| 引用本文: |
林湘东,向茗,陈新宇.温阳振衰颗粒对TGF-β1诱导肾小管上皮细胞转分化状态下ERK/STAT3信号转导通路的影响[J].湖南中医药大学学报,2022,42(9):1419-1425[点击复制] |
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| 温阳振衰颗粒对TGF-β1诱导肾小管上皮细胞转分化状态下ERK/STAT3信号转导通路的影响 |
| 林湘东,向茗,陈新宇 |
| (湖南中医药大学第一附属医院, 湖南 长沙 410007;湖南中医药大学中医诊断学教研室, 湖南 长沙 410208) |
| 摘要: |
| 目的 探讨温阳振衰颗粒对转化生长因子－β1（transforming growth factor-β1,TGF-β1）诱导肾小管上皮细胞转分化状态下细胞外信号调节激酶（extracellular signal-regulated kinase,ERK）/信号转导及转录激活因子3（signal transduction and activator of transcription 3,STAT3）信号转导通路的影响。方法 体外培养人肾小管上皮(HK-2)细胞，将HK-2细胞分为空白对照组、TGF-β1组（10 ng/mL TGF-β1）、TGF-β1+空白血清组（10 ng/mL TGF-β1）、TGF-β1+温阳振衰颗粒组（10 ng/mL TGF-β1+10%含药血清）、TGF-β1+U0126组（MEK1/2抑制剂，母液1∶10 000）、TGF-β1+厄贝沙坦组（10 ng/mL TGF-β1+10%含药血清），处理时间为24 h。倒置显微镜下观察细胞的形态改变，采用Western blot法和RT-qPCR法检测钙黏附蛋白E（E-cadherin）、波形蛋白（Vimentin）、α-平滑肌肌动蛋白（α-SMA）、ERK1/2、磷酸化ERK1/2（p-ERK1/2）、STAT3、磷酸化STAT3（p-STAT3）蛋白和mRNA的表达。结果 与空白对照组比较，TGF-β1组和TGF-β1+空白血清组细胞形态发生明显改变，呈长梭形或形态多样无规则，细胞排列稀疏，类似成纤维细胞形态，生长缓慢，贴壁差，多见漂浮细胞；与TGF-β1组和TGF-β1+空白血清组比较，TGF-β1+温阳振衰颗粒组、TGF-β1+U0126组、TGF-β1+厄贝沙坦组形态显著改善，大部分仍保持鹅卵石样，贴壁尚可，漂浮细胞明显减少。与空白对照组比较，TGF-β1组和TGF-β1+空白血清组E-cadherin蛋白和mRNA表达下调，Vimentin、α-SMA蛋白和mRNA表达上调，p-ERK1/2、p-STAT3蛋白表达上调（P＜0.05）；与TGF-β1组和TGF-β1+空白血清组比较，TGF-β1+温阳振衰颗粒组、TGF-β1+U0126组和TGF-β1+厄贝沙坦组E-cadherin蛋白和mRNA表达上调，Vimentin、α-SMA蛋白和mRNA表达下调，p-ERK1/2、p-STAT3蛋白表达下调（P＜0.05）,且3组效果相当；实验所有组别中ERK1/2、STAT3蛋白和mRNA表达差异无统计学意义（P＞0.05）。结论 温阳振衰颗粒可能通过下调ERK1/2和STAT3蛋白的磷酸化，从而部分阻断ERK/STAT3信号转导通路的激活，对肾小管上皮细胞转分化起到延缓作用。 |
| 关键词: 温阳振衰颗粒 转化生长因子－β1 肾小管 上皮细胞转分化 ERK/STAT3 机制研究 |
| DOI：10.3969/j.issn.1674-070X.2022.09.002 |
| 投稿时间：2021-07-20 |
| 基金项目:湖南省科学技术厅重点领域研发计划(2019SK2321);湖南省发改委创新引导专项(湘发改投资[2019]412号);湖南省教育厅优秀青年项目(18B240)。 |
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| Effects of Wenyang Zhenshuai Granule on ERK/STAT3 signal transduction pathway with epithelial mesenchymal transition of renal tubules induced by TGF-β1 |
| LIN Xiangdong,XIANG Ming,CHEN Xinyu |
| (The First Hospital of Hunan University of Chinese Medicine, Changsha, Hunan 410007, China;Department of Diagnostics in Chinese Medcine, Hunan University of Chinese Medicine, Changsha, Hunan 410208, China) |
| Abstract: |
| Objective To investigate the effects of Wenyang Zhenshuai Granule on extracellular signal-regulated kinase (ERK)/signal transduction and activator of transcription 3 (STAT3) signal transduction pathway with epithelial mesenchymal transition of renal tubules induced by transforming growth factor-β1 (TGF-β1). Methods Human renal tubular epithelial (HK-2) cells were cultured in vitro, and HK-2 cells were divided into blank control group, TGF-β1 group (10 ng/mL TGF-β1), TGF-β1+blank serum group (10 ng/mL TGF-β1), TGF-β1+Wenyang Zhenshuai Granule group (10 ng/mL TGF-β1+10% serum containing drug), TGF-β1+U0126 group (MEK1/2 inhibitor, stock solution was 1∶10 000), TGF-β1+irbesartan group (10 ng/mL TGF-β1+10% serum containing drug); the treatment time was 24 h. The morphological changes of cells were observed under an inverted microscope. Western blot and RT-qPCR were used to detect E-cadherin, Vimentin, α-smooth muscle actin (α-SMA), ERK1/2, phosphorylated ERK1/2 (p-ERK1/2), STAT3, phosphorylated STAT3 (p-STAT3) protein and mRNA expression. Results Compared with the blank control group, the morphology of the cells in the TGF-β1 group and the TGF-β1+blank serum group changed significantly. They showed a long spindle shape, or a variety of irregular shapes. Cells were sparsely arranged, similar to fibroblasts with slow growth and poor adherence, and mostly floating. Compared with TGF-β1 group and TGF-β1+blank serum group, the morphology of TGF-β1+Wenyang Zhenshui Granule group, TGF-β1+U0126 group and TGF-β1+irbesartan group was significantly improved. Some of them still remained cobblestone-like, with acceptable adherence to the wall, and the floating cells were significantly reduced. Compared with the blank control group, the expression of E-cadherin protein and mRNA in TGF-β1 group and TGF-β1+blank serum group was down-regulated; the protein and mRNA expression of Vimentin and α-SMA were up-regulated, and the protein expression levels of p-ERK1/2 and p-STAT3 were up-regulated (P＜0.05); compared with TGF-β1 group and TGF-β1+blank serum group, E-cadherin protein and mRNA in TGF-β1+Wenyang Zhenshui Granule group, TGF-β1+U0126 group and TGF-β1+irbesartan group were up-regulated; the expression levels of Vimentin, α-SMA protein and mRNA were down-regulated; the expression levels of p-ERK1/2 and p-STAT3 proteins were down-regulated (P<0.05), and the effects of the three groups were equivalent. The difference of the expression levels of ERK1/2, STAT3 protein and mRNA in all groups was not statistically significant (P>0.05). Conclusion Wenyang Zhenshuai Granule may partially block the activation of ERK/STAT3 signal transduction pathway by down-regulating the phosphorylation of ERK1/2 and STAT3 proteins, and delay the epithelial mesenchymal transition of renal tubules. |
| Key words: Wenyang Zhenshuai Granule transforming growth factor-β1 renal tubules epithelial mesenchymal transition ERK/STAT3 mechanism research |
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