Objective To explore the mechanism underlying the effects of miR-195/497 on the expression of TGF-β in human bronchial epithelial cells exposed to cigarette smoke extract(CSE).Methods BEAS-2B cells were exposed to 3％CSE to simulate the pathogenesis of chronic obstructive pulmonary disease(COPD),and the expression levels of miR-195,miR-497,SMAD7 and TGF-β genes were detected by quantitative real-time polymerase chain reaction(qRT-PCR).The targeting relationship of miR-195 and miR-497 with SMAD7 was validated via the dual luciferase reporter assay.After transfection of miR-195 mimic and miR-497 mimic into cells treated with 3％CSE,the expression of SMAD7 was detected by qRT-PCR.Following transfection of miR-195 mimic,miR-497 mimic and si-SMAD7 into cells treated with 3％CSE,the gene expressions of miR-195,miR-497,SMAD7 and TGF-β were detected by qRT-PCR,and the protein expressions of SMAD7 and TGF-β were measured via Western blotting.Results Compared with the normal cells,the relative expression levels of miR-195 and miR-497 were lower(P＜0.01),and the relative expression levels of SMAD7 and TGF-β were higher in cells treated with 3％CSE(P＜0.05).The relative expression of Wt-SMAD7 in the miR-195 mimic group was lower than that in the mimic NC group(P＜0.05).There was no difference in the relative expression of Mut-SMAD7 between the miR-195 mimic group and the mimic NC group(P＞0.05).The relative expression of Wt-SMAD7 in the miR-497 mimic group was lower than that in the mimic NC group(P＜0.05).There was no difference in the relative expression of Mut-SMAD7 between the miR-497 mimic group and the mimic NC group(P＞0.05).The relative mRNA expression of SMAD7 in the miR-195 mimic group was lower than that in the mimic NC group(P＜0.05).The mRNA relative expression of SMAD7 in the miR-497 mimic group was lower than that in the mimic NC group(P＜0.05).The expressions of miR-195 and miR-497 were lower(P＜0.05),and the expressions of SMAD7 and TGF-βwere higher in cells treated with 3％CSE than those in cells of the control group(P＜0.05).Compared with the 3％CSE group,the expression of miR-195 was higher(P＜0.05),the expressions of SMAD7 and TGF-β were lower(P＜0.05),and the expression of miR-497 was not significantly altered(P＞0.05)in the miR-195 mimic+3％CSE group.Compared with the 3％CSE group,the expression of miR-497 was higher(P＜0.05),the expressions of SMAD7 and TGF-β were lower(P＜0.05),and the expression of miR-195 was not significantly altered(P＞0.05)in the miR-497 mimic+3％CSE group.The expressions of SMAD7 and TGF-β in the si-SMAD7+3％CSE group were lower than those in the 3％CSE group(P＜0.05),while there was no difference in the expressions of miR-195 and miR-497 between the two groups(P＞0.05).The relative protein expressions of SMAD7 and TGF-β in the 3％CSE group were higher than those in the control group(P＜0.05),whereas those in the miR-195 mimic+3％CSE group,miR-497 mimic+3％CSE group,and si-SMAD7+3％CSE group were lower compared to those in the 3％CSE group(P＜0.05).Conclusions The miR-195/497 could affect the expression of TGF-β in human bronchial epithelial cells exposed to CSE by regulating the expression of SMAD7.
国家科技期刊平台
NSTL
万方数据
