The Study Of Association Of The Elastin And Its Metabolic Pathway Associated Genes SNPs And Haplotypes With Intracerebral Hemorrhage In Chinese Han Of Changsha Area

Background:Stroke is the second most common cause of death and a leading cause of adult disability worldwide. Intracerebral hemorrhage (ICH) is one of the most serious types of stroke with extraordinary high morbidity and mortality rate. Due to the lack of effective alternative therapies, accurate risk factors identification and preventive interventions have recently become the key issues in addressing the public health burden of ICH. ICH is a late-onset multifactorial disease that results from the contributions of environmental and a large number of minor genetic risk factors. Studies on candidate gene single nuclear polymorphisms (SNPs) will help us to uncover the genetic architectures of ICH and provide valid scientific basis for prevention and treatment of ICH.Our epidemiological study conducted on ICH has shown that Changsha had an extraordinarily high proportion of ICH with obvious family aggregation. In addition, hypertension and atherosclerosis were the major risk factors for ICH. Moreover, studies suggested that elastin family played a crucial role in the development of hypertension, atherosclerosis and micro-intracranial aneurysms, which further increased the susceptibility for ICH. However, there is dearth of information concerning the relationship between these genes'SNPs and ICH, especially ICH with family history. The aim of this study was to investigate the relationship between ELN and its metabolic pathway related MMP-9, TIMP-1genes'SNPs in Han people with ICH in Changsha, Hunan.Methods: This study enrolled ICH patients and relatives of Chinese Han in Changsha who were consecutively admitted to the Neurology Department of Xiangya Hospital, Central South University, between January2006and January2011. The control group was recruited from healthy Changsha Han Chinese for check-up in Xiangya Hospital. All subjects were divided into three groups:1. Intracerebral hemorrhage with family history group (ICHFH):339members from82pedigrees,189men and150women, including:82ICH patients,165first-degree relatives,30second-degree relatives,12third-degree relatives and50relatives with no kinship with proband.2. Sporadic intracerebral hemorrhage (SICH) group:418patients with sporadic intracerebral hemorrhage,280men and138women.3. Control group:206healthy controls,130men and76women. ICHFH group, SICH group and control group were comparable in age and sex.Selection of candidate gene's SNPs were based on the following criteria:①SNPs that have been proved to be associated with artery diseases, especially hypertension, atherosclerosis and aneurysms;②SNPs located at exons that cause amino acid changes or located at transcriptional regulation regions;③SNPs with minor allele frequency>5%to improve genetic calculator power;④The use of Haploview software to screen tag SNPs. With respect to the above basis, Multiplex SNaPshot technique and DNA sequencing were used to genotype8SNPs of3genes associated with ELN metabolism of all subjects, including: rs2071307, rs34208922, rs2856728and rs41350445of ELN gene; rs17576and rs2250889of MMP-9gene; rs4898and rs2070584of TIMP-1gene.We took a two-step method to analyze the genotyping results:①In step one, we analyzed the SICH and control group based on case-control study. The frequencies of alleles and genotypes between the two groups were analyzed using χ2test. Blood pressure levels were compared among genotypes using ANOVA analysis (t test for alleles). We performed all these analyses with SPSS software (Version16.0);②In step two, based on familial association study, we carried out transmission disequilibrium test (TDT) and association analysis by using FBAT software to screen out the causative mutation for familial intracerebral hemorrhage.Results:1. The distribution features of the candidate genes'SNPs:There were ELN gene rs207137,rs34208922,rs2856728,rs41350445polymorphisms, MMP-9gene rs17576,rs2250889polymorphisms and TIMP-1gene rs4898,rs2070584polymorphisms in Changsha Han population. The distributions of these SNPs'genotypes and/or alleles showed obvious racial and geographic differences.2. The results of the case-control study in SICH and normal controls were as follows:①There was significant difference of ELN gene rs2856728genotypes distribution between SICH group and control group (χ2=15.414, p<0.001). The frequency of rs2856728T allele was significantly higher in SICH group than in controls (χ2=13.123, p<0.001). There was significant difference of ELN gene rs41350445genotypes'distribution between SICHs and controls (χ2=4.241, p=0.039). However, there were no differences in genotype and allele frequencies of ELN gene rs2071307and rs34208922between SICH and control groups.②There was significant difference of MMP-9gene rs17576genotypes distribution between SICHs and controls (χ2=7.522, p=0.023), while no significant differences were observed in MMP-9gene rs2250889distribution (P>0.05).③There was significant difference of TIMP-1gene rs2070584allele distribution between male SICHs and male controls. The frequency of the G allele of rs2070584was significantly higher in male SICHs than in male controls (χ2=5.814, P=0.016). There were no significant differences in genotype or allele frequencies of TIMP-1gene rs2070584between the two groups (P>0.05).3. The results of FBAT of ICHFH pedigrees showed the following characteristics:①TDT analysis revealed that C alleles of ELN gene rs41350445was over-transmitted, and was associated with ICHFH susceptibility (Z=2.180, P=0.029). Both single SNP per test and haplotype analysis showed that ELN gene rs2071307, rs34208922and rs2856728were not over-transmitted in ICHFH pedigrees (P>0.05).②MMP-9gene rs17576,rs2250889revealed no evidence of transmission disequilibrium with ICHFH (P>0.05). Haplotyping and Haplotype Frequency Estimation of the two sites didn't reveal any significant results, either (P>0.05).4. Analysis of ENL, MMP-9and TINP-1genes'polymorphisms with blood pressure levels in Changsha Chinese Han:①The levels of systolic pressure, diastolic pressure and mean arterial blood pressure were statistically different among genotypes of ELN gene rs2856728in both SICH and control groups. However, there were no significant differences of blood pressure levels among genotypes of rs2071307, rs34208922and rs41350445polymorphisms of ELN gene either in SICHs or in control groups.②The levels of systolic pressure and mean arterial blood pressure were different among genotypes of MMP-9gene rs17576in controls while there were no significant differences of blood levels among genotypes of MMP-9gene rs2250889polymorphisms.③The levels of systolic pressure and mean arterial blood pressure of TIMP-1gene rs2070584TG genotype were significantly higher than those of the GG genotype in female SICHs. There were no significant differences of systolic blood pressure, diastolic pressure and mean arterial blood pressure levels among rs4898genotypes of TIMP-1gene (P>0.05). Conclusions:1. The ELN gene rs2856728,rs41350445, MMP-9gene rs17576and TIMP-1gene rs2070584polymorphism could be associated with increased susceptibility of ICH in Changsha Han Chinese population.2. There is no association of ELN gene rs207137,rs34208922, MMP-9gene rs2250889and TIMP-1gene rs4898polymorphisms with ICH in Changsha Han Chinese population.3. There is no association between haplotypes comprised by ELN gene rs2071307,rs34208922,rs2856728,rs41350445, MMP-9gene rs17576,rs2250889polymorphisms and ICHFH in Changsha Han Chinese population.4. The polymorphisms of ELN gene rs2856728, MMP-9gene rs17576and TIMP-1rs2070584could be associated with high blood pressure in Changsha Han population. Furthermore, rs2856728polymorphism may increase the susceptibility of ICH by affecting blood pressure.