基于Wnt/β-catenin信号通路研究扶肝化纤汤抑制HSC-T6活化的作用

doi:10.6040/j.issn.1671-7554.0.2023.0283

摘要/Abstract

摘要： 目的探讨扶肝化纤汤抑制大鼠肝星状细胞系HSC-T6增殖的作用机制。方法健康清洁级雄性SD大鼠40只,予扶肝化纤汤等效量灌胃,制备含药血清及正常血清。CCK8法检测不同含药血清浓度(5%、8%、10%、15%、20%)对HSC-T6增殖率的影响,选择最佳作用浓度。将HSC-T6分为空白组(15%胎牛血清)、模型组［转化生长因子-β1(TGF-β1)+15%正常血清)］和治疗组(TGF-β1+15%含药血清),Western blotting、qt-PCR 检测Wnt1、β-catenin、Cyclin D1蛋白和mRNA表达,细胞免疫荧光染色法检测α-SMA表达。结果镜下观察治疗组HSC-T6细胞数量减少,形态变圆,体积缩小,悬浮细胞增多。CCK8法检测结果显示,扶肝化纤汤含药血清抑制TGF-β1诱导活化的HSC-T6细胞增殖(P<0.01),其中15%、20%浓度效果最佳,但15%与20%两组间比较差异无统计学意义(P>0.05)。Western blotting、qt-PCR 检测结果显示,与模型组比较,治疗组Wnt1、β-catenin、Cyclin D1的mRNA和蛋白表达下降(P<0.01)。细胞免疫荧光染色法结果显示,模型组胞质内α-SMA表达增加,治疗组与模型组比较胞质内α-SMA表达减少。结论扶肝化纤汤含药血清可抑制HSC-T6活化增殖,降低活化后HSC-T6中Wnt1、β-catenin、Cyclin D1的蛋白及mRNA表达;扶肝化纤汤可能通过抑制Wnt/β-catenin信号通路的激活从而防治HF。

关键词: 扶肝化纤汤, 肝纤维化, 肝星状细胞, Wnt1蛋白, β-catenin蛋白

Abstract: Objective To explore the inhibitory effect of Fugan Huaqian Decoction on hepatic stellate cell line HSC-T6. Methods A total of 40 healthy and clean male SD rats were given equivalent amount of Fugan Huaxian Decoction by gavage to prepare drug-containing serum and normal serum. The effects of different drug-containing serum concentrations(5%, 8%, 10%, 15%, 20%)on the proliferation rate of HSC-T6 cells were detected with CCK8 assay to determine the optimal acting concentration. HSC-T6 cells were divided into blank group(15% fetal bovine serum), model group ［transforming growth factor-β1(TGF-β1)+15% normal serum］ and treatment group(TGF-β1+15% medicated serum). Protein and mRNA expressions of Wnt1, β-catenin and Cyclin D1 were detected with Western blotting and q-PCR, respectively. Expression of α-SMA was determined with immunofluorescence staining. Results In the treatment group, the number of cells decreased, the morphology turned round, the volume shrank, and the number of suspension cells increased. CCK8 assay showed that drug-containing serum inhibited TGF-β1-induced proliferation of HSC-T6 cells(P<0.01), and the best effect was observed at 15% and 20% concentration, but there was no significant difference between the two groups(P>0.05). Western blotting and q-PCR showed that compared with the model group, the treatment group had decreased mRNA and protein expressions of Wnt1, β-catenin and Cyclin D1(P<0.01). Immunofluorescence staining showed that the cytoplasmic expression of α-SMA increased in the model group, but decreased in the treatment group. Conclusion Serum containing Fugan Huaqian Decoction can inhibit HSC-T6 activation and proliferation, and decrease the protein and mRNA expressions of Wnt1, β-catenin and Cyclin D1 in activated HSC-T6 cells. Fugan Huaqian Decoction may prevent and treat hepatic fibrosis by inhibiting the activation of Wnt/β-catenin signaling pathway.

Key words: Fugan Huaqian Decoction, Hepatic fibrosis, Hepatic stellate cells, Wnt1, β-catenin

中图分类号:

R575

肖政华,石以石则,杨庆万,李婷婷,李珊珊,钟燕. 基于Wnt/β-catenin信号通路研究扶肝化纤汤抑制HSC-T6活化的作用[J]. 山东大学学报 (医学版), 2023, 61(6): 41-46.

XIAO Zhenghua, SHIYI Shize, YANG Qingwan, LI Tingting, LI Shanshan, ZHONG Yan. Mechanism of Fugan Huaqian Decoction inhibiting HSC-T6 activation via Wnt/β-catenin signaling pathway[J]. Journal of Shandong University (Health Sciences), 2023, 61(6): 41-46.

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