肝纤维化大鼠肝细胞内质网形态及GRP78表达的研究

Morphology of endoplasmic reticulum and expression of GRP78 in rat fibrotic liver

目的:观察四氯化碳(CCl4)致大鼠肝纤维化过程中内质网形态及内质网应激标志性蛋白———葡萄糖调节蛋白78(GRP78)的表达变化,探讨内质网应激在肝纤维化发病机制中可能的作用。方法:雄性Wistar大鼠皮下注射CCl4制备肝纤维化模型,分别在4周及8周处死大鼠测定肝脏指数、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和肝细胞内质网形态,免疫组化和real-time PCR分别检测肝组织GRP78蛋白及mRNA表达变化。结果:肝纤维化组大鼠肝脏指数、血清ALT和AST活性显著高于正常对照组(P<0.01),肝纤维化明显,电镜下见肝细胞内质网扩张,数量明显减少;肝细胞胞浆中GRP78蛋白表达量及mRNA表达量较正常组显著增加(P<0.01)。结论:在CCl4诱导的肝纤维化发生过程中肝细胞内质网形态有明显损伤性变化,内质网应激蛋白GRP78蛋白及基因表达水平明显增加,提示内质网应激参与肝纤维化发生发展。

AIM： To observe the changes of endoplasmic reticulum and the biomarker of endoplasmic retidum stress（ERS）,glucose-regulated protein 78（GRP78）,in the process of liver fibrosis induced by carbon tetrachloride（CCl4）.METHODS： Male Wistar rats weighing 180 g to 220 g were divided into control group and liver fibrosis group.The rats in liver fibrosis group were induced by hypodermic injection of CCl4 for 4 weeks or 8 weeks.The liver index and the serumactivity of alanine aminotransferase（ALT） and aspartate aminotransferase（AST） were analyzed.The liver fibrosis and the morphological changes of endoplasmic reticulum were observed under light and electronic microscopes,respectively.Additionally,the expression of GRP78 at mRNA and protein levels was determined by real-time PCR and the method of immunohistochemistry,respectively.RESULTS： The liver index,serum ALT and AST activity in liver fibrosis group were obviously higher than those in control group.Swelling and reduced number of endoplasmic reticulum were observed in the hepatocytes of fibrotic rats compare to the controls.The levels of GRP78 protein and GRP78 mRNA in the liver of hepatic fibrotic rats were obviously higher than those in the control rats.CONCLUSION： In the process of liver fibrosis induced by CCl4,the obvious morphological changes of endoplasmic reticulum and increased expression of ERS protein indicate that ERS plays an important role in the liver fibrosis.