The development of multidrug resistance is often associated with the over-expression of P-glycoprotein (P-gp). This protein prevents drug accumulation and extrudes them out of the cell before they reach the intended target. The aim of this study was to develop an in vitro MCF-7 cell line with increased expression of P-gp and test the phototoxicity of a novel photoactivated zinc phthalocyanine tetrasulfonic acid (ZnPcS4) on these cells. The over-expressed P-gp MCF-7 cells (MCF-7/DOX) were developed from wildtype (WT) MCF-7 cells by a stepwise continuous exposure of the WT cells to different concentrations of Doxorubicin (DOX) (0.1 - 1 μM) over a period of 4 months. The P-gp expression was measured using flow cytometry, immunofluorescence and enzyme immunoassay. To verify whether zinc phthalocyanine-mediated photodynamic therapy (ZnPcS4 - PDT) is effective in MCF-7/DOX, we studied the subcellular localization, phototoxicity and nuclear damage. The flow cytometry result showed two distinct peaks of P-gp positive and negative expression in MCF-7/DOX cell population, which correlates with the ELISA-based assay (p˂0.001). The ME16C (Normal breast cells) was used as control. The localization studies showed that ZnPcS4 have greater affinity for lysosome than mitochondria. Phototoxicity results indicated that photoactivated zinc phthalocyanine decreased the cell proliferation and viability as the drug and laser light dosages increased to 16 μM and 20 J/cm2 respectively. PDT-induced cytotoxicity using lactose dehydrogenase (LDH) enzyme leakage as measure did not increase likewise. The ZnPcS4-induced PDT was less effective for MCF-7/DOX cells which could be attributed to decreased retention of ZnPcS4 in major cellular organelles due to the presence of increased drug efflux P-gp. The current findings suggest that, increased P-gp expression, a characteristic of multidrug resistance together with other related intrinsic mechanisms might contribute to render MCF-7/DOX cells less sensitive to ZnPcS4-induced phototoxicity.

中文翻译：

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酞菁锌四磺酸锌对过表达P-糖蛋白的MCF-7细胞的光毒性作用。

多药耐药性的发展通常与P-糖蛋白（P-gp）的过表达有关。这种蛋白质可防止药物积聚，并在它们达到预定目标之前将其从细胞中挤出。这项研究的目的是开发具有增加的P-gp表达的体外MCF-7细胞系，并测试新型光活化锌酞菁锌四磺酸（ZnPcS4）对这些细胞的光毒性。通过将野生型（WT）MCF-7细胞逐步连续暴露于不同浓度的阿霉素（DOX）（0.1-1），从野生型（WT）MCF-7细胞发育出过表达的P-gp MCF-7细胞（MCF-7 / DOX） μM），持续4个月。使用流式细胞术，免疫荧光和酶免疫测定法测量P-gp表达。为了验证锌酞菁介导的光动力疗法（ZnPcS4-PDT）在MCF-7 / DOX中是否有效，我们研究了亚细胞定位，光毒性和核损伤。流式细胞仪检测结果显示MCF-7 / DOX细胞群中P-gp阳性和阴性表达有两个明显的峰值，这与基于ELISA的测定相关（p˂0.001）。使用ME16C（正常乳腺细胞）作为对照。本地化研究表明，ZnPcS4对溶酶体的亲和力比线粒体更大。光毒性结果表明，随着药物剂量和激光剂量分别增加至16μM和20 J / cm2，光活化的酞菁锌会降低细胞增殖和活力。使用乳糖脱氢酶（LDH）酶泄漏作为测量结果，PDT诱导的细胞毒性没有同样增加。ZnPcS4诱导的PDT对MCF-7 / DOX细胞的有效性较差，这可能归因于ZnPcS4在主要细胞器中的保留减少，这是由于药物外排P-gp增加所致。目前的发现表明，增加的P-gp表达，多药耐药性的特征以及其他相关的内在机制可能导致MCF-7 / DOX细胞对ZnPcS4诱导的光毒性不那么敏感。