抑制受体相互作用蛋白3改善实验性自身免疫性肝炎  

Inhibition of receptor-interacting protein 3 improves experimental autoimmune hepatitis

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

机构地区：[1]天津医科大学总医院消化科,天津300052

出　　处：《中华肝脏病杂志》2022年第11期1231-1236,共6页Chinese Journal of Hepatology

基　　金：国家自然科学基金(81860109)。

摘　　要：目的评估受体相互作用蛋白3(RIP3)作为自身免疫性肝炎(AIH)治疗靶点的潜力。方法使用免疫荧光法观察AIH肝组织和肝囊肿患者囊肿旁肝组织中RIP3及其下游信号混合谱系蛋白激酶样结构域(MLKL)的活化表达水平。经尾静脉注射刀豆蛋白A(ConA)诱导小鼠急性免疫性肝炎,并通过腹腔注射RIP3抑制剂GSK872或载体溶剂进行干预。收集外周血和肝组织,进行血清转氨酶水平测定、qPCR检测和流式细胞分析。组间比较采用独立样本t检验。结果p-RIP3和p-MLKL是RIP3及其下游信号MLKL磷酸化后的活性形式,其在AIH患者肝组织中的表达水平显著高于对照者。与对照组相比,AIH患者肝组织内RIP3和MLKL mRNA的表达水平也显著升高(相对表达量3.28±0.29对比0.98±0.09,4.55±0.51对比1.06±0.11),差异有统计学意义(t值分别为6.71、6.77,P值均<0.01)。ConA诱导的免疫性肝炎小鼠肝组织中RIP3和MLKL mRNA的表达水平也显著高于对照组(相对表达水平2.35±0.09对比0.89±0.11,2.77±0.22对比0.73±0.16,t值分别为10.4、6.33,P值均<0.01)。RIP3抑制剂GSK872显著减轻ConA诱导的免疫性肝损伤,抑制肝脏肿瘤坏死因子-α、白细胞介素(IL)-6、IL-1β和NLRP3的表达。与对照组相比,ConA+Vehicle组小鼠肝脏CD45+F4/80+巨噬细胞、CD4+IL-17+Th17细胞、CD4+CD25+调节性T(Treg)细胞和CD11b+Gr-1+骨髓来源抑制性细胞(MDSCs)比例均显著升高。与ConA+Vehicle组相比,ConA+GSK872组小鼠肝脏CD45+F4/80+巨噬细胞和CD4+IL-17+Th17细胞比例显著下降而具有免疫调节功能的CD4+CD25+Treg细胞和CD11b+Gr-1+MDSCs比例显著升高。结论AIH患者和ConA诱导的免疫性肝炎小鼠肝组织RIP3信号激活。抑制RIP3降低免疫性肝炎小鼠肝脏促炎因子的表达、减少炎性细胞比例、促进具有免疫调节功能的CD4+CD25+Treg细胞和CD11b+Gr-1+MDSCs在肝脏的积累,从而减轻肝脏炎症和损伤。因此,抑制RIP3有望成为治疗AIH的一种新方法。Objective To evaluate the potential of receptor-interacting protein 3(RIP3)as a therapeutic target for autoimmune hepatitis(AIH).Methods Immunofluorescence assay was used to observe the activated expression levels of RIP3 and its downstream signal mixed lineage protein kinase domain-like protein(MLKL)in the liver tissues of patients with AIH and hepatic cyst.Concanavalin A(ConA)was injected into the tail vein to induce acute immune-mediated hepatitis in mice.Intervention was performed by intraperitoneal injection of RIP3 inhibitor GSK872 or solvent carrier.Peripheral blood and liver tissues were collected.Serum transaminases level,qPCR and flow cytometry were analyzed.The intergroup comparison was performed with an independent sample t-test.Results The expression level of p-RIP3(the activated forms of RIP3)and phosphorylated p-MLKL(MLKL after phosphorylation)downstream signal were significantly higher in the liver tissue of AIH patients than those of controls.Compared with the control group,the expression levels of RIP3 and MLKL mRNA were significantly increased in the liver tissue of AIH patients(relative expression levels 3.28±0.29 vs.0.98±0.09,4.55±0.51 vs.1.06±0.11),and the differences were statistically significant(t=6.71 and 6.77,respectively,and P<0.01).The expression levels of RIP3 and MLKL mRNA were significantly higher in the mice liver tissue of ConA-induced immune hepatitis than those in the control group(relative expression levels 2.35±0.09 vs.0.89±0.11,2.77±0.22 vs.0.73±0.16,t=10.4,6.33,P<0.01).RIP3 inhibitor GSK872 had significantly attenuated ConA-induced immune liver injury and inhibited the expression of tumor necrosis factor-α,interleukin-6,interleukin-1βand NLRP3 in liver.Compared with the control group,the proportions of CD45+F4/80+macrophages,CD4+IL-17+Th17 cells,CD4+CD25+regulatory T(Treg)cells and CD11b+Gr-1+myeloid derived suppressor cells(MDSCs)were significantly increased in the liver of ConA+Vehicle group.Compared with ConA+Vehicle group,the proportion of CD45+F4/80+macrophag

关 键 词：自身免疫性肝炎 刀豆蛋白A 受体相互作用蛋白3 免疫细胞 细胞因子 

分 类 号：R575.1[医药卫生—消化系统]