Colorectal cancer cell adhesion attenuates Ad-E2F-1 mediated apoptosis.
Colorectal cancer cell adhesion attenuates Ad-E2F-1 mediated apoptosis.
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DOI：
10.1016/s0022-4804(03)00137-9
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发表时间：
2003-07
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影响因子：
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通讯作者：
C. Chao;Azemat Jamshidi‐Parsian;Warner W Wang;K. McMasters
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文献类型：
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作者：
C. Chao;Azemat Jamshidi‐Parsian;Warner W Wang;K. McMasters
细胞粘附分子、细胞内信号传导和肿瘤生长行为的复杂相互作用对大多数传统癌症治疗的失败具有重要意义。细胞与基底膜的粘附已被证明可以促进肿瘤细胞的存活。我们假设基质底物的存在通过细胞粘附分子的信号传导促进了化学耐药。材料与方法srko结直肠癌细胞表达整合素β1粘附于底物。在巨细胞病毒启动子的控制下，我们检测了含有转基因E2F-1 （Ad-E2F-1）和Ad-LacZ（作为对照）的腺病毒载体感染巨细胞病毒后细胞的凋亡情况。细胞被镀在Matrigel上，一种类似于基底膜的细胞外基质，并与组织培养塑料进行比较。流式细胞术TUNEL检测细胞凋亡，WST-1检测细胞增殖。Western blot分析证实E2F-1表达。使用功能阻断抗β1整合素抗体来评估β1对细胞存活的贡献。结果感染50次的RKO细胞感染120 h后，与Ad-LacZ对照处理的细胞相比，塑料处理的细胞在Ad-E2F-1的作用下发生了明显的凋亡（分别为53%和1%）。然而，当细胞涂于Matrigel时，相同剂量的E2F-1对诱导细胞凋亡无效（将Ad-E2F-1与Ad-LacZ对照进行比较，凋亡率为3% vs 1%）。细胞增殖试验显示，e2f -1感染的细胞在Matrigel上的细胞存活率是塑料上的3倍（P < 0.004）。通过Western blot分析，细胞凋亡的衰减可能是Matrigel的转导效率降低的结果，功能阻断型抗β1整合素抗体并不能消除Matrigel对细胞凋亡的抑制作用。结论腺病毒介导的E2F-1细胞凋亡逃逸，至少部分与细胞内E2F-1表达降低有关。通过β1整合素与基质蛋白的细胞粘附的相互作用似乎不会导致基因治疗耐药性。进一步的研究将探讨基因和/或化疗后其他特异性受体-配体相互作用。
BACKGROUNDThe complex interplay of cell adhesion molecules, intracellular signaling, and tumor growth behavior have important implications for the failure of most conventional cancer therapies. Cell adhesion to the basement membrane has been shown to promote tumor cell survival. We hypothesize that the presence of matrix substrate contributes to chemoresistance through signaling via cell adhesion molecule.MATERIALS AND METHODSRKO colorectal cancer cells express integrin β1 to adhere to substrate. We measured apoptosis of the cells after infection with adenovirus vector containing the transgene E2F-1 (Ad-E2F-1), a potent tumor suppressor gene, and Ad-LacZ (as control), both under the control of the cytomegalovirus promoter. Cells were plated on Matrigel, an extracellular substrate similar to basement membrane and compared to tissue culture plastic. Apoptosis was assessed by flow cytometry-based TUNEL assay and cell proliferation was assessed by WST-1 assay. E2F-1 expression was confirmed by Western blot analysis. A function-blocking anti-β1 integrin antibody was used to assess the contribution of β1 on cell survival.RESULTSAt 120 h postinfection of RKO cells with 50 multiplicity of infection, cells plated on plastic underwent marked apoptosis in response to Ad-E2F-1 compared with Ad-LacZ control-treated cells (53% vs 1% apoptosis, respectively). However, when cells were plated on Matrigel, the same dose of E2F-1 was ineffective at inducing apoptosis (3% vs 1% apoptosis, comparing Ad-E2F-1 with Ad-LacZ control). The cell proliferation assay showed >3-fold cell survival in E2F-1-infected cells on Matrigel vs plastic (P < 0.004). By Western blot analysis, attenuation of apoptosis may be a result of reduction in transduction efficiency on Matrigel and function-blocking anti-β1 integrin antibody does not abolish the decrease in apoptosis afforded by Matrigel.CONCLUSIONSThese data suggest that escape from adenoviral E2F-1-mediated apoptosis, at least in part, is related to reduction of intracellular E2F-1 expression. Interactions involving cellular adhesion via β1 integrin to matrix proteins does not seem to contribute toward gene therapy resistance. Further studies will investigate other specific receptor-ligand interactions after gene and/or chemotherapy.