肝纤维化病理过程中ZEB1和ZEB2的动态表达变化及意义

Dynamic changes of ZEB1 and ZEB2 during development of liver fibrosis induced by chronic liver injury in rats

目的:观察慢性肝损伤致肝纤维化过程中上皮-间质转化(EMT)调节蛋白锌指E盒结合同源盒蛋白(ZEB)1和ZEB2的动态表达变化并探讨其调节机制。方法:雄性Wistar大鼠40只,随机分为正常对照组、2周、4周、6周和8周模型组,每组各8只,模型组按3 mL/kg体重的剂量皮下注射60%CCl4,每隔3 d注射1次,处死大鼠后测定肝脏指数、血清丙氨酸氨基转移酶(ALT)和天冬氨酸氨基转移酶(AST)活性,观察肝组织病理改变;免疫组织化学法检测肝组织中ZEB1、ZEB2、E-钙黏蛋白(E-cadherin)和α-平滑肌肌动蛋白(α-SMA)的表达情况;real-time RT-PCR方法检测肝脏组织中ZEB1及ZEB2 mRNA的表达变化。结果:模型各组大鼠肝脏指数、血清ALT和AST活性显著高于正常对照组(P<0.01),8周模型组肝纤维化明显;随着肝脏损伤逐渐加重,纤维化程度加深,E-cadherin蛋白的表达显著降低,α-SMA蛋白表达显著升高,ZEB1和ZEB2蛋白和mRNA表达量也逐渐增加,8周模型组肝组织中ZEB1和ZEB2蛋白(46.42±14.36和57.71±13.32)与mRNA(189.00±47.39和277.28±48.55)表达较正常组显著增加(P<0.01)。结论:ZEB1和ZEB2蛋白及mRNA表达量随纤维化程度的加重而增加,提示EMT可能通过ZEB1和ZEB2参与肝纤维化的发生发展。

AIM：To observe the dynamic changes of zinc finger E-box binding homeobox 1（ZEB1） and zinc finger E-box binding homeobox 2（ZEB2） during the development of liver fibrosis induced by chronic liver injury in rats. METHODS：Forty male Wistar rats weighing 180~220 g were divided intonormal control group and 4 model groups. The rats in model groups were induced by hypodermic injection of CCl4 at a dose of 3 mL/kg and once per 3 days for 2 weeks, 4 weeks, 6 weeks and 8 weeks, respectively. The liver index and serum levels of alanine aminotransferase（ALT） and aspartate aminotransferase（AST） were analyzed. The liver fibrosis was observed under microscope. Additionally, the expression of ZEB1, ZEB2, E-cadherin and α-smooth muscle actin（α-SMA） SMA at protein level was determined by immunohistochemistry. The mRNA expression of ZEB1 and ZEB2 was detected by real-time RT-PCR. RESULTS：The liver index and the serum levels of ALT and AST in the 4 model groups were obviously higher than those in normal control group. The apparent liver fibrosis was observed in 8-week model group. The protein levels of α-SMA（37.84±7.04）, ZEB1（46.42±14.36） and ZEB2（57.71±13.32）, and mRNA expression of ZEB1（189.00±47.39） and ZEB2（277.28±48.55） in the livers of 8-week model rats were obviously higher than those in the control rats（P〈0.01）. The protein level of E-cadherin in the liver of 8-week model rats was obviously lower than that in the control rats（P〈0.01）. CONCLUSION：In the process of liver fibrosis induced by CCl4, the obvious changes of ZEB1/ZEB2 may play an important role in chronic liver injury and liver fibrosis.