大鼠肢体缺血/再灌注后肝脏iNOS表达的变化及其意义

CHANGES OF iNOS GENES EXPRESSION IN LIVER FOLLOWING ISCHEMIA AND REPERFUSION OF LIMBS AND THEIR SIGNIFICANCE IN RATS

目的 :探讨肢体缺血 /再灌注 (I/R)致肝损伤时肝组织iNOS表达的变化及其意义。方法 :夹闭大鼠双侧股动脉根部 4h、开放 2～ 2 4h ,制备肢体I/R模型。RT PCR检测肝组织iNOSmRNA表达的改变 ,免疫组化染色法观察iNOS蛋白及过氧亚硝基阴离子 (ONOO-)的生成与分布 ,比色法测定肝组织MDA含量及SOD活性 ;对肢体I R大鼠用氨基胍抑制iNOS活性后 ,观察其肝组织的病理学变化。结果 :肢体I R后 ,肝组织iNOSmRNA的表达水平较对照组显著上调 (P <0 .0 5 ) ,肝组织内出现大量iNOS及ONOO-阳性肝细胞 ,肝组织MDA含量升高及SOD活性降低均与对照组有显著性差异 (P <0 .0 1)。应用氨基胍抑制iNOS活性 ,使肢体I R所致肝组织病变减轻。结论 :肢体I/R后 ,肝组织iNOSmRNA及蛋白表达显著上调 ,所诱生的高浓度NO参与介导了肢体I/R引发的肝脏损伤。

Aim: To detect the changes of inducible nitric oxide synthase (iNOS) expression in liver following ischemia/reperfusion(I/R) of hindlimbs and to elucidate their significance. Methods: I/R was established using the occlusion of the femoral arteries for 4 h and re-opening for 2～24 h in rats. The expression of iNOS mRNA, and iNOS protein and the nitrotyrosine (NT), a marker of peroxynitrite (ONOO-), in liver tissue were detected with reverse transcription-polymerase chain reaction (RT-PCR) and immunohistochemical technique, respectively. The liver superoxide dismutase (SOD) activity and malondialdehyde (MDA) contents were spectraphoto-metrically measured. The observation of pathologic changes of liver was made following the inhibition of iNOS by aminoguanidine (AG). Results: Compared with control groups, the relative expression level of iNOS mRNA significantly increased in I/R group. There were more iNOS positive hepatocytes and more NT positive hepatocytes in I/R group than control groups. The contents of MDA markedly increased, while the activity of SOD significantly decreased in I/R group, compared with those in the control groups. The pathologic changes of rat liver became milder in I/R group following the inhibition of iNOS by AG. Conclusion: The expressions of iNOS mRNA and protein in liver are significantly upregulated, excess induction of iNOS-NO is contributed to the liver injury du- ring the I/R of hindlimbs.