| 引用本文:肖贵梅，蔡向志，龙海洮，田丽霞，任燕.白术内酯Ⅲ介导的肌动蛋白相关蛋白2/3抑制肠上皮间质转化以缓解溃疡性结肠炎的研究[J].世界中医药,2024,(15):. |
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| 白术内酯Ⅲ介导的肌动蛋白相关蛋白2/3抑制肠上皮间质转化以缓解溃疡性结肠炎的研究 |
| Atractylenolide Ⅲ-mediated Actin-related Protein 2/3 Alleviates Ulcerative Colitis by Inhibiting Intestinal Epithelial-mesenchymal Transition |
| 投稿时间：2024-01-08 |
| DOI：10.3969/j.issn.1673-7202.2024.15.004 |
| 中文关键词: 白术内酯Ⅲ 大鼠小肠隐窝上皮细胞 溃疡性结肠炎 肌动蛋白相关蛋白2/3 有限酶切-质谱法 上皮细胞-间充质转化 紧密连接通路 脂多糖 |
| English Keywords:Atractylenolide Ⅲ IEC-6 Ulcerative colitis Actin-related protein 2/3 Limited proteolysis-mass spectrometry Epithelial-mesenchymal transition Tight junction pathway Lipopolysaccharide |
| 基金项目:国家自然科学基金项目（82360754）；贵州省科技计划项目（黔科合LH字〔2017〕7295号） |
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| 目的：探寻白术内酯Ⅲ（AT-Ⅲ）的关键靶点，探究肌动蛋白相关蛋白2/3（Arp2/3）抑制溃疡性结肠炎（UC）的作用机制。方法：采用有限酶切-质谱法（lip-SMap）筛选大鼠小肠隐窝上皮细胞（IEC-6）特征蛋白，进行基因本体（GO）和京都基因与基因组百科全书（KEGG）富集分析，确定AT-Ⅲ的关键靶点。脂多糖（LPS）与IEC-6共孵育模拟UC模型，酶联免疫吸附实验（ELISA）检测白细胞介素-6（IL-6），肿瘤坏死因子-α（TNF-α）含量；划痕实验测定IEC-6迁移速率；蛋白质印迹法测定上皮钙黏素（E-cadherin）、波形蛋白（Vimentin）和Arp2/3含量；分子对接评估AT-Ⅲ与关键靶点的结合能力。结果：基于lip-SMap获得的特征蛋白参与调控磷脂酰肌醇3激酶-丝苏氨酸激酶（PI3K-AKT）信号通路和紧密连接（TJ）信号通路；与对照组比较，LPS组中IL-6和TNF-α释放量升高（均P<0.05），迁移速率变快（P<0.05），E-cadherin和Vimentin的表达都有不同程度的影响（均P<0.05）；与LPS组比较，LPS+CK666+AT-Ⅲ组显著降低IL-6和TNF-α含量（均P<0.01），有效逆转LPS诱导的细胞迁移变化及E-cadherin和Vimentin表达（均P<0.05）；分子对接显示，Arp2/3与AT-Ⅲ有较好的结合活性。结论：Arp2/3作为AT-Ⅲ的关键靶点，可通过上皮细胞-间充质转化（EMT）缓解LPS诱导的UC。 |
| English Summary: |
| To identify the key targets of atractylenolide Ⅲ(AT-Ⅲ) and to explore the mechanism of actin-related protein 2/3(Arp2/3) in inhibiting ulcerative colitis(UC).Methods:Characteristic proteins of rat small intestinal crypt epithelial cells(IEC-6) were screened using limited proteolysis-mass spectrometry(lip-SMap).Gene Ontology(GO) and Kyoto Encyclopedia of Genes and Genomes(KEGG) enrichment analyses were conducted to determine the key targets of AT-Ⅲ.An UC model was simulated by co-incubating lipopolysaccharide(LPS) with IEC-6 cells.Enzyme-linked immunosorbent assay(ELISA) was used to measure the levels of interleukin-6(IL-6) and tumor necrosis factor-alpha(TNF-α).A scratch assay was performed to measure the migration rate of IEC-6 cells.Western blot was used to measure the levels of E-cadherin，vimentin，and Arp2/3.Molecular docking was conducted to evaluate the binding ability of AT-Ⅲ to its key targets.Results:The characteristic proteins obtained from lip-SMap were involved in regulating the phosphatidylinositol 3-kinase-serine/threonine kinase(PI3K-AKT) signaling pathway and the tight junction(TJ) signaling pathway.Compared with the control group，the LPS group showed increased levels of IL-6 and TNF-α(both P<0.05)，an increased migration rate(P<0.05)，and various impacts on the expression of E-cadherin and vimentin(both P<0.05).Compared with the LPS group，the LPS+CK666+AT-Ⅲ group significantly reduced IL-6 and TNF-α levels(both P<0.01)，effectively reversed LPS-induced changes in cell migration and the expression of E-cadherin and vimentin(both P<0.05).Molecular docking showed that Arp2/3 had good binding activity with AT-Ⅲ.Conclusion:Arp2/3 is a key target of AT-Ⅲ and can alleviate LPS-induced UC through epithelial-mesenchymal transition(EMT). |
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