Homocysteine, blood pressure and gene-diet interactions in relation to vascular function measures of black South Africans.

BACKGROUND AND AIMS: We investigated circulating homocysteine (Hcy), a cardiovascular disease (CVD) risk factor, examining its dietary associations to provide personalized nutrition advice. This study addressed the inadequacy of current dietary interventions to ultimately address the disproportionately high incidence of CVD in Black populations. METHODS AND RESULTS: Cross-sectional analyses of 1,867 Black individuals of the PURE-SA study allowed the identification of dietary intake and cardiovascular measure interactions on three sub-categories: (1) normal blood pressure (BP), hypertension or Hcy-related hypertension (H-type), (2) low, normal or high Hcy concentrations, and (3) Hcy-related genetic combinations. Favorable body composition, but adverse dietary intake and cardiovascular determinants, were observed in higher Hcy categories. H-types, compared to regular hypertensives, had higher alcohol and lower macronutrient and micronutrient consumption. Inverse associations with carotid-radial pulse wave velocity were evident between monounsaturated fatty acid (FA) consumption and H-type hypertension as well as polyunsaturated FA and CBS883/ins68 TT carriers. Energy intake was positively associated with vascular cell adhesion molecule-1 (VCAM-1) in variant CBST883C/ins68 and CBS9276 GG carriers. VCAM-1 was also positively associated with plant protein intake in CBS9276 GG and MTR2756 AA carriers and negatively with total protein intake and CBS9276 GG carriers. Alcohol intake was positively associated with intercellular adhesion molecule-1 in MTR2756 minor allele carriers. CONCLUSION: Because Hcy gene-diet interactions are evident, personalized nutrition, by adjusting diets based on genetic profiles (e.g., CBS and MTR variations) and dietary interactions (e.g., FAs and proteins), can enhance cardiovascular outcomes by managing Hcy and related hypertension in genetically susceptible individuals.

Understanding and applying gene-environment interactions: a guide for nutrition professionals with an emphasis on integration in African research settings.

Noncommunicable diseases (NCDs) are influenced by the interplay between genetics and environmental exposures, particularly diet. However, many healthcare professionals, including nutritionists and dietitians, have limited genetic background and, therefore, they may lack understanding of gene-environment interactions (GxEs) studies. Even researchers deeply involved in nutrition studies, but with a focus elsewhere, can struggle to interpret, evaluate, and conduct GxE studies. There is an urgent need to study African populations that bear a heavy burden of NCDs, demonstrate unique genetic variability, and have cultural practices resulting in distinctive environmental exposures compared with Europeans or Americans, who are studied more. Although diverse and rapidly changing environments, as well as the high genetic variability of Africans and difference in linkage disequilibrium (ie, certain gene variants are inherited together more often than expected by chance), provide unparalleled potential to investigate the omics fields, only a small percentage of studies come from Africa. Furthermore, research evidence lags behind the practices of companies offering genetic testing for personalized medicine and nutrition. We need to generate more evidence on GxEs that also considers continental African populations to be able to prevent unethical practices and enable tailored treatments. This review aims to introduce nutrition professionals to genetics terms and valid methods to investigate GxEs and their challenges, and proposes ways to improve quality and reproducibility. The review also provides insight into the potential contributions of nutrigenetics and nutrigenomics to the healthcare sphere, addresses direct-to-consumer genetic testing, and concludes by offering insights into the field's future, including advanced technologies like artificial intelligence and machine learning.

H-Type Hypertension among Black South Africans and the Relationship between Homocysteine, Its Genetic Determinants and Estimates of Vascular Function.

Elevated homocysteine (Hcy) increases cardiovascular disease (CVD) risk. Our objective was to emphasize Hcy's contribution in hypertension and CVD management by determining H-type hypertension (hypertension with Hcy ≥ 10 µmol/L) and associations between Hcy, blood pressure (BP) and estimates of vascular function among Black South Africans. We included 1995 adults (63% female). Plasma Hcy and cardiovascular measures (systolic and diastolic BP (SBP, DBP), pulse pressure, heart rate (HR), carotid-radialis pulse wave velocity (cr-PWV), intercellular adhesion molecule-1 (ICAM-1) and vascular cell adhesion molecule-1) were quantified. Five Hcy-related polymorphisms (cystathionine ß-synthase (CBS 844ins68, T833C, G9276A); methylenetetrahydrofolate reductase (MTHFR C677T) and methionine synthase (MTR A2756G)) were genotyped. Hcy was >10 µmol/L in 41% (n = 762), and of the 47% (n = 951) hypertensives, 45% (n = 425) presented with H-type. Hcy was higher in hypertensives vs. normotensives (9.86 vs. 8.78 µmol/L, p < 0.0001, effect size 0.56) and correlated positively with SBP, DBP, cr-PWV and ICAM-1 (r > 0.19, p < 0.0001). Over Hcy quartiles, SBP, DBP, HR, cr-PWV and ICAM-1 increased progressively (all p-trends ≤ 0.001). In multiple regression models, Hcy contributed to the variance of SBP, DBP, HR, cr-PWV and ICAM-1. H-type hypertensives also had the lowest MTHFR 677 CC frequency (p = 0.03). Hcy is positively and independently associated with markers of vascular function and raised BP.

Associations Between 25-Hydroxyvitamin D and Total and γ' Fibrinogen and Plasma Clot Properties and Gene Interactions in a Group of Healthy Black South African Women.

The role of 25-hydroxyvitamin D [25(OH)D] in reducing the risk of cardiovascular disease (CVD) has been recognized, but the mechanisms involved are unclear. Researchers have discovered a link between vitamin D and fibrinogen. Until now, data on the relationship between vitamin D and the γ' splice variant of fibrinogen and fibrin clot characteristics remain unexplored. In this study, 25(OH)D, total and γ' fibrinogen, as well as turbidimetrically determined plasma clot properties, were quantified, and fibrinogen and FXIII SNPs were genotyped in 660 Black, apparently healthy South African women. Alarmingly, 16 and 45% of the women presented with deficient and insufficient 25(OH)D, respectively. Total fibrinogen and maximum absorbance (as a measure of clot density) correlated inversely, whereas γ' fibrinogen correlated positively with 25(OH)D. γ' fibrinogen increased whereas maximum absorbance decreased over the deficient, insufficient, and sufficient 25(OH)D categories before and after adjustment for confounders. 25(OH)D modulated the association of the SNPs regarding fibrinogen concentration and clot structure/properties, but did not stand after correction for false discovery rate. Because only weak relationships were detected, the clinical significance of the findings are questionable and remain to be determined. However, we recommend vitamin D fortification and supplementation to reduce the high prevalence of this micronutrient deficiency and possibly to improve fibrinogen and plasma clot structure if the relationships are indeed clinically significant. There is a need for large cohort studies to demonstrate the relationship between vitamin D and cardiovascular and inflammatory risk factors as well as to uncover the molecular mechanisms responsible.

Certain Associations Between Iron Biomarkers and Total and γ' Fibrinogen and Plasma Clot Properties Are Mediated by Fibrinogen Genotypes.

Introduction: Evidence for the relationship between body iron and cardiovascular disease (CVD) is inconsistent and mechanisms involved remain poorly understood. Therefore, we first investigated whether there are linear or non-linear relationships between iron status and total and γ' fibrinogen as well as plasma fibrin clot properties and, second, determined whether there are interactions with iron biomarkers and fibrinogen and FXIII single nucleotide polymorphisms (SNPs) in relation to fibrinogen concentration and functionality. Methods: In this cross-sectional analysis of 2,010 apparently healthy Black South Africans we quantified total and γ' fibrinogen, serum iron, ferritin and transferrin using standardized methods and calculated transferrin saturation (TS). Clot architecture and lysis were explored with a global analytical turbidity assay. The SNPs were determined through an Illumina BeadXpress® platform. Results: Total, but not %γ', fibrinogen negatively correlated with serum iron concentrations, although both decreased over iron tertiles. %γ' fibrinogen correlated negatively with transferrin and decreased over the transferrin tertiles. A weak negative association between total fibrinogen and TS was detected with fibrinogen decreasing over the TS tertiles and categories based on TS. Lag time correlated positively with transferrin and increased over transferrin tertiles, when adjusting for fibrinogen. Before adjusting for fibrinogen, lag time was shorter in those with adequate iron status based on TS than other iron subcategories. Clot lysis time (CLT) negatively correlated with ferritin and was longer in the first than in the third ferritin tertile. Among iron status categories based on ferritin, only CLT differed and was longer in those with adequate iron than with iron-overload. CLT negatively correlated with TS, albeit weakly, shortened over the TS tertiles and was shorter in those with adequate iron based on TS categories. Interactions were observed between FGB SNPs and some of the markers of iron status investigated, in relation to the clot properties with the most prominent associations detected in homozygous carriers of the variant alleles for whom increased iron status was more beneficial than for those harboring the wild-type alleles. Iron modulated the influence of the SNPs so that for the majority iron was beneficial in respect of clot properties, but even more so for a minority group harboring specific variant alleles. Conclusion: This is the first large-scale epidemiological study to relate fibrinogen concentration and functionality to markers of iron status and to take genetic factors into consideration. We have detected a relationship between iron biomarkers and fibrinogen as well as clot characteristics that are influenced by the genetic make-up of an individual.

Comparison of DNA methylation clocks in Black South African men.

Aims: DNA methylation clocks are widely used to estimate biological age, although limited data are available on non-European ethnicities. This manuscript characterizes the behavior of five DNA methylation clocks in 120 older Black South African men. Methods: The age estimation accuracy of the Horvath, Hannum and skin and blood clocks and the relative age-related mortality risk and predicted time to death portrayed by the PhenoAge and GrimAge biomarkers are investigated, respectively. Results: The results confirm the tendency of DNA methylation clocks to underestimate the biological age of older individuals. GrimAge more accurately characterizes biological decline in this African cohort compared with PhenoAge owing to the unique inclusion of smoking-related damage in the GrimAge estimate. Conclusions: Each clock provides a different fraction of information regarding the aging body. It is essential to continue studying under-represented population groups to ensure methylation-derived indicators are robust and useful in all populations.

The Relationship of Circulating Homocysteine with Fibrinogen, Blood Pressure, and Other Cardiovascular Measures in African Adolescents.

OBJECTIVES: To evaluate the associations between homocysteine (Hcy) and cardiovascular health in South African adolescents. STUDY DESIGN: Circulating Hcy concentrations of 172 South African adolescents (105 girls, ages 13 to <18 years) were measured. Anthropometric and cardiovascular factors were also included and cross-sectionally analyzed through general linear models. RESULTS: Hcy correlated positively with body weight (P = .03; after adjusting for multiple testing, it was not regarded as significant) and muscle mass (P = .01), but negatively with fibrinogen concentrations (P = .001). Across Hcy tertiles, blood pressure produced approximating U-shaped curves, with differences between the middle and upper tertiles (all P < .02). Forty percent of the adolescents had elevated blood pressure, of whom 37% fell in the lowest and 38% in the highest Hcy tertiles. Hcy differed between the sexes (with boys having higher Hcy), but not between subgroups based on puberty, weight, stunting, smoking, or alcohol consumption. CONCLUSIONS: Both high and low Hcy could be early contributing risk factors to cardiovascular health. The associations between Hcy and blood pressure suggest that dietary and lifestyle manipulation, to achieve the optimal range of Hcy, may be beneficial in preventing Hcy-related hypertension in adulthood. The inverse relationship between Hcy and fibrinogen remains to be clarified.

The association of alcohol with circulating total fibrinogen and plasma clot density is mediated by fibrinogen and FXIII genotypes.

BACKGROUND: Alcohol consumption is associated with haemostasis and so may influence cardiovascular conditions. It is unknown whether the association of alcohol with total and γ' fibrinogen concentrations, as well as clot structure, are modulated by fibrinogen and factor (F) XIII single nucleotide polymorphisms (SNPs). METHODS: Total fibrinogen, γ' fibrinogen and clot properties of 2010 healthy Africans residing in South Africa were measured in relation to alcohol intake as well as its markers - gamma-glutamyltransferase (GGT), percentage carbohydrate deficient transferrin (%CDT), aspartate aminotransferase (AST), and alanine aminotransferase (ALT). Fourteen fibrinogen and two SNPs in the FXIII gene were genotyped to determine their influence. RESULTS: Alcohol intake and its markers correlated negatively with fibrinogen and clot lysis time (CLT) as well as with most of the clot properties. Percentage γ' fibrinogen correlated positively with AST and negatively with alcohol intake. We then stratified for alcohol intake and found inverse associations between γ' fibrinogen and both %CDT and GGT-CDT in consumers, but the positive association with AST remained only in abstainers. Alcohol intake and its markers modulated the influence of fibrinogen SNPs on total fibrinogen concentrations and the fibrinogen SNPs as well as an FXIII SNP on clot density (all p < 0.004). CONCLUSION/S: We show for the first time that some individuals harbour certain genotypes that, in combination with alcohol consumption, might predispose or protect them from haemostatic factors that might lead to the development of cardiovascular disease. Studies are needed to clarify the mechanisms related to the interplay between alcohol and the gene variants observed here.

Education, Smoking and CRP Genetics in Relation to C-Reactive Protein Concentrations in Black South Africans.

Because elevated circulating C-reactive protein (CRP) and low socio-economic status (SES), have both been implicated in cardiovascular disease development, we investigated whether SES factors associate with and interact with CRP polymorphisms in relation to the phenotype. Included in the study were 1569 black South Africans for whom CRP concentrations, 12 CRP single nucleotide polymorphisms (SNPs), cardiovascular health markers, and SES factors were known. None of the investigated SES aspects was found to associate with CRP concentrations when measured individually; however, in adjusted analyses, attaining twelve or more years of formal education resulted in a hypothetically predicted 18.9% lower CRP concentration. We also present the first evidence that active smokers with a C-allele at rs3093068 are at an increased risk of presenting with elevated CRP concentrations. Apart from education level, most SES factors on their own are not associated with the elevated CRP phenotype observed in black South Africans. However, these factors may collectively with other environmental, genetic, and behavioral aspects such as smoking, contribute to the elevated inflammation levels observed in this population. The gene-smoking status interaction in relation to inflammation observed here is of interest and if replicated could be used in at-risk individuals to serve as an additional motivation to quit.

Methylation vs. Protein Inflammatory Biomarkers and Their Associations With Cardiovascular Function.

DNA methylation data can be used to estimate proportions of leukocyte subsets retrospectively, when directly measured cell counts are unavailable. The methylation-derived neutrophil-to-lymphocyte and lymphocyte-to-monocyte ratios (mdNLRs and mdLMRs) have proven to be particularly useful as indicators of systemic inflammation. As with directly measured NLRs and LMRs, these methylation-derived ratios have been used as prognostic markers for cancer, although little is known about them in relation to other disorders with inflammatory components, such as cardiovascular disease (CVD). Recently, methylation of five genomic cytosine-phosphate-guanine sites (CpGs) was suggested as proxies for mdNLRs, potentially providing a cost-effective alternative when whole-genome methylation data are not available. This study compares seven methylation-derived inflammatory markers (mdNLR, mdLMR, and individual CpG sites) with five conventionally used protein-based inflammatory markers (C-reactive protein, interleukins 6 and 10, tumor-necrosis factor alpha, and interferon-gamma) and a protein-based inflammation score, in their associations with cardiovascular function (CVF) and risk. We found that markers of CVF were more strongly associated with methylation-derived than protein-based markers. In addition, the protein-based and methylation-derived inflammatory markers complemented rather than proxied one another in their contribution to the variance in CVF. There were no strong correlations between the methylation and protein markers either. Therefore, the methylation markers could offer unique information on the inflammatory process and are not just surrogate markers for inflammatory proteins. Although the five CpGs mirrored the mdNLR well in their capacity as proxies, they contributed to CVF above and beyond the mdNLR, suggesting possible added functional relevance. We conclude that methylation-derived indicators of inflammation enable individuals with increased CVD risk to be identified without measurement of protein-based inflammatory markers. In addition, the five CpGs investigated here could be useful surrogates for the NLR when the cost of array data cannot be met. Used in tandem, methylation-derived and protein-based inflammatory markers explain more variance than protein-based inflammatory markers alone.

Leveraging the urban-rural divide for epigenetic research.

Urbanization coincides with a complex change in environmental exposure and a rapid increase in noncommunicable diseases (NCDs). Epigenetics, including DNA methylation (DNAm), is thought to mediate part of the association between genetic/environmental exposure and NCDs. The urban-rural divide provides a unique opportunity to investigate the effect of the combined presence of multiple forms of environmental exposure on DNAm and the related increase in disease risk. This review evaluates the ability of three epidemiological study designs (migration, income-comparative and urban-rural designs) to investigate the role of DNAm in the association between urbanization and the rise in NCD prevalence. We also discuss the ability of each study design to address the gaps in the current literature, including the complex methylation-mediated risk attributable to the cluster of forms of exposure characterizing urban and rural living, while providing a platform for developing countries to leverage their demographic discrepancies in future research ventures.

Gene interactions observed with the HDL-c blood lipid, intakes of protein, sugar and biotin in relation to circulating homocysteine concentrations in a group of black South Africans.

BACKGROUND: Elevated homocysteine (Hcy) is associated with several pathologies. Gene-diet interactions related to Hcy might be used to customize dietary advice to reduce disease incidence. To explore this possibility, we investigated interactions between anthropometry, biochemical markers and diet and single-nucleotide polymorphisms (SNPs) in relation to Hcy concentrations. Five SNPs of Hcy-metabolizing enzymes were analyzed in 2010 black South Africans. RESULTS: Hcy was higher with each additional methylenetetrahydrofolate reductase (MTHFR) C677T minor allele copy, but was lower in methionine synthase (MTR) 2756AA homozygotes than heterozygotes. Individuals harboring cystathionine ß synthase (CBS) 833 T/844ins68 had lower Hcy concentrations than others. No interactive effects were observed with any of the anthropometrical markers. MTHFR C677T and CBS T833C/844ins68 homozygote minor allele carriers presented with lower Hcy as high density lipoprotein cholesterol (HDL-c) increased. Hcy concentrations were negatively associated with dietary protein and animal protein intake in the TT and TC genotypes, but positively in the CC genotype of CBS T833C/844ins68. Hcy was markedly higher in TT homozygotes of MTHFR C677T as added sugar intake increased. In CBS T833C/844ins68 major allele carriers, biotin intake was negatively associated with Hcy; but positively in those harboring the homozygous minor allele. CONCLUSIONS: The Hcy-SNP associations are modulated by diet and open up the possibility of invoking dietary interventions to treat hyperhomocysteinemia. Future intervention trials should further explore the observed gene-diet and gene-blood lipid interactions.

Replication and expansion of epigenome-wide association literature in a black South African population.

BACKGROUND: DNA methylation is associated with non-communicable diseases (NCDs) and related traits. Methylation data on continental African ancestries are currently scarce, even though there are known genetic and epigenetic differences between ancestral groups and a high burden of NCDs in Africans. Furthermore, the degree to which current literature can be extrapolated to the understudied African populations, who have limited resources to conduct independent large-scale analysis, is not yet known. To this end, this study examines the reproducibility of previously published epigenome-wide association studies of DNA methylation conducted in different ethinicities, on factors related to NCDs, by replicating findings in 120 South African Batswana men aged 45 to 88 years. In addition, novel associations between methylation and NCD-related factors are investigated using the Illumina EPIC BeadChip. RESULTS: Up to 86% of previously identified epigenome-wide associations with NCD-related traits (alcohol consumption, smoking, body mass index, waist circumference, C-reactive protein, blood lipids and age) overlapped with those observed here and a further 13% were directionally consistent. Only 1% of the replicated associations presented with effects opposite to findings in other ancestral groups. The majority of these inconcistencies were associated with population-specific genomic variance. In addition, we identified eight new 450K array CpG associations not previously reported in other ancestries, and 11 novel EPIC CpG associations with alcohol consumption. CONCLUSIONS: The successful replication of existing EWAS findings in this African population demonstrates that blood-based 450K EWAS findings from commonly investigated ancestries can largely be extrapolated to ethnicities for which epigenetic data are not yet available. Possible population-specific differences in 14% of the tested associations do, however, motivate the need to include a diversity of ethnic groups in future epigenetic research. The novel associations found with the enhanced coverage of the Illumina EPIC array support its usefulness to expand epigenetic literature.

CRP Genotypes Predict Increased Risk to Co-Present with Low Vitamin D and Elevated CRP in a Group of Healthy Black South African Women.

Low 25-hydroxyvitamin D (25(OH)D) and elevated C-reactive protein (CRP) concentrations are independently associated with adverse health outcomes, including cardiovascular disease (CVD). Although an inverse association between these factors has been described, the underlying mechanisms remain unknown. We postulate that environment-gene interactions, through which 25(OH)D interacts with single nucleotide polymorphisms (SNPs) within the CRP gene, modulate CRP; that certain CRP genotypes predispose individuals to a co-phenotype of low 25(OH)D and elevated CRP concentrations; and that this co-phenotype is associated with higher CVD risk. Twelve CRP SNPs were genotyped, and both 25(OH)D and CRP were quantified, in 505 black South African women. Alarmingly, 66% and 60% of the women presented with deficient/insufficient 25(OH)D and elevated CRP concentrations, respectively. CRP concentrations were higher in individuals with lower 25(OH)D concentrations. However, no 25(OH)D-CRP genotype interactions were evident. Several genotypes were associated with an altered risk of presenting with the co-phenotype, indicating a genetic predisposition. Women presenting with this co-phenotype had higher blood pressure and increased anthropometric measures, which may predispose them to develop CVD. We recommend increasing vitamin D fortification and supplementation efforts to reduce inflammation among black women with vitamin D deficiency, thereby possibly curbing diseases contingent on the co-phenotype described here.

Fibrinogen and clot-related phenotypes determined by fibrinogen polymorphisms: Independent and IL-6-interactive associations.

Interleukin-6 (IL-6) induces the expression of fibrinogen, and polymorphic variation within the fibrinogen genes is believed to alter the magnitude of this expression. The identification of the functional relevance of individual fibrinogen single nucleotide polymorphisms (SNPs) has been hindered by the high linkage disequilibrium (LD) reported in the European fibrinogen gene locus. This study investigated two novel and 12 known fibrinogen SNPs of potential functional relevance, in 2010 Tswana individuals known to have low LD. We aimed to identify functional polymorphisms that contribute to clot-related phenotypes and total and γ' fibrinogen concentrations independently and through their interaction with IL-6, by taking advantage of the high fibrinogen and IL-6 concentrations and the low LD reported in black South Africans. Fibrinogen was significantly associated with IL-6, thereby mediating associations of IL-6 with clot formation and structure, although attenuating the association of IL-6 with clot lysis time. None of the common European fibrinogen haplotypes was present in this study population. Putative functional fibrinogen SNPs FGB-rs7439150, rs1800789 (-1420G/A) and rs1800787 (-148C/T) were significantly associated with fibrinogen concentration and altered clot properties, with several associations significantly influenced by IL-6 concentrations. The impact of harbouring several minor fibrinogen SNP alleles on the association of IL-6 and fibrinogen concentration was cumulative, with possession of each additional minor allele showing a stronger relationship of IL-6 with fibrinogen. This was also reflected in differences in clot properties, suggesting potential clinical relevance. Therefore, when investigating the effect of fibrinogen genetics on fibrinogen concentrations and CVD outcome, the possible interactions with modulating factors and the fact that SNP effects seem to be additive should be taken into account.

Socio-Demographic and Lifestyle Factors Predict 5-Year Changes in Adiposity among a Group of Black South African Adults.

The rising prevalence of obesity and excessive adiposity are global public health concerns. Understanding determinants of changes in adiposity over time is critical for informing effective evidence-based prevention or treatment. However, limited information is available to achieve this objective. Cultural, demographic, environmental, and behavioral factors including socio-economic status (SES) likely account for obesity development. To this end, we related these variables to anthropometric measures in 1058 black adult Tswana-speaking South Africans who were HIV negative in a prospective study over five years. Body mass index (BMI) and waist circumference increased in both sexes, whereas triceps skinfold thickness remained the same. Over the five years, women moved to higher BMI categories and more were diagnosed with central obesity. Age correlated negatively, whereas SES, physical activity, energy, and fat intake correlated positively with adiposity markers in women. In men, SES, marital status, physical activity, and being urban predicted increases in adiposity. For women, SES and urbanicity increased, whereas menopause and smoking decreased adiposity. Among men, smokers had less change in BMI than those that never smoked over five years. Our findings suggest that interventions, focusing on the urban living, the married and those with the highest SES-the high-risk groups identified herein-are of primary importance to contain morbidity and premature mortality due to obesity in black South Africans.

Candidate gene analysis of the fibrinogen phenotype reveals the importance of polygenic co-regulation.

Fibrinogen and its functional aspects have been linked to cardiovascular disease. There is vast discrepancy between the heritability of fibrinogen concentrations observed in twin studies and the heritability uncovered by genome wide association studies. We postulate that some of the missing heritability might be explained by the pleiotropic and polygenic co-regulation of fibrinogen through multiple targeted genes, apart from the fibrinogen genes themselves. To this end we investigated single nucleotide polymorphisms (SNPs) in genes coding for phenotypes associated with total and γ' fibrinogen concentrations and clot properties. Their individual and accumulative associations with the fibrinogen variables were explored together with possible co-regulatory processes as a result of the gain and loss of transcription factor binding sites (TFBS). Seventy-eight SNPs spanning the APOB, APOE, CBS, CRP, F13A1, FGA, FGB, FGG, LDL-R, MTHFR, MTR, PCSK-9 and SERPINE-1 genes were included in the final analysis. A novel PCSK-9 SNP (rs369066144) was identified in this population, which associated significantly (p=0.04) with clot lysis time (CLT). Apart from SNPs in the fibrinogen (FGA, FGB and FGG) and FXIII (F13A1) genes, the fibrinogen phenotypes were also associated with SNPs in genes playing a role in lipid homeostasis (LDL-R, PCSK-9) together with CBS and CRP polymorphisms (particularly, CRP-rs3093068). The genetic risk scores, presenting accumulative genetic risk, were significantly associated (p≤0.007) with total and γ' fibrinogen concentrations, lag time, slope and CLT, highlighting the importance of a polygenetic approach in determining complex phenotypes. SNPs significantly associated with the fibrinogen phenotypes, resulted in a total of 75 TFBS changes, of which 35 resulted in a loss and 40 in a gain of TFBS. In terms of co-regulation, V$IRF4.02, V$E2FF and V$HIFF were of particular importance. The investigation into TFBS provided valuable insight as to how sequence divergences in seemingly unrelated genes can result in transcriptional co-regulation of the fibrinogen phenotypes. The observed associations between the identified SNPs and the fibrinogen phenotypes therefore do not imply direct effects on cardiovascular disease outcomes, but may prove useful in explaining more of the genetic regulation of the investigated fibrinogen phenotypes.

Degree of obesity influences the relationship of PAI-1 with body fat distribution and metabolic variables in African women.

INTRODUCTION: Although the relationship of plasminogen activator inhibitor-1 (PAI-1) with obesity has been well established, the relationship of PAI-1 with different body fat distribution patterns is less clear particularly in non-white ethnicities. METHODS: We investigated the cross-sectional association of PAI-1act with body fat % and two different body fat distribution patterns, namely sarcopenic obesity (SO) and visceral (VAT) compared to subcutaneous (SCAT) abdominal obesity, in 246 healthy African women by creating sub-groups according to different body fat distribution patterns. RESULTS: The PAI-1act level of the SO group did not differ significantly from that of the excessive % body fat, non-sarcopenic group (p=0.8). The relationship of PAI-1act, with body fat %, insulin, triglycerides and appendicular skeletal mass (ASM) was influenced by body fat distribution patterns and degree of obesity. PAI-1act was higher (1.65 vs 0.16U/ml; p=0.001) in women with a proportionally higher abdominal VAT compared to higher abdominal SCAT compartment in the total study population, but not in the centrally obese sub-group (1.72 vs 0.83U/ml; p=0.5). Multiple regression models indicated that body fat % per se did not contribute significantly to PAI-1act variance in women with increased fat mass. CONCLUSION: Fat distribution patterns and degree of obesity influenced the association of PAI-1act with insulin, triglycerides, ASM and body fat % in African women. In centrally obese women, abdominal VAT no longer contributed more to plasma PAI-1act, than abdominal SCAT. Inflammation and endothelial dysfunction contributed more to PAI-1act variance in obese African women than did body fat % per se.

Homocysteine influences blood clot properties alone and in combination with total fibrinogen but not with fibrinogen γ' in Africans.

Simultaneously increased fibrinogen and homocysteine (Hcy) in blood are believed to elevate the risk of cardiovascular disease mortality. However, the pathophysiological mechanisms involved are unknown. We sought to determine whether Hcy or its genetic determinants influence blood clot properties alone or in combination with fibrinogen. In addition, we investigated, for the first time, the gamma prime (γ') isoform of fibrinogen with Hcy in relation to clot architecture and lysis. Single-nucleotide polymorphisms, Hcy and hemostatic variables, including clot lysis, determined with a global fibrinolytic assay [giving lag time, slope, maximum absorbance and clot lysis time (CLT)], were measured in 1867 healthy black South Africans and cross-sectionally analyzed. Increasing Hcy did not affect fiber cross-sectional area (maximum absorbance). However, it decreased the time needed to initiate the coagulation cascade and for fibrin fibers to grow (lag time), it increased the tempo of lateral aggregation (slope) and reduced CLT. None of the single-nucleotide polymorphisms measured had effects on clot properties. Combined effects were observed between Hcy and total fibrinogen in predicting CLT. Fibrinogen γ', which affected markers of the fibrinolytic assay, did not have conjoint effects with Hcy. We believe that there is value in recognizing the combined effects of Hcy and fibrinogen, but not its γ' isoform in relation to clot structure and lysis. The enhanced fibrinolysis rate observed in patients with low fibrinogen and high Hcy may have adverse consequences for health if it disturbs hemostasis and results in a bleeding tendency.

The contribution of genetic and environmental factors to changes in total and γ' fibrinogen over 5 years.

INTRODUCTION: Increased fibrinogen is associated with cardiovascular disease risk. It is, however, not known to what extend environmental and genetic factors and/or their interaction influence changes in total and γ' fibrinogen over time. We aimed to determine how variation within the fibrinogen gene as well as environmental factors influence the change in total and γ' fibrinogen over time, and also whether gene-environment interactions influence total and γ' fibrinogen on a cross-sectional and prospective level in Africans. MATERIALS AND METHODS: This prospective study consisted of 2010 participants at baseline and 1288 participants at follow-up (5 years). RESULTS: The gene-environment interactions that were associated with fibrinogen concentration on a cross-sectional level were: FGA 2224 G>A (rs2070011) with age (p=0.005), FGB Arg448Lys (rs4220) with HIV status (p<0.0001) and FGB 1038 G>A (rs1800791) with HbA1c (p=0.01). The only factor that independently influenced the change in total fibrinogen levels over time, was baseline CRP (p<0.0001) and FGG 10034 C>T (rs2066865) was the only single nucleotide polymorphism that independently influenced the change in fibrinogen γ' levels over time (p=0.02). Only the FGG 9340 T>C (rs1049636) with HbA1c interaction was found to predict change in total fibrinogen concentrations over time (p=0.005). CONCLUSIONS: Gene-environment interactions influenced fibrinogen levels cross-sectionally and also mediated changes in levels over time.