CAJ | 학술논문

目的观察促红细胞生成素(EPO)对缺氧/复氧心肌细胞凋亡的影响,并初步探讨蛋白激酶C(PKC)和线粒体ATP敏感性钾通道(mitoKATP)与EPO在抗凋亡信号通路中的相互关系.方法分离培养SD大鼠乳鼠心肌细胞,并分为对照组、缺氧/复氧组、EPO组和PKC抑制剂白屈菜红碱组,建立缺氧/复氧模型;用流式细胞术检测心肌细胞凋亡率,激光共聚焦显微镜扫描观察细胞黄素蛋白自体荧光强度变化,监测钾通道开放情况.结果缺氧/复氧组心肌细胞凋亡率明显高于对照组[(42.56±8.00)%比(17.88±2.00)%,P<0.053,黄素蛋白自体荧光强度值与对照组比较差异无统计学意义[(0.278±0.170)×10-2比(0.149±0.050)×10-2,P>0.05];EPO组心肌细胞凋亡率明显低于缺氧/复氧组[(22.73±5.00)%比(42.56±8.00)%,P<0.05],黄素蛋白自体荧光强度值则较缺氧/复氧组明显增强[(2.201±1.090)×10-2比(0.278±0.170)×10-2,P<0.01];白屈菜红碱对EPO抗凋亡和增强黄素蛋白自体荧光强度有阻断作用[细胞凋亡率:(46.72±17.00)%比(22.73±5.00)%,荧光强度:(0.986±0.320)×10-2比(2.201±1.090)×10-2,P<0.01和P<0.05].结论通过激活PKC继而开放mitoKATP通道可能是EPO抗缺氧/复氧心肌细胞凋亡的信号通路之一.
목적 관찰촉홍세포생성소(EPO)대결양/복양심기세포조망적영향,병초보탐토단백격매C(PKC)화선립체ATP민감성갑통도(mitoKATP)여EPO재항조망신호통로중적상호관계.방법 분리배양SD대서유서심기세포,병분위대조조、결양/복양조、EPO조화PKC억제제백굴채홍감조,건립결양/복양모형;용류식세포술검측심기세포조망솔,격광공취초현미경소묘관찰세포황소단백자체형광강도변화,감측갑통도개방정황.결과 결양/복양조심기세포조망솔명현고우대조조[(42.56±8.00)%비(17.88±2.00)%,P<0.053,황소단백자체형광강도치여대조조비교차이무통계학의의[(0.278±0.170)×10-2비(0.149±0.050)×10-2,P>0.05];EPO조심기세포조망솔명현저우결양/복양조[(22.73±5.00)%비(42.56±8.00)%,P<0.05],황소단백자체형광강도치칙교결양/복양조명현증강[(2.201±1.090)×10-2비(0.278±0.170)×10-2,P<0.01];백굴채홍감대EPO항조망화증강황소단백자체형광강도유조단작용[세포조망솔:(46.72±17.00)%비(22.73±5.00)%,형광강도:(0.986±0.320)×10-2비(2.201±1.090)×10-2,P<0.01화P<0.05].결론 통과격활PKC계이개방mitoKATP통도가능시EPO항결양/복양심기세포조망적신호통로지일.
Objective To investigate the anti-apoptosis effect of erythropoietin (EPO) on myocardial cells after hypoxia/reoxygenation in vitro, and the relationship among protein kinase C (PKC), the mitochondrial ATP-sensitive potassium (mitoKATP) channel and EPO in the anti-apoptotic signaling pathways.Methods Cardiocytes were harvested from neonatal rats and cultured.Cultured myocardial cells were divided into the control group, the hypoxia/reoxygenation group, the EPO group and the chelerythrine group, and a hypoxia/reoxygenation model of cardiocytes was reproduced.Apoptosis rate was assayed by flow cytometry.Flavoprotein fluorescence was scanned by confocal laser microscope to assess the mitoKATP channel activity.Results Apoptosis rate was significantly higher in hypoxia/reoxygenation group than that of control group [(42.56±8.00)% vs.(17.88±2.00)%, P<0.05].There was no statistically significant difference in flavoprotein fluorescence between this group and the control group [(0.278±0.170) ×10-2 vs.(0.149±0.050)× 10-2, P>0.05].Myocardial cell apoptosis rate in EPO group was lower than that in hypoxia/reoxygenation group [(22.73 ±5.00) % vs.(42.56 ± 8.00)%, P < 0.05], and flavoprotein fluorescence intensity was significantly enhanced when compared with hypoxia/reoxygenation group [(2.201±1.090)×10-2 vs.(0.278±0.170) ×10-2, P<0.01].However, when chelerythrine was added,the anti-apoptosis effect of EPO was blocked, and the intensity of cardiocytes flavoprotein fluorescence was decreased [the apoptosis rate was (46.72± 17.00)% and the flavoprotein fluorescence intensity was(0.986±0.320)× 10-2].When compared with EPO group there was statistically significant difference (P<0.01 and P< 0.05).Conclusion Myocardial cell apoptosis occurs in hypoxia/reoxygenation injury, and EPO can protect rat cardiomyocytes from hypoxia/reoxygenation induced apoptosis.The protective effect is partly associated with the PKC/mitoKATP pathway.