磷酸二酯酶5的短发夹RNA对肌细胞纤维化的作用抑制研究

目的 通过短发夹RNA(shRNA)延长对磷酸二酯酶5(PDE5)的抑制作用,探索shRNA-PDE5对细胞存活率的影响以及对心肌梗死后心功能及心室重构的作用及其机制.方法 (1)细胞实验:分离SD大鼠骨髓间充质干细胞(MSCs),将Ad-shRNA-PDE5和Ad-Null分别感染体外培养的MSCs,嘌呤霉素筛选稳定...

Inhibition of phosphodiesterase 5 short hairpin RNA on myocyte fibrosis

Objective To explore the effect of shRNA-PDE5 on cell viability,cardiac function and ventricular remodeling after myocardial infarction through the inhibition of phosphodiesterase 5(PDE5)by short hairpin RNA(shRNA).Methods(1)Cell experiment:The mesenchymal stem cells(MSCs)of SD rats were isolated,and the MSCs cultured in vitro were infected with Ad-shRNA-PDE5 and Ad-Null,respectively.Puromycin cell lines were screened for stable expression and were divided into shPDE5 group,blank control vector(Ad-Null)group,and normal group was set up.TdT-mediated dUTP nick-end labeling staining was used to detect the apoptosis of stabilized MSCs induced by oxygen and sugar deprivation(OGD)in each group.(2)Animal experiment:Male SD rats were selected and the myocardial infarction model was established by ligating the anterior descending branch of the left coronary artery.Rats were randomly divided into 6 groups:sham operation group,model group,control group,transfection-1 group,transfection-2 group,and transfection-3 group.After descending branches before ligation,MSCs with different treatments were injected at each of 4 points in the myocardial infarction area and surrounding areas immediately after ligation.Western blot was used to detect the expression levels(optical density values)of transforming growth factor-β(TGF-β)and fibroblast growth factor 2(FGF2)in rat myocardium.Masson staining was used to detect fibrosis in myocardial infarction area and surrounding area.Results(1)The number of apoptotic cells in shPDE5 group,Ad-Null group and normal group were(6.00±1.00),(14.33±1.52)and(5.33±1.52),respectively.The number of apoptotic cells in shPDE5 group and Ad-Null group was significantly reduced,and the difference was statistically significant(P<0.05).(2)The expression of FGF2 protein in the sham operation group,model group,control group,transfection-1 group,transfection-2 group,and transfection-3 group were 0.11±0.02,0.22±0.05,0.27±0.08,0.28±0.04,0.33±0.05 and 0.50±0.03,respectively;the expression of TGF-βprotein in the 6 groups were 0.18±0.02,0.91±0.03,0.72±0.01,0.72±0.03,0.48±0.02 and 0.27±0.09,respectively.Comparison between transfection-2 group,transfection-3 and sham operation group,model group,control group,transfection-1 group,the differences of the factors were statistically significant(all P<0.05).Compared with the model group,the degree of cell fibrosis in the myocardial tissue around infarction area of rats in control group,transfection-1 group,transfection-2 group,and transfection-3 were reduced successively.Conclusion shRNA-PDE5 can improve cardiac function by regulating the expression of related factors and inhibiting myocardial fibrosis through oligocytic apoptosis.