Institute of Tropical Diseases, Guangzhou University of Chinese Medicine, Guangzhou, Guangdong, 510000, People's Republic of China.
Department of Spleen and Stomach Diseases, Second Hospital of Traditional Chinese Medicine of Guangdong, Guangzhou, Guangdong, 510000, People's Republic of China.
Invest New Drugs. 2019 Oct;37(5):865-875. doi: 10.1007/s10637-018-0702-x. Epub 2018 Nov 29.
Purpose Transdifferentiation exists within stromal cells in the tumour microenvironment. Transforming growth factor-β (TGF-β) secreted by tumour-associated fibroblasts (TAFs) affects the differentiation states of epithelial cells, including epithelial-mesenchymal transition (EMT). Evodiamine, a natural drug, can regulate differentiation. However, the specific effects and relative mechanisms of evodiamine remain unknown. Design We used four models to observe the influence of TAF-like CCD-18Co cells on the colon epithelial cell line HCoEpiC: the 3D- and 2D-mono-culture system, Transwell and direct co-culture model. Additionally, we established conditioned medium from CCD-18Co cells. The TGF-β pathway inhibitor LY364947 and evodiamine were added. Morphological changes and classical EMT markers were observed and detected using phase contrast microscopy and immunofluorescence. Cell migration was measured by the wound-healing assay. Western blotting was performed to detect the TGF-β/Smad signalling pathway. Results CCD-18Co cells induced EMT-like changes in the 2D- and 3D-cultured epithelial cell line HCoEpiC, accompanied by high expression of ZEB1 and Snail and the enhancement of migration. Moreover, CCD-18Co-derived conditioned medium caused dysfunction of TGF-β/Smad signalling in EMT. Evodiamine inhibited these EMT-like HCoEpiC and their migration. Additionally, evodiamine down-regulated the expression of ZEB1/Snail and up-regulated the expression of phosphorylated Smad2/3 (pSmad2/3). Evodiamine also increased the ratios of pSmad2/Smad2 and pSmad3/Smad3. Conclusion Based on our observations, evodiamine can reverse the TAF-induced EMT-like phenotype in colon epithelial cells, which may be associated with its mediation of phosphorylated Smad2 and Smad3 expression.
肿瘤微环境中的基质细胞存在转分化现象。肿瘤相关成纤维细胞（TAFs）分泌的转化生长因子-β（TGF-β）影响上皮细胞的分化状态，包括上皮-间充质转化（EMT）。吴茱萸碱是一种天然药物，可调节分化。然而，吴茱萸碱的具体作用和相对机制尚不清楚。
我们使用四个模型来观察 TAF 样 CCD-18Co 细胞对结肠上皮细胞系 HCoEpiC 的影响：3D 和 2D 单核培养系统、Transwell 和直接共培养模型。此外，我们还建立了 CCD-18Co 细胞的条件培养基。加入 TGF-β 通路抑制剂 LY364947 和吴茱萸碱。通过相差显微镜和免疫荧光观察和检测形态变化和经典 EMT 标志物。通过划痕愈合试验测量细胞迁移。通过 Western blot 检测 TGF-β/Smad 信号通路。
CCD-18Co 细胞在 2D 和 3D 培养的上皮细胞系 HCoEpiC 中诱导 EMT 样变化，伴随着 ZEB1 和 Snail 的高表达和迁移增强。此外，CCD-18Co 衍生的条件培养基导致 EMT 中 TGF-β/Smad 信号通路功能障碍。吴茱萸碱抑制这些 EMT 样 HCoEpiC 及其迁移。此外，吴茱萸碱下调 ZEB1/Snail 的表达，上调磷酸化 Smad2/3（pSmad2/3）的表达。吴茱萸碱还增加了 pSmad2/Smad2 和 pSmad3/Smad3 的比值。
根据我们的观察，吴茱萸碱可以逆转 TAF 诱导的结肠上皮细胞 EMT 样表型，这可能与其介导的磷酸化 Smad2 和 Smad3 表达有关。
