Objective: To investigate the therapeutic effects and mechanisms of interleukin-10 (IL-10) gene-modified dendritic cells (DC-IL-10) in mice with liver fibrosis. Methods: DC-IL-10 was constructed in vitro, the phenotype and function of which were evaluated by flow cytometry. BALB/c mice were treated with intraperitoneal injection of carbon tetrachloride (CCl4) to establish liver fibrotic model. DC-IL-10 was administrated via tail vein. Animals were divided into 4 groups including normal dendritic cell(DC) control, liver fibrosis only, negative lentiviral transfection DC (DC-mock) and DC-IL-10. Liver function, cytokine secretion, T lymphocyte differentiation and liver histomorphology were tested. Real-time PCR and western blot were used to analyze the effect of DC-IL-10 on Wnt/β-catenin signaling pathway and its role in liver fibrosis. Results: When compared with DC control and DC-mock, the expression of DC-IL-10 surface stimulating molecules (major histocompatibity complex-Ⅱ, CD(80), CD(86)) were significantly decreased (F=14.708, 22.503, 12.595, respectively, all P<0.05), and DC-IL-10 significantly inhibited T lymphocyte proliferation (F=50.295, P<0.05). When compared with liver fibrosis group, serum alanine aminotransferase and aspartate transaminase were decreased in DC-IL-10 treated group (all P<0.05), other parameters including inflammatory factors (tumor necrosis factor α, IL-6, IL-1β) reduced (all P <0.05), the proportion of regulatory T cells (Treg) increased (F=6.742, P<0.05), pathological damage improved, the expression of Wnt3a, α-SMA and β-catenin mRNA and protein significantly reduced in DC-IL-10 treatment group (all P<0.001) . Conclusions: DC-IL-10 induces elevation of Treg for immune tolerance, as well as inhibition of inflammatory response, block of Wnt/β-catenin signaling pathway, which translates into improvement of liver fibrosis.
目的： 探讨白细胞介素（IL）10基因修饰的树突状细胞（DC）对肝纤维化小鼠的治疗作用及其机制。 方法： 体外构建IL-10基因修饰的DC（DC-IL-10），流式细胞术检测DC-IL-10表型及功能的变化。6~8周龄健康雄性BALB/c小鼠按随机数字表法随机分成空白对照组（DC组）、肝纤维化组、阴性对照慢病毒转染树突状细胞（DC-mock）处理组和DC-IL-10治疗组。比较各组小鼠肝功能炎症因子、免疫功能变化，以及肝脏组织形态学情况；实时PCR及免疫印迹法研究DC-IL-10对Wnt/β-连环蛋白信号通路的影响及在肝纤维化中的作用。 结果： 体外实验显示，组间表面分子（主要组织相容性复合物、CD(80)、CD(86)）表达差异有统计学意义（F值分别为14.708、22.503、12.595，P值均<0.05）；DC-IL-10组明显降低于DC组和DC-mock组（P值均<0.05）；T淋巴细胞增殖组间比较差异有统计学意义（F=50.295，P<0.05），DC-IL-10组T淋巴细胞增殖率明显低于DC组和DC-mock组（P值均<0.001）。体内实验显示，DC-IL-10治疗组丙氨酸转氨酶（ALT）和天冬氨酸转氨酶（AST）水平明显低于肝纤维化组（P值均<0.05）；肿瘤坏死因子（TNF）α、IL-6、IL-1β的表达明显低于肝纤维化组（P值均<0.05）；调节性T细胞（Treg）比例明显高于肝纤维化组（P<0.05）；病理显示肝组织损伤轻于肝纤维化组；Wnt3a、α-平滑肌肌动蛋白、β-连环蛋白mRNA表达量及蛋白表达量均明显低于肝纤维化组（P值均<0.001）。 结论： DC-IL-10诱导Treg产生免疫耐受，抑制过度的炎症反应，阻断Wnt/β-连环蛋白信号转导通路，进而抑制肝纤维化的进展。.
Keywords: Dendritic cells; Interleukin 10; Liver fibrosis; Regulatory T cells; Wnt/β-catenin signaling pathway.