CX3CR1沉默介导的巨噬细胞极化减轻CCl_(4)诱导的小鼠肝纤维化  

CX3CR1 silencing-mediated macrophage polarization attenuates CCl_(4)-induced liver fibrosis in mice

作　　者：黄莎莎 邹艳丽 谭洁[1] 刘蒙[1] 陈巍[1] HUANG Shasha;ZOU Yanli;TAN Jie;LIU Meng;CHEN Wei(Department of Gastroenterology,Tongren Hospital Affiliated to Wuhan University(Wuhan Third Hospital),Wuhan,Hubei Province,430074,China)

机构地区：[1]武汉大学附属同仁医院(武汉市第三医院)消化内科,武汉430074

出　　处：《陆军军医大学学报》2022年第13期1314-1321,共8页Journal of Army Medical University

基　　金：武汉市卫生和计划生育委员会科研项目(WX18Q12)。

摘　　要：目的探讨CX3CR1在CCl_(4)诱导的小鼠肝纤维化中的作用。方法将小鼠分为正常组(n=24)、CCl_(4)组(n=36)、CCl_(4)+sh-NC组(n=36)、CCl_(4)+sh-CX3CR1组(n=36)。除正常组外,其余组通过腹腔注射CCl_(4)诱导肝纤维化。从CCl_(4)诱导的肝纤维化4周开始,每周分别向CCl_(4)+sh-NC组和CCl_(4)+sh-CX3CR1组小鼠尾静脉注射sh-NC或sh-CX3CR1,持续2周。通过免疫组织化学确定小鼠肝脏组织中CX3CR1的表达情况,免疫荧光染色分析肝脏组织中M1-巨噬细胞(F4/80^(+)iNOS^(+))或M2-巨噬细胞(F4/80^(+)CD206^(+))表达。ELISA试剂盒法检测小鼠血清IL-1β、IL-6和TGF-β1水平。使用流式细胞仪测定原代肝脏巨噬细胞与活化HSC共培养后巨噬细胞M1和M2表型变化。结果在健康的非纤维化对照肝脏中几乎检测不到CX3CR1,而在CCl_(4)处理4周和6周后,CX3CR1强烈上调。通过沉默CCl_(4)小鼠CX3CR1表达,小鼠肝炎症细胞募集和纤维组织沉积化的严重程度均明显减轻,并且α-SMA、N-cadherin和Vimentin的蛋白水平降低,E-cadherin的蛋白表达增加。与CCl_(4)+sh-NC组相比,CCl_(4)+sh-CX3CR1组小鼠血清IL-1β、IL-6和TGF-β1水平均下调4.6~8.1倍(P<0.01),且M2-巨噬细胞的百分率和M2/M1比值均显著增加(P<0.01)。在用CX3CR1沉默的小鼠肝星状细胞和巨噬细胞的共培养中,高M2/M1-巨噬细胞比值抑制胶原的产生。结论CX3CR1沉默通过促进M1向M2-巨噬细胞的极化发挥抗肝纤维化作用。ObjectiveTo investigate the role of CX3CR1 in liver fibrosis induced by CCl_(4) in mice.MethodsThe mice were divided into normal group(n=24),CCl_(4) group(n=36),CCl_(4)+sh-NC group(n=36),and CCl_(4)+sh-CX3CR1 group(n=36),and all the groups were induced liver fibrosis by intraperitoneal injection of CCl_(4) except the normal group.From the 4th week of CCl_(4) injection,the mice of CCl_(4)+sh-NC group and CCl_(4)+sh-CX3CR1 group were respectively injected with shRNA or shRNA-CX3CR1 by tail vein weekly,for 2 weeks.The expression of CX3CR1 in the liver tissues was determined by immunohistochemistry,and the number of M1 macrophages(F4/80^(+)iNOS^(+))or M2 macrophages(F4/80^(+)CD206^(+))in the liver tissues was measured by immunofluorescence staining.ELISA was used to detect the serum levels of IL-1β,IL-6 and TGF-β1 in mice,and flow cytometry was adopted to analyze the phenotypic changes of M1 and M2 macrophages after co-culture of primary liver macrophages with activated hepatic stellate cells(HSC).ResultsCX3CR1 was hardly detected in healthy non-fibrotic control livers,but strongly up-regulated 4~6 weeks after CCl_(4) treatment.By silencing CX3CR1 expression in CCl_(4)-induced liver fibrosis mice,the severity of inflammatory cell recruitment and fibrous tissue deposition in liver tissues was significantly reduced,and the protein levels ofα-SMA,N-cadherin and Vimentin were decreased,while the expression of E-cadherin increased.As compared with the CCl_(4)+sh-NC group,the serum levels of IL-1β,IL-6 and TGF-β1 were down-regulated by 4.6-8.1 times in the CCl_(4)+sh-CX3CR1 group(P<0.01),and the percentage of M2-macrophages as well as the M2/M1 ratio were remarkably elevated(P<0.01).In the co-cultures of HSC and macrophages extracted from CX3CR1 silencing mice,collagen production was inhibited due to the high M2/M1 ratio.ConclusionCX3CR1 silencing plays an anti-hepatic fibrosis role by promoting polarization of M1 to M2 macrophage.

关 键 词：CX3CR1 巨噬细胞极化 小鼠 肝纤维化 

分 类 号：R322.47[医药卫生—人体解剖和组织胚胎学] R329.24[医药卫生—基础医学] R392.2