摘要
目的研究外源Smad7基因对四氯化碳(CCl4)诱导的大鼠实验性肝纤维化的影响。方法将40只雄性SD大鼠随机分为4组。第一组为正常对照组,其余3组采用CCl4皮下注射法建立大鼠肝纤维化模型,并在实验第2周和第4周分别经尾静脉导入生理盐水、重组复制缺陷型腺病毒AdSmad7或对照病毒AdGFP。标准酶法测定各组大鼠肝功能情况;放射免疫法测定血清Ⅲ型前胶原(PCⅢ)、层粘连蛋白(LN)、Ⅳ型胶原(CⅣ)水平;RT-PCR法检测大鼠肝组织Smad7 mRNA表达。免疫组织化学法测定α-肌动蛋白(α-SMA)、Smad7表达;肝组织切片采用HE和VG染色进行肝纤维化程度分级。结果与模型对照组相比,AdSmad7导入组大鼠血清谷丙转氨酶(GPT)和碱性磷酸酶(AKP)水平有不同程度的降低,GPT降低更明显;血清PCⅢ和LN水平也显著下降(P<0.05)。RT-PCR和免疫组织化学染色证实,AdSmad7导入组大鼠肝组织Smad7表达显著上调,而α-SMA表达则明显下降(P<0.05)。病理学检查显示:AdSmad7导入组大鼠肝组织结缔组织增生程度减轻,纤维间隔变细,假小叶形成不明显,与模型对照组比较有显著性差异(P<0.05)。结论重组复制缺陷型腺病毒AdSmad7经尾静脉注射后可在大鼠肝组织中表达;外源Smad7基因可改善纤维化大鼠肝功能,降低血清PCⅢ水平,下调α-SMA表达,减轻大鼠肝纤维化程度。
Objective To investigate the effect of aline Smad7 gene on experimental liver fibrosis in rats.MethodsThe experimental liver fibrosis in SD rats was induced by subcutaneous injection of CCl4.Forty male SD rats were randomized to 4 groups.Group 1 was served as normal control group,and the rats in other three groups were given subcutaneous injection of 40% CCl4 at indicated time points.At the end of the second and fourth week,rats in group 2,3,4 were injected with saline,4×109 pfu AdGFP,4×109 pfu AdSmad7 via tail veins,respectively.In the end（CCl4 injection for 15 times）,all animals were sacrificed.Standard enzyme method was used to detect the liver function.Meanwhile,the radioimmuno-assay method was used to determine the serum levels of procollagen Ⅲ（PCⅢ）,laminin（LN） and collagen IV（CIV）.Expression of Smad7 mRNA was evaluated by RT-PCR.At the same time,tissue sections were stained with HE and VG methods and also subjected to immunohistostaining using antibodies against Smad7 and α-SMA.Results Compared with the model group,the serum levels of GPT,PCⅢ and LN in AdSmad7 group decreased conspicuously.RT-PCR indicated the expression of Smad7 mRNA in AdSmad7 group was much higher than that in other three groups.Hepatic pathology was milder in AdSmad7 group amd VG staining demonstrated that liver fibrosis degree in the Smad7 group was obviously milder than that in model group.Liver tissue immunohistostaining showed that there was a high expression of Smad7 in the AdSmad7 group.Meanwhile,the expression of α-SMA decreased significantly.Conclusion Smad7 gene delivery in vivo could improve liver functions,cut down the serum level of PCIII,reduce extracellular matrix deposition, up-regulate the expression of Smad7,down-regulate the expression of SMA, and thereby slow down the process of liver fibrosis.
出处
《胃肠病学和肝病学杂志》
CAS
2010年第9期783-786,共4页
Chinese Journal of Gastroenterology and Hepatology
基金
国家自然科学基金资助项目(30170412)
上海市科技发展基金项目(03ZR14050)
