John L. VandeBerg

Genetic and environmental contributions to cardiovascular risk factors in Mexican Americans: The San Antonio Family Heart Study

BACKGROUND The familial aggregation of coronary heart disease can be in large part accounted for by a clustering of cardiovascular disease risk factors. To elucidate the determinants of cardiovascular disease, many epidemiological studies have focused on the behavioral and lifestyle determinants of these risk factors, whereas others have examined whether specific candidate genes influence quantitative variation in these phenotypes. METHODS AND RESULTS Among Mexican Americans from San Antonio (Tex), we quantified the relative contributions of both genetic and environmental influences to a large panel of cardiovascular risk factors, including serum levels of lipids, lipoproteins, glucose, hormones, adiposity, and blood pressure. Members of 42 extended families were studied, including 1236 first-, second-, and third-degree relatives of randomly ascertained probands and their spouses. In addition to the phenotypic assessments, information was obtained regarding usual dietary and physical activity patterns, medication use, smoking habits, alcohol consumption, and other lifestyle behaviors and medical factors. Maximum likelihood methods were used to partition the variance of each phenotype into components attributable to the measured covariates, additive genetic effects (heritability), household effects, and an unmeasured environmental residual. For the lipid and lipoprotein phenotypes, age, gender, and other environmental covariates accounted in general for < 15% of the total phenotypic variance, whereas genes accounted for 30% to 45% of the phenotypic variation. Similarly, genes accounted for 15% to 30% of the phenotypic variation in measures of glucose, hormones, adiposity, and blood pressure. CONCLUSIONS These results highlight the importance of considering genetic factors in studies of risk factors for cardiovascular disease.

Chemerin Is Associated with Metabolic Syndrome Phenotypes in a Mexican-American Population

CONTEXT Chemerin is a novel adipokine previously associated with metabolic syndrome phenotypes in a small sample of subjects from Mauritius. OBJECTIVE The aim of the study was to determine whether plasma chemerin levels were associated with metabolic syndrome phenotypes in a larger sample from a second, unrelated human population. DESIGN, SETTING, PATIENTS, AND INTERVENTION Plasma samples were obtained from the San Antonio Family Heart Study (SAFHS), a large family-based genetic epidemiological study including 1431 Mexican-American individuals. Individuals were randomly sampled without regard to phenotype or disease status. This sample is well-characterized for a variety of phenotypes related to the metabolic syndrome. MAIN OUTCOMES Plasma chemerin levels were measured by sandwich ELISA. Linear regression and correlation analyses were used to determine associations between plasma chemerin levels and metabolic syndrome phenotypes. RESULTS Circulating chemerin levels were significantly higher in nondiabetic subjects with body mass index (BMI) greater than 30 kg/m(2) compared with those with a BMI below 25 kg/m(2) (P < 0.0001). Plasma chemerin levels were significantly associated with metabolic syndrome-related parameters, including BMI (P < 0.0001), fasting serum insulin (P < 0.0001), triglycerides (P < 0.0001), and high-density lipoprotein cholesterol (P = 0.00014), independent of age and sex in nondiabetic subjects. CONCLUSION Circulating chemerin levels were associated with metabolic syndrome phenotypes in a second, unrelated human population. This replicated result using a large human sample suggests that chemerin may be involved in the development of the metabolic syndrome.

Conservation of the H19 noncoding RNA and H19-IGF2 imprinting mechanism in therians

Comparisons between eutherians and marsupials suggest limited conservation of the molecular mechanisms that control genomic imprinting in mammals. We have studied the evolution of the imprinted IGF2-H19 locus in therians. Although marsupial orthologs of protein-coding exons were easily identified, the use of evolutionarily conserved regions and low-stringency Bl2seq comparisons was required to delineate a candidate H19 noncoding RNA sequence. The therian H19 orthologs show miR-675 and exon structure conservation, suggesting functional selection on both features. Transcription start site sequences and poly(A) signals are also conserved. As in eutherians, marsupial H19 is maternally expressed and paternal methylation upstream of the gene originates in the male germline, encompasses a CTCF insulator, and spreads somatically into the H19 gene. The conservation in all therians of the mechanism controlling imprinting of the IGF2-H19 locus suggests a sequential model of imprinting evolution.

A microassay for ATPase

A newly developed microtechnique for quantitating activity of myosin ATPase (EC 3.6.1.32) is more sensitive and less time-consuming than existing spectrophotometric methods. Measurement of ATPase activity using the new method can be accomplished in a final volume of 0.25 ml, allowing the assay to be conducted in individual wells of 96-well microplates commonly used for the enzyme-linked immunosorbent assay (ELISA). The microassay is performed by adding purified myosin to microplate wells followed by addition of ATP to initiate the enzymatic reaction. The reaction is subsequently terminated by addition of an acidic solution containing malachite green and ammonium molybdate. The level of inorganic phosphate produced by enzymatic hydrolysis of ATP is measured by scanning the microplates using a microELISA plate reader. An entire 96-well microplate can be scanned in less than 2 min, and data from the microassay can be transferred directly to a microprocessor for statistical analysis. The microassay is capable of detecting between 0.2 and 3 nmol of inorganic phosphate in a reaction volume of 50 microliter, and the ATPase activity of as little as 10 ng of rat cardiac myosin can be measured. The increased sensitivity compared with that of other spectrophotometric assays and ease of performing the microassay enable a detailed analysis of the enzymatic properties of cardiac myosin to be conducted on large numbers of small tissue specimens. Several kinetic properties of rat cardiac myosin were determined using this technique.

Genes on chromosomes 1 and 13 have significant effects on Ascaris infection

Nematode parasites show a characteristic aggregated distribution among hosts. This observation has important implications for pathogenesis, immunology, and control of these infections, but the relative roles of environment and genetics in determining these patterns have remained uncertain. This paper presents the results of the first genome scan for susceptibility to infection with roundworm (Ascaris lumbricoides). Data on 375 genetic markers were generated for each of 444 members of a genetically isolated Nepalese population, the Jirels. Ascaris worm burden as assessed by egg counts was measured in these same individuals by using the Kato Katz thick smear method. The extensive genealogical data available for the population allowed assignment of all 444 individuals to a single pedigree that contained 6,209 pairs of relatives that were informative for genetic analysis. A variance components linkage analysis resulted in the unequivocal localization of two genes (one on chromosome 1 and another on chromosome 13) with clear, significant effects on susceptibility to Ascaris infection. This is the first evidence that individual quantitative trait loci influence variation in Ascaris burden in humans.

Identification of autoantigens recognized by the 2F5 and 4E10 broadly neutralizing HIV-1 antibodies

Immunological tolerance to self-antigen impairs humoral responses to HIV-1.

Plasma HDL Cholesterol, Triglycerides, and Adiposity A Quantitative Genetic Test of the Conjoint Trait Hypothesis in the San Antonio Family Heart Study

BACKGROUND The conjoint trait hypothesis proposes that combined low HDL cholesterol (HDL-C) and high triglyceride (TG) levels represent a single, inherited phenotype that adiposity may influence in an unspecified manner. We conducted formal statistical genetic tests of the conjoint trait hypothesis and the relation of the conjoint trait to adiposity using data for 569 subjects in 25 pedigrees from the San Antonio Family Heart Study. METHODS AND RESULTS We conducted multivariate genetic analyses to detect the effects of genes and environmental factors on variation in plasma concentrations of HDL-C and TG, fat mass (as percent body weight [FM%], determined by bioelectric impedance), and body mass index (BMI). We used maximum-likelihood methods to simultaneously estimate the phenotypic means and SDs, heritabilities (h2), effects of sex, age-by-sex, eight dietary and medical covariates, and genetic and environmental correlations. Likelihood ratio tests disclosed significant heritabilities (P < .001) for all traits (h2HDL-C = 0.55, h2TG = 0.53, h2FM% = 0.37, h2BMI = 0.44) but significant genetic correlations (P < .001), indicating pleiotropy, between two trait pairs only: HDL-C and TG (PG = -0.52) and fat mass and BMI (PG = 0.86). We obtained significant environmental correlations between all trait pairs except HDL-C and BMI (P > .05). CONCLUSIONS Both shared genes (pleiotropy) and shared environmental factors contribute to the commonly observed inverse phenotypic association between plasma levels of HDL-C and TG. Rather than low HDL-C and high TG being a single, genetically transmissible entity, it is the inverse relation between these two phenotypes throughout their normal ranges of variation as well as at the extremes that is influenced by shared genes and shared environments. However, common environmental factors, not shared genes, account for reported associations of plasma HDL-C and TG levels with measures of adiposity.

Human Pedigree-Based Quantitative-Trait–Locus Mapping: Localization of Two Genes Influencing HDL-Cholesterol Metabolism

Common disorders with genetic susceptibilities involve the action of multiple genes interacting with each other and with environmental factors, making it difficult to localize the specific genetic loci responsible. An important route to the disentangling of this complex inheritance is through the study of normal physiological variation in quantitative risk factors that may underlie liability to disease. We present an analysis of HDL-cholesterol (HDL-C), which is inversely correlated with risk of heart disease. A variety of HDL subphenotypes were analyzed, including HDL particle-size classes and the concentrations and proportions of esterified and unesterified HDL-C. Results of a complete genomic screen in large, randomly ascertained pedigrees implicated two loci, one on chromosome 8 and the other on chromosome 15, that influence a component of HDL-C-namely, unesterified HDL2a-C. Multivariate analyses of multiple HDL phenotypes and simultaneous multilocus analysis of the quantitative-trait loci identified permit further characterization of the genetic effects on HDL-C. These analyses suggest that the action of the chromosome 8 locus is specific to unesterified cholesterol levels, whereas the chromosome 15 locus appears to influence both HDL-C concentration and distribution of cholesterol among HDL particle sizes.

A Genome Search Identifies Major Quantitative Trait Loci on Human Chromosomes 3 and 4 That Influence Cholesterol Concentrations in Small LDL Particles

Small, dense LDL particles are associated with increased risk of cardiovascular disease. To identify the genes that influence LDL size variation, we performed a genome-wide screen for cholesterol concentrations in 4 LDL size fractions. Samples from 470 members of randomly ascertained families were typed for 331 microsatellite markers spaced at approximately 15 cM intervals. Plasma LDLs were resolved by using nondenaturing gradient gel electrophoresis into 4 fraction sizes (LDL-1, 26.4 to 29.0 nm; LDL-2, 25.5 to 26.4 nm; LDL-3, 24.2 to 25.5 nm; and LDL-4, 21.0 to 24.2 nm) and cholesterol concentrations were estimated by staining with Sudan Black B. Linkage analyses used variance component methods that exploited all of the genotypic and phenotypic information in the large extended pedigrees. In multipoint linkage analyses with quantitative trait loci for the 4 fraction sizes, only LDL-3, a fraction containing small LDL particles, gave peak multipoint log10 odds in favor of linkage (LOD) scores that exceeded 3.0, a nominal criterion for evidence of significant linkage. The highest LOD scores for LDL-3 were found on chromosomes 3 (LOD=4.1), 4 (LOD=4.1), and 6 (LOD=2.9). In oligogenic analyses, the 2-locus LOD score (for chromosomes 3 and 4) increased significantly (P=0.0012) to 6.1, but including the third locus on chromosome 6 did not significantly improve the LOD score (P=0.064). Thus, we have localized 2 major quantitative trait loci that influence variation in cholesterol concentrations of small LDL particles. The 2 quantitative trait loci on chromosomes 3 and 4 are located in regions that contain the genes for apoD and the large subunit of the microsomal triglyceride transfer protein, respectively.

Rsx is a metatherian RNA with Xist-like properties in X-chromosome inactivation

In female (XX) mammals one of the two X chromosomes is inactivated to ensure an equal dose of X-linked genes with males (XY)1. X-inactivation in eutherian mammals is mediated by the non-coding RNA Xist2. Xist is not found in metatherians3 and how X-inactivation is initiated in these mammals has been the subject of speculation for decades4. Using the marsupial Monodelphis domestica we identify Rsx (RNA-on-the-silent X), an RNA that exhibits properties consistent with a role in X-inactivation. Rsx is a large, repeat-rich RNA that is expressed only in females and is transcribed from, and coats, the inactive X chromosome. In female germ cells, where both X chromosomes are active, Rsx is silenced, linking Rsx expression to X-inactivation and reactivation. Integration of an Rsx transgene on an autosome in mouse embryonic stem cells leads to gene silencing in cis. Our findings permit comparative studies of X-inactivation in mammals and pose questions about the mechanisms by which X-inactivation is achieved in eutherians.In female (XX) mammals, one of the two X chromosomes is inactivated to ensure an equal dose of X-linked genes with males (XY). X-chromosome inactivation in eutherian mammals is mediated by the non-coding RNA Xist. Xist is not found in metatherians (marsupials), and how X-chromosome inactivation is initiated in these mammals has been the subject of speculation for decades. Using the marsupial Monodelphis domestica, here we identify Rsx (RNA-on-the-silent X), an RNA that has properties consistent with a role in X-chromosome inactivation. Rsx is a large, repeat-rich RNA that is expressed only in females and is transcribed from, and coats, the inactive X chromosome. In female germ cells, in which both X chromosomes are active, Rsx is silenced, linking Rsx expression to X-chromosome inactivation and reactivation. Integration of an Rsx transgene on an autosome in mouse embryonic stem cells leads to gene silencing in cis. Our findings permit comparative studies of X-chromosome inactivation in mammals and pose questions about the mechanisms by which X-chromosome inactivation is achieved in eutherians.