The Study On MiR-21-5p Via The PI3K/Akt Pathway Regulating AEC Ⅱ Apoptosis Of Rats Induced By H₂O₂

Objective: To verify that miR-21-5p can regulate the PI3K/Akt pathway to antagonize AECⅡ apoptosis,and play a protective role through studying that miR-21-5p regulate the apoptosis of alveolar type II epithelial cells（AECⅡ）induced by hydrogen peroxide（H₂O₂）.Methods: The subculture AECⅡ were divided into 6 groups（n=7）: group A（control group:Using PBS liquid treatment）,group B（injury group: 0.5mmol/L H₂O₂ processing）,group C（miR-21-5p overexpression group: miR-21-5p lentivirus overexpression vector was transfected into AECⅡ 24 h,and induced by 0.5mmol/L H₂O₂ established apoptosis model）,group D（miR-21-5p empty group: without miR-21-5p lentivirus vector was transfected into AECⅡ 24 h,and induced by 0.5mmol/L H₂O₂ established apoptosis model）,group E（miR-21-5p inhibition vector group: miR-21-5p lentivirus inhibited that vector was transfected into AECⅡ 24 h,and induced by 0.5mmol/L H₂O₂ established apoptosis model）,group F（PI3K/Akt blocker group: miR-21-5p lentivirus overexpression vector was transfected into AECⅡ 24 h,adding 25 μmol /L PI3K/Akt blocking agent LY294002,and induced by 0.5mmol/L H₂O₂ established apoptosis model）.CCK8 detected cell proliferation activity of 0h,12 h,24h,48 h and observed cell proliferation activity of group A,B,C,D,E,F;Flow cytometry detected the apoptosis rate of 0h,12 h,24h,48 h in each group;Real-time fluorescence quantitative PCR detected the miR-21-5p expression of AECⅡ in each group;Western Blot-Western blot tested the expression of p Akt protein in each group.Results: 1.The cell proliferation activity of CCK8: with the extension of time,the cell proliferation activity of group A gradually increased.The cell proliferation activity in group B,C,D,E,F decreased（P < 0.05）,and the cell proliferation activity of group C in the five groups decreased obviously slowly than the other four groups（P < 0.05）.2.Flow cytometry detecting the early apoptosis rate（FCM）: the apoptosis rate increases with the extension of injury time in each group（P < 0.05）;In 12 h,24 h,48 h test,the cell apoptosis rate group B is higher than that in group A（P < 0.05）,the apoptosis rate in group C is obviously lower than that in group D,E and F（P < 0.05）;At the identical time point,the apoptosis rate in group E and F is higher than that in group D（P < 0.05）.3.Real-time fluorescence quantitative PCR detecting the miR-21-5p m RNA expression of AECⅡ in each group: miR-21-5 P expression level in group B is lower than that in group A（P < 0.05）;in group C,D,E and F,miR-21-5 P expression level in group C,F is highest（P < 0.05）,and that in group E is the lowest（P < 0.05）.4.Western blot testing the expression of p Akt protein in each group: B group,p Akt protein expression level in group B is higher than that in group A（P <0.05）;in group C,D,E and F,p Akt protein expression level group C is the highest（P <0.05）,that in group F is the lowest（P <0.05）,and that in group D is higher than that in group E（P <0.05）.Conclusion:（1）miR-21-5p can play an anti apoptotic effect on AECⅡ induced by H₂O₂.（2）miR-21-5p can inhibit AECⅡ apoptosis through regulating the PI3K/Akt pathway,and play a protective role.