多黏菌素B对大鼠移植肝的保护作用

Protective effect of polymyxin B in liver transplantation in rats

目的探讨多黏菌素B（PMB）抗内毒素处理对大鼠移植肝的保护作用及相关机制。方法取SD大鼠，采用双套管法建立原位肝移植模型。采用随机数字表法将受鼠分为对照组（移植前肌注生理盐水0．5ml）和实验组（移植前肌注含1mg／ml PMB的生理盐水0．5ml）。肝移植术后，监测受鼠肝肾功能，采用动态浊度法检测门静脉血内毒素水平，采用酶联免疫吸附试验检测血清α肿瘤坏死因子（TNF-α）和白细胞介素6（IL-6）水平，采用聚合酶链反应和蛋白质印迹法检测移植肝组织CD14、Toll样受体4（TLR4）、核因子（NF）-κB、转录因子激活蛋白1（AP-1）等内毒素通路相关指标及热休克蛋白（Hsp）60的蛋白和mRNA水平，观察移植肝组织的病理学改变。结果再灌注后1h，实验组受鼠门静脉血内毒素水平较对照组明显下降；再灌注后6h，两组均接近正常水平（P〉0．05）。术后实验组丙氨酸转氨酶、TNF-α和IL-6的水平较对照组明显下降（P〈0．05），肝组织CD14和TLR4 mRNA的表达明显减少（P〈0．05），肝组织Hsp60 mRNA和蛋白及NF-κB、AP-1蛋白的表达降低（P〈0．05），肝组织病理损伤明显减轻。结论PMB可降低大鼠肝移植术后门静脉血内毒素水平，改善受者肝功能，减轻血清炎症反应，降低肝组织内毒素通路指标，减轻病理损伤。

Objective To investigate the protective effect of polymyxin B （PMB） to the liver graft after liver transplantation and the underlying mechanism in rats. Methods Male SD rats were selected as the donors and recipients. Non-artery whole liver transplantation model was established in rats according to Kamada＇s two-cuff method. The rats were divided into two groups by the way of random number table method： control group （normal saline, 0. 5 ml） and PMB group （PMB, 1 mg/ml, 0. 4 mg/kg＋ normal saline 0. 5 ml）. The levels of portal vein plasma endtotoxin （EU/ml） were determined by endotoxin-analyzing machine of BET-24A. ALT, BUN, and TNF-α, IL-6 in serum were measured by using machine of Automatic Analyzer and ELISA, respectively. The CD14, TLR4, NF-κB and AP-1 in the grafts were measured by RT-PCR and Western blotting, and pathological changes were observed. Results PMB decreased the levels of portal vein plasma endotoxin 1 h after reperfusion in PMB group as compared with control group （P〈0. 05）, and the levels of portal vein plasma endotoxin returned to the normal levels 6 h after reperfusion in both two groups （P〉0. 05）. After operation, the levels of ALT, TNF-α and IL-6 in serum were significantly reduced （P〈 0. 05）, the expression of CD14 and TLR4 mRNA in the grafts was significantly decreased （P〈0. 05）, the expression of Hsp60 protein and mRNA, and NF-κB and AP1 proteins in the grafts were reduced （P〈 0. 05）, and the pathological damage to the grafts was significantly alleviated in PMB group as compared with control group. Conclusion PMB reduced the levels of portal vein plasma endotoxin after reperfusion in liver transplantation in rats. PMB improved liver function, reduced the injury of inflammatory response, decreased the levels of endotoxin signal pathway markers and alleviated the pathological damage to the grafts.