环氧合酶-2在大鼠肝移植缺血再灌注损伤中对IP/TP的影响

Effect of Cyclooxygenase-2 on Prostaglandin I_2 Receptor/Thromboxane A_2 Receptor in Ischemia Reperfusion Injury after Liver Transplantation in Rats

目的探讨前列腺素I2受体(IP)、血栓素A2受体(TP)与环氧合酶-2(COX-2)在肝移植缺血再灌注损伤中的相互作用及其机理。方法雄性SD大鼠随机分为3组:正常对照组(n=16)、原位肝移植组(n=32)和尼美舒利(nimesulide)干预组(n=32),分别于手术后3、6、12和24 h取血液和肝组织标本备检。RT-PCR检测肝组织中IP、TP及COX-2 mRNA表达;免疫组化法检测COX-2在肝组织中的定位及表达;HE染色确定组织损伤程度;检测血清ALT和AST变化。结果免疫组化染色结果显示,原位肝移植组COX-2蛋白表达比正常对照组明显增强,主要分布于汇管区肝窦内皮细胞、肝细胞及巨噬细胞内,尼美舒利干预组COX-2蛋白表达较原位肝移植组明显减弱。原位肝移植组中IP mRNA、TP mRN及COX-2 mRNA表达水平均比正常对照组明显升高,IP/TP比值也明显升高(P<0.05)。尼美舒利干预组IP mRNA和TP mRNA表达水平(术后6及12 h)比原位肝移植组明显降低(P<0.05),IP/TP比值手术后3、6及24 h较原位肝移植组降低(P<0.05);COX-2 mRNA表达水平在术后6、12及24 h低于原位肝移植组(P<0.05)。HE染色见原位肝移植组肝损伤明显,尼美舒利干预组肝损害较原位肝移植组明显减轻;血清中各时点AST和3、6及12 h的ALT在原位肝移植组明显增高于其他2组(P<0.05),且均在再灌注后6 h达峰值。结论IP/TP的平衡在肝移植缺血再灌注损伤中具有重要作用,抑制COX-2的表达可通过改善IP/TP的失衡从而减轻肝移植缺血再灌注损伤。

Objective To study the interaction and mechanism of prostaglandin I2 （PGI2） receptor/thromboxane A2 （TxA2） receptor （IP/TP） and cyclooxygenase-2 （COX-2） in ischemia reperfusion injury after liver transplantation of rat. Methods Rats were randomly divided into three groups： control group （n=16）, orthotropic liver transplantation group （n=32） and nimesulide intervention group （n=32）. The samples were obtained at 3 h, 6 h, 12 h and 24 h after operation. The expressions of COX-2, IP and TP mRNA were detected by RT-PCR. Immunohistochemistry was used to detect the localization and expression of COX-2. Hematoxylin Eosin staining was used to classify the injury extent of liver. Serum ALT and AST levels were detected to evaluate the changes of liver enzyme. Results COX-2 protein expression detected by immunohistochemistry in orthotropic liver transplantation group mainly distributed in the district of liver sinusoidal endothelial cells, liver cells and macrophage cells, which was significantly higher than control group and nimesulide intervention group. Expressions of IP mRNA, TP mRNA and COX-2 mRNA in the ortbotropic liver transplantation group were significantly increased than those in control group （P〈0.05）, and the ratio of IP/TP increased （P〈0.05）. Expressions of IP mRNA and TP mRNA in nimesulide intervention group were significantly lower than that in the orthotropic liver transplantation group at 6 h and 12 h after operation （P〈0.05）, and the ratio of IP/TP decreased at 3 h, 6 h and 24 h after operation （P〈0.05）. The expression of COX-2 mRNA in nimesulide intervention group was significantly lower than that in the orthotropic liver transplantation group at 6 h, 12 h and 24 h after operation. In orthotropic liver transplantation group liver injury was obvious by HE staining, and more severve than that in nimesulide intervention group. Serum AST （each time） and ALT （3 h, 6 h and 12 h） levels in the orthotropic liver transplantation group were significantly higher than that in control group and nimesulide intervention group （P〈0.05） and peaked at 6 h after operation. Conclusion The balance of IP/TP takes part in and plays an important role in the ischemia reperfusion injury of liver transplantation. Changing imbalance of IP/TP may reduce liver transplantation ischemia reperfusion injury by inhibiting COX-2 expression.