The Research Into High Risk Factors And NFATC1 Gene Polymorphism Associated With Primary Cerebral Hemorrhage

ObjectiveCerebral hemorrhage refers to the non-invasive spontaneous bleeding Of the blood vessels within the brain parenchyma. Bleeding can also be broken into the brain ventricles or subarachnoid space.ICH accounts for 10%-15% of all stroke, Cerebral hemorrhage can occur in any part of the brain parenchyma,not only can be single, but also can be multiple.Primary ICH is usually due to a long-term hypertension or cerebral amyloid angiopathy or cerebral atherosclerosis which lead the small blood vessels or small arteries to spontaneous rupture.Cerebral hemorrhage as a multifactorial disease affect by environmental factors and genetic factors epidemiological studies have shown that high blood pressure, hyperlipidemia, age, excessive alcohol consumption, smoking, Body Mass Index, diabetes are risk factors for ICH. In recent years, with the phenomenon of familial aggregation of cerebral hemorrhage was found, genetic factors in the pathogenesis of cerebral hemorrhage caused by people's concern and become a research hotspot.In recent years, with the phenomenon of familial aggregation of cerebral hemorrhage was found, genetic factors in the pathogenesis of cerebral hemorrhage caused by people's concern, become a research hotspot. Into the 21st century an increasing number of studies have found that NFAT family, as a based factor of many intracellular signal transduction pathway that can regulate expression levels of many factors in T cells, B cells and many of cytokines (IL-2, IL-3, IL-4, IL-5, IL-6, granulocyte macrophage colony-stimulating factor, tumor necrosis factor-a, Cyclooxygenase-2, Apob,nur77 etc.) Because TNF-a is closely related to essential hypertension and abnormal lipid metabolism and atherosclerosis, COX-2 is closely related to atherosclerosis, Apob,IL-6, nur77, SREBP-1 are closely related to abnormal lipid metabolism, NFATC1 mediates the activation of TNF-a. There are different transcription factor potential binding site of AP-1/Ets/NFAT in the TNF-a initiation factor. These transcription factors play an important role on the activation of TNF-a gene, NFATC1 mediates COX-2 expression, NFATC1 expression and COX-2 expression is positively correlated, There are SREBP-1 binding sites on exon-2 of NFATC1, So NFATC1 directly involved in SREBP-1 negative regulation.NFATC1 also mediates expression of the nur77. NFATC1 makes T cells extensively infiltrate into the intima and causes to inflammation, through mediating expression of a number of factors in T cells.Hypertension is an important independent risk factors of intracerebral hemorrhage Abnormal lipid metabolism and cerebral artery atherosclerosis are important pathological basis of cerebral hemorrhage, NFATC1 control the factors which are closely related to high blood pressure, arteriosclerosis and dyslipidemia.NFATCl is involved in intimal lesions of the inflammatory response through regulating the factoris of T cells, So we have reasons to believe that transcription factor NFATC1 is closely related to cerebral hemorrhagic stroke. At present the expression of NFATC1 in the cerebral hemorrhage and its mechanism is not yet clear, Especially there is no study about effect of polymorphisms of NFATC1 on cerebral hemorrhage at home and abroad. Thus requires us to do a lot of works to make it clear that the effect of polymorphisms of NFATC1 on cerebral hemorrhage and to explore the pathogenesis of cerebral hemorrhage from the molecular level, If this correlation exists, We can find high-risk group of cerebral hemorrhage early and execute individualized treatment through finding out the gene in the population, which can more effectively prevent and control the occurrence of cerebral hemorrhage and will bring new hope to the prevention and treatment of cerebral hemorrhage.Material and menthodA case-control design was intruduced, The subject of ICH group were consecutively recruited from northern China region patients with a total of 230 cases.male 144 and female 86, aged 33-85 years old, Han nationality, no consanguinity with each other. The controls matched by gender, age,nationality and ethnic orign were recruited from the same patients geographic region, with a total of 233 cases, male 146 and female 87. This study methods includes questionnaire survey and investigation etc. are the basic situations of the subject were recorded(gender, age, BMI, systolic blood pressure, diastolic blood pressure, addicted to tobacco and alcohol history, etc.) Hitachi 7600 automatic biochemical analyzers were used to detect CHO,TG,LDL,HDL,Apoe-a,Apoe-b of people in patients group and controls group. The functional SNPS of NFATC1 exon-2 were genotyped by Dideoxy-termination sequencing method,The variable number tandem repeat(VNTR) of intron-7 was genotyped by polymerase chain reaction analysis, The snp rs754093(Cys751Gly) of NFATC1 exon-9 was genotyped by polymerase chain reaction and restriction fragment length polymorphism (PCR-RFLP) analysis. Genotyping results were confirmed by dideoxy-termination sequencing method Thex2 goodness of fit chi-square was used to test for deviation of genotype distribution from Hardy-Weinberg equilibrium, Statistical Packages for Social Sciences (SPSS 13.0) and EXCEL was used to the establish databases. The measurement data were tested for normal distribution, All statistics of normal distribution were expressed by mean±standard. Statistics of Skewed distribution were performed logarithmic conversion and then data processed.T test was used to assess difference about blood lipids and other measurement data between case group and control group,χ2 test was used to assess difference about Count data of basic background information between case group and control group Four grid table and the R×C listx2 test was used to assess difference about allele and genotype frequency between case group and control group. One-Way ANOVA was used to assess difference about serum lipid of each genotype in the case group Significant difference of statistic was considered by P< 0.05. Logistic regression analysis was used to judge risk degree of various risk factors of cerebral hemorrhage. Their respective odds ratio (OR) and 95% confidence interval (CI) were calculated. Statistical significant difference was considered by OR of 95% CI which does not contain 1.Result1.There was no statistical difference in gender, age, body mass index (BMI) between two groups(P> 0.05). It was significantly higher comparing hypertension, Diabetesmellitus, hyperlipidemia, smoking, drinking in cerebral hemorrhage group with that in the the control group,(P<0.05). There was no significant statistical difference in triglyceride (TG), apolipoprotein a (Apoa) between acerebral hemorrhage group and the control group(P> 0.05); It was significantly higher comparing the high-density lipoprotein (HDL), low-density lipoprotein (LDL), apolipoprotein b (Apob), systolic blood pressure (SBP), diastolic blood pressure (DBP) in the cerebral hemorrhage group with that in the control group, while It was significantly lower comparing CHO lesterol (CHO) in the cerebral hemorrhage group with that in the control group. There was significant difference of statistic in two groups (P<0.05).2.The genotype frequencies of all polymorphism conformed to the expectations of Hardy-Weinberg equilibrium (P> 0.05),It was suggested that the selected samples could represent the population and were suitable for genetic analysis.3.The SNPS as rs1051978, rs62096875, rsl2605457, rs1063669, rs61731547 of NFATC1 exon-2 were genotyped by Dideoxy-termination sequencing method The rs 1051978 is A/C two-states SNP, It was lower comparing A/A genotype frequencies in cerebral hemorrhage group with that in the the control group.while It was higher comparing the A/C genotype frequency in cerebral hemorrhage group with that in the the control group. Genotype frequenciesχ2=0.349, P=0.555, Allele frequenciesχ2=0.35, P= 0.661. There was no statistical difference in genotype frequencies and allele frequencies between two groups. rs62096875 is G/G homozygotes, rs 12605457 is C/C homozygous, rs 1063669 is G/G homozygotes, rs61731547 is C/C homozygotes. No new mutations were found.4.It was lower comparing variable number tandem repeat (VNTR) L/L, S/S genotype frequencies of intron-7 in cerebral hemorrhage group with that in the the control group.while It was higher comparing the L/S genotype frequency in cerebral hemorrhage group with that in the the control group. Genotype frequenciesχ2= 0.957, P= 0.620, Allele frequenciesχ2= 0.12, P= 0.727. There was no statistical difference in genotype frequencies and allele frequencies between two groups.5.It was lower comparing SNP rs754093 T/T, T/G genotype frequencies of NFATC1 exon-9 in cerebral hemorrhage group with that in the the control group, while It was significantly higher comparing the the G/G genotype frequency in cerebral hemorrhage group with that in the the control group. Genotype frequenciesχ2= 6.343, df= 2, P= 0.042; G and T/G allele frequenciesχ2= 6.39, df= 1, P= 0.011,OR= 1.411,95% CI= 1.080～1.843. There was significant difference of statistic in genotype frequencies and allele frequencies between two groups (P<0.05). 6.There was significant difference of statistic in CHO, LDL-C, Apob, among RS754093 T/G genotypes in Cerebral hemorrhage group by ANOVA. Pairwise comparison between the two groups showed that it was significantly higher comparing CHO, LDL-C, Apob in the GG genotype group with that in T/T and T/G genotypes group. There was significant difference of statistic in CHO between GG genotype and T /T and T/G genotype F=10.423, P=0.000. There was significant difference of statistic in LDL between GG genotype and T/T and T/G genotype F=23.398, P= 0.000. There was significant difference of statistic in Apob between GG genotype and T /T and T/G genotype F=32.176 P=0.000. It was higher comparing SBP in GG genotype with that in T/G genotype. It was significantly higher comparing SBP in GG genotype with that in T/T genotype. There was significant difference of statistic in SBP between GG genotype and TT genotype F=2.129, P=0.042. There was no statistical difference in diabetes, tobacco, alcohol; BMI etc among RS754093 T/G genotypes in Cerebral hemorrhage group.7.Logistic regression analysis showed NFATC1 rs754093 GG genotype is one of independent risk factors for cerebral hemorrhage (OR=3.902,95% CI 1.735～8.775, P =0.001). In addition, cerebral hemorrhage associated withThe other risk factors for cerebral hemorrhage are, LDL, Apob and systolic blood pressure.Conclution1.Hypertension, Diabetesmellitus, hyperlipidemia, Smoking, drinking are significantly associated with cerebral hemorrhage, high density lipoprotein, low-density lipoprotein, apolipoprotein b, systolic blood pressure, diastolic blood pressure, total cholesterol are significantly associated with cerebral hemorrhage.There is no significant association of Triglyceride, apolipoprotein a with cerebral hemorrhage.2.There is no significant association of the functional SNPS rs1051978, rs62096875, rs12605457, rs1063669, rs61731547 of The exon-2 with cerebral hemorrhage in Han population of Liaoning Province China.3.There is no significant association of (VNTR) L/S genotype with cerebral hemorrhage in Han population of Liaoning Province China.4.The rs754093 G/G genotype of NFATC1 exon-9 might be the independent risk factor of cerebral hemorrhage in Han population of Liaoning Province China, and its risk mainly come from G allele.5.NFATC1 gene is the susceptible gene significantly associate with cerebral hemorrhage in Han population of Liaoning Province China,in NFATC1 gene SNP rs754093 T/G allele may be a susceptible and mutational site of cerebral hemorrhage, G allele is associated with high risk of cerebral hemorrhage.6.NFATC1 rs754093 G/G genotype is significantly associated with high risk factors of cerebral hemorrhage which are CHO LDL, Apob, high systolic blood pressure.7.LDL, Apob, high systolic blood pressure are independent risk factors of cerebral hemorrhage too.