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目的研究IL-22对ox-LDL诱导的CRL-1730细胞(人脐静脉内皮细胞)凋亡的影响及其机制。方法 100μg/ml ox-LDL刺激CRL-1730细胞,不同时间点经实时荧光定量RT-PCR检测CRL-1730细胞中IL-22受体R1(IL-22R1)mRNA的表达状况。然后在ox-LDL诱导的CRL-1730细胞凋亡模型中加入20 ng/ml外源性IL-22,通过AnnexinⅤ-PI凋亡试剂盒检测其对细胞凋亡的影响,经实时荧光定量反转录PCR检测Bcl-2、Bcl-xl、Bcl-w、caspase 3及STAT3mRNA的表达,采用免疫印迹法分析STAT3磷酸化和Bcl-2蛋白的水平。结果 IL-22R1 mRNA在ox-LDL刺激后表达明显增高(P<0.05),且于12 h达峰值(P<0.01);加入IL-22可明显抑制ox-LDL刺激诱导的内皮细胞凋亡(P<0.01),促进抗凋亡基因Bcl-2、Bcl-xl和Bcl-w表达,而降低caspase 3 mRNA表达(P<0.01)。此外,IL-22作用1～2 h STAT3发生磷酸化,4 h后Bcl-2蛋白表达升高(P<0.01)。结论 IL-22可抑制ox-LDL诱导的CRL-1730细胞凋亡,这可能与其促使STAT3磷酸化,上调抗凋亡分子Bcl-2、Bcl-xl和Bcl-w的表达,抑制促凋亡分子caspase 3的表达有关。
Objective To investigate the effect of IL-22 on the apoptosis of CRL-1730 cells (human umbilical vein endothelial cells) induced by ox-LDL and its mechanism. Methods CRL-1730 cells were stimulated with ox-LDL at 100μg / ml, and the expression of IL-22R1 mRNA in CRL-1730 cells was detected by real-time fluorescence quantitative RT-PCR at different time points. Then, 20 ng / ml exogenous IL-22 was added into ox-LDL-induced apoptosis model of CRL-1730 cells. The apoptosis rate was detected by Annexin Ⅴ-PI apoptosis kit. Real-time fluorescence quantitative reverse The expression of Bcl-2, Bcl-xl, Bcl-w, caspase 3 and STAT3 mRNA were detected by PCR. The levels of STAT3 phosphorylation and Bcl-2 protein were analyzed by Western blotting. Results The expression of IL-22R1 mRNA was significantly increased after ox-LDL stimulation (P <0.05), and reached its peak at 12 h (P <0.01). The addition of IL-22 significantly inhibited the apoptosis of endothelial cells induced by ox- P <0.01), promote the expression of anti-apoptotic genes Bcl-2, Bcl-xl and Bcl-w and decrease the expression of caspase 3 mRNA (P <0.01). In addition, STAT3 was phosphorylated at 1 ~ 2 h after IL-22 treatment and increased at 4 h (P <0.01). Conclusion IL-22 can inhibit the apoptosis of CRL-1730 cells induced by ox-LDL, which may be related to its role in promoting the phosphorylation of STAT3, up-regulating the expressions of Bcl-2, Bcl-xl and Bcl- caspase 3 expression.