Study On The Relationship Between CpG Methylation Level And Transmembrane Signaling Pathway And Essential Hypertension In The Promoter Region Of IL-6 Gene And Its Related Mechanism

Objective:To investigate the relationship between CpG methylation level in promoter region of IL-6 gene and essential hypertension,and to analyze the role and mechanism of IL-6 and its transmembrane signal transduction related receptor(s IL-6R,gp130)in essential hypertension.Methods:Part I: From October 2017 to May 2018,100 patients with essential hypertension were enrolled in Luoyang Central Hospital affiliated to Zhengzhou University,and100 healthy people were recruited as control group.The plasma TG(TG),uric acid(UA),total cholesterol(TC),high density lipoprotein(HDL),low density lipoprotein(LDL),serum creatine(Scr),leukocyte count(WBC)was measured by AU2700 automatic analyzer.The CpG site methylation in the promoter region of IL-6 gene was detected by pyrophosphate sequencing.The difference of biochemical indexes and CpG site methylation in the promoter region of IL-6 gene in essential hypertension patients and the control group.Part II: 60 patients with simple hypertension,60 patients with hypertension with left ventricular hypertrophy,60 patients with hypertension with ischemic stroke and60 healthy controls were collected to compare the expression of pro-inflammatory factor IL-6(IL-6),soluble RANK ligand(s RANKL),osteoprotegerin(OPG),tumor necrosis factor-α(TNF-α),IL-1β and IL-21 in peripheral blood of patients with simple hypertension.The concentration difference of s IL-6R and sgp130 in the serumof each group were compared.patients with simple hypertension were followed up for 1 year to record the prognosis and analyze the related risk factors of prognosis.Results:Part I:1 The indexes of triglyceride(TG),uric acid(UA),total cholesterol(TC),high-density lipoprotein(HDL),low-density lipoprotein(LDL),serum creatinine(Scr)and white blood cell count(WBC)and fasting blood sugar,There was no significant difference between the patients with essential hypertension and the control group(P>0.05).2 The methylation level of CpG1 site in essential hypertension group was 62.17± 7.29,and that in control group was 63.88 ± 8.88,there was no significant difference between the two groups(P>0.05),but the level of CpG2 methylation in essential hypertension group was 58.43±7.53 and 62.34±9.65 in control group,and the level of CpG2 methylation in essential hypertension group was significantly lower than that in control group(P<0.05).The methylated level of CpG3 site was 51.52±6.18 in essential hypertension group and 57.45±8.29 in control group.The level of CpG3 methylation in essential hypertension group was significantly lower than that in control group(P<0.05).3 The methylation levels of CpG1～3 in male patients were 64.84±7.06,62.04±7.40,55.42±8.50,and the methylation levels of CpG1～3 in female patients were 61.84±8.61,59.30±9.57,53.87±7.41,and the level of CpG1 and CpG2 in male patients was significantly higher than that of female patients(P<0.05).There was no statistical significance between the two groups of CpG3 methylation levels(P>0.05).4 The levels of CpG1～3 methylation in patients with smoking history were 63.89±7.28,60.41±7.74,53.70±8.62,and those without smoking history were 66.16±6.63,64.28±6.36 and 57.78±7.87,respectively.the levels of CpG2 and CpG3 in patients with smoking history were significantly lower than those in patients without smoking history(P<0.05).There was no significant difference in CpG1 methylation level between the two groups(P>0.05).5 The levels of CpG1～3 methylation in patients with history of drinking were63.81±7.29,60.89±7.32 and 53.23±7.99.Respectively,the levels of CpG1～3methylation in patients without drinking history were 67.22±5.98,64.70±7.03 and60.48±7.58.The level of CpG2～3 methylation in patients with drinking history was significantly lower than that in patients without drinking history,the difference was statistically significant(P<0.05).6 CpG2(area under the curve: 0.638,95%CI: 0.560～0.716,P=0.001)and CpG3(area under the curve: 0.706,95% CI: 0.633～0.779,P<0.001)may have an application value for predicting the risk of essential hypertension.CpG1 has no diagnostic value.Part II:1 There were significant differences in s RANKL(2.05±0.15;5.2±0.28;5.61±0.31 vs 5.90±0.29 pmol/L),OPG(4.07±1.9;4.13±1.8;4.24±2.1 VS 3.27±0.95pmol/L),TNF-α(8.17±1.8;9.07±2.1;8.77±1.9 VS 5.24±0.85pg/ml),IL-1β(7.06±1.6;8.07±2.3;7.07±1.8 VS 3.37±0.96 pg/ml)and IL-21(6.67±1.3;7.07±2.0;6.67±1.2 VS 3.56±0.55 pg/ml)in peripheral blood of patients with simple hypertension,hypertension with left ventricular hypertrophy and hypertension with ischemic stroke(P<0.05),but there was no significant difference among the three groups with hypertension(P>0.05).2 Compared with the healthy control group,the serum levels of IL-6 and s IL-6R,in patients with essential hypertension were higher than those in the healthy control group,but the concentration of sgp130 did not change significantly.3 The concentrations of IL-6 and s IL-6R in serum of patients with hypertensionwith left ventricular hypertrophy and hypertension with ischemic stroke were higher than those of simple hypertension.4 One year later,6 patients had clinical progress,including 4 cases of myocardial hypertrophy,2 cases of cerebrovascular accidents(cerebral infarction and cerebral hemorrhage),and 54 cases remained stable.5 The levels of IL-6,BNP and uric acid in the progressive group were 3.25±0.75pg/m L,470±12 pg/m L,and 7.8±2.4 mg/dl,respectively.The levels of IL-6,BNP and uric acid in the non-progressive group were 1.68±0.39 pg/m L,118 ±19 pg/m L,and5.1 ±1.6 mg/dl.,respectively.The levels of IL-6,BNP and uric acid in the progressive group were significantly higher than those in the non-progressive group(P<0.05).6 ROC curve analysis showed that the area under the curve of IL-6 was 0.906(95%CI 0.682～0.991),and the IL-6 cut-off value was 2.3 pg/ml.The sensitivity and specificity were 91% and 73%,respectively.7 Kaplan-Meier analysis showed that the incidence of hypertension in patients with IL-6<2.3 pg/ml was significantly lower than that in patients with IL-6≥2.3 pg/ml,and there was significant difference between the two groups(P<0.05).Conclusion:1 The low methylation of the 1IL-6 promoter CpG site may be related to the occurrence of essential hypertension.2 The detection of the CpG site methylation of the IL-6 gene may be of potential diagnostic value for essential hypertension.3 The increase of s IL-6R may be related to the occurrence and development of essential hypertension.4 The concentration of IL-6 greater than or equal to 2.3 pg/ ml may suggest that the prognosis of essential hypertension is poor.