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目的研究苦参素对大鼠肝缺血再灌注(IRI)中肝细胞的保护作用及其相关机制。方法采用阻断和恢复大鼠肝动脉和门静脉血供,制作全肝缺血30min再灌注90min模型。雄性Wistar大鼠随机数字法分为假手术组(sham组)、苦参素组(oxymatrine组)和对照组(IRI组),每组10只。血清天冬氨酸氨基转移酶(AST)、丙氨酸氨基转移酶(ALT)水平检测肝功状况,苏木素-伊红(HE)染色观察病理形态学改变,原位末端标记法(TUNEL)观察肝细胞凋亡情况,流式细胞术(FCM)测定肝细胞凋亡率及细胞周期,Western印迹法检测肝Fas基因表达情况。结果Oxymatrine组AST和ALT水平(513U/L±96U/L,352U/L±72U/L)明显低于IRI组(1326U/L±211U/L,768U/L±175U/L),差异有统计学意义(t值分别为4·21、4·13,P均<0·01);IRI组AST和ALT水平明显高于sham组(112U/L±53U/L、55U/L±17U/L),差异有统计学意义(t值分别为4·72、4·36,P均<0·01)。HE染色显示Oxymatrine组大鼠肝组织病理改变较IRI组轻,TUNEL检测显示oxymatrine组大鼠肝组织凋亡细胞数量较IRI组少。FCM检测显示oxymatrine组肝细胞凋亡明显少于IRI组(t=4·53,P<0·01);oxymatrine组较IRI组处于G0/G1期的肝细胞少,而S期肝细胞明显增多(t=4·52,P<0·01)。结论苦参素通过保护肝细胞和促进肝细胞更新,可明显减轻大鼠肝细胞在肝缺血再灌注中的损伤。
Objective To study the protective effect of oxymatrine on hepatocytes in hepatic ischemia-reperfusion (IRI) and its related mechanisms. Methods The hepatic arterial and portal venous blood supply were blocked and restored, and the model of total hepatic ischemia 30 min and reperfusion 90 min was made. Male Wistar rats were randomly divided into sham operation group (sham group), matrine group (oxymatrine group) and control group (IRI group), 10 rats in each group. Serum aspartate aminotransferase (AST) and alanine aminotransferase (ALT) levels were used to measure liver function. Hematoxylin-eosin (HE) staining was used to observe the pathological changes. In situ end labeling (TUNEL) observation was performed. Apoptosis of hepatocytes was detected by flow cytometry (FCM), and hepatocyte apoptosis rate and cell cycle were measured. Expression of hepatic Fas gene was detected by Western blot. Results The levels of AST and ALT in oxymatrine group (513U/L±96U/L, 352U/L±72U/L) were significantly lower than those in IRI group (1326U/L±211U/L, 768U/L±175U/L). Significance (t values were 4.21, 4.13, P all <0.01); AST and ALT levels in IRI group were significantly higher than those in sham group (112 U/L ± 53 U/L, 55 U/L ± 17 U/L ), The difference was statistically significant (t values were 4.72, 4.36, P all <0.01). HE staining showed that the pathological changes of liver tissue in Oxymatrine group were lighter than those in IRI group. TUNEL assay showed that the number of apoptotic cells in the oxymatrine group was less than that in the IRI group. FCM showed that hepatocyte apoptosis was significantly less in oxymatrine group than in IRI group (t=4.33, P<0.01); oxymatrine group had less hepatocytes in G0/G1 phase than IRI group, and hepatocytes in S phase increased significantly. (t=4.52, P<0.01). Conclusion Oxymatrine can significantly reduce the damage of rat hepatocytes during liver ischemia and reperfusion by protecting hepatocytes and promoting hepatocyte renewal.