熊果酸通过调节miR-373-3p对肺癌A549细胞凋亡和迁移的影响

Effects of ursolic acid on apoptosis and migration of A549 lung cancer cells by regulating miR-373-3p

目的探讨熊果酸通过调节mi R-373-3p对肺癌A549细胞凋亡和迁移的影响。方法 MTT法检测熊果酸对肺癌A549细胞的毒性作用;AnnexinⅤ-FITC/PI双染法、流式细胞术检测熊果酸作用于转染mi R-373-3p的A549细胞后对A549细胞凋亡影响;实时荧光定量PCR法分别检测熊果酸加药组和转染mi R-373-3p mimics组中A549细胞mi R-373-3p表达程度;RT-PCR和免疫印迹法检测熊果酸作用下mi R-373-3p调控的下游靶基因TXNIP m RNA及蛋白的表达;细胞划痕愈合实验检测mi R-373-3p细胞迁移能力。结果熊果酸显著降低肺癌A549细胞的生长活力(P<0.05),IC 50为60μmol/L;瞬时转染mi R-373-3p mimics可以上调A549细胞中mi R-373-3p的表达(24 h,11.367±2.120;48 h,12.167±1.108);熊果酸作用于转染mi R-373-3p mimics的A549细胞后,能够显著提高细胞中mi R-373-3p mimics的表达水平(24 h,16.787±3.109;48 h,16.980±3.106);并且mi R-373-3p mimics下游预测的目标基因TXNIP m RNA(10.757±0.79)及蛋白表达量(0.8210±0.043)均明显上升;FCM结果显示:mi R-373-3p能够促进肺癌A549细胞凋亡(46.20±5.970)。细胞转染mi R-373-3p mimics后,迁移能力明显受到抑制(P<0.001)。结论熊果酸能够通过促进肺癌A549细胞凋亡抑制其迁移,其作用机制可能是通过上调mi R-373-3p表达来完成的。

Objective To investigate the effect of ursolic acid on apoptosis and migration of lung cancer A549 cells by regulating miR-373-3 p.Methods We used an MTT assay to detect the toxic effects of ursolic acid on A549 lung cancer cells.AnnexinⅤ-FITC/PI double staining and flow cytometry were used to detect the effect of ursolic acid on A549 cells transfected with miR-373-3 p.Real-time fluorescence quantitative PCR was used to detect the expression level of miR-373-3 p in A549 cells in the ursolic acid dosing group and the transfected miR-373-3 p mimics group.RT-PCR and Western blot were used to detect m RNA and the protein expression of the downstream target gene,TXNIP,which is regulated by miR-373-3 p under the action of ursolic acid.The scratch text was used to evaluate miR-373-3 p cell migration ability.Results Ursolic acid significantly decreased the growth viability of A549 lung cancer cells(P<0.05),with an IC50 of 60μmol/L.Transiently transfected miR-373-3 p mimics up-regulated the expression of miR-373-3 p in A549 cells(24 h,11.367±2.120;48 h,12.167±1.108),and ursolic acid significantly increased the expression level of miR-373-3 p mimics in A549 cells transfected with miR-373-3 p mimics(24 h,16.787±3.109;48 h,16.980±3.106).m RNA for the target gene,TXNIP,(10.757±0.79)and protein expression(0.8210±0.043)predicted by miR-373-3 p mimics were significantly increased.FCM result showed that miR-373-3 p promoted apoptosis of A549 lung cancer cells(46.20±5.970),and after being transfected with miR-373-3 p mimics,the cells’migration ability was significantly inhibited(P<0.001).Conclusions Ursolic acid can promote the apoptosis of A549 lung cancer cells and inhibit their migration.The mechanism may involve up-regulation of miR-373-3 p expression.