Hypertension Susceptibility Gene In Chinese Han Population And Blood Pressure Reactivity In The Heritability Analysis

Hypertension is an important worldwide public-health challenge because of it is one of the most important modifiable risk factors for CVD and renal disease.With changes in lifestyle and diet and an increase in life expectancy,the morbidity and mortality of hypertension have greatly increased.According to previous national surveys conducted in China,the prevalence and absolute numbers of hypertension have increased dramatically during the past several decades.It was estimated that 18.8%of the adult population in 2002 had hypertension,and the total number of adults with hypertension was 160 million.Essential hypertension is considered to be a typical complex disease and is influenced by both genetic and environmental factors.The genetic contribution to blood pressure(BP) variation ranges from 30 to 50%.It is likely that a number of genes with smaller effects account for the heritability of this complex disorder.Since the biological process of hypertension involves multiple physiological pathways,each of which may be affected by multiple gene products,the underlying genetics of hypertension is not only based on multiple genes with minor effects,but also on gene-gene interactions.To test this hypothesis,we focused on 11 representive candidate genes involved in multiple biochemical pathways that have been implicated in the development and progression of hypertension(ACE,AGTR1,CYP11B2, ADRA1A,ADRB2,TH,LPL,GNB3,NOS3,GRK4,WNK4),performed PCR-PFLP or direct sequencing for the genotyping,and tested their associations with hypertension in a case-control study(in this paper,the associations of three genes, ADRA1A,CYP11B2 and TH,with hypertension were shown).We also evaluated the influence of gene-gene interactions on the risk of hypertension.Furthermore,the biological relevance of the significantly associated rs2070762 in TH gene was studied by various functional assays in vitro.Genetic epidemiology studies have identified many biological candidate genes that were related to BP.However,it is generally found that association of any of these genes accounts for a very little variance.Thus,genetic factors may not independently affect BP,and the effect on BP is influenced by environmental factors.The increased dietary intake of salt as an importantly environmental factor plays a pathogenic role in essential hypertension.Epidemiological studies have shown that dietary sodium intake was positively associated with blood pressure.However,BP response to dietary sodium intake varies considerably among individual subjects.Genetic factors might play an important role in determining the BP responses of an individual subject to dietary sodium intake.To evaluate the effect of genetic factors,the Genetic Epidemiology Network of Salt Sensitivity(GenSalt) Study was designed to examine the genetic influence on BP responses to dietary sodium and potassium intake in Chinese families.Our study improve our understanding of mechanisms of hypertension and multiple gene-gene/gene-environment interactions.The findings support the multigenic nature of the etiology of essential hypertension and propose a potential gene-gene interactive model for future studies.Our study also measures the heritability of BP response to dietary sodium intake.Understanding the genetic effects has important implications for identifying important novel genes for salt sensitivity of BP,and contributes to individualized prevention and therapy on hypertension.SECTION ONE ADRA1A gene variants and hypertensionα₁-adrenergic receptors(α_1A-,α_1B-,α_1D-) are a family of G protein-coupled transmembrane receptors that mediate actions in the sympathetic nervous system through binding the catecholamines,epinephrine and norepinephrine,α₁-adrenergic receptors regulate cardiovascular function and may play a role on the development of hypertension.Our previous report linked human chromosome 8p22,which is near the location of humanα_1Aadrenergic receptor(ADRA1A) gene,to essential hypertension and blood pressure.Upon this,here we present a case-control study on the relationship between ADRA1A gene variants and hypertension in northern Han Chinese.All DNA samples and clinical data were collected from the International Collaborative Study of Cardiovascular Disease in Asia(InterASIA).The measurements and interviews were taken under standard conditions.We enrolled 480 unrelated stage-2 hypertensive patients and their individually age-(within 2 years), gender-and area-matched healthy control subjects.Stage-2 hypertension was defined as an average SBP≥160 mm Hg and/or DBP≥100 mm Hg.Control subjects had SBP and DBP＜140 mm Hg and＜90 mm Hg.Subjects with a clinical history of secondary hypertension,coronary heart disease and diabetes were excluded from the study.The interview included questions related to the diagnosis and treatment of hypertension.Anthropometrical measurements,including height,weight,waist and hip circumferences were taken.Concentrations of serum lipids were also determined by standard protocols.Seven polymorphisms were identified by direct sequencing of genomic DNA derived from 48 randomly recruited hypertensives and 48 healthy subjects.We used McNemar's test to estimate the odds ratio(OR) for hypertension for each polymorphism.Conditional logistic regression analysis was used to assess whether the genetic variation was associated independently with hypertension after adjustment for covariates.The extent of pairwise linkage disequilibrium was expressed in terms of D',and the pairwise correlation was presented as r².Haplotype Trend Regression(HTR) was used to test the association between each haplotype and hypertension..We observed significantly higher frequencies of the 347Arg allele and 2547G alleles in the cases compared to their controls(P=0.04 and 0.007,respectively). McNemar's test revealed carriers of 2547G alleles were at a higher risk for EH with an OR of 3.00(95%CI:1.23-8.35).We then performed a conditional logistic regression to adjust the effects of conventional risk factors,revealing an OR of 2.84 for carriers of 2547G allele(95%CI:1.15-6.99).With the haplotypic probabilities estimated using the PHASE software,we performed the Haplotype Trend Regression analysis,showing a significant association between haplotype 7 and EH(P=0.02), after adjustment for conventional risk factors.Our findings suggest that the genetic variations in the ADRA1A gene are significantly associated with EH and may play an important role in the development of essential hypertension in this Chinese population. SECTION TWO CYP11B2 gene haplotypes and hypertensionHuman aldosterone synthase(CYP11B2),a steroid 11β-hydroxylase as well as an 18-hydroxylase and an 18-oxidase,catalyzes the terminal steps of aldosterone biosynthesis in the zona glomerulosa cells of the adrenal.The objective of this study was to investigate the association of polymorphisms in the aldosterone synthase gene CYP11B2(T-344C,Lys173Arg,and an intronic conversion[IC]) with stage-2 hypertension in northern Han Chinese.A total of 503 hypertensives and their age-, gender- and area-matched controls were included in this study.The female hypertensives had significantly higher frequencies of the -344T,173Lys and IC-conversion alleles(p=0.002,0.002 and 0.014,respectively).The estimated frequency of haplotype composed of the -344T,173Lys and IC-conversion alleles (haplotype 4) was significantly higher in the female hypertensives compared with their controls(p=0.007).Using a multivariate score test,we found that haplotype 4 remained associated with female hypertension after the adjustment for covariates (p=0.003),while the haplotype 3 of T-Arg-WT showed a protective effect both in the males and in the females(p=0.03 and 0.006,respectively).These results indicate that the 173Lys and the IC-conversion allele of the CYP11B2 gene confer an increased risk for stage-2 hypertension in northern Han Chinese women.SECTION THREE Association study of TH Gene with Hypertension and Primary Functional AnalysisThe tyrosine hydroxylase(TH) gene encodes the rate-limiting enzyme of catecholamine biosynthesis,thus it plays a pivotal role in the physiology of sympathetic nervous system.However,as one of the candidate genes of essential hypertension,the relation between the variants of TH gene and hypertension had not been extensively studied.We designed a case-control study consisting of 490 normotensive(NT) controls and 503 hypertensive(HT) cases matched in area,age and gender to systematically investigate the relationship between TH gene and hypertension.Based on the HapMap and dbSNP data,four SNPs,rs6356,rs6357, rs2070762 and rs1800033 in the TH gene were selected for genotyping.The SNP rs1800033 did not present polymorphic in our studied population.No significant distribution differences were observed for rs6356 and rs6357 between HT and NT groups.However,both the genotype and allele frequencies of rs2070762 showed significant differences between cases and controls(P＜0.001 and P=0.005, respectively).The SNP rs6356 and rs6357 were both in weak linkage disequilibrium with rs2070762.In haplotype analysis,total eight haplotypes are observed in the entire population and the overall frequency distributions differ significantly between HT group and NT group.Specifically,haplotype A-A-C(in the order of rs6356-rs6357-rs2070762) occurs only in HT group and A-G-C occurs more often in HT subjects than in NT subjects(P=0.003 and P=0.013,respectively).Moreover, haplotype G-G-C carriers get about 1.83-fold higher risk for hypertension than non-carriers(p=0.0049) after adjustment for covariates.Functional analyses showed that C allele confered a two-fold higher promoter activity when fused to a heterologous promoter than the T allele,and unidentified nuclear factor(s) binded specifically to T allele,but not C allele.This suggested the existence of transcriptional repressor(s) responsible for the lower promoter activity regulated by T allele.These results provide evidence for an association of the functional SNP rs2070762 of TH gene with essential hypertension and suggest that the up-regulated expression of TH gene by rs2070762 C allele without binding transcriptional repressor(s) might be involved in the pathogenesis of hypertension.SECTION FOUR Association Study with 33 SNPs in 11 Candidate Genes for Hypertension Essential hypertension is considered to be a typical complex disease with multifactorial etiology,which leads to inconsistent findings in genetic studies.One possibility of failure to replicate some single-locus results is that the underlying genetics of hypertension is not only based on multiple genes with minor effects,but also on gene-gene interactions.To test this hypothesis,a case-control study was constructed in Chinese,detecting both single locus and multilocus effects.Eleven candidate genes were selected from related genes that have been implicated in the development and progression of hypertension and thirty-three polymorphisms were evaluated in 503 hypertension patients and 490 age-,gender-matched controls. Single-locus associations,using traditional logistic regression analyses,and multi-locus associations,using classification and regression trees(CART) and multivariate adaptive regression splines(MARS) were both explored in this study. Final models were selected using either Bonferroni correction or cross-validation. Three polymorphisms,TH^*rs2070762,ADRB2^*Q27E and GRK4^*A486V,were found to be independently associated with essential hypertension in Chinese.In addition to these individual predictors,a potential interaction of CYP11B2-AGTR1 is also involved in the etiology of hypertension.These findings support the multigenic nature of the etiology of essential hypertension and propose a potential gene-gene interactive model for future studies.SECTION FIVE Heritability of Blood Pressure Responses to Dietary Sodium and Potassium IntakeThe heritability of blood pressure responses to dietary intervention has not been well studied.We examined the heritability of blood pressure responses to dietary sodium and potassium intake in a family feeding study among 1906 study participants living in rural North China.The dietary intervention included a 7-day low-sodium feeding(51.3mmol per day),a 7-day high-sodium feeding(307.8 mmol per day),and a 7-day high-sodium plus potassium supplementation(60 mmol per day).Blood pressure was measured 9 times during the 3-day baseline period preceding the intervention and also during the last 3 days of each intervention phase using a random-0 sphygmomanometer.Heritability was computed using maximum likelihood methods under a variance components model as implemented in the computer program SOLAR.The heritabilities of baseline blood pressure were 0.31 for systolic, 0.32 for diastolic,and 0.34 for mean arterial pressure.The heritabilities increased significantly under dietary intervention and were 0.49,0.49,and 0.51 during low sodium;0.47,0.49,and 0.51 during high sodium;and 0.51,0.52,and 0.53 during potassium supplementation for systolic,diastolic,and mean arterial pressure, respectively.The heritabilities for percentage of blood pressure responses to low sodium were 0.20,0.21,and 0.23;to high-sodium were 0.22,0.33,and 0.33;and to potassium supplementation were 0.24,0.21,and 0.25 for systolic,diastolic,and mean arterial pressure,respectively.Our study indicated that the heritabilities of blood pressure under controlled dietary sodium and potassium intake were significantly higher than those under a usual diet.In addition,the heritabilities of blood pressure responses to dietary sodium and potassium intake were moderate in this study population.