受体相互作用蛋白3介导自身免疫性肝炎肝脏单核细胞来源巨噬细胞募集  

Receptor interaction protein 3 mediated the recruitment of hepatic monocytes/macrophages in autoimmune hepatitis

作　　者：刘曼[1] 张红霞[1] 周璐[1] 王邦茂[1] Liu Man;Zhang Hongxia;Zhou Lu;Wang Bangmao(Department of Gastroenterology and Hepatology,Tianjin Medical University General Hospital,Tianjin 300052,China)

机构地区：[1]天津医科大学总医院消化科,300052

出　　处：《中华消化杂志》2021年第1期35-42,共8页Chinese Journal of Digestion

基　　金：国家自然科学基金(81860109)。

摘　　要：目的探索受体相互作用蛋白3(RIP3)对自身免疫性肝炎(AIH)肝脏单核细胞来源巨噬细胞浸润的调控作用。方法纳入2018年1至6月于天津医科大学总医院消化科行肝穿刺活组织病理学检查的AIH患者10例,同期选择年龄和性别均匹配且无肝功能异常的5例肝囊肿患者作为对照,应用免疫荧光染色观察AIH患者和对照者肝组织单核细胞来源巨噬细胞浸润情况。将Raw264.7巨噬细胞分为对照组、脂多糖组、脂多糖+RIP3抑制剂GSK872(GSK872)组,采用定量聚合酶链反应(qPCR)检测巨噬细胞RIP3、混合谱系蛋白激酶样结构域(MLKL)、TNF-α、IL-6、IL-1β、细胞炎性小体Nod样蛋白3(NLRP3)、CC趋化因子配体(CCL)2和CCL5的mRNA水平;将Raw264.7巨噬细胞分为对照组、脂多糖组、脂多糖+地塞米松组,采用qPCR检测巨噬细胞TNF-α、NLRP3、RIP3和MLKL的mRNA水平。选择24只6周龄雌性C57BL/6小鼠建立急性AIH小鼠模型,并将其分为对照组、刀豆蛋白A(ConA)组、ConA+地塞米松组和ConA+GSK872组(每组6只),处死小鼠后收集外周血和肝组织,观察小鼠肝脏病理学表现,测定血清ALT和AST水平,采用qPCR检测CCL2和CC趋化因子受体2(CCR2)的mRNA水平,采用流式细胞术分析小鼠肝脏巨噬细胞比例。统计学方法采用独立样本t检验和单因素方差分析。结果AIH患者肝脏CD68阳性组织驻留巨噬细胞(库普弗细胞)和MAC387阳性单核细胞来源巨噬细胞比例均高于对照者[(0.84±0.21)%比(0.09±0.03)%、(0.79±0.13)%比(0.03±0.01)%],差异均有统计学意义(t=3.00、4.84,P均<0.05)。脂多糖组巨噬细胞内RIP3、MLKL、TNF-α、IL-6、IL-1β、NLRP3、CCL2、CCL5的mRNA水平均高于对照组和脂多糖+GSK872组(1.64±0.16比1.07±0.07和0.63±0.11,10.45±1.37比1.10±0.33和1.51±0.63,5.43±0.59比0.94±0.06和2.59±0.45,204.20±30.73比1.26±0.19和111.40±11.62,20848.00±362.00比1.09±0.26和10940.00±566.60,7.47±1.17比1.09±0.09和3.79±0.89,68.03±5.15比1.14±0.19�Objective To explore the role of receptor-interaction protein 3(RIP3)in regulating the infiltration of monocytes/macrophages into the liver in autoimmune hepatitis(AIH).Methods From January to June in 2018,at Department of Gastroenterology and Hepatology,Tianjin Medical University General Hospital,10 AIH patients who underwent liver biopsy were enrolled,and at the same time,5 age and gender matched individuals with normal liver function and hepatic cyst were selected as control.The infiltration of monocytes/macrophages in the liver tissues was observed by immunofluorescence detection in the patients with AIH and controls.Raw264.7 macrophages were divided into control group,lipopolysaccharide group and lipopolysaccharide+RIP3 inhibitor GSK872(GSK872)group.The expression of RIP3,mixed lineage kinase domain like pseudokinase(MLKL),tumor necrosis factor(TNF)-α,interleukin(IL)-6,IL-1β,nod-like receptor protein 3(NLRP3),CC motif chemokine ligand(CCL)2 and CCL5 at mRNA levels were detected by quantitative polymerase chain reaction(qPCR).Raw264.7 macrophages were also divided into control group,lipopolysaccharide group and lipopolysaccharide+dexamethasone group.The relative expression of TNF-α,NLRP3,RIP3 and MLKL at mRNA level in macrophage were detected by qPCR.Twenty-four 6-week-old female C57BL/6 mice were chosen to establish AIH mice model and were randomly divided into control group,concanavalin A(ConA)group,ConA+dexamethasone group and ConA+GSK872 group(6 mice in each group).After the mice were executed,the peripheral blood and liver tissues were collected.The histopathology of mice liver were observed and the serum alanine aminotransferase(ALT)and aspartate aminotransferase(AST)levels were measured.The expression of CCL2 and CC motif chemokine receptor 2(CCR2)at mRNA level were detected by qPCR.The proportion of macrophages in mice livers were analyzed by flow cytometry.The independent sample t test and one-way analysis of variance were performed for statistical analysis.Results The percentages of CD68 positive

关 键 词：肝炎 自身免疫性 巨噬细胞 受体相互作用蛋白3 细胞因子类 趋化因子 

分 类 号：R575.1[医药卫生—消化系统]