miR-23b-3p靶向PALM3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

Down-regualtion of miR-23b-3p protects vascular endothelial cell from injury induced by Streptococcus pneumoniae through targeting PALM3

目的研究miR-23b-3p对肺炎链球菌诱导的肺泡上皮细胞凋亡和炎症的作用及机制。方法将肺泡上皮细胞A549细胞分为对照组、感染组、转染组、感染组+转染组;对照组A549细胞常规培养,感染组用1×108 CFU/mL肺炎链球菌培养A549细胞,转染组转染不同载体,感染组+转染组在A549细胞被肺炎链球菌处理前48h进行转染。qRT-PCR检测细胞中miR-23b-3p和paralemmin-3(PALM3)mRNA的水平,流式细胞术检测细胞凋亡率,Western blot检测细胞中PALM3、B淋巴细胞瘤-2基因(Bcl-2)和Bcl-2相关X蛋白(Bax),酶联免疫法(ELISA)检测肺炎链球菌诱导后细胞培养上清中白介素-6(interleukin 6,IL-6)和白介素-10(interleukin 10,IL-10)的含量;双荧光素酶报告系统验证miR-23b-3p与PALM3的关系。结果与对照组相比,感染组A549细胞中miR-23b-3p、IL-10、Bcl-2含量明显降低(P<0.05),PALM3、IL-6、Bax水平及细胞凋亡率明显升高(P<0.05);过表达miR-23b-3p和干扰PALM3表达均可抑制肺炎链球菌诱导的A549细胞凋亡和炎症;miR-23b-3p靶向负调控PALM3的表达;过表达PALM3逆转了过表达miR-23b-3p对肺炎链球菌诱导的A549细胞凋亡和炎症的作用。结论 miR-23b-3p靶向PALM3抑制肺炎链球菌诱导的A549炎症和细胞凋亡。

Objective To investigate the effect and mechanism of miR-23 b-3 p on apoptosis and inflammation of alveolar epithelial cells induced by Streptococcus pneumoniae.Methods Alveolar epithelial cells A549 were divided into control group,infection group,transfection group and infection group+transfection group.A549 cells in the control group were routinely cultured,and A549 cells in the infected group were cultured with Streptococcus pneumoniae of 1×108 CFU/mL.Transfection groups were transfected with different vectors.A549 cells in infection group+transfection group were transfected 48 hours before being treated by Streptococcus pneumoniae.The levels of miR-23 b-3 p and paralemmin-3(PALM3)mRNA were detected by qRT-PCR.Apoptotic rate was detected by flow cytometry.The levels of PALM3,B-cell lymphoma-2(Bcl-2)and Bcl-2 related X protein(Bax)were detected by Western blot.The levels of interleukin-6(IL-6)and interleukin-10(IL-10)in cell culture supernatants were detected by enzyme-linked immunosorbent assay(ELISA).The relationship between miR-23 b-3 p and PALM3 was validated by dual-luciferase reporter assay system.Results Compared with the control group,the levels of miR-23 b-3 p,IL-10 and Bcl-2 in A549 cells in the infected group were significantly decreased(P<0.05),the levels of PALM3,IL-6 and Bax and the apoptotic rate were significantly increased(P<0.05).Overexpression of miR-23 b-3 p and interference the expression of PALM3 both inhibited the apoptosis and inflammation of A549 cells induced by Streptococcus pneumoniae.Overexpression of PALM3 reversed the effect of overexpression of miR-23 b-3 p on apoptosis and inflammation of A549 cells induced by Streptococcus pneumoniae.Conclusion miR-23 b-3 p alleviates the inflammation and inhibits apoptosis of A549 induced by Streptococcus pneumoniae by targeting PALM3.