白细胞介素-10对抗增殖蛋白在小鼠肠纤维化中表达的影响

Prohibitin has anti-inflammatory and antifibrotic effects in the interleukin-10 knockout mice model of Crohn's Disease

目的探讨白细胞介素-10(IL-10)对抗增殖蛋白(Prohibitin)在小鼠肠纤维化进程中表达的影响。方法采用IL-10基因敲除(IL-10KO)小鼠作肠纤维化模型。将30只IL-10KO小鼠随机分组,分别建立肠纤维化模型组(模型组)和IL-10治疗组(治疗组),另取18只野生型小鼠作为阴性对照组(正常组)。造模后模型组和正常组分别于第12、14、16周各处死6只小鼠,治疗组于第12周开始腹腔内注射IL-10,3次/周,于第14、16周各处死6只。采用HE染色及Masson胶原三色染色观察小鼠结肠组织损伤和纤维化变化,采用实时荧光定量PCR法检测小鼠结肠组织中肠纤维化相关细胞因子如胶原蛋白Ⅰ、转化生长因子-β1(TGF-β1)、Prohibitin和NF-E2相关因子2(Nrf2)的mRNA的表达,采用定量免疫组化染色检测小鼠结肠组织中Prohibitin蛋白表达。结果 14、16周模型组、治疗组小鼠组织学损伤评分、胶原面积比均较显著高于正常组(P<0.05),16周治疗组小鼠组织学损伤评分显著低于模型组(P<0.05),14、16周治疗组小鼠胶原面积比显著低于模型组(P<0.05),16周治疗组小鼠胶原面积比较正常组无统计学差异(P>0.05)。12、14、16周模型组Prohibitin的mRNA和蛋白表达显著少于正常组(P<0.05),纤维化时间越长,表达量越低;模型组胶原蛋白Ⅰ、TGF-β1表达量均显著高于正常组(均P<0.05)。模型组小鼠结肠黏膜组织Prohibitin的蛋白表达与纤维化指数呈高度负相关(r=-0.6332,P<0.01)。结论 IL-10可通过提高Prohibitin表达量达到抑制肠纤维化的进展,从而在肠纤维化模型小鼠中起到抗炎及抗纤维化的作用,为IL-10治疗炎症性肠病肠纤维化的机制上提供了理论依据。

Objective To detect and investigate the effects of interleukin-10 （IL-10）on the expression of prohibitin in mice with intestinal fibrosis induced by interleukin-10 knockout（IL-10KO） mice.Methods thirty IL-10KO female mice（12-weeks-old） were randomly allocated into two groups：model group（18 IL-10KO mice）and IL-10 treatment group （12 IL-10KO mice）and another eighteen wild-type female mice （12-weeks-old）were used as normal group.The mice in model group and normal group received no treatment and sixth mice of each group were killed at 12,14 and 16 weeks.The mice in treatment were established by intraperitoneal injection of IL-10（5 μg/kg body weight,three times per week）and sixth mice of were killed at week 14 and 16 respective.Histopathological abnormalities of colon were calculated with HE staining and Masson＇s trichrome stain.The expressions of mRNA and protein of prohibitin were assayed by real time PCR and immunohistochemistry.The expressions of intestinal fibrosis related cytokines such as collagen Ⅰ,nuclear factor-erythroid 2-related factor （nrf2）,transforming growth factor （TGF）-β1 mRNA were determined by FQ-PCR.Results The group of IL-10KO mice had resulted in a marked decrease in their body weight as compared to the normal group at 12,14 and 16 weeks（P 〈 0.05）.Compared with normal group,the histologic colitis score and collagen area were increased significantly in model group at 12,14 and 16 weeks （P 〈 0.05）.In treatment group,the histopathological abdominal scores were decreased significantly than those in model group at week 16（P 〈0.05）,the collagen area were decreased significantly than those in model group at week 14,16（P 〈0.05）.The model group had higher increased level of mRNA of collagen Ⅰ and TGF-β1 （P 〈0.05）in a time-dependent manner.Moreover,the model group had significantly decreased expression of mRNA and protein of prohibitin compared with normal group（P 〈 0.05）and the course of disease for a longer period showed the lower expression.The expression of prohibitin and Nrf2 in treatment group was higher than model group at week 14and 16.Correlation analysis showed that the index of intestinal fibrosis was negatively correlated with prohibitin（r =-0.6332,P 〈 0.05）.Conclusions The lower expression of prohibitin in intestinal of IL-10KO mice suggest that it might play an important role in intestinal fibrosis.We hypothesized that IL-10 treatment is associated with increased prohibitin would decrease inflammation and fibrosisin in an animal model of Crohn＇s disease,and may be a potential target for intestinal fibrosis associated with inflammatory bowel disease.