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目的探讨山楂酸处理对高糖介导大鼠心肌细胞H9c2损伤和凋亡分子机制影响。方法以高糖诱导大鼠心肌细胞H9c2为模型,采用乳酸脱氢酶(LDH)法检测细胞损伤情况,TUNEL化学染色检测细胞凋亡率,Western blot检测AMPK的磷酸化水平以及Bcl-2、Bax的表达水平,免疫荧光检测细胞活性氧(ROS)的生成。结果 (1)与对照组相比,高糖组细胞LDH活性和细胞凋亡比率显著增高(P<0.05),山楂酸处理却能抑制高糖诱导的H9c2细胞LDH的积累,降低细胞凋亡(P<0.05)。(2)Western blot检测显示,与对照组相比,高糖组p-AMPK、Bcl-2下调(P<0.05),Bax上调(P<0.05);与高糖组相比,山楂酸处理组p-AMPK、Bcl-2上调(P<0.05),Bax下调(P<0.05),给予AMPK阻断剂Compound C后可以抑制山楂酸诱发的这些效应(P<0.05)。(3)ROS检测显示,山楂酸能够抑制高糖刺激的ROS的产生(P<0.05),而AMPK拮抗剂compound c却能够拮抗山楂酸对ROS的抑制(P<0.05)。结论山楂酸能通过激活AMPK信号通路,从而增强BCl-2表达、抑制Bax和ROS生成,拮抗高糖诱导的大鼠心肌细胞H9c2凋亡,对心肌细胞起保护作用。
Objective To investigate the effects of maslinic acid treatment on H9c2 injury and apoptosis in rat cardiomyocytes induced by high glucose. Methods H9c2 was induced by high glucose in rat cardiomyocytes. The cell injury was detected by lactate dehydrogenase (LDH) assay. The apoptosis rate was detected by TUNEL staining. The phosphorylation of AMPK and the expressions of Bcl-2, Bax The expression of reactive oxygen species (ROS) was detected by immunofluorescence. Results (1) Compared with the control group, LDH activity and apoptosis rate of H9c2 cells in high glucose group were significantly increased (P <0.05), while hawthorn acid treatment could inhibit the accumulation of LDH in H9c2 cells induced by high glucose, P <0.05). (2) Western blot showed that compared with the control group, p-AMPK and Bcl-2 were down-regulated and Bax was up-regulated in high glucose group (P <0.05) P-AMPK, Bcl-2 up-regulated (P <0.05) and Bax down-regulated (P <0.05). These effects were inhibited by maslinic acid (P <0.05). (3) ROS assay showed that maslinic acid could inhibit the ROS production induced by high glucose (P <0.05), while AMPK antagonist compound C could antagonize the inhibition of ROS by maslinic acid (P <0.05). Conclusion Maslinic acid can inhibit the expression of Bcl-2 and Bax, activate AMPK signal pathway, inhibit the production of Bax and ROS, and antagonize the high glucose-induced apoptosis of rat cardiac myocytes H9c2, which can protect cardiomyocytes.