Mitochondrial toxicity (Mito-Tox) risk has increased due to the administration of several classes of drugs, particularly some life-long antiretroviral drugs for HIV⁺ individuals. However, no suitable in vitro assays are available to test long-term Mito-Tox (≥4 weeks). The goal of this study is to develop a 3D spheroid system of human primary urine-derived stem cells (USC) for the prediction of drug-induced delayed Mito-Tox. The cytotoxicity and Mito-Tox were assessed in 3D USC spheroids 4 weeks after treatment with antiretroviral drugs: zalcitabine (ddC; 0.1, 1 and 10 µM), tenofovir (TFV; 3, 30 and 300 µM) or Raltegravir (RAL; 2, 20 and 200 µM). Rotenone (RTNN, 10 µM) and 0.1% DMSO served as positive and negative controls. Despite only mild cytotoxicity, ddC significantly inhibited the expression of oxidative phosphorylation enzyme Complexes I, III, and IV; and RAL transiently reduced the level of Complex IV. A significant increase in caspase 3 and ROS/RNS level but a decrease in total ATP were observed in USC treated with ddC, TFV, RAL, and RTNN. Levels of mtDNA content and mitochondrial mass were decreased in ddC but minimally or not in TFV- and RAL-treated spheroids. Thus, 3D USC spheroid using antiretroviral drugs as a model offers an alternative platform to assess drug-induced late Mito-Tox.

中文翻译：

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人原代尿液干细胞的 3D 球体在药物诱导的线粒体毒性评估中的应用

由于使用几类药物，特别是一些针对 HIV ⁺个体的终生抗逆转录病毒药物，线粒体毒性 (Mito-Tox) 风险有所增加。然而，没有合适的体外测定可用于测试长期 Mito-Tox（≥4 周）。本研究的目标是开发人类原代尿源干细胞 (USC) 的 3D 球体系统，用于预测药物诱导的延迟 Mito-Tox。使用抗逆转录病毒药物治疗 4 周后，在 3D USC 球体中评估细胞毒性和 Mito-Tox：扎西他滨（ddC；0.1、1 和 10 µM）、替诺福韦（TFV；3、30 和 300 µM）或拉特拉韦（RAL；2、 20 和 200 µM）。鱼藤酮 (RTNN, 10 µM) 和 0.1% DMSO 作为阳性和阴性对照。尽管只有轻微的细胞毒性，ddC 仍显着抑制氧化磷酸化酶复合物 I、III 和 IV 的表达；RAL 暂时降低了复合物 IV 的水平。在用 ddC、TFV、RAL 和 RTNN 处理的 USC 中观察到 caspase 3 和 ROS/RNS 水平显着增加，但总 ATP 减少。ddC 中 mtDNA 含量和线粒体质量水平下降，但 TFV 和 RAL 处理的球体中的下降幅度很小或没有。因此，使用抗逆转录病毒药物作为模型的 3D USC 球体提供了评估药物诱导的晚期 Mito-Tox 的替代平台。