氧化低密度脂蛋白（ox-LDL）抑制急性冠脉综合征病人体内转录因子BACH2的表达

发布时间：2019-02-11 11:55

【摘要】：目的：研究氧化低密度脂蛋白(ox-LDL)对转录因子BACH2和Toll样受体4(TLR4)表达量的影响,以及促炎标记物和抑炎标记物在急性冠脉综合征患者或稳定型心绞痛患者和正常对照组三组受试者中的相应变化。探讨转录因子BACH2在冠心病发病中的作用和机制。实验方法：使用RT-PCR和Western Blot技术检测急性冠脉综合征患者(35例)、稳定型心绞痛患者(35例)和正常对照组(40例)三组受试者外周血单个核细胞(以及分离的淋巴细胞和单核细胞)中的Bach2和TLR4的表达水平。采用酶联免疫吸附法(ELISA)检测三组受试者血清标本中的ox-LDL及TH1, TH2, TH17and Treg细胞各自标志性细胞因子(IFN-γ, IL-4, IL-17和TGF-β1)的水平。在体外实验中,在不同浓度ox-LDL条件下与正常人外周淋巴细胞和单核细胞共培养,采用基因干扰技术敲低BACH2的表达,或者使用阻断性抗体阻断TLR4的表达后,检测对促炎标记物和抑炎标记物表达量的影响。另外,还使用来自急性冠脉综合征患者和正常对照组的血清与正常人外周淋巴细胞和单核细胞共培养,检测其对BACH2的表达影响。结果：RT-PCR和Western blot检测三组受试者PBMC中的BACH2及TLR4的表达趋势一致。和稳定型心绞痛组及正常对照组病人相比较,急性冠脉综合征组病人外周血单个核细胞中的BACH2表达量显著下调,TLR4表达量显著上调。ACS组病人血清中的ox-LDL及TH1, TH2和TH17细胞的细胞因子水平较SA组和CONTROL组显著升高,而Treg细胞的细胞因子显著降低。BACH2在淋巴细胞中的表达量明显高于单核细胞。在体外试验中,敲低BACH2的表达导致细胞因子IFN-γ, IL-4和IL-17的表达显著升高,以及TGF-β1显著降低。阻断TLR4导致BACH2表达显著升高。ox-LDL浓度增加下调BACH2的表达,阻断性TLR4抗体显著阻断了不同浓度ox-LDL诱导的淋巴细胞和单核细胞细胞因子的表达。与来自CONTROL组血清相比,淋巴细胞与来自ACS组病人血清共培养导致BACH2表达量显著下调。差异具有统计学意义。结论：急性冠脉综合征病人外周血中高浓度的ox-LDL下调BACH2的表达。BACH2的低表达可能促进了促炎的细胞因子IFN-γ、IL-4和IL17的产生,并进而造成急性冠脉综合征不稳定动脉粥样斑块的破裂。
[Abstract]:Objective: to study the effect of oxidized low density lipoprotein (ox-LDL) on the expression of transcription factor BACH2 and Toll like receptor 4 (TLR4). And the corresponding changes of proinflammatory markers and anti-inflammatory markers in patients with acute coronary syndrome or stable angina pectoris and normal control group. To investigate the role and mechanism of transcription factor BACH2 in the pathogenesis of coronary heart disease. Methods: RT-PCR and Western Blot techniques were used to detect acute coronary syndrome (ACS) in 35 patients. The expression of Bach2 and TLR4 in peripheral blood mononuclear cells (PBMC) of 35 patients with stable angina pectoris and 40 normal controls. Enzyme linked immunosorbent assay (ELISA) was used to detect the levels of IFN- 纬, IL-4, IL-17 and TGF- 尾 1 in ox-LDL and TH1, TH2, TH17and Treg cells in the serum samples of the three groups. In vitro, after co-culture with peripheral lymphocytes and monocytes of different concentrations of ox-LDL, gene interference technique was used to knock down the expression of BACH2, or blocking antibody was used to block the expression of TLR4. The expression of proinflammatory markers and anti-inflammatory markers were detected. In addition, serum from patients with acute coronary syndrome (ACS) and normal control group were co-cultured with peripheral lymphocytes and monocytes of normal subjects to detect the expression of BACH2. Results: the expression trend of BACH2 and TLR4 in PBMC by RT-PCR and Western blot was the same. Compared with the patients with stable angina pectoris and normal control, the expression of BACH2 and TLR4 in peripheral blood mononuclear cells of patients with acute coronary syndrome were significantly down-regulated, and the expression of TLR4 was significantly up-regulated. Ox-LDL and TH1, in serum of patients with ACS were significantly increased. The levels of cytokines in TH2 and TH17 cells were significantly higher than those in SA and CONTROL groups, while the cytokines of Treg cells were significantly decreased. The expression of BACH2 in lymphocytes was significantly higher than that in monocytes. In vitro, low expression of BACH2 resulted in a significant increase in the expression of cytokines IFN- 纬, IL-4 and IL-17, and a significant decrease in TGF- 尾 1. Blocking TLR4 increased the expression of BACH2, increased the concentration of ox-LDL and down-regulated the expression of BACH2, and blocked the expression of lymphocyte and monocyte cytokines induced by ox-LDL at different concentrations. Compared with serum from CONTROL group, lymphocyte coculture with serum from ACS group resulted in a significant down-regulation of BACH2 expression. The difference is statistically significant. Conclusion: high concentration of ox-LDL in peripheral blood of patients with acute coronary syndrome can down-regulate the expression of BACH2. The low expression of BACH2 may promote the production of pro-inflammatory cytokines IFN- 纬, IL-4 and IL17. And then lead to the rupture of unstable atherosclerotic plaque in acute coronary syndrome.
【学位授予单位】：华中科技大学
【学位级别】：博士
【学位授予年份】：2015
【分类号】：R541.4

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