LncRNA SOX2OT对肺炎链球菌诱导的肺泡上皮细胞凋亡和炎症反应的影响及机制

Effect of LncRNA SOX2OT on the apoptosis and inflammation of alveolar epithelial cells induced by streptococcus pneumoniae and its mechanism

目的探讨LncRNA SOX2OT对肺炎链球菌诱导的肺泡上皮细胞凋亡和炎症反应的影响及机制。方法以29例健康体检者为对照,采用qRT-PCR法检测51例肺炎患者外周血中LncRNA SOX2OT和miR-146a-5p表达。体外培养肺泡上皮细胞HEPApiC,分为对照(con)组和肺炎链球菌诱导组(model组),qRT-PCR法检测细胞中LncRNA SOX2OT和miR-146a-5p表达。双荧光素酶报告基因实验验证LncRNA SOX2OT和miR-146a-5p的调控关系。分别转染si-LncRNA SOX2OT或共转染si-LncRNA SOX2OT与anti-miR-146a-5p至HEPApiC细胞中,再用肺炎链球菌处理,流式细胞术检测细胞凋亡,蛋白质印迹法检测细胞中Bax和Bcl-2蛋白表达,酶联免疫吸附法检测细胞培养上清液中IL-6、TNF-α和IL-10含量。结果肺炎患者外周血中LncRNA SOX2OT表达量高于健康对照组(P<0.05),miR-146a-5p表达量低于健康对照组(P<0.05)。model组HEPApiC细胞中LncRNA SOX2OT表达量高于con组(P<0.05),miR-146a-5p表达量低于con组(P<0.05)。LncRNA SOX2OT靶向结合miR-146a-5p,且下调LncRNA SOX2OT的HEPApiC细胞中miR-146a-5p表达升高(P<0.05)。下调LncRNA SOX2OT后,肺炎链球菌诱导的HEPApiC细胞凋亡率、细胞中Bax蛋白表达及细胞培养上清液中IL-6和TNF-α水平均降低(P<0.05),而Bcl-2蛋白表达和IL-10水平均升高(P<0.05)。下调miR-146a-5p逆转了下调LncRNA SOX2OT对肺炎链球菌诱导的HEPApiC细胞凋亡和炎症反应的影响。结论下调LncRNA SOX2OT可能通过靶向上调miR-146a-5p抑制肺炎链球菌诱导的肺泡上皮细胞HEPApiC凋亡和炎症反应。

Objective To investigate the effect and mechanism of LncRNA SOX2OT on the apoptosis and inflammation of alveolar epithelial cells induced by streptococcus pneumoniae.Methods Taking 29 healthy subjects as controls,qRT-PCR was used to detect the expression of LncRNA SOX2OT and miR-146a-5p in the peripheral blood of 51 patients with pneumonia.The alveolar epithelial cells HEPApiC were cultured in vitro and divided into the control group and the streptococcus pneumoniae induction group(the model group),and then the expression of LncRNA SOX2OT and miR-146a-5p in the cells was detected by qRT-PCR.The dual luciferase reporter gene experiment verified the regulatory relationship between LncRNA SOX2OT and miR-146a-5p.After HEPApiC cells were transfected with si-LncRNA SOX2OT or co-transfected with si-LncRNA SOX2OT and anti-miR-146a-5p,they were treated with streptococcus pneumoniae.And then the cell apoptosis was detected by flow cytometry,and the protein expression of Bax and Bcl-2 was detected by western blotting.Enzyme-linked immunosorbent assay were used to detect the content of IL-6,TNF-αand IL-10 in the cell culture supernatant.Results The expression of LncRNA SOX2OT in the peripheral blood of patients with pneumonia was higher than that of the healthy control group(P<0.05),but the expression of miR-146a-5p was lower than that of the healthy control group(P<0.05).The expression of LncRNA SOX2OT in HEPApiC cells in the model group was higher than that in the control group(P<0.05),but the expression of miR-146a-5p was lower than that in the control group(P<0.05).LncRNA SOX2OT could target to bind miR-146a-5p,and the expression of miR-146a-5p was increased in HEPApiC cells that down-regulated LncRNA SOX2OT(P<0.05).After down-regulating LncRNA SOX2OT,the apoptosis rate of HEPApiC cells induced by streptococcus pneumoniae,the protein expression of Bax in cells,and the content of IL-6,TNF-αand IL-10 in the cell culture supernatant were decreased(P<0.05),but the protein expression of Bcl-2 and the content of IL-10 were increased(P<0.05).Down-regulating miR-146a-5p reversed the effect of down-regulating LncRNA SOX2OT on apoptosis and inflammation of HEPApiC cells induced by streptococcus pneumoniae.Conclusion Down-regulation of LncRNA SOX2OT may inhibit the apoptosis and inflammation of streptococcus pneumoniae-induced HEPApiC cells by targeting up-regulation of miR-146a-5p.