Department of Internal Medicine I, Cardiology Division, Friedrich Schiller University, Jena, Germany.
J Cell Biochem. 2010 Feb 1;109(2):434-46. doi: 10.1002/jcb.22422.
ABC transporters like P-glycoprotein (P-gp/ABCB1) are membrane proteins responsible for the transport of toxic compounds out of non-malignant cells and tumor tissue.
To investigate the effect of glycolysis and the tissue redox state on P-gp expression in multicellular tumor spheroids derived from prostate adenocarcinoma cells (DU-145), glioma cells (Gli36), and the human cervix carcinoma cell line KB-3-1 transfected with a P-gp-EGFP fusion gene that allows monitoring of P-gp expression in living cells. During cell culture of DU-145, Gli36, and KB-3-1 tumor spheroids P-gp expression was observed as well as increased lactate and decreased pyruvate levels and expression of glycolytic enzymes. Inhibition of glycolysis for 24 h by either iodoacetate (IA) or 2-deoxy-D-glucose (2-DDG) downregulated P-gp expression which was reversed upon coincubation with the radical scavenger ebselen as shown by semi-quantitative immunohistochemisty in DU-145 and Gli36 tumor spheroids, and by EGFP fluorescence in KB-3-1 tumor spheroids. Consequently endogenous ROS generation in DU-145 tumor spheroids was increased in the presence of either IA or 2-DDG, which was abolished upon coincubation with ebselen. Exogenous addition of pyruvate significantly reduced ROS generation, increased P-gp expression as well as efflux of the P-gp substrate doxorubicin. Doxorubicin transport was significantly blunted by 2-DDG and IA, indicating that inhibition of glycolysis reversed the multidrug resistance phenotype. In summary our data demonstrate that P-gp expression in tumor spheroids is closely related to the glycolytic metabolism of tumor cells and can be downregulated by glycolysis inhibitors via mechanisms that involve changes in the cellular redox state.
ABC 转运蛋白，如 P-糖蛋白（P-gp/ABCB1），是负责将有毒化合物从非恶性细胞和肿瘤组织中排出的膜蛋白。
研究糖酵解和组织氧化还原状态对多细胞肿瘤球体中 P-糖蛋白表达的影响，这些肿瘤球体源自前列腺腺癌细胞（DU-145）、神经胶质瘤细胞（Gli36）和转染 P-糖蛋白-EGFP 融合基因的人宫颈癌细胞系 KB-3-1，该融合基因允许在活细胞中监测 P-糖蛋白的表达。在 DU-145、Gli36 和 KB-3-1 肿瘤球体的细胞培养过程中观察到 P-糖蛋白表达增加，同时乳酸水平升高，丙酮酸水平降低，糖酵解酶表达增加。用碘乙酸（IA）或 2-脱氧-D-葡萄糖（2-DDG）抑制糖酵解 24 小时，下调 P-糖蛋白表达，用半定量免疫组织化学法在 DU-145 和 Gli36 肿瘤球体中，以及用 KB-3-1 肿瘤球体中的 EGFP 荧光显示，与自由基清除剂 ebselen 共孵育后可逆转 P-糖蛋白表达。因此，在存在 IA 或 2-DDG 的情况下，DU-145 肿瘤球体中的内源性 ROS 生成增加，而在用 ebselen 共孵育时则被消除。外源性添加丙酮酸可显著减少 ROS 生成，增加 P-糖蛋白表达以及 P-糖蛋白底物阿霉素的外排。2-DDG 和 IA 显著抑制阿霉素的转运，表明糖酵解抑制剂的抑制作用逆转了多药耐药表型。总之，我们的数据表明，肿瘤球体中的 P-糖蛋白表达与肿瘤细胞的糖酵解代谢密切相关，糖酵解抑制剂可以通过改变细胞氧化还原状态来下调 P-糖蛋白表达。
