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目的以乳鼠心肌细胞缺氧/复氧模型模拟在体心肌缺血再灌注损伤,观察附子多糖后处理对缺氧/复氧后心肌细胞的作用及机制。方法建立乳鼠心肌细胞缺氧/复氧模型,将乳鼠心肌细胞分为正常对照组、缺氧/复氧组、缺氧后适应组和附子多糖组。缺氧/复氧组给予心肌细胞缺氧3 h后复氧6 h;缺氧后适应组在细胞缺氧3 h后,复氧前即给予3个循环的5 min复氧/5 min缺氧,随后复氧6 h;附子多糖组在缺氧3 h后,将心肌细胞换入含附子多糖浓度为10 mg.mL-1的培养液中,常规培养6 h。MTT法检测细胞活力,流式细胞仪测定细胞内钙离子浓度和心肌细胞凋亡率,检测细胞培养液中乳酸脱氢酶(LDH)和肌酸激酶(CK)的活性。结果与缺氧/复氧组比较,经附子多糖后处理,可以增加心肌细胞存活率(P<0.01),减少LDH(P<0.01)和CK(P<0.05)的释放,降低细胞内钙离子浓度(P<0.05),有效抑制心肌细胞的凋亡(P<0.05)。结论附子多糖对缺氧/复氧后心肌细胞产生保护效应,机制与其抑制钙超载、减轻线粒体的损伤有关。
OBJECTIVE: To simulate myocardial ischemia-reperfusion injury in neonatal rat cardiomyocytes subjected to hypoxia / reoxygenation to observe the effect and mechanism of monkshood polysaccharide postconditioning on hypoxia / reoxygenation cardiomyocytes. Methods The neonatal rat cardiomyocytes hypoxia / reoxygenation model was established. The cardiomyocytes were divided into normal control group, hypoxia / reoxygenation group, hypoxia adaptation group and monkshood polysaccharide group. Hypoxia / reoxygenation group was given cardiomyocytes 3h after reoxygenation 6h; After hypoxia adaptation group 3h after cell hypoxia, reoxygenation before giving 3 cycles of 5min reoxygenation / 5min hypoxia , Followed by reoxygenation for 6 h. After treated with hypoxia for 3 h, the mononuclear polysaccharide group was used to replace cardiomyocytes with medium containing monkshood polysaccharide (10 mg.mL-1) for 6 h. Cell viability was measured by MTT assay. Intracellular calcium concentration and apoptotic rate were measured by flow cytometry. The activity of lactate dehydrogenase (LDH) and creatine kinase (CK) in cell culture medium was detected. Results Compared with hypoxia / reoxygenation group, aflatoxin postconditioning could increase the survival rate of cardiomyocytes (P <0.01), decrease the release of LDH (P <0.01) and CK (P <0.05) and decrease the intracellular calcium Concentration (P <0.05), effectively inhibiting cardiomyocyte apoptosis (P <0.05). Conclusion Aconite polysaccharides have a protective effect on cardiomyocytes after hypoxia / reoxygenation, and its mechanism is related to inhibiting calcium overload and mitigating mitochondrial damage.