益气逐瘀方对缺氧诱导乳鼠心肌细胞凋亡及miR-24/Bim通路的影响

目的研究益气逐瘀方(参元丹)对缺氧诱导乳鼠心肌细胞凋亡及miR-24/Bim通路的影响。方法分离并培养乳鼠心肌细胞,建立心肌细胞缺氧模型,Lipofectamine2000转染miR-24 mimic/inhibitor至心肌细胞,实验分为正常组、缺氧组、参元丹组、参元丹+miR-24 mimic组、参元丹+miR-24 inhibitor组,治疗组予参元丹含药血清干预。比色法检测心肌细胞培养基上清液LDH、CPK活性,MTT法检测心肌细胞存活率,Annexin V/PI双染流式细胞术检测心肌细胞凋亡,qRT-PCR检测心肌细胞miR-24、Bim mRNA表达。结果与低氧组相比,参元丹组、参元丹+miR-24 mimic组、参元丹+miR-24 inhibitor组均能显著降低细胞培养基上清液LDH、CPK活性(P<0.05),提高心肌细胞存活率(P<0.05),降低心肌细胞凋亡(P<0.05),升高心肌细胞miR-24 mRNA表达(P<0.05),降低Bim mRNA表达(P<0.05);治疗组之间比较,参元丹+miR-24 mimic组作用更显著(P<0.05)。结论益气逐瘀方参元丹能够减轻缺氧诱导乳鼠心肌细胞损伤及细胞凋亡,其作用机制可能与其上调miR-24表达,同时抑制其靶基因Bim mRNA表达有关。

Influence of Yiqi Zhuyu Decoction on Cardiomyocyte Apoptosis and miR-24/Bim Pathway in Hypoxia Induced Neonatal Rat Myocardial Cells

Objective:To observe the influence of Yiqi Zhuyu Decoction(Shenyuandan,SYD)on miR-24/Bim pathway on cardiomyocyte apoptosis induced by hypoxia in suckling mice.Methods:Myocardial cells were isolated and cultured to establish hypoxia model.Lipofectamine2000 was transfected with miR-24 mimic/inhibitor to cardiomyocytes.The experiment was divided into the normal group,hypoxia group,Shenyuandan group,Shenyuandan+miR-24 mimic group and Shenyuandan+miR-24 inhibitor group.The treatment groups were treated with serum containing Shenyuandan.The activity of LDH and CPK in the supernatant of cardiomyocyte culture medium was detected by colorimetry,the survival rate of cardiomyocytes by MTT,the apoptosis of cardiomyocytes by Annexin V/PI double staining flow cytometry,the expression of miR-24 and Bim mRNA in cardiomyocytes by qRT-PCR.Results:Compared with hypoxia group,all Shenyuandan groups could significantly reduce the activity of LDH and CPK in the supernatant of cell culture medium,improve the survival rate of cardiomyocytes,reduce the apoptosis of cardiomyocytes,increase the expression of miR-24 mRNA and decrease the expression of Bim mRNA in cardiomyocytes(P<0.05).Among all Shenyuandan groups,Shenyuandan+miR-24 mimic group had more significant effect.Conclusion:Yiqi Zhuyu Decoction Shenyuandan can reduce the injury and apoptosis of cardiomyocytes induced by hypoxia.The mechanism may be related to the up regulation of miR-24 expression and the inhibition of BIM mRNA expression.