摘要
目的 探讨人脐带间充质干细胞(hUC-MSC)对肝纤维化小鼠的治疗作用及其机制。方法 选取18只SPF级6周龄C57BL/6小鼠,使用随机数字表法随机分为3组,分别为对照组、CCl_(4)模型组(CCl_(4)组)和h UC-MSC治疗组(MSC组),每组6只。CCl_(4)组和MSC组腹腔注射CCl_(4)溶液构建小鼠肝纤维化模型,对照组同时注射相同剂量玉米油,MSC组在注射CCl_(4)的过程经尾静脉注射hUC-MSC。于第8周末采取小鼠血清,处死小鼠,取小鼠肝脏固定。使用酶联免疫吸附法测定炎症因子水平,自动生化检测仪检测肝功能相关指标,HE染色、Masson染色、天狼星红染色及α-SMA免疫荧光用于评估肝纤维化情况;TGF-β刺激的肝星状细胞(HSC)分别在有无几丁质酶3样蛋白1(CHI3L1)添加的培养基中与hUC-MSC共培养,Western Blot试验检测蛋白表达水平。计量资料多组间比较采用单因素方差分析,进一步两两比较采用Dunnett-t检验。结果 Masson染色、天狼星红染色提示CCl_(4)组小鼠纤维化较对照组明显(P值均<0.05),MSC组小鼠纤维化较CCl_(4)组减轻(P值均<0.05)。CCl_(4)组IL-1β、IL-6、AST、ALT和ALP水平较对照组明显升高(P值均<0.05),MSC组IL-6、AST、ALT及ALP水平较CCl_(4)组明显降低(P值均<0.05)。CCl_(4)组CHI3L1和α-SMA的表达均高于对照组和MSC组(P值均<0.05)。细胞培养实验显示,MSC+HSC组Bax表达高于HSC组和MSC+CHI3L1组(P值均<0.05),提示CHI3L1逆转了MSC对活化的HSC的促凋亡作用。结论 hUC-MSC治疗可以改善小鼠肝纤维化,其作用机制可能与抑制CHI3L1从而促进HSC的凋亡相关。
Objective To investigate the effect of human umbilical cord mesenchymal stem cells(hUCMSCs)in the treatment of mice with liver fibrosis and its mechanism.Methods A total of 18 specific pathogen-free C57BL/6 mice,aged 6 weeks,were selected and divided into control group(n=6),carbon tetrachloride(CCl_(4))model group(CCl_(4) group,n=6),and hUCMSCs treatment group(MSC group,n=6)using a random number table.The mice in the CCl_(4) group and the MSC group were given intraperitoneal injection of CCl_(4) solution to establish a mouse model of liver fibrosis,while those in the control group were injected with the same dose of corn oil,and the mice in the MSC group were injected with hUCMSCs via the caudal vein during the injection of CCl_(4).At the end of week 8,mouse serum was collected,and the mice were sacrificed to collect and fix the liver.Enzyme-linked immunosorbent assay was used to measure the levels of inflammatory factors;an automatic biochemical detector was used to measure liver function parameters;HE staining,Masson staining,Sirius Red staining,andα-SMA immunofluorescence assay were used to evaluate liver fibrosis.Hepatic stellate cells(HSCs)stimulated by TGF-βwere co-cultured with hUCMSCs in the medium with or without chitinase-3 like-protein-1(CHI3L1),and Western blot was used to measure the expression levels of proteins.A one-way analysis of variance was used for comparison of continuous data between multiple groups,and the Dunnett’s t-test was used for further comparison between two groups.Results Masson staining and Sirius Red staining showed that the CCl_(4) group had a significantly higher degree of fibrosis than the control group(both P<0.05),and the MSC group had significant alleviation of fibrosis compared with the CCl_(4) group(both P<0.05).Compared with the control group,the CCl_(4) group had significant increases in the levels of interleukin-1β,interleukin-6(IL-6),aspartate aminotransferase(AST),alanine aminotransferase(ALT),and alkaline phosphatase(ALP)(all P<0.05),and compared with the CCl_(4) group,the MSC group had significant reductions in the levels of IL-6,AST,ALT,and ALP(all P<0.05).The CCl_(4) group had significantly higher expression levels of CHI3L1 andα-SMA than the control group and the MSC group(all P<0.05).The cell culture experiment showed that the MSC+HSC group had a significantly higher expression level of Bax than the HSC group and the MSC+CHI3L1 group(both P<0.05),suggesting that CHI3L1 reversed the pro-apoptotic effect of MSC on activated HSCs.Conclusion This study shows that hUCMSCs can improve liver fibrosis in mice,possibly by inhibiting CHI3L1 to promote the apoptosis of HSCs.
作者
刘平箕
姚黎超
胡雪
王铮
熊芷玉
江应安
LIU Pingji;YAO Lichao;HU Xue;WANG Zheng;XIONG Zhiyu;JIANG Yingan(Department of Infectious Diseases,Renmin Hospital of Wuhan University,Wuhan 430060,China)
出处
《临床肝胆病杂志》
CAS
北大核心
2024年第3期527-532,共6页
Journal of Clinical Hepatology
基金
武汉大学教育发展基金会-抗衰老研究中心专款(2002330)
中央高校基本科研业务费专项资金(2042022kf1115)。
