白芍总苷通过miR-23b-3p/FoxA1对肺炎链球菌诱导的肺泡上皮细胞凋亡的影响

目的探讨白芍总苷及对肺炎链球菌诱导的肺泡上皮细胞凋亡的影响分子机制。方法肺泡上皮细胞A549随机分为对照(Con)组、模型(Model)组(1×10⁸CFU/mL的肺炎链球菌感染)、Model+白芍总苷低、中、高剂量组(17.5 mg/mL、35 mg/mL、70 mg/mL的白芍总苷+1×10⁸CFU/mL的肺炎链球菌感染)、Model+白芍总苷+anti-miR-23b-3p组(转染miR-23b-3p抑制剂+70 mg/mL白芍总苷+1×10⁸CFU/mL肺炎链球菌感染);细胞计数试剂盒8(CCK-8)检测细胞活性;流式细胞术检测细胞凋亡;酶联免疫吸附试验(ELISA)检测IL-6、IL-10水平;实时荧光定量PCR(RT-qPCR)检测miR-23b-3p表达水平;蛋白质印迹法检测FoxA1蛋白表达;双荧光素酶报告实验检测miR-23b-3p和FoxA1的靶向关系。结果低、中、高剂量白芍总苷处理后,肺炎链球菌诱导的肺泡上皮细胞的活性升高,细胞凋亡率降低,IL-6含量降低,IL-10含量升高,miR-23b-3p表达水平升高,FoxA1蛋白表达水平降低,呈剂量依赖性(P<0.05)。过表达miR-23b-3p后,FoxA1蛋白表达水平降低,细胞活性升高,细胞凋亡率降低,IL-6含量降低,IL-10含量升高(P<0.05)。抑制miR-23b-3p可逆转白芍总苷对肺炎链球菌诱导A549细胞凋亡、活性和IL-6和IL-10含量的影响。miR-23b-3p靶向调控FoxA1。结论白芍总苷通过调控miR-23b-3p/FoxA1轴抑制肺炎链球菌诱导的肺泡上皮细胞凋亡和炎症。

Effect of total glucosides of paeony on alveolar epithelial cell apoptosis induced by Streptococcus pneumoniae via miR-23b-3p/FoxA1

Objective To explore the total glucosides of paeony and the molecular mechanism of its effect on the apoptosis of alveolar epithelial cells induced by Streptococcus pneumoniae.Methods Alveolar epithelial cells A549 were randomly divided into the control group,the model group(1×10⁸CFU/mL of Streptococcus pneumoniae infection),the model+total glucosides of paeony low,medium,and high dose groups(17.5 mg/mL,35 mg/mL,70 mg/mL total glucosides of paeony+1×10⁸CFU/mL of Streptococcus pneumoniae infection),the model+total glucosides of paeony+anti-miR-23 b-3 p group(transfected with miR-23 b-3 p inhibitor+70 mg/mL total glucosides of paeony+1×10⁸CFU/mL Streptococcus pneumoniae infection).It used cell counting kit 8(CCK-8)to detect cell viability,flow cytometry to detect cell apoptosis,enzyme-linked immunosorbent assay(ELISA)to detect the levels of IL-6 and IL-10,real-time fluorescent quantitative PCR(RT-qPCR)to detect the expression level of miR-23 b-3 p,Western blotting to detect the expression of FoxA1 protein,and dual luciferase reporter experiment to detect targeting relationship of miR-23 b-3 p and FoxA1.Results After treatment with low,medium and high doses of total glucosides of paeony,the activity of alveolar epithelial cells induced by Streptococcus pneumoniae was increased,the rate of apoptosis was decreased,the content of IL-6 was decreased,the content of IL-10 was increased,the expression of miR-23 b-3 p was increased,and the FoxA1 protein expression was decreased,in a dose-dependent manner(P<0.05).After over-expression of miR-23 b-3 p,FoxA1 protein expression was decreased,cell viability was increased,cell apoptosis rate was decreased,IL-6 content was decreased,and IL-10 content was increased(P<0.05).The inhibition of miR-23 b-3 p could reverse the effect of total glucosides of paeony on A549 cell apoptosis,activity and IL-6 and IL-10 content induced by Streptococcus pneumoniae.miR-23 b-3 p targeted Fox A1.Conclusion The total glucosides of paeony can inhibit the apoptosis and inflammation of alveolar epithelial cells induced by Streptococcus pneumoniae by regulating the miR-23 b-3 p/Fox A1 axis.