中性粒细胞弹性蛋白酶抑制剂sivelestat对非酒精性脂肪性肝炎的防治作用及其机制  

Sivelestat alleviates nonalcoholic steatohepatitis in mice through inhibiting activation of Kupffer cells

作　　者：臧淑妃[1] 马晓洁[1] 王磊[1] 朱葛丽 杨文君[1] 柳银兰[1] 严健[1] 罗燕[1] 庄振杰[1] 陈建玉[1] 荀运浩 施军平[1] 

机构地区：[1]杭州师范大学附属医院,310015 [2]杭州市西溪医院,310023

出　　处：《中华肝脏病杂志》2017年第5期371-376,共6页Chinese Journal of Hepatology

基　　金：国家自然科学基金项目（81570524）;浙江省自然科学基金项目（LY14H070004、LY15H070004、LQ15H070006）;杭州市科学技术委员会基金项目（20140633809）

摘　　要：目的研究中性粒细胞弹性蛋白酶抑制剂sivelestat对非酒精性脂肪性肝炎的防治作用及其机制。方法将4周龄雄性C57BL／6J ApoE基因敲除小鼠40只平均分为：（1）正常饮食组：正常饮食＋等渗盐水；（2）正常饮食＋干预组：正常饮食＋sivelestat；（3）模型组：高脂高胆固醇饮食＋等渗盐水；（4）模型＋干预组：高脂高胆固醇饮食＋sivelestat。连续喂养12周后取血和肝脏检测血清生物化学指标，用肝组织油红O染色评价脂肪变程度，用HE染色进行非酒精性脂肪性肝病活动度评分评价炎症反应。用荧光定量RT—PCR及免疫组织化学方法检测肝脏炎症因子及CD68和F4／80水平。4组间均数比较采用单因素方差分析，组问两两比较采用LSD或SNK法（方差齐）和Tamhane或Dunnett法（方差不齐）。结果高脂高胆固醇饮食12周能诱导ApoE基因敲除小鼠出现典型非酒精性脂肪性肝炎表现。sivelestat干预组空腹血糖、血胆固醇和肝生物化学指标较模型组明显降低，胰岛素敏感性增加，非酒精性脂肪性肝病活动度积分降低（5．71±1．11比3．16±1．16，P〈0．05），肿瘤坏死因子和单核细胞趋化蛋白-1也降低。sivelestat干预后CD68和F4／80的mRNA水平明显降低，免疫组织化学结果显示CD68在肝脏内表达减少。结论中性粒细胞弹性蛋白酶抑制剂sivelestat对非酒精性脂肪性肝炎的防治作用明确，减少枯否细胞活化可能是其防治非酒精性脂肪性肝炎的重要机制。Objective To investigate the role of neutrophil elastase inhibitor, sivelestat, in preventing and treating nonalcoholic steatohepatitis （NASH） and its underling mechanisms. Methods A total of forty 4-week-old male C57BL/6J ApoE-/- mice were equally divided into the following four groups： standard chow （SC）＋isotonic saline; SC＋sivelestat; high-fat, high-cholesterol （HFHC） diet＋isotonic saline; and HFHC＋sivelestat. These mice were treated with above methods for 12 weeks. Blood and liver tissue samples were collected to measure biochemical parameters, hepatic steatosis and non-alcoholic fatty liver disease （NAFLD） activity score （inflammation） were evaluated by oil red O staining and HE staining, respectively. The mRNA and protein expression levels of hepatic inflammatory cytokines, CD68, and F4/80 were determined by quantitative RT-PCR and immunohistochemistry, respectively. Comparison of means between the four groups was made by one-way analysis of variance, and comparison between any two groups was made by the LSD or SNK method （for data with homogeneity of variance） or the Tamhane or Dunnett method （for data with heterogeneity of variance）. Results Mice fed with an HFHC diet for 12 weeks developed typical pathological features of NASH compared with those fed with SC. Compared with mice fed with HFHC diet without sivelestat, those treated with HFHC and sivelestat exhibited the following features： （1） significantly reduced fast blood glucose, blood cholesterol, and hepatic biochemical parameters, as well as increased insulin sensitivity; （2） significantly reduced NAFLD activity score （5.71±1.11 vs 3.16±1.16, P 〈 0.05）; （3） reduced monocyte chemoattractant protein-1 and tumor necrosis factor -α; （4） significantly reduced mRNA levels of CD68 and F4/80; and （5） reduced expression of CD68 in the liver. Conclusion Sivelestat alleviates the hepatic steatosis and inflammation of NASH in mice by inhibiting the activation of Kupffer cells.

关 键 词：脂肪性肝炎 非酒精性 中性粒细胞 中性粒细胞弹性蛋白酶 

分 类 号：R575.1[医药卫生—消化系统]