IL-6／STAT3通路在冷缺血-再灌注损伤后胆管上皮细胞再生中的意义

Role of the activation of interleukin-6/STAT3 in cholangiocyte proliferation induced by cold ischemia reperfusion and injury

目的探讨白介素6／信号转导及转录激活因子3（IL-6／STAT3）通路在冷缺血-再灌注损伤诱导的胆管上皮细胞（BEC）再生过程中的意义。方法将大鼠随机分为4组：对照组、CP1h组、CP12h组（供肝分别冷保存1、12h后行原位肝移植术）和anti—IL-6组（CP12h组术前1h给予抗IL-6中和抗体0．5mg／kg，术后每日给予相同剂量直至观察结束）。术后1、3、7、14d，分别采用ELISA法、免疫组织化学法测定肝组织IL-6浓度及BEC再生情况；实时荧光定量PCR法、Westernblot法检测各组BEC内IL-6mRNA以及磷酸化STAT3（p-STAT3）、细胞周期蛋白（cyclin）D1表达水平，测定血清碱性磷酸酶（ALP）、谷氨酰转肽酶（GGT）浓度，并进行肝脏组织学检查。结果与对照组相比，CP1h组术后IL-6、p-STAT3及cyclinD1表达水平略增加，血清GGT、ALP仅有轻度、短暂的升高，BEC损伤及再生均不明显。CP12h组术后BEC损伤严重，血清GGT、ALP明显升高，至术后14d恢复正常，肝组织及BEC内IL-6含量增加、BEC内p-STAT3和cyclinD1蛋白表达上调，BEC再生明显。经anti—IL-6中和抗体处理后，CP12h组IL-6以及p-STAT3和cyclinD1表达降低，BEC再生明显抑制，术后14d仍可见细胞损伤及ALP、GGT的升高。结论IL-6／STAT3信号转导通路可能参与缺血-再灌注损伤后胆管上皮细胞的再生过程，对肝移植术后的胆道功能恢复有利。

Objective To investigate the role of IL-6/STAT3 pathway in the proliferation of cholangiocyte induced by cold ischemia and reperfusion injury. Methods Rats were randomized into CP 1 h and CP 12 h groups （supplied livers were preserved for 1 or 12 h） ,anti-IL-6R （rats in CP 12 h group were administrated with anti-rat soluble IL-6 receptor antibody）, and control group. At 1,3,7,14 d postoperative, IL-6 concentration in liver homogenate and cholangiocyte proliferation were detected by enzyme linked immunosorbent assay and histochemistry respectively. Expressions of IL-6 mRNA,phophorylated-STAT3 and cyclin D1 protein in cholangiocytes were determined by real-time PCR or Western blot analysis. Serum concentrations of ALP and GGT and histology analysis were also performed. Results Minimal expressions of IL-6 ,p-STAT3 and cyclin D1 were detected in CP 1 h group, with a slight cholangiocytes proliferation. Cholangiocytes responded to extended cold preservation with severe bile duct injures and marked increase in IL-6 secretion, p-STAT3 and cyclinD1 protein expression, followed by compensatory cholangiocytes regeneration. Parallel to this observation, biochemical index and morphology indicated that bile duct injury was recovery at 14 d postoperative. However, anti-sIL-6R inhibited cholangiocytes proliferation and reduced the expressions of IL-6, STAT3 and cyclin D, with the cellular injury and increase of serum ALP or GGT. Conclusions IL-6/STAT3 pathway might participate to initiate cholangiocytes regeneration after cold ischemia and preservation injury, which might benefit biliary recovery after liver transplantation.