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目的：探讨芪参汤对FXR/LKB1/NF-κB信号通路介导的树突状细胞成熟分化的影响，阐明其抑制肝星状细胞活化治疗肝纤维化的分子机制。方法：分离小鼠骨髓源性树突状细胞，采用脂多糖诱导构建成熟分化模型，并分别给予ω-鼠胆酸或芪参汤含药血清处理。通过流式细胞术检测各组树突状细胞CD40、CD80和MHC-II的表达水平，Western blot和RT-qPCR检测FXR/LKB1/NF-κB信号通路相关分子的表达。在此基础上，通过树突状细胞-初始CD4~+T细胞与初始CD4~+T细胞-肝星状细胞(JS1)共培养，采用流式细胞术检测分别检测各组初始CD4~+T细胞Treg分化和JS1细胞凋亡水平、酶联免疫吸附试验检测初始CD4~+T细胞上清中IL-10和TGF-β的含量、CCK8法及Western blot检测检测JS1细胞增殖活性和α-SMA蛋白的表达。结果：与模型组相比，药物干预显著抑制了树突状细胞CD40、CD80和MHC-II的表达，上调了树突状细胞中FXR和LKB1蛋白和mRNA的表达，降低了p-P65/total-P65的比值，增加了细胞质中P65蛋白的表达，抑制了细胞核中P65蛋白的分布，差异均具有统计学意义(P<0.01)。此外，细胞共培养结果表明，与模型组相比，ω-MCA、芪参汤和芪参汤联合ω-MCA组CD4~+T细胞Treg的百分比、IL-10和TGF-β的含量显著增加，JS1细胞凋亡水平显著增加，细胞活性和α-SMA蛋白的表达显著降低，差异均具有统计学意义(P<0.01)。结论：芪参汤通过抑制FXR/LKB1/NF-κB通路的激活，抑制树突状细胞的成熟分化，从而抑制肝星状细胞活化，达到治疗肝纤维化的目的。
Abstract：Objective: This study aims to investigate the effects of Qishen decoction on the FXR/LKB1/NF-κB signaling pathway-mediated maturation and differentiation of dendritic cells and to elucidate its molecular mechanism in inhibiting the activation of hepatic stellate cells for the treatment of liver fibrosis. Methods: Mouse bone marrow-derived dendritic cells were isolated, and a maturation model was constructed using lipopolysaccharide. The cells were treated with either ω-muricholic acid (ω-MCA) or Qishen decoction-containing serum. Flow cytometry was employed to detect the expression levels of CD40, CD80, and MHC-II on dendritic cells. Western blot and RT-qPCR were used to analyze the expression of FXR/LKB1/NF-κB signaling pathway related molecules. Furthermore, co-culture experiments were conducted between dendritic cells and naive CD4+ T cells, as well as between naive CD4+ T cells and hepatic stellate cells(JS1). Flow cytometry was used to assess the differentiation of naive CD4+ T cells into regulatory T cells(Tregs) and the apoptosis of JS1 cells. Enzyme-linked immunosorbent assays(ELISA) were performed to measure the levels of interleukin-10(IL-10) and transforming growth factor-β(TGF-β) in the supernatants of naive CD4+ T cells. CCK-8 assays and Western blot were used to evaluate the proliferation activity and α-smooth muscle actin(α-SMA) protein expression of JS1 cells.Results: Compared to the model group, drug intervention significantly inhibited the expression of CD40, CD80, and MHC-II, upregulated the expression of FXR and LKB1 proteins and mRNA, reduced the ratio of phosphorylated P65 to total P65, increased the expression of P65 protein in the cytoplasm, and inhibited the distribution of P65 protein in the nucleus in dendritic cells, with all differences being statistically significant(P<0.01). Additionally, the co-culture results showed that compared to the model group, the percentage of Tregs, IL-10, and TGF-β levels in the ω-MCA, Qishen, and Qishen decoction combined with ω-MCA groups were significantly increased, while the apoptosis of JS1 cells, cell viability, and α-SMA protein expression were significantly reduced, with all differences being statistically significant(P<0.01). Conclusion: Qishen decoction inhibits the activation of the FXR/LKB1/NF-κB signaling pathway, thereby suppressing the maturation of dendritic cells, which in turn inhibits the activation of hepatic stellate cells and ameliorates liver fibrosis. This study provides new insights and potential therapeutic strategies for the treatment of liver fibrosis.
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基本信息:
DOI：10.13210/j.cnki.jhmu.20250609.002
中图分类号:R285.5
引用信息:
[1]杨柳欣,高婷婷,王炳予,等.基于FXR/LKB1/NF-κB信号通路介导的树突状细胞成熟分化探讨芪参汤抑制肝纤维化的机制研究[J].海南医科大学学报,2025,31(14):1076-1084+1093.DOI:10.13210/j.cnki.jhmu.20250609.002.
基金信息:
黑龙江省自然科学基金优秀青年项目（YQ2022H015）~~
2025-06-10
2025-06-10
2025-06-10