苦参碱通过调节Bcl-2/Bax的平衡抑制大鼠缺血再灌注引起的肝细胞凋亡

Matrine inhibits hepatocyte apoptosis by regulating the balance of Bcl-2/Bax in rat liver ischemic reperfusion injury

目的肝脏缺血再灌注损伤是肝脏切除手术中最常见的并发症,由于激发细胞异常凋亡,常导致肝功能不全甚至引起死亡。本研究旨在揭示肝脏缺血再灌注损伤发生过程中苦参碱参与调节凋亡蛋白Bcl-2/Bax平衡的分子机制。方法健康成年SD大鼠45只,随机分为3组:一组作为空白对照组(SO组,n=15),不阻断肝脏血流;另外2组大鼠行夹闭门静脉和肝动脉缺血70 min后进行再灌注方法建立HIRI模型(缺血再灌注损伤模型),以缺血/再灌注+生理盐水组为对照组(NS组),缺血/再灌注+苦参碱组为实验组(MT组)。MT组在夹闭门静脉和肝动脉前经门静脉主干注入苦参碱。NS组注入生理盐水,阻断供血1h后恢复灌注,在恢复灌注2 h后采集标本行mRNA及Western blot检测。结果 qRT-PCR结果显示,正常对照SO组Bcl-2 mRNA表达水平显著高于Bax,在肝脏缺血再灌注模型组NS组中,高Bcl-2表达水平受到抑制,Bax表达水平显著高于Bcl-2。当行苦参碱干预后,Bax mRNA水平显著下降(MT组),基本接近SO组Bax水平。Western blot验证结果与mRNA水平结果相似。结论苦参碱显著抑制大鼠缺血再灌注引起的肝细胞凋亡,该途径可能由于苦参碱参与调节Bcl-2/Bax的平衡所致。

Objective Ischemia reperfusion injury （IRI） is the normal complication in hepatectomy, which always results in liver dysfunction by promoting cellular apoptosis. The present study aimed to explore the molecular pattern of matrine regulating the bal- ance of Bel-2/Bax in hepatic IRI process. Methods SD rats were randomly divided into 3 groups based on different treatment. Hepatic IRI model was established by clip-closed portal vein and hepatic artery followed by reperfusion, and then the rats were divided into 3 groups （n= 15 each） ： normal group （SO group）, model group （NS group） and MT group. In MT group, matrine was injected through main portal vein before closing portal vein and hepatic artery, while normal saline was injected in the same method in NS group. The blood perfusion was recovered 1 h minutes after blocking blood supply, and the related samples were collected 2 hours after recovering perfusion to be measured by qRT-PCR and western blot. Results qRT-PCR data showed that Bcl-2 mRNA in SO group is highest. Bax was activated in liver IRI model （NS group）. Bcl-2 level was rescued and Bax level was inhibited when matrine treated liver IRI model （MT group）. This results were also validated by Western blot. Conclusion In hepatic IRI model, matrine inhibits hepatocyte apopto- sis might be though regulating the balance of Bcl-2/Bax, which may rescue the level of Bcl-2 and suppress the level of Bax.