盐酸小檗碱对肝纤维化大鼠肠道黏膜机械屏障的影响

Effect of berberine hydrochloride on mechanical barrier of intestinal mucosa in rats with liver fibrosis

目的探讨盐酸小檗碱对肝纤维化大鼠肠黏膜机械屏障的影响及其机制。方法将雄性SD大鼠60只随机分为正常对照组、模型组和盐酸小檗碱组,每组20只。模型组和盐酸小檗碱组采用40%CCl_(4)橄榄油溶液皮下注射建立大鼠肝纤维化模型,盐酸小檗碱组于模型建立第4周予盐酸小檗碱150 mg/(kg·d)灌胃干预动物。各组大鼠肝组织HE及Masson三色染色观察肝组织炎症活动度及纤维化程度;各组大鼠小肠组织HE染色观察黏膜上皮绒毛结构及排列情况;全自动生物化学检测仪测定各组大鼠血清丙氨酸氨基转移酶(ALT)、天冬氨酸氨基转移酶(AST),总胆红素(TBIL)及白蛋白(ALB)含量;分光光度法测定各组大鼠血二胺氧化酶(DAO)活性;酶联免疫法测定各组大鼠血清肿瘤坏死因子α(TNF-α);免疫组织化学法及实时荧光定量PCR检测各组大鼠小肠黏膜紧密连接蛋白Occludin蛋白及mRNA的表达。结果与正常对照组比较,模型组和盐酸小檗碱组肝组织G评分及S评分均显著升高(P<0.05);盐酸小檗碱组显著低于模型组(P<0.05)。与正常对照组比较,模型组和盐酸小檗碱组小肠黏膜结构不完整,肠绒毛排列不整齐;盐酸小檗碱组较模型组小肠黏膜结构完整,肠绒毛排列整齐。与正常对照组比较,模型组及盐酸小檗碱组血清ALT、AST、TBIL、DAO、TNF-α均升高(P<0.05);盐酸小檗碱组血清ALT、TBIL、DAO、TNF-α均低于模型组(P<0.05)。与正常对照组比较,模型组和盐酸小檗碱组血清ALB、小肠Occludin蛋白及mRNA的表达降低(P<0.05),盐酸小檗碱组高于模型组(P<0.05)。结论肝纤维化时肠黏膜机械屏障受损,盐酸小檗碱能减轻肝纤维化时肠黏膜机械屏障的损伤,其机制可能与降低TNF-α水平有关。

Objective To explore the effect and its mechanism of berberine hydrochloride on the mechanical barrier of intestinal mucosa in rats with liver fibrosis.Methods Sixty male SD rats were randomly divided into normal control group(n=20),model group(n=20)and berberine hydrochloride group(n=20).The liver fibrosis model was established by subcutaneous injection of 40%CCl_(4)in olive oil solution in model group and berberine hydrochloride group.The rats in berberine hydrochloride group were treated with berberine hydrochloride 150 mg/(kg·d)by gavage at week 4 after model establishment.Inflammatory activity and fibrosis degree of liver tissue were scored by HE and Masson staining in each group.The structure and the arrangement of mucosal epithelial villi were observed by HE staining in each group.The serum contents of alanine aminotransferase(ALT),aspartate aminotransferase(AST),total bilirubin(TBIL)and albumin(ALB)were measured by automatic biochemical assay in each group.Blood diamine oxidase(DAO)activity was measured by spectrophotometry in each group.Serum tumor necrosis factorα(TNF-α)was determined by enzyme-linked immunoassay in each group.Immunohistochemistry and real-time PCR were used to detect the protein and mRNA levels of Occludin in small intestinal mucosa of rats in each group.Results G score and S score of liver tissue in model group and berberine hydrochloride group were significantly increased compared with normal control group(P<0.05).G score and S score of liver tissue in berberine hydrochloride group were significantly lower than those in model group(P<0.05).The intestinal mucosal structure was not complete and the intestinal villi were not neatly arranged in model group and berberine hydrochloride group.The structure of the small intestinal mucosa was intact and the intestinal villi were arranged neatly in berberine hydrochloride group.The serum levels of ALT,AST,TBIL,DAO and TNF-αin model group and berberine hydrochloride group were higher than those in normal control group(P<0.05),and the serum levels of ALT,TBIL,DAO and TNF-αin berberine hydrochloride group were lower than those in model group(P<0.05).The serum ALB level and the expressions of Occludin protein and mRNA in the small intestine were lower in model group and berberine hydrochloride group than those in normal control group(P<0.05),and the levels of the above indexes in berberine hydrochloride group were higher than those in model group(P<0.05).Conclusion The intestinal mucosa mechanical barrier is damaged during the liver fibrosis.Berberine hydrochloride can alleviate the damage of intestinal mucosa mechanical barrier during the liver fibrosis,which may be related to the reduction of TNF-αlevel.