泽兰水提取物对H_9C_2心肌细胞缺氧/复氧损伤的保护作用研究

Protective effect of aqueous extract of Lycopus lycidus Turcz. on hypoxia /reoxygenation Injury in H_9C_2 Cardiomyocytes

目的观察泽兰水提取物对H_9C_2心肌细胞缺氧/复氧(hypoxia-reoxygenation,H/R)损伤的保护作用,并探讨其机制。方法制备H_9C_2心肌细胞H/R损伤的模型,用噻唑蓝(MTT)法观察泽兰与H_9C_2心肌细胞活力的量效关系,明确泽兰的保护作用。实验分为正常对照组(control),模型组(H/R),H/R+泽兰组(100,200,400μg·mL^(-1))。试剂盒检测乳酸脱氢酶(LDH)、超氧化物歧化酶(SOD)、丙二醛(MDA)、谷胱甘肽过氧化物酶(GSH-Px)含量的改变。Western Blot检测泽兰抗H_9C_2心肌细胞H/R损伤中对HIF-1,HO-1,VEGF等蛋白表达的影响。结果与control组相比,H/R组细胞活力、SOD及GSH-Px含量水平显著降低,LDH、MDA释放量明显增多(P<0.01);与H/R组相比,泽兰能够显著逆转上述变化(P<0.05)。Western Blot显示,与control组相比,H/R组HIF-1a,HO-1,VEGF蛋白表达均显著增加(P<0.01);与H/R组相比,H/R+泽兰中、高剂量组显著改善上述变化(P<0.05)。此外,H/R时Akt磷酸化水平显著降低,而泽兰能明显增加p-Akt表达,但此作用被LY294002所阻断(P<0.01)。结论泽兰水提取物对H/R诱导的H_9C_2心肌细胞发挥保护作用,其机制可能与调控PI3K/Akt信号通路,调节HIF-1a及其下游蛋白表达相关。

Objective To observe the protective effect of aqueous extract of Lycopus lycidus Turcz.(ALT)on hypoxia/reoxygenation(H/R)injury in H9C2 cardiomyocytes,and to explore its possible mechanism.Methods the H/R model of H9C2 cardiomyocytes was prepared.The dose effect relationship between the activity of ALT and H9C2 cardiomyocytes was observed by MTT assay,and the protective effect of ALT was determined.The experiment set:control group,H/R group,H/R+ALT groups including the different dose groups(100,200,400μg·mL-1).The changes in the content of lactate dehydrogenase(LDH),superoxide dismutase(SOD),malondialdehyde(MDA)and glutathione peroxidase(GSH-Px)were detected by these kits.Western blot detect the effect of protein expression of HIF-1 a,HO-1,VEGF on H/R injury in H9C2 myocardial cells.Results Compared with the control group,the viability of cell,SOD and GSH-Px contents in H/R group significantly decreased,and the release of LDH and MDA increased significantly(P<0.01);Compared with H/R group,Lycopus significantly reverse the changes(P<0.05).Western blot showed:Compared with the control group,the protein expression of HIF-1 a,HO-1,VEGF in H/R group significantly increased(P<0.01);Compared with H/R group,H/R+ALT groups were significantly improve the changes(P<0.05).In addition,the phosphorylation level of Akt decreased significantly in H/R group,while the expression of p-Akt in ALT groups was significantly increased,but this effect was blocked by LY294002(P<0.01).Conclusion ALT exerts protective effect on H9C2 cardiomyocytes injury induced by H/R,its mechanism may be related to regulation of PI3 K/Akt signaling pathway,regulated HIF-1 a and its downstream proteins.