CAJ | 학술논문

目的：探讨间充质干细胞（ MSC）条件培养基（ MSCCM）对氧化应激损伤的心肌细胞的保护作用及机制。方法：用流式细胞术对MSC进行鉴定，再用MTT实验确定MSCCM对抗H2 O2氧化应激损伤的最佳孵育时间。将H9c2细胞分为正常组、模型组、模型＋MSCCM组和MSCCM组。模型＋MSCCM组和MSCCM组均用MSC－CM进行预孵育24 h，模型组和模型＋MSCCM组用浓度为300μmol／L的H2 O2作用4 h模拟心肌细胞的氧化应激损伤。利用流式细胞术检测损伤后心肌细胞的线粒体膜电位变化、凋亡比率等指标，通过荧光显微镜检测细胞ROS产生的变化，Western blot检测Nrf2／ARE通路的相关蛋白表达。结果：MSCCM组心肌细胞的线粒体膜电位、凋亡比率和ROS产生与正常组相比均无显著差异（ P＞0．05）；模型＋MSCCM组的线粒体膜电位、凋亡比例和ROS产生与模型组相比差异显著（P＜0．01）；在0 h到24 h这段时间内，心肌细胞Nrf2／ARE通路中的Nrf2核转位与HO－1表达均随着MSCCM孵育时间延长而增加。结论：MSCCM可以保护心肌细胞，对抗H2 O2诱导的氧化应激损伤，其机制可能与激活Nrf2／ARE通路有关。
목적：탐토간충질간세포（ MSC）조건배양기（ MSCCM）대양화응격손상적심기세포적보호작용급궤제。방법：용류식세포술대MSC진행감정，재용MTT실험학정MSCCM대항H2 O2양화응격손상적최가부육시간。장H9c2세포분위정상조、모형조、모형＋MSCCM조화MSCCM조。모형＋MSCCM조화MSCCM조균용MSC－CM진행예부육24 h，모형조화모형＋MSCCM조용농도위300μmol／L적H2 O2작용4 h모의심기세포적양화응격손상。이용류식세포술검측손상후심기세포적선립체막전위변화、조망비솔등지표，통과형광현미경검측세포ROS산생적변화，Western blot검측Nrf2／ARE통로적상관단백표체。결과：MSCCM조심기세포적선립체막전위、조망비솔화ROS산생여정상조상비균무현저차이（ P＞0．05）；모형＋MSCCM조적선립체막전위、조망비례화ROS산생여모형조상비차이현저（P＜0．01）；재0 h도24 h저단시간내，심기세포Nrf2／ARE통로중적Nrf2핵전위여HO－1표체균수착MSCCM부육시간연장이증가。결론：MSCCM가이보호심기세포，대항H2 O2유도적양화응격손상，기궤제가능여격활Nrf2／ARE통로유관。
[ ABSTRACT] AIM: To investigate the protective effect of mesenchymal stem cell ( MSC)-conditioned medium (MSCCM) on myocardial cell line H9c2 and its mechanism.METHODS:Verification of MSC was performed by flow cy-tometry analysis, followed by MTT assay to determine the optimal incubation time of MSCCM with myocardial cells.The cells were divided into 4 groups:normal ( N) group, model ( M) group, M+MSCCM group and MSCCM group.The cells in M+MSCCM group and MSCCM group were pre-incubated with MSCCM for 24 h.The cells in M group and M+MSCCM group were treated with 300 μmol/L H2 O2 for 4 h to imitate oxidative injury of myocardial cells.Mitochondrial membrane potential and apoptotic rate of injured myocardial cells were detected by flow cytometry.The ROS production was measured by fluorescence microscopy.The nuclear translocation of Nrf2 and expression of HO-1 was examined by Western blot.RE-SULTS:No difference of mitochondrial membrane potential, apoptotic rate or ROS production between MSCCM group and N group was observed (P>0.05).The mitochondrial membrane potential depolarization, apoptotic rate and ROS produc-tion in M+MSCCM group were significantly lower than those in M group ( P<0.01 ) .The nuclear translocation of Nrf2 and expression of HO-1 in the myocardial cells were increased with MSCCM incubation time prolonged.CONCLUSION:MSCCM protects the myocardial cells against oxidative injury induced by H2 O2 .The anti-oxidative mechanism would be as-sociated with the activation of Nrf2/ARE pathway.