模拟低氧微环境中基质硬度对结肠癌细胞上皮-间质转化的影响
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国家自然科学基金项目(81770058),济宁医学院教师科研扶持基金项目(JYFC2018KJ071)


Effects of Substrate Stiffness on Epithelial-Mesenchymal Transition of Colon Cancer Cells under Simulated Hypoxia Environment
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    目的 通过模拟人结肠癌组织微环境,研究基质硬度和低氧协同对结肠癌细胞 SW480 上皮-间质转化(epithelial-mesenchymal transition, EMT) 的影响。 方法 制备硬度分别为 4. 5、 20、 40 kPa 聚乙烯醇 ( polyvinylalcohol, PVA)凝胶模拟结肠癌组织各部位的刚度范围。 以氯化钴(CoCl2 )模拟低氧微环境,在不同硬度基底上检测细胞形态变化;利用 Western blot 检测低氧诱导因子(hypoxia inducible factor, HIF-1α)以及 EMT 相关蛋白 E-钙黏蛋白(E-cadherin)、波形蛋白(Vimentin)、转录因子 Snail 1 表达;通过实时荧光定量 PCR 检测 HIF-1α、E-cadherin、Vimentin、Snail 1、基质金属蛋白酶 2(matrix metalloproteinase-2, MMP-2)和 MMP-9 基因表达。 结果 模拟低氧微环境中,随基质硬度增加,细胞铺展面积增大,由圆形逐渐变为纺锤形;通过上调 Vimentin、Snail-1、MMP-2、MMP-9 表达以及下调 E-cadherin 表达,可以促进结肠癌细胞 SW480 EMT 的发生。 结论 研究结果对进一步探索基质硬度和低氧对结肠癌细胞 EMT 协同作用及分子机制具有重要意义。

    Abstract:

    Objective To explore the effects from the synergy of substrate stiffness and hypoxia on epithelialmesenchymal transition (EMT) of colon cancer cells SW480 by simulating the microenvironment of human colon cancer tissues. Methods Polyvinyl alcohol gels with different stiffness ( 4. 5, 20, 40 kPa) were prepared to simulate the stiffness of each part of colon cancer tissues. The morphological change of cells on substrate with different stiffness was detected under simulated hypoxia ( CoCl2 ) environment. The expression of hypoxiainducible factor (HIF-1α), and EMT markers E-cadherin, Vimentin, Snail 1 were detected by Western blot. The mRNA expression of E-cadherin, Vimentin, Snail 1, matrix metalloproteinase-2 ( MMP-2), and MMP-9 was detected by quantitative real-time PCR ( qRT-PCR). Results Under simulated hypoxia environment, with the increase of substrate stiffness, the SW480 cells spreading area increased, and transformed from round shape into irregular polygon. The EMT of SW480 could be enhanced through up-regulating expression of Vimentin, Snail 1, MMP-2, MMP-9, and down-regulating expression of E-cadherin. Conclusions This study is important for exploring the synergistic effect of substrate stiffness and hypoxia on the EMT of colon cancer cells as well as the molecular mechanism.

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张 静,周家建,党青亚,许欣欣,林英豪,李勋晓,李海滨.模拟低氧微环境中基质硬度对结肠癌细胞上皮-间质转化的影响[J].医用生物力学,2023,38(2):261-267

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  • 收稿日期:2021-11-15
  • 最后修改日期:2022-01-16
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  • 在线发布日期: 2023-04-25
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