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Purpose: In a case control study, we examined the allelic frequencies and genotype distributions of two restricted fragment length polymorphisms (RFLP) in the alpha-2A-adrenoceptor gene (ADRA2A) and beta-2-adrenoceptor gene (ADRB2) among elite endurance athletes (EEA) and sedentary controls (SC)., Methods: The EEA group included 148 Caucasian male subjects recruited on the basis that they had a VO2max > 74 mL O2 x kg(-1) x min(-1). The SC group comprised 149 unrelated sedentary male subjects, all Caucasians, from the Quebec Family Study. After digestion with the restriction enzymes Dra I (ADRA2A) and Ban I (ADRB2), Southern blotting and hybridization techniques were used to detect the mutations in the two ADR genes, which are encoded on chromosomes 10 (q24-26) and 5 (q31-32), respectively., Results: For the Dra I ADRA2A RFLP, we observed a significant difference in genotype distributions between the two groups (P = 0.037). A higher frequency of the 6.7-kb allele was observed in the EEA group compared with the SC group (P = 0.013). No statistically significant difference was found between groups for the Ban I ADRB2 polymorphic site. Genotype frequencies for both genes in both groups were in Hardy-Weinberg equilibrium., Conclusions: In summary, we found evidence that ADRA2A gene variability detected with Dra I is weakly associated with elite endurance athlete status, and we conclude that genetic variation in the ADRA2A gene or a locus in close proximity may play a role in being able to sustain the endurance training regimen necessary to attain a high level of maximal aerobic power.

Purpose: We have reported a significant association between VO2max in the sedentary state and its response (delta VO2max) to an endurance training program with a muscle-specific creatine kinase (CKMM) gene polymorphism. The purpose of this study was to test the hypothesis of genetic linkage between the same CKMM marker and VO2max in the sedentary state as well as delta VO2max., Methods: Sib-pair linkage analysis was performed on 277 full sib-pairs from 98 Caucasian nuclear families of the HERITAGE Family Study. VO2max was measured during cycle ergometry tests before and after 20 wk of endurance training. The CKMM polymorphism was detected by the polymerase chain reaction and digestion with the Ncol restriction enzyme., Results: Frequencies for the rare (1170 base pairs) and common (985 + 185 base pairs) alleles were 0.32 and 0.68, respectively. No significant linkage (t = -0.02, P = 0.49) was detected between the CKMM marker and the age and sex adjusted VO2max (mL x kg(-1) x min(-1)) in the sedentary state. However, after adjustment of delta VO2max for the effects of age, sex, initial VO2max, and body mass, evidence for linkage between the CKMM locus and delta VO2max was suggestive (P = 0.04)., Conclusion: The present results provide further support for the notion that the CKMM gene, or some gene in close linkage disequilibrium with it, may contribute to individual differences in the VO2max response to endurance training.

This study examined the associations between elite endurance athlete (EEA) status and three mitochondrial DNA (mtDNA) restriction fragment length polymorphisms (RFLPs) in the subunit 5 of the NADH dehydrogenase (MTND5) locus and one in the D-loop region. A group of 125 Caucasian male EEA well endowed with the phenotypic expression of VO2max (78.9 +/- 3.8 mL x kg(-1) x min(-1), mean +/- SD) and 65 sedentary controls (SCON: VO2max = 39.8 +/- 8.2 mL x kg(-1) x min(-1)) participated in the study. VO2max was determined during an incremental exercise test on a cycle ergometer or a motor-driven treadmill. mtDNA was extracted from white blood cells or lymphoblastoid cell lines and specific regions were amplified by the polymerase chain reaction. The Pearson Chi-square statistic test and Fisher exact test revealed no significant association (P > 0.05) between any of the three mtDNA RFLPs and EEA status. The MTND5-BamHI RFLP at bp 13,470 (morph 3) was found in 12.8% of the EEA and 12.3% of the SCON (chi2 = 0.009, P = 0.92). The prevalence of the MTND5-Ncil RFLP at bp 13,364 (morph 2) was 12.9% and 14% for the EEA and SCON, respectively (chi2 = 0.043, P = 0.83). The D-loop-KpnI RFLP at bp 16,133 (morph 1) was found in 5.8% of the EEA and in 1.6% of the SCON (Fisher exact test = 1.80, P = 0.18). The MTND5-HincII RFLP at bp 12,406 (morph 1) was not present in this study sample. These results indicate no evidence for a difference in the frequency of two polymorphic restriction sites in the subunit 5 of the NADH dehydrogenase gene of mtDNA and one in the D-loop region between elite endurance athletes and sedentary controls.

The purpose of this study was to investigate the association between elite endurance athlete (EEA) status and two restriction fragment length polymorphisms (RFLPs) at the muscle-specific creatine kinase (CKMM) gene locus. Genomic DNA was extracted from white blood cells or lymphoblastoid cell lines of 124 unrelated Caucasian male EEA (VO2max > 73 mL.kg-1.min-1) and 115 unrelated Caucasian sedentary male controls (SCON). The genetic polymorphism at the CKMM locus was detected by the polymerase chain reaction and DNA digestion with the NcoI and TaqI restriction endonucleases. The allelic frequencies for the NcoI and TaqI RFLPs were not different (P > 0.05) between EEA and SCON subjects. The three expected genotypes for CKMM-NcoI (1170/1170 bp, 1170/985 + 185 bp, and 985 + 185/985 + 185 bp) and CKMM-TaqI (1170/1170 bp, 1170/1020 + 150 bp, and 1020 + 50/1020 + 150 bp) were observed in the EEA and SCON groups. These genotype frequencies were in Hardy-Weinberg equilibrium, but they were not significantly (P > 0.05) different between the EEA and SCON. A strong (P < 0.001) linkage disequilibrium was detected among the NcoI and TaqI RFLPs in both EEA and SCON. These findings indicate that the skeletal muscle CK-NcoI and CK-TaqI gene polymorphisms are not associated with the elite endurance athlete status.

This study examined the association between a DNA polymorphism in the muscle-specific creatine kinase (CKMM) gene and VO2max in the sedentary state, as well as its response (deltaVO2max) to a standardized 20-wk endurance training program. The subjects were 160 biologically unrelated Caucasian parents (80 women, 80 men) and 80 biologically unrelated adult offspring of the HERITAGE Family Study. The CKMM polymorphism was detected by PCR and digestion with the NcoI restriction enzyme. VO2max was measured during maximal cycle ergometer tests. VO2max was 2119 +/- 45 mL x min(-1) (mean +/- SE) or 26 +/- 0.4 mL x kg(-1) x min(-1). Both sexes had a significant (P < 0.05) increase in the deltaVO2max (women = 283 +/- 20 mL x min[-1] and men = 363 +/- 25 mL x min[-1]). Allele and genotype frequencies were not significantly different (P > 0.05) between sexes. Age and sex adjusted VO2max was significantly (P = 0.007) associated with the CKMM genotype in the parents, whereas no association (P > 0.05) was observed in the offspring. DeltaVO2max values adjusted for age, sex, VO2max, and body mass were characterized by genotype differences in both parents (P = 0.0004) and offspring (P = 0.0025). A significantly (P < 0.05) lower deltaVO2max to endurance training was detected in both parents and offspring homozygotes for the rare allele. The genotype accounted for at least 9% of the variance in deltaVO2max. These results indicate that the NcoI polymorphism in the 3' untranslated region of the muscle-specific creatine kinase gene is associated with the deltaVO2max to endurance training.

A single-gene rodent mutation (diabetes) and a quantitative trait locus (dietary obese 1) mapped to the mid portion of mouse chromosome 4 have been related to obesity and/or insulin levels. Synteny relationships place their putative human homologs on 1p31 and 1p35-p31, respectively. In 137 sibships of adult brothers and sisters from the Québec Family Study, genetic linkages between seven microsatellite markers from 1p32-p22 and various obesity- and diabetes-related quantitative phenotypes were examined using single locus sibpair linkage analysis. Suggestive linkages were observed between markers D1S476 and body mass index (p = 0.05), fat mass (p = 0.02), the sum of six skinfolds (p = 0.02), the insulin area after an oral glucose tolerance test (p = 0.02), and between the neighboring marker D1S200 and body mass index (p = 0.03), and fat mass (p = 0.009). Suggestive linkages were also observed between the more telomeric markers D1S193 and body mass index (p = 0.03), and between the neighboring marker D1S197 and fasting insulin level (p = 0.05). No linkage was observed with the trunk to extremity skinfolds ratio. These linkages suggest that human homologs of the mouse diabetes or dietary obese 1 and/or other genes in this interval on chromosome 1 play a role in the regulation of body mass, body composition, and insulin levels, but not of subcutaneous fat distribution.

An update of the human obesity gene map up to October 1996 is presented. Evidence from Mendelian disorders exhibiting obesity as a clinical feature, single-gene mutation rodent models, quantitative trait loci uncovered in crossbreeding experiments with mouse, rat, and pig models, association and case-control studies with candidate genes, and linkage studies with genes and other markers is reviewed. All chromosomal locations of the animal loci are converted into human genome locations based on syntenic relationships between the genomes. A complete listing of all these loci reveals that only 4 of the 24 human chromosomes are not yet represented, i.e., 9, 18, 21, and Y. Several chromosome arms are characterized by the presence of several putative loci. The following arms include at least three such loci: 1p, 1q, 3p, 4q, 6p, 7q, 8p, 8q, 11p, 11q, 15q, 20q, and Xq. Studies with negative association and linkage results are also reviewed.

The polymorphic Bcl I site in the human ucp gene associated to percentage fat gain over time in the Québec Family Study cohort (Oppert et al. Int J Obesity 1994; 18: 526-531) has been positioned to the 5'-flanking region. This polymorphism results from a unique A/G mutation. Oligonucleotides used to amplify the polymorphic region, and allowing future studies of any cohort of patients, are described.

The aim of this study was to investigate the relationships between DNA restriction fragment length polymorphisms (RFLP) in the alpha 2- and beta 2-adrenoceptor genes and body fat distribution in humans. Skinfold thickness measurements and genetic analyses (Southern blot) were performed on 280 individuals (142 parents and 138 offsprings) from the Québec Family Study. Using the association study design in unrelated adults, women but not men carrying the 6.3-kb allele of an alpha 2A-adrenoceptor/DraI RFLP had a significantly higher trunk to extremity skinfold ratio (= sum of subscapular+suprailiac+abdominal skinfolds/sum of biceps+triceps+medial calf skinfolds) compared to women without the allele (1.44 +/- 0.52 vs. 1.12 +/- 0.33; p < 0.005 after adjustment for age, p < 0.002 after adjustment for age and body mass index or for age and subcutaneous fat). Using the sib-pair linkage procedure, a significant inverse relationship was found between the proportion of alleles identical by descent shared by sibs at the alpha 2A RFLP marker locus and the squared differences of the trunk to extremity skinfold ratio (p = 0.02 after adjustment for age or for age and body mass index or for age and subcutaneous fat). For a beta 2-adrenoceptor/BanI RFLP, no significant association or linkage was found between fat distribution indicators and the marker. These results suggest that alpha 2A-adrenoceptor gene variability detected with DraI is associated with a relative subcutaneous fat pattern favoring accumulation of truncal-abdominal fat in women, and that the alpha 2A-adrenoceptor gene, or a locus in close proximity, may be linked to body fat distribution in humans independently of the overall level of fatness.

The aim of this study was to investigate the association between a restriction fragment length polymorphism (RFLP) at the 3beta-hydroxysteroid dehydrogenase locus and adipose tissue distribution phenotypes. A total of 132 unrelated individuals from the Quebec Family Study were followed prospectively for an average period of 11.3 years. The BglII polymorphism in exon 4 of the 3beta-HSD gene was detected by PCR. Body mass, body fat, and regional fat distribution indicators were adjusted for age and age2 within each gender. Associations were assessed in unrelated adults with ANOVA across three genotypes. No association was found for the indicators of body mass, body fat, and regional distribution of adipose tissue measured in 1992. In women, the changes (difference between data collected in 1992 and at entry) in the sum of six skinfolds (p=0.04), abdominal skinfold (p=0.01), and abdominal skinfold adjusted (p=0.03) for the sum of six skinfolds at entry were related to the BglII polymorphism at the 3beta-HSD locus. These relations were not found in men, but they gained less body mass and body fat over the 11.3-year period. This suggests that sequence variation at the 3beta-HSD locus or in neighboring genes on chromosome 1 may contribute to individual differences in body fat content and adipose tissue distribution in adult women, particularly in abdominal adipose tissue deposition as they grow older and gain body fat.

The objective of this study was to identify DNA sequence variation in the UCP gene and to investigate its relationship with some obesity phenotypes. Two studies were carried out: (1) association study in unrelated subjects, and (2) sib-pair linkage analysis study in brothers and sisters. The subjects were 261 individuals from the Québec Family Study (123 parents and 138 offsprings from 64 families). The following were measured: Body mass index, percent body fat (measured by hydrostatic weighing), and subcutaneous fat (estimated by the sum of 6 skinfolds) were measured in 1978-81 and again 12 years later. Resting metabolic rate (RMR) was measured only in 1989-93. Genetic analyses were performed using Southern blotting technique and a human UCP genomic probe. (1) A BcII restriction fragment length polymorphism was identified with two alleles of 8.3 and 4.5 kb in length, and respective frequencies of 0.28 and 0.72. (2) In unrelated adults from the parental generation, a cross-sectional analysis of the 1989-93 data showed no difference in body fat and RMR between the UCP genotypes. No significant difference for the absolute changes in body fat over the 12-year period among the UCP genotypes was observed. However, a higher frequency (P < 0.05) of the 8.3-kb allele was found in high gainers compared to low gainers (i.e., above and below the median value) for percent body fat over the 12-year period. (3) No evidence of linkage between any of the obesity phenotypes and the UCP BcII marker was found. For the first time, the presence of DNA polymorphism in the human UCP gene is reported. Although, no significant association and linkage were found between the UCP BcII gene marker and body fat in the cohort of the Quebec Family Study, a higher frequency of the 8.3-kb allele was found in individuals who gained more body fat over time.

The aim of this study was to investigate whether the EcoRI restriction fragment length polymorphism (RFLP) of the apolipoprotein (apo) B-100 gene influences the associations described among obesity, regional adipose tissue distribution, and plasma lipoprotein levels. For this purpose, blood samples were collected from 56 healthy men for whom we had extensive measurements of regional adipose tissue distribution (both anthropometric and computed tomography-derived measurements) and data on the plasma lipoprotein-lipid profile. DNA was extracted from white blood cells, and RFLP analysis was performed. Subjects were classified into two groups on the basis of their apoB-100 EcoRI genotype: subjects homozygous for the major 11-kb allele, the 11/11 group (n = 40), and subjects carrying the minor 13-kb allele, the 13/11 group (n = 16). Subjects carrying the 13-kb allele had lower percent body fat and abdominal adipose tissue accumulation than subjects homozygous for the 11-kb allele (P < .05). Although leaner, the 13/11 group did not show a more favorable plasma lipoprotein-lipid profile than the group homozygous for the 11-kb allele. In fact, after statistical control for the difference in percent body fat between the two genotype groups, the 13/11 group showed significantly higher plasma cholesterol levels (P < .05) and nearly significantly higher apoB levels than the 11/11 group (P = .06). The association patterns between indices of regional adiposity and plasma cholesterol and apoB levels were also different between the two EcoRI genotype groups. Only in the 13/11 group was the abdominal visceral adipose tissue area significantly associated with these plasma variables.(ABSTRACT TRUNCATED AT 250 WORDS)

A DNA polymorphism of the nuclear-encoded subunit Va of the human cytochrome c oxidase (COX), a mitochondrial respiratory enzyme, is reported. No polymorphism was detected in genes for the subunits IV and Vb of the same enzyme.

A rare polymorphism in the human sex hormone-binding globulin (SHBG) gene was detected using a human SHBG cDNA probe. It is the first DNA sequence variation reported in this gene.

Mitochondrial DNA sequence variation was determined in 46 sedentary young adult males who took part in ergocycle endurance training programs in two laboratories to assess whether mitochondrial DNA variants were associated with individual differences in maximal oxygen uptake (VO2max) and its response to training. VO2max was obtained from a progressive ergocycle test to exhaustion. White blood cell mitochondrial DNA was characterized with the restriction fragment length polymorphism (RFLP) technique using 22 restriction enzymes and human mitochondrial DNA as a probe for hybridization. Multiple mitochondrial DNA variants were detected with 15 of the enzymes. Some subjects exhibited many RFLPs, while others showed no variation. These RFLPs (morphs) were generated by base substitutions located in gene regions coding for mitochondrial proteins as well as in the noncoding regions. Carriers of three mitochondrial DNA morphs, two in the subunit 5 of the NADH dehydrogenase gene and one in the tRNA for threonine, had a VO2max (ml.kg-1.min-1) in the untrained state significantly higher than noncarriers, while carriers of one mitochondrial DNA morph in subunit 2 of NADH dehydrogenase had a lower initial VO2max. Endurance training increased VO2max by a mean of 0.5 l of O2, with individual differences ranging from gains of 0.06 to 1.03. After adjustment for training site and initial VO2max, a lower response was observed for three carriers of a variant in subunit 5 of the NADH dehydrogenase detected with HincII (mean gain of 0.28 l; P < 0.05). These results suggest that sequence variation in mitochondrial DNA may contribute to individual difference in VO2max and its response to training.

Mitochondrial DNA sequence variation was determined in 46 sedentary young adult males who took part in ergocycle endurance training programs in two laboratories to assess whether mitochondrial DNA variants were associated with individual differences in maximal oxygen uptake (VO2max) and its response to training. VO2max was obtained from a progressive ergocycle test to exhaustion. White blood cell mitochondrial DNA was characterized with the restriction fragment length polymorphism (RFLP) technique using 22 restriction enzymes and human mitochondrial DNA as a probe for hybridization. Multiple mitochondrial DNA variants were detected with 15 of the enzymes. Some subjects exhibited many RFLPs, while others showed no variation. These RFLPs (morphs) were generated by base substitutions located in gene regions coding for mitochondrial proteins as well as in the noncoding regions. Carriers of three mitochondrial DNA morphs, two in the subunit 5 of the NADH dehydrogenase gene and one in the tRNA for threonine, had a VO2max (ml.kg-1.min-1) in the untrained state significantly higher than noncarriers, while carriers of one mitochondrial DNA morph in subunit 2 of NADH dehydrogenase had a lower initial VO2max. Endurance training increased VO2max by a mean of 0.51 of O2, with individual differences ranging from gains of 0.06 to 1.03. After adjustment for training site and initial VO2max, a lower response was observed for three carriers of a variant in subunit 5 of the NADH dehydrogenase detected with HincII (mean gain of 0.28 I; P less than 0.05). These results suggest that sequence variation in mitochondrial DNA may contribute to individual difference in VO2max and its response to training.

A general method for isolating biologically active messenger RNA (mRNA) from agarose gels is reported. Purified cellular RNA is resolved by preparative agarose gel electrophoresis and recovered in high yields (80%) by passive diffusion. Polyadenylated mRNA isolated from the eluted RNA is functionally intact based on the ability of the RNA to serve as a template in cell-free translation systems and complementary DNA synthesis reactions. The entire procedure is simple and rapid. A substantial purification of the mRNAs coding for skeletal muscle myosin heavy chain, light chain subunits and carbonic anhydrase III has been achieved employing this method.

We have found a specific binding protein for synthetic progestins 6,7-[3H]methyltrienolone (R1881) and 17,21-dimethyl-19-norpregna-4,9-diene-3,20-dione (R5020) and in the testis cytosol from three "sisters" with the complete form of the testicular feminization syndrome. The binding component sediments in the 8S region of sucrose gradients. It is saturable. The apparent affinity constant (Ka) for R5020 was determined in two cases and found to be 1.8 and 0.6 X 10(8) M-1. The number of binding sites calculated from Scatchard plots is relatively high: 572 and 826 fmol/mg protein. Competition studies indicate that this putative receptor is specific for natural and synthetic progestins but not for 5 alpha-dihydrotestosterone and cortisol. Similar progestin binding could not be found in normal human and rat testes.

Some characteristics of the dorsal skin cytoplasmic androgen receptor (AR) have been studied in male rats. The affinity constant, the binding specificity, and the sedimentation profile of the receptor have been found to be similar to the rat prostate AR. The measurement of the number of binding sites in various hormonal conditions (deprivation) led to the conclusion that this receptor was largely occupied by endogeneous hormones from gonadal and (or) adrenal sources. Administration of spironolactone or canrenone to 7-day-castrated rats was accompanied by a rapid and drastic decrease of available binding sites. This diminution was ascribed to the competitive inhibition of canrenone, the active in vivo metabolite of spironolactone. It is postulated that the antiandrogenic action of spironolactone, at the skin level, is mediated by canrenone which inhibits the formation of specific testosterone and (or) 5 alpha-dihydrotestosterone receptor complex in cytoplasm and consequently in nuclei.

In our studies of the binding of steroids in rat skeletal muscles, we have shown the existence of a saturable '4S' dihydrotestosterone-binding protein of low affinity (Kd = 1.16 X 10(-6) M). Competition studies led us to believe that the binding of dihydrotesterone (DHT) was due to 3 alpha-hydroxysteroid dehydrogenase (3 alpha-OHSD) enzyme responsible for the conversion of DHT to androstanediol (Adiol). Indeed, both the binding and the enzymatic activity are inhibited by various 3-keto steroids. In addition, the dissociation constant (Kd) and the Michaelis constant (Km) of the enzyme are similar. However, experiments with ammonium sulfate fractions of the cytosol have shown a partial separation of the binding and of the enzymatic activity. On the other hand, we have established that DHT (3 micrometer) is almost completely metabolized to Adiol during a 2-h incubation of 0 degrees C even in the absence of added coenzymes. Furthermore the '4S' protein binds Adiol more strongly than DHT and this binding is not saturable. Finally the binding behaviour of both DHT and Adiol with either muscle cytosol or rat albumin is similar when subjected to ammonium sulfate fractionation and sucrose density gradient centrifugation. In conclusion, the skeletal muscle 3 alpha-OHSD rapidly metabolizes DHT into Adiol which then binds strongly to a nonspecific binding protein, presumably rat serum albumin. Thus it can be said that the observed saturability of DHT binding is only apparent.