Inhibition of Na+/K+-ATPase induces hybrid cell death and enhanced sensitivity to chemotherapy in human glioblastoma cells.
Inhibition of Na+/K+-ATPase induces hybrid cell death and enhanced sensitivity to chemotherapy in human glioblastoma cells.
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DOI：
10.1186/1471-2407-14-716
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发表时间：
2014-09-26
期刊：
影响因子：
3.8
通讯作者：
Yu SP
中科院分区：
文献类型：
--
作者：
Chen D;Song M;Mohamad O;Yu SP
多形性胶质母细胞瘤（GBM）很难用传统的抗癌/抗凋亡药物治疗。我们测试了这样的假设：Na+/K+-ATP酶的抑制会导致同时发生的细胞凋亡和坏死的混合或混合形式，因此应该增强化疗对胶质母细胞瘤细胞的抗癌作用。在人 LN229 和耐药 T98G 胶质母细胞瘤细胞培养物中，使用免疫细胞化学和蛋白质印迹法测量细胞死亡和信号通路。应用荧光染料测量细胞内Ca2+、Na+和K+的变化。特定的 Na+/K+-ATPase 阻断剂哇巴因 (0.1 - 10 μM) 以时间和浓度依赖性方式诱导细胞死亡和 K+ 稳态破坏。 annexin-V 易位和 caspase-3 激活表明哇巴因细胞毒性中存在细胞凋亡成分，同时伴随着 Bcl-2 表达和线粒体膜电位的降低。哇巴因诱导的细胞死亡可被半胱天冬酶抑制剂 Z-VAD (100 μM) 部分减弱。一致地，K+离子载体缬氨霉素以K+流出依赖性方式启动LN229细胞的细胞凋亡。哇巴因最初引起细胞膨胀，随后细胞体积持续减少。电子显微镜揭示了同一细胞中凋亡和坏死改变的超微结构特征。最后，与 TMZ 敏感细胞系 LN229 和正常人星形胶质细胞相比，对化疗药物替莫唑胺 (TMZ) 耐药的人 T98G 胶质母细胞瘤细胞表现出独特的 Na+/K+-ATPase α2 和 α3 亚基高表达。在低浓度下，哇巴因选择性杀死 T98G 细胞。敲低 α3 亚基会使 T98G 细胞对 TMZ 敏感，并导致更多细胞死亡。这项研究表明，抑制 Na+/K+-ATP 酶会引发混合细胞死亡，并作为增强胶质母细胞瘤细胞化疗效果的潜在机制。本文的在线版本 (doi:10.1186/1471-2407-14-716) 包含补充材料，可供授权用户使用。
Glioblastoma multiforme (GBM) is very difficult to treat with conventional anti-cancer/anti-apoptotic drugs. We tested the hypothesis that inhibition of Na+/K+-ATPase causes a mixed or hybrid form of concurrent apoptosis and necrosis and therefore should enhance anti-cancer effects of chemotherapy on glioblastoma cells. In human LN229 and drug-resistant T98G glioblastoma cell cultures, cell death and signal pathways were measured using immunocytochemistry and Western blotting. Fluorescent dyes were applied to measure intracellular Ca2+, Na+ and K+ changes. The specific Na+/K+-ATPase blocker ouabain (0.1 - 10 μM) induced cell death and disruption of K+ homeostasis in a time- and concentration-dependent manner. Annexin-V translocation and caspase-3 activation indicated an apoptotic component in ouabain cytoxicity, which was accompanied with reduced Bcl-2 expression and mitochondrial membrane potential. Ouabain-induced cell death was partially attenuated by the caspase inhibitor Z-VAD (100 μM). Consistently, the K+ ionophore valinomycin initiated apoptosis in LN229 cells in a K+ efflux-dependent manner. Ouabain caused an initial cell swell, which was followed by a sustained cell volume decrease. Electron microscopy revealed ultrastructural features of both apoptotic and necrotic alterations in the same cells. Finally, human T98G glioblastoma cells that are resistant to the chemotherapy drug temozolomide (TMZ) showed a unique high expression of the Na+/K+-ATPase α2 and α3 subunits compared to the TMZ-sensitive cell line LN229 and normal human astrocytes. At low concentrations, ouabain selectively killed T98G cells. Knocking down the α3 subunit sensitized T98G cells to TMZ and caused more cell death. This study suggests that inhibition of Na+/K+-ATPase triggers hybrid cell death and serves as an underlying mechanism for an enhanced chemotherapy effect on glioblastoma cells. The online version of this article (doi:10.1186/1471-2407-14-716) contains supplementary material, which is available to authorized users.
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影响因子：
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作者：
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