCR device for 50 cycles and afterwards into the
BioTrove OpenArrayTM NT Cycler where 3 plates can be analyzed at the same
time, which means that more than 9000 individual reactions can be run. The
intensity of fluorescence is detected automatically at every point and can be
analyzed afterwards.
Pic. 1.
Pic 2.
Discussion:
Our future objective is to evaluate the results of the complex DNA testing on a great number of sample and establish a complementary screening method which would filter out
the individuals with cumulative risk factors and subject them to further medical control.
Continuing our sportgenomic research project we would like to supplement sport selection, exercise physiological, anthropometric and basic sport medicine measurements.
Contact: [email protected]