| 引用本文: |
肖雪飞, 刘石, 符艳, 游柏稳.黄芪甲苷通过调控Wnt/β-catenin信号通路对人成纤维细胞凋亡和肌成纤维细胞转化的作用[J].湖南中医药大学学报,2024,44(12):2183-2192[点击复制] |
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| 黄芪甲苷通过调控Wnt/β-catenin信号通路对人成纤维细胞凋亡和肌成纤维细胞转化的作用 |
| 肖雪飞,刘石,符艳,游柏稳 |
| (湖南中医药大学第一附属医院, 湖南 长沙 410007;湖南中医药大学第二附属医院, 湖南 长沙 410005) |
| 摘要: |
| 目的 利用转化生长因子β1（transforming growth factor-β1,TGF-β1）刺激人成纤维细胞HFL-1模拟特发性肺纤维化的病理过程，探讨黄芪甲苷（astragaloside-Ⅳ,AS-Ⅳ）对人成纤维细胞凋亡和肌成纤维细胞转化的作用和机制。方法 采用10 ng/mL TGF-β1刺激HFL-1细胞，给予不同浓度（0、1.25、2.5、5、10 μmol/L）的AS-Ⅳ溶液干预24 h后，利用RT-PCR检测纤连蛋白（fibronectin,FN）mRNA表达量。随后细胞分为空白组、TGF-β1组、TGF-β1+AS-Ⅳ组、TGF-β1+XAV939组。采用流式细胞术检测细胞凋亡率；采用RT-PCR法检测α-平滑肌肌动蛋白（α-smooth muscle actin,α-SMA）、FN、重组人胶原蛋白-1（collagenⅠ）、B淋巴细胞瘤2（B-cell lymphoma 2,Bcl-2）及Bcl-2相关X蛋白（Bcl-2 associated X protein,Bax） mRNA表达情况；采用免疫荧光法检测α-SMA、上皮细胞钙黏蛋白（E-Cadherin）、collagenⅠ、FN、β-联蛋白（β-catenin）及半胱氨酸蛋白酶蛋白-3（Cleaved Caspase-3）的蛋白表达；Western blot检测FN、β-catenin、Cleaved Caspase-3蛋白表达水平。结果 与空白组比较，TGF-β1组细胞凋亡率上升（P<0.05），FN、collagenⅠ、α-SMA和Bcl-2 mRNA表达增加（P<0.05），Bax mRNA表达减少（P<0.05），α-SMA、collagenⅠ、FN、Cleaved Caspase-3及β-catenin蛋白表达增加（P<0.05）；与TGF-β1组比较，TGF-β1+AS-Ⅳ组和TGF-β1+XAV939组细胞凋亡率下降（P<0.05），FN、collagenⅠ、α-SMA和Bcl-2 mRNA表达减少（P<0.05），Bax mRNA表达增加（P<0.05），α-SMA、collagenⅠ、FN、Cleaved Caspase-3及β-catenin蛋白表达减少（P<0.05）；与TGF-β1+XAV939组相比较，TGF-β1+AS-Ⅳ组细胞凋亡率下降（P<0.05），FN、collagenⅠ、Bcl-2和α-SMA mRNA表达量降低（P<0.05），Bax mRNA表达量增加（P<0.05）。结论 AS-Ⅳ可抑制TGF-β1诱导的HFL-1细胞凋亡和肌成纤维细胞转化，该作用可能与Wnt/β-catenin信号通路有关。 |
| 关键词: 黄芪甲苷 特发性肺纤维化 成纤维细胞 肌成纤维细胞转化 Wnt β-联蛋白 凋亡 |
| DOI：10.3969/j.issn.1674-070X.2024.12.006 |
| 投稿时间：2024-06-09 |
| 基金项目:湖南省中医药科研计划项目（2021165）。 |
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| Effects of astragaloside-Ⅳ on human fibroblast apoptosis and myofibroblast transformation by regulating Wnt/β-catenin signaling pathway |
| XIAO Xuefei, LIU Shi, FU Yan, YOU Baiwen |
| (The First Hospital of Hunan University of Chinese Medicine, Changsha, Hunan 410007, China;The Second Hospital of Hunan University of Chinese Medicine, Changsha, Hunan 410005, China) |
| Abstract: |
| Objective To simulate the pathological process of idiopathic pulmonary fibrosis (IPF) by stimulating human fibroblast (HFL-1) cells with transforming growth factor-β1 (TGF-β1), and to explore the effects and mechanism of astragaloside-Ⅳ (AS-Ⅳ) on human fibroblast apoptosis and myofibroblast transformation. Methods HFL-1 cells were stimulated with 10 ng/mL TGF-β1 and then treated with different concentrations (0, 1.25, 2.5, 5, 10 μmol/L) of AS-Ⅳ solution for 24 h. The mRNA expression of fibronectin (FN) was determined by RT-PCR. Subsequently, the cells were divided into blank group, TGF-β1 group, TGF-β1+AS-Ⅳ group, and TGF-β1+XAV939 group. Flow cytometry was used to measure cell apoptosis rate, RT-PCR to examine the mRNA expressions of α-smooth muscle actin (α-SMA), FN, recombinant human collagen-1 (collagen Ⅰ), B-cell lymphoma 2 (Bcl-2), and Bcl-2 associated X protein (Bax), immunofluorescence to determine the protein expressions of α-SMA, E-Cadherin, collagen Ⅰ, FN, β-catenin, and Cleaved Caspase-3, and Western blot to check the protein expression levels of FN, β-catenin, and Cleaved Caspase-3. Results The optimal concentration of AS-Ⅳ for inhibiting FN expression was 5 μmol/L. Compared with the blank group, the cell apoptosis rate in the TGF-β1 group increased (P<0.05), the mRNA expressions of FN, collagen Ⅰ, α-SMA, and Bcl-2 were elevated (P<0.05), while Bax mRNA expression decreased (P<0.05). Additionally, the protein expressions of α-SMA, collagen Ⅰ, FN, Cleaved Caspase-3, and β-catenin also increased (P<0.05). Compared with the TGF-β1 group, both the TGF-β1+AS-Ⅳ group and TGF-β1+XAV939 group showed decreased cell apoptosis rates (P<0.05), reduced mRNA expressions of FN, collagen Ⅰ, α-SMA, and Bcl-2 (P<0.05), increased Bax mRNA expression (P<0.05), and decreased protein expressions of α-SMA, collagen Ⅰ, FN, Cleaved Caspase-3, and β-catenin (P<0.05). Compared with the TGF-β1+XAV939 group, the cell apoptosis rate in the TGF-β1+AS-Ⅳ group decreased (P<0.05), the mRNA expressions of FN, collagen Ⅰ, Bcl-2, and α-SMA decreased (P<0.05), while that of Bax increased (P<0.05). Conclusion AS-Ⅳ can inhibit TGF-β1-induced HFL-1 cell apoptosis and myofibroblast transformation, which may be related to the Wnt/β-catenin signaling pathway. |
| Key words: astragaloside-Ⅳ idiopathic pulmonary fibrosis fibroblast myofibroblast transformation Wnt β-catenin apoptosis |
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