目的:探讨突变型人肝细胞生长因子(HGF-(K132E,R134E),tvNK1)对四氯化碳(CCl4)诱导的SD大鼠肝纤维化的影响。方法:生物发酵大量制备tvNK1,并经腹腔注射于CCl4诱导的纤维化SD大鼠体内,6周后取肝脏组织,通过real-time PCR和Western blot检测tvNK1对纤维化SD大鼠肝脏中I型胶原蛋白(Collagen type I,Col1A1)、IV型胶原蛋白(Collagen type IV,Col4A1)和α-平滑肌肌动蛋白(α-smooth muscle actin,α-SMA)在mRNA和蛋白水平表达的影响,并进一步通过HE和Masson染色观察tvNK1对纤维化SD大鼠肝脏形态和肝组织胶原纤维的影响。结果:SDSPAGE检测结果显示,获得纯度≥95%的tvNK1。Real-time PCR和Western blot结果显示,tvNK1降低纤维化大鼠肝脏中Col1A1、Col4A1和α-SMA在mRNA和蛋白水平上的表达。HE和Masson染色结果显示,tvNK1减缓纤维化大鼠肝结构的病理性改变,并降低肝脏中胶原纤维含量。结论:tvNK1能抑制CCl4诱导的大鼠肝纤维化,为预防肝纤维化和其他器官纤维化疾病提供实验依据。
Objective:In order to determine the effects of human truncated variant of hepatocyte growth factor( tvNK1) on carbon tetrachloride( CCl4)-induced liver fibrosis in SD rat.Methods:The recombinant tvNK1 was high-level expression in Escherichia coli BL21 by bio-fermentation and rapid purification of tvNK1 protein using Ni^2+-affinity chromatograph.The SD rat model of CCl4-induced hepatic fibrosis was received intraperitoneal injection of tvNK1 for 6 weeks.The mRNA and protein expression levels of Col1A1、Col4A1 andα-SMA in hepatic tissue of different groups were assayed by real-time PCR and Western blot,respectively.The morphology and the collagen fiber of liver sections in different groups were determined by Hematoxylin and eosin( HE) and Masson staining,respectively.Results:The high purity tvNK1 was assayed by SDS-PAGE.The recombinant tvNK1 reduced the mRNA and protein expression levels of Col1A1、Col4A1 and α-SMA in CCl4-induced hepatic tissue in Rat.Moreover,injection with tvNK1 restored the morphology and the collagen fiber in CCl4-induced hepatic tissue in Rat.Conclusion:The recombinant tvNK1 protein attenuated progression of hepatic fibrosis in rat and provided a potential treatment for liver fibrosis and other fibrosis diseases.