多巴胺受体(DR2)激活对乳鼠心肌细胞缺氧/再灌注损伤的保护作用及其机制

The protective effect of DR2 activation on hypoxia/reperfusion injury in the neonatal rat cardiomyocytes and related mechanism

目的:观察多巴胺受体(DR2)激活对乳鼠心肌细胞缺氧/再灌注损伤的影响,并探讨其机制。方法:复制原代培养乳鼠心肌细胞缺氧/再灌注损伤模型,细胞随机分为正常组(Control)、缺氧/再灌注组(H/R)、DR2激动剂组(溴麦角环肽,Bro)、抑制剂组(氟哌啶醇,Hal)。倒置显微镜、透射电镜、流式细胞仪检测细胞凋亡情况;检测细胞培养液乳酸脱氢酶(LDH)、超氧化物歧化酶(SOD)活性和丙二醛(MDA)含量;RT-PCR和Western blot方法检测心肌细胞促凋亡因子(Cyt C、caspase-3、caspase-8、caspase-9、Fas及Fas-L)及抑凋亡因子(Bc-l 2)mRNA和蛋白表达。结果:与正常组相比,H/R组细胞凋亡率、LDH活性、MDA含量、促凋亡因子及抑凋亡因子表达均增加,唯有SOD活性降低;与H/R组比较,Bro组可减轻或逆转上述指标的变化;Hal组上述指标变化不明显。结论:DR2激活可抑制缺氧/再灌注所致的乳鼠心肌细胞损伤和凋亡,机制与减少氧自由基有关。

Objective： To observe the effect of dapamine receptor（DR2）activation on hypoxia/reperfusion injury（HRI） in the neonatal rat cardiomyocytes,and to explore its mechanism.Methods： The hypoxia/reperfusion（H/R） injury model was established in primarily cultured neonatal rat cardiomyocytes,and randomly assigned： control,H/R,bromocriptine（Bro）and haloperidol（Hal）groups.The cell apoptosis was detected using inverted microscope,transmission electron microscope and flow cytometry（FCM）.The lactate dehydrogenase（LDH） and suporoxide dismutase（SOD）activity and malondialdehyde（MDA）content in cell medium were analyzed.The expression of mRNA and protein of caspase-3,caspase-8,caspase-9,Fas,Fas-L,Cyt C and Bcl-2 were detected by RT-PCR and Western blot,respectively.Results： Compared with the control group,apoptosis rate,LDH activity,MDA content and the expression of pro-apoptotic factors and anti-apoptotic factors were increased,but SOD activity was decreased in H/R group.Compared with the H/R group,all index above-mentioned were down-regulated or reversed in Bro-group,and had no obvious differences in Hal-group.Conclusion： The neonatal rat cardiomyocytes injury and apoptosis caused by hypoxia/reperfusion can be inhibited with DR2 activation,which mechanism is related to scavenging oxygen radical.