硫化氢抑制NF-κBp65因子的表达在抗动脉粥样硬化中的作用

The inhibitory effects of hydrogen sulfide on the expression of NF-κBp65 in the treatment of atherosclerosis

目的 建立动脉粥样硬化（AS）细胞模型,探讨硫化氢供体（NaHS）对氧化低密度脂蛋白（ox-LDL）诱导的动脉粥样硬化中NF-κBp65因子表达的影响.方法 应用浓度为80μg/mL ox-LDL诱导人肝癌细胞HepG2建立AS细胞模型,设置对照组、治疗组（10μmol/L、15μmol/L、30μmol/L、100μmol/L、500μmol/L、1000μmol/L）,观察硫化氢作用此细胞模型时的细胞增殖活力以及测定TNF-α和IL-10因子的表达,通过RT-qPCR和Western-blot法测定NF-κBp65的表达情况.结果 ox-LDL诱导24h开始,HepG2细胞内出现大量红染颗粒,游离胆固醇和总胆固醇均比对照组增加,细胞游离胆固醇含量大于总胆固醇的50%,表明AS细胞模型建立成功;CCK8法细胞增殖活力检测结果表明：NaHS在30μmol/L,作用时间为48h时细胞增殖活力最大;运用ELISA法对TNF-α细胞因子进行检测结果表明,治疗组在15μmol/L、30μmol/L浓度时与对照组相比显著降低（P〈0.05）,而IL-10的表达明显升高,具有统计学意义（P〈0.05）.RT-qPCR和Western-blot法结果表明NF-κBp65因子的表达与对照组相比显著降低,具有统计学意义（P〈0.05）.结论 H2S可通过降低NF-κBp65因子的表达对动脉粥样硬化病变起到一定的保护作用,为H2S供体型药物的研发与临床治疗提供有价值的参考.

Objective Established the model of atherosclerosis （AS） cells.To explore Hydrogen sulfide donor （NaHS） in atherosclerosis model by oxidized low density lipoprotein （ox-LDL） induced of the NF-κBp65 factor expression.MethodsApplication concentration of 80ug/mL ox-LDL induced liver cancer cell HepG2 cells AS model, set up a control group and treatment group （10μmol/L、15μmol/L、30μmol/L、100μmol/L、500μmol/L、1000μmol/L）, to observe hydrogen sulfide this cell model of the cell proliferation activity and the determination of TNF-α and IL-10 factors of the expression, through the RT-qPCR and Western-blot method for determining the expression of the NF-κBp65 expression situation.ResultsAt 24h the ox-LDL induced a large number of red-stained granules, free cholesterol and total cholesterol in HepG2 cells increased, and free cholesterol content was more than 50% of total cholesterol in HepG2 cells, indicating that AS cell model was established successfully.CCK8 method of cell proliferation activity test results showed that the NaHS in 30umol/L, duration of 48 h when the largest cell proliferation activity.The TNF-α cytokine by ELISA method for testing the results showed that the treatment group in 15umol/L and 30umol/L concentration compared with the control group decreased significantly （P〈0.05）, while the expression of IL-10 increased significantly （P〈0.05）, with statistical significance.RT-qPCR and Western-blot method results show that the NF-κBp65 factor expression significantly reduced compared with control group （P〈0.05）, with statistical significance.ConclusionH2S play a protective role in atherosclerotic lesions by reducing the NF-κBp65 factor expression, and provide a valuable reference for the development and clinical treatment of H2S donor-type drugs.