miR-127-5p靶向IRAK4对肺炎链球菌诱导的肺泡上皮细胞凋亡及炎症因子表达的影响

目的探究miR-127-5p靶向白细胞介素1受体相关激酶4(IRAK4)对肺炎链球菌诱导的肺泡上皮细胞凋亡及炎症因子表达的影响方法首先分为对照组和感染组,感染组用肺炎链球菌感染A549细胞,感染组细胞分别转染miR-127-5p mimic、si-IRAK4及阴性对照质粒,qRT-PCR检测A549细胞中miR-127-5p的表达水平,western blot检测IRAK4、Bax、Bcl-2蛋白水平,流式细胞仪检测细胞凋亡情况,ELISA检测白介素-10(IL-10)、白介素-6(IL-6)水平,双荧光素酶报告基因检测miR-127-5p与IRAK4的靶向关系。结果与对照组比较,肺炎链球菌感染后A549细胞中miR-127-5p、Bcl-2、IL-10水平显著降低,细胞凋亡率、IRAK4、Bax、IL-6水平显著升高(P<0.05);与感染+miR-NC组比较,过表达miR-127-5p后,A549细胞中miR-127-5p、Bcl-2、IL-10水平显著升高,细胞凋亡率、IRAK4、Bax、IL-6显著降低;与感染+si-NC组比较,抑制IRAK4表达后,A549细胞中细胞凋亡率、IRAK4、Bax、IL-6水平显著降低,Bcl-2、IL-10水平显著升高;双荧光素酶报告基因及Western blot结果显示,miR-127-5p可通过与IRAK4特异性结合负向调控IRAK4的表达;与感染+miR-127-5p+pcDNA组比较,感染+miR-127-5p+pcDNA-IRAK4组A549细胞中凋亡率、IRAK4、Bax、IL-6水平显著升高,Bcl-2、IL-10水平显著降低(P<0.05)。结论miR-127-5p靶向IRAK4调控肺炎链球菌诱导的肺泡上皮细胞凋亡及炎症因子IL-10、IL-6的表达。

Effect of miR-127-5p targeting IRAK4 on alveolar epithelial cells apoptosis induced by Streptococcus pneumoniae and inflammatory factor expression

Objective To investigate the effect of interleukin-1 receptor-related kinase 4 targeting at microRNA-127-5p on the apoptosis of alveolar epithelial cells induced by Streptococcus pneumoniae and the expression of inflammatory factors.Methods The A549 cells were infected with Streptococcus pneumoniae.The infected cells were transfected with microRNA-127-5p mimic,si-IRAK4 and negative control plasmids.The expression of microRNA-127-5p in A549 cells was detected by qRT-PCR,the protein levels of IRAK4,Bax and Bcl-2 were detected by Western blot,the apoptosis was detected by flow cytometry,and the interleukin-10 and interleukin-6 water were detected by ELISA.The targeting relationship between mir-127-5p and IRAK4 was detected by double luciferase reporter gene.Results Compared with the control group,the levels of microRNA-127-5p,Bcl-2 and IL-10 in A549 cells infected with Streptococcus pneumoniae decreased significantly,and the apoptotic rate,IRAK4,Bax and IL-6 levels increased significantly(P<0.05).Compared with the infection+microRNA-NC group,the levels of microRNA-127-5p,Bcl-2 and IL-10 in A549 cells increased significantly,and the apoptotic rate,IRAK4,Bax,IL-6 decreased significantly.Compared with the i-NC group after inhibiting the expression of IRAK4,the apoptotic rate,IRAK4,Bax and IL-6 levels of A549 cells decreased significantly,while the levels of Bcl-2 and IL-10 increased significantly.The results of double luciferase reporter gene and Western blot showed that the expression of IRAK4 could be negatively regulated by the specific binding of mir-127-5p to IRAK4.Compared with the infection+mir-127-5p+pcDNA group,the infection+mir-127-5p+pcDNA-IRAK4 group A549 was fine.Their apoptotic rate,IRAK4,Bax and IL-6 levels were significantly increased,while Bcl-2 and IL-10 levels were significantly decreased(P<0.05).Conclusion Mir-127-5p targeting IRAK4 regulates the apoptosis of alveolar epithelial cells induced by Streptococcus pneumoniae and the expression of inflammatory factors IL-10 and IL-6.