炎癥因子基因DNA甲基化與原發(fā)性高血壓的關聯(lián)性研究
發(fā)布時間：2018-09-12 07:01
【摘要】：目的:1、探討原發(fā)性高血壓患者與正常對照間炎癥因子基因TLR2、IL6和IFN-γ啟動子DNA甲基化水平的差異;2、研究性別、吸煙和飲酒等環(huán)境因素對炎癥因子基因DNA甲基化差異的影響;3、探討炎癥因子基因啟動子Cp G位點的DNA甲基化差異對原發(fā)性高血壓的臨床診斷價值。方法:1、運用多階段抽樣的方法納入研究對象,進行人口學特征、環(huán)境因素的問卷調查、體格檢查及實驗室檢查,最終確定按性別、年齡1:1匹配的96個新發(fā)病例和96個對照,并獲得192個研究對象的基線資料、血液樣本及血生化指標;2、將采集的血樣進行DNA的提取、亞硫酸鹽修飾以及運用焦磷酸測序法檢測血液樣本中炎癥因子基因TLR2、IL6和IFN-γ啟動子DNA甲基化水平;3、通過分別比較檢測的炎癥因子基因在兩組間的甲基化程度,分析甲基化程度的差異,了解環(huán)境因素對甲基化及原發(fā)性高血壓發(fā)病的影響,并用Logistic回歸校正混雜因素。結果:1、各基因甲基化水平比較:(1)TLR2基因:新發(fā)病例組與正常對照組相比,新發(fā)病例組TLR2啟動子Cp G6位點甲基化水平降低(對照組VS新發(fā)病例組(%):Cp G6:8.30±4.13 VS 3.58±3.64,OR(95%CI)=0.91(0.861~0.979),adjusted P=0.009);(2)IL6基因:新發(fā)病例組與正常對照組相比,新發(fā)病例組IL6啟動子Cp G3位點甲基化水平降低(對照組VS新發(fā)病例組(%):Cp G3:57.45±8.29VS 51.52±6.18,OR(95%CI)=0.90(0.838~0.965),adjusted P=0.004);(3)IFN-γ基因:新發(fā)病例組與正常對照組相比,新發(fā)病例組IFN-γ啟動子Cp G2位點甲基化水平降低(對照組VS新發(fā)病例組(%):Cp G2:10.50±0.92VS 9.23±0.83,OR(95%CI)=0.64(0.480~0.856),adjusted P=0.003)。2、對照組中性別間比較:(1)TLR2基因:女性TLR2啟動子Cp G6位點甲基化水平偏高(男性VS女性(%):Cp G6:7.31±1.49 VS 8.94±5.09,adjusted P=0.013);(2)IL6基因:女性IL6啟動子Cp G2位點甲基化水平偏低(男性VS女性(%):Cp G2:65.53±7.31 VS 60.26±10.46,adjusted P=0.046);(3)IFN-γ基因:符號秩和檢驗分析發(fā)現(xiàn),女性IFN-γ啟動子Cp G1位點甲基化水平偏低(男性VS女性(%):Cp G1:37.58±1.31 VS 36.52±1.64,adjusted P=0.020)。3、男性中,吸煙、飲酒與炎癥因子基因DNA甲基化關聯(lián)分析:(1)IL6基因:男性中,吸煙者的IL6啟動子Cp G2-3甲基化水平降低(不吸煙者VS吸煙者(%):Cp G2:64.28±6.36 VS 60.41±7.74,adjusted P=0.023;Cp G3:57.78±7.87 VS 53.70±8.62,adjusted P=0.038);飲酒者的IL6啟動子Cp G2-3甲基化水平降低(不飲酒者VS飲酒者(%):Cp G2:64.70±7.03 VS 60.89±7.32,adjusted P=0.038;Cp G3:60.48±7.58 VS 53.23±7.99,adjusted P=4.24×10-4);(2)TLR2和IFN-γ基因:未發(fā)現(xiàn)差異化位點。4、相關性分析:(1)TLR2基因:TLR2啟動子Cp G6位點甲基化水平與血壓成負相關(Cp G6 and SBP:r=-0.329,adjusted P=3.95×10-5;Cp G6 and DBP:r=-0.304,adjusted P=1.80×10-4);(2)IL6基因:IL6啟動子Cp G2-3位點甲基化水平與血壓成負相關(Cp G2and SBP:r=-0.274,adjusted P=0.009;Cp G2 and DBP:r=-0.183,adjusted P=0.011;Cp G3 and SBP:r=-0.321,adjusted P=5.60×10-6;Cp G3 and DBP:r=-0.274,adjusted P=1.17×10-4);(3)IFN-γ基因:IFN-γ啟動子Cp G2位點甲基化水平與血壓成負相關(Cp G2 and SBP:rs=-0.546,adjusted P=2.60×10-16;Cp G2 and DBP:rs=-0.539,adjusted P=7.60×10-16)。5、受試者工作特征曲線(Receiver operating characteristic curve,簡稱ROC曲線)分析:(1)TLR2基因:分析發(fā)現(xiàn)TLR2啟動子Cp G1位點和Cp G6具有診斷價值(Cp G1:AUC(area under curve)=0.612,P=0.007;Cp G6:AUC(area under curve)=0.834,P=1.31×10-15);(2)IL6基因:分析發(fā)現(xiàn)IL6啟動子Cp G2-3位點具有診斷學價值(Cp G2:AUC(area under curve)=0.638,P=0.001;Cp G3:AUC(area under curve)=0.704,P=8.12×10-7);(3)IFN-γ基因:分析發(fā)現(xiàn)IFN-γ啟動子Cp G2位點具有診斷學價值(Cp G2:AUC(area under curve)=0.834,P=1.40×10-15)。結論:1、TLR2啟動子Cp G6位點,IL6啟動子Cp G3位點和IFN-γ啟動子Cp G2位點的低甲基化是原發(fā)性高血壓的危險因素。2、女性TLR2啟動子Cp G6位點甲基化水平偏高,女性IL6啟動子Cp G2位點和IFN-γ啟動子Cp G1位點甲基化水平偏低。男性和女性基因甲基化水平存在差異。男性中,吸煙者和飲酒者的IL6啟動子Cp G2-3甲基化水平降低,吸煙和飲酒作為EH的危險因素可能通過影響基因Cp G位點甲基化水平影響原發(fā)性高血壓的發(fā)生發(fā)展。3、相關性分析發(fā)現(xiàn)TLR2啟動子Cp G6位點、IL6啟動子Cp G2-3位點甲基化水平與血壓成負相關,IFN-γ啟動子Cp G2位點甲基化水平與血壓成負相關。4、TLR2啟動子Cp G1和Cp G6位點,IL6啟動子Cp G2-3位點和IFN-γ啟動子Cp G2位點具有診斷價值。
[Abstract]:Objective: 1. To investigate the difference of DNA methylation of inflammatory factor gene TLR2, IL6 and IFN-gamma promoters between essential hypertension patients and normal controls; 2. To study the effect of gender, smoking and drinking on DNA methylation of inflammatory factor gene promoter Cp G site; 3. To explore the difference of DNA methylation of inflammatory factor gene promoter Cp G site on primary hypertension. Methods: 1. Using the method of multi-stage sampling, the subjects were included in the study. The demographic characteristics, environmental factors, physical examination and laboratory examination were conducted. Finally 96 new cases and 96 controls matched by sex and age 1:1 were determined. The baseline data of 192 subjects and blood samples were obtained. 2. DNA extraction, sulfite modification and pyrophosphate sequencing were used to detect the methylation level of TLR2, IL6 and IFN-gamma promoters in blood samples; 3. The methylation degree of inflammatory factor genes between the two groups was compared and the difference of methylation degree was analyzed. Results: 1. Comparison of the methylation levels of each gene: (1) TLR2 gene: The methylation level of TLR2 promoter Cp G6 in the new case group was lower than that in the normal control group (control group, VS new case group (%): (2) IL 6 gene: The methylation level of IL 6 promoter Cp G3 in the new case group was lower than that in the normal control group (control group: Cp G3: 57.45 [8.29VS] 51.52 [6.18], OR (95% CI) = 0.91 (0.861-0.979), adjusted P = 0.009); (2) IL 6 gene: The methylation level of IL 6 promoter Cp G3 in the new case group was lower than that in the normal control group (control group: Cp G3: 57.45 [8.29VS] 51.52 [6.18, OR (95% CI) = 0.90 (0.838-0.965), adjusted P = 0.004);(control group N-gamma gene: The methylation level of Cp G2 site of IFN-gamma promoter in the new case group was lower than that in the normal control group (control group vs new case group (%): Cp G2: 10.50 (%) 0.92VS 9.23 (%) 0.83, OR (95% CI) = 0.64 (0.480-0.856), adjusted P = 0.003). 2, sex comparison in the control group: (1) TLR2 gene: female TLR2 promoter Cp G6 site A (2) IL6 gene: female IL6 promoter Cp G2 site methylation level is low (male VS female (%): Cp G2: 65.53 [7.31] VS 60.26 [10.46], adjusted P = 0.046; (3) IFN - gamma gene: symbol rank and test analysis found that the female IFN - gamma promoter Cp G1 site Low methylation level (male vs female (%): Cp G1: 37.58 (%) vs 1.31 vs 36.52 (%) vs 1.64, adjusted P = 0.020). Correlation analysis between smoking, drinking and DNA methylation of inflammatory factor genes in male: (1) IL 6 gene: In male, the methylation level of IL 6 promoter Cp G2-3 in smokers decreased (non-smokers vs smokers (%): Cp G2: 64.28 (%) vs 6.36 vs 60.41 (%) vs 7.74, a Djusted P = 0.023; Cp G3: 57.78 [7.87 VS = 53.70 [(adjusted P = 0.038); CpG 2-3 methylation in IL 6 promoter Cp G2-3 decreased in drinkers (non-drinkers (%): Cp G2: 64.70 [7.03 VS 60.89 [.89] [7.89 [.32, adjusted P = 0.038; Cp G3: 0.03; Cp G3: 57.78 [: 60.48 [7.48 [7.58 VS 53.53.23 [.23 [53] [7.23 [7.99, adjusted P = adjusted P = 4.24-4); (2) differentiation site Point 4, Correlation analysis: (1) TLR2 gene: TLR2 promoter Cp G6 site methylation level was negatively correlated with blood pressure (Cp G6 and SBP: r = - 0.329, adjusted P = 3.95 *10-5; Cp G6 and DBP: r = - 0.304, adjusted P = 1.80 *10-4); IL6 gene: IL6 promoter Cp G2-3 site methylation level was negatively correlated with blood pressure (Cp G2 and SBP: r = - 0.274, adjusted P = 0.0.0). 09; Cp G2 and DBP: r = - 0.183, adjusted P = 0.011; Cp G3 and SBP: r = - 0.321, adjusted P = 5.60 x 10-6; Cp G3 and DBP: r = - 0.274, adjusted P: adjusted P: r = - 0.274, adjusted P = 1.17 x 10-4); (3) IFN-gamgene: IFN-gampromoter CpG2methylation level of CpG2promoter CpG2promoter CpG2promoter methylation level was negatively correlwith blood pressure (Cp G2 and SBP: rs = - 0.546, adjusted P = - 0.546, adjusted P = 2.60 x 10-16; Cp G3 and DBP: r = - 0.274: r = - 0 9, adjusted P = 7.60 5, Receiver operating characteristic curve (ROC curve) analysis: (1) TLR2 gene: analysis found that TLR2 promoter Cp G1 locus and Cp G6 have diagnostic value (Cp G1: AUC (area under curve) = 0.612, P = 0.007; Cp G6: AUC (area under curve) = 0.834, P = 1.31 *10-15); (2) IL6 gene: analysis found that IL6: IL6 gene Promoter Cp G2-3 locus has diagnostic value (Cp G2: AUC (area under curve) = 0.638, P = 0.001; Cp G3: AUC (area under curve) = 0.704, P = 8.12 *10-7); (3) IFN-gamma gene: analysis found that IFN-gamma promoter Cp G2 locus has diagnostic value (Cp G2: AUC (area under curve) = 0.834, P = 1.40 *10-15). Hypomethylation at Cp G3 and Cp G2 loci of IFN-gamma promoters is a risk factor for essential hypertension. 2. High methylation at Cp G6 locus in female TLR2 promoter, low methylation at Cp G2 locus and Cp G1 locus in female IL6 promoter. Differences in gene methylation levels between men and women. Smoking in men The methylation level of IL 6 promoter Cp G2-3 was decreased in both subjects and drinkers. Smoking and drinking as risk factors for EH may affect the occurrence and development of essential hypertension by affecting the methylation level of Cp G locus. 3. Correlation analysis showed that the methylation level of TLR2 promoter Cp G6 locus and IL 6 promoter Cp G2-3 locus was negatively correlated with blood pressure. The methylation level at Cp G2 site of - gamma promoter was negatively correlated with blood pressure. 4. TLR2 promoter Cp G1 and Cp G6 sites, IL6 promoter Cp G2-3 sites and IFN-gamma promoter Cp G2 sites were of diagnostic value.
【學位授予單位】：寧波大學
【學位級別】：碩士
【學位授予年份】：2017
【分類號】：R544.11
本文編號：2238259
[Abstract]:Objective: 1. To investigate the difference of DNA methylation of inflammatory factor gene TLR2, IL6 and IFN-gamma promoters between essential hypertension patients and normal controls; 2. To study the effect of gender, smoking and drinking on DNA methylation of inflammatory factor gene promoter Cp G site; 3. To explore the difference of DNA methylation of inflammatory factor gene promoter Cp G site on primary hypertension. Methods: 1. Using the method of multi-stage sampling, the subjects were included in the study. The demographic characteristics, environmental factors, physical examination and laboratory examination were conducted. Finally 96 new cases and 96 controls matched by sex and age 1:1 were determined. The baseline data of 192 subjects and blood samples were obtained. 2. DNA extraction, sulfite modification and pyrophosphate sequencing were used to detect the methylation level of TLR2, IL6 and IFN-gamma promoters in blood samples; 3. The methylation degree of inflammatory factor genes between the two groups was compared and the difference of methylation degree was analyzed. Results: 1. Comparison of the methylation levels of each gene: (1) TLR2 gene: The methylation level of TLR2 promoter Cp G6 in the new case group was lower than that in the normal control group (control group, VS new case group (%): (2) IL 6 gene: The methylation level of IL 6 promoter Cp G3 in the new case group was lower than that in the normal control group (control group: Cp G3: 57.45 [8.29VS] 51.52 [6.18], OR (95% CI) = 0.91 (0.861-0.979), adjusted P = 0.009); (2) IL 6 gene: The methylation level of IL 6 promoter Cp G3 in the new case group was lower than that in the normal control group (control group: Cp G3: 57.45 [8.29VS] 51.52 [6.18, OR (95% CI) = 0.90 (0.838-0.965), adjusted P = 0.004);(control group N-gamma gene: The methylation level of Cp G2 site of IFN-gamma promoter in the new case group was lower than that in the normal control group (control group vs new case group (%): Cp G2: 10.50 (%) 0.92VS 9.23 (%) 0.83, OR (95% CI) = 0.64 (0.480-0.856), adjusted P = 0.003). 2, sex comparison in the control group: (1) TLR2 gene: female TLR2 promoter Cp G6 site A (2) IL6 gene: female IL6 promoter Cp G2 site methylation level is low (male VS female (%): Cp G2: 65.53 [7.31] VS 60.26 [10.46], adjusted P = 0.046; (3) IFN - gamma gene: symbol rank and test analysis found that the female IFN - gamma promoter Cp G1 site Low methylation level (male vs female (%): Cp G1: 37.58 (%) vs 1.31 vs 36.52 (%) vs 1.64, adjusted P = 0.020). Correlation analysis between smoking, drinking and DNA methylation of inflammatory factor genes in male: (1) IL 6 gene: In male, the methylation level of IL 6 promoter Cp G2-3 in smokers decreased (non-smokers vs smokers (%): Cp G2: 64.28 (%) vs 6.36 vs 60.41 (%) vs 7.74, a Djusted P = 0.023; Cp G3: 57.78 [7.87 VS = 53.70 [(adjusted P = 0.038); CpG 2-3 methylation in IL 6 promoter Cp G2-3 decreased in drinkers (non-drinkers (%): Cp G2: 64.70 [7.03 VS 60.89 [.89] [7.89 [.32, adjusted P = 0.038; Cp G3: 0.03; Cp G3: 57.78 [: 60.48 [7.48 [7.58 VS 53.53.23 [.23 [53] [7.23 [7.99, adjusted P = adjusted P = 4.24-4); (2) differentiation site Point 4, Correlation analysis: (1) TLR2 gene: TLR2 promoter Cp G6 site methylation level was negatively correlated with blood pressure (Cp G6 and SBP: r = - 0.329, adjusted P = 3.95 *10-5; Cp G6 and DBP: r = - 0.304, adjusted P = 1.80 *10-4); IL6 gene: IL6 promoter Cp G2-3 site methylation level was negatively correlated with blood pressure (Cp G2 and SBP: r = - 0.274, adjusted P = 0.0.0). 09; Cp G2 and DBP: r = - 0.183, adjusted P = 0.011; Cp G3 and SBP: r = - 0.321, adjusted P = 5.60 x 10-6; Cp G3 and DBP: r = - 0.274, adjusted P: adjusted P: r = - 0.274, adjusted P = 1.17 x 10-4); (3) IFN-gamgene: IFN-gampromoter CpG2methylation level of CpG2promoter CpG2promoter CpG2promoter methylation level was negatively correlwith blood pressure (Cp G2 and SBP: rs = - 0.546, adjusted P = - 0.546, adjusted P = 2.60 x 10-16; Cp G3 and DBP: r = - 0.274: r = - 0 9, adjusted P = 7.60 5, Receiver operating characteristic curve (ROC curve) analysis: (1) TLR2 gene: analysis found that TLR2 promoter Cp G1 locus and Cp G6 have diagnostic value (Cp G1: AUC (area under curve) = 0.612, P = 0.007; Cp G6: AUC (area under curve) = 0.834, P = 1.31 *10-15); (2) IL6 gene: analysis found that IL6: IL6 gene Promoter Cp G2-3 locus has diagnostic value (Cp G2: AUC (area under curve) = 0.638, P = 0.001; Cp G3: AUC (area under curve) = 0.704, P = 8.12 *10-7); (3) IFN-gamma gene: analysis found that IFN-gamma promoter Cp G2 locus has diagnostic value (Cp G2: AUC (area under curve) = 0.834, P = 1.40 *10-15). Hypomethylation at Cp G3 and Cp G2 loci of IFN-gamma promoters is a risk factor for essential hypertension. 2. High methylation at Cp G6 locus in female TLR2 promoter, low methylation at Cp G2 locus and Cp G1 locus in female IL6 promoter. Differences in gene methylation levels between men and women. Smoking in men The methylation level of IL 6 promoter Cp G2-3 was decreased in both subjects and drinkers. Smoking and drinking as risk factors for EH may affect the occurrence and development of essential hypertension by affecting the methylation level of Cp G locus. 3. Correlation analysis showed that the methylation level of TLR2 promoter Cp G6 locus and IL 6 promoter Cp G2-3 locus was negatively correlated with blood pressure. The methylation level at Cp G2 site of - gamma promoter was negatively correlated with blood pressure. 4. TLR2 promoter Cp G1 and Cp G6 sites, IL6 promoter Cp G2-3 sites and IFN-gamma promoter Cp G2 sites were of diagnostic value.
【學位授予單位】：寧波大學
【學位級別】：碩士
【學位授予年份】：2017
【分類號】：R544.11
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