S-nitrosocysteine and glutathione depletion synergize to induce cell death in human tumor cells: Insights into the redox and cytotoxic mechanisms.
S-nitrosocysteine and glutathione depletion synergize to induce cell death in human tumor cells: Insights into the redox and cytotoxic mechanisms.
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DOI：
10.1016/j.freeradbiomed.2020.08.026
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发表时间：
2020-11-20
影响因子：
7.4
通讯作者：
Benhar M
中科院分区：
文献类型：
--
作者：
Knany A;Engelman R;Hariri HA;Biswal S;Wolfenson H;Benhar M
一氧化氮（NO）依赖性信号传导和细胞毒性作用部分通过蛋白质S-亚硝基化介导。S-亚硝基化的幅度和持续时间由两个主要的巯基还原系统，谷胱甘肽（GSH）和硫氧还蛋白（Trx）抗氧化系统控制。近年来，已经开发了利用NO/亚硝基化的细胞毒性潜力来抑制肿瘤细胞生长的方法。然而，在这一领域的进展受到阻碍的平衡和细胞亚硝基化，其他氧化过程和GSH/Trx系统之间的相互作用的认识不足。此外，巯基氧化还原失衡与癌细胞死亡之间的机制关系尚未完全了解。在此，我们探讨了S-亚硝基化剂，S-亚硝基半胱氨酸（CysNO），在GSH充足和缺乏的人肿瘤细胞诱导的氧化还原和细胞效应。我们使用L-丁硫氨酸-亚砜亚胺（BSO）诱导GSH缺乏，并采用氧化还原，生化和细胞分析来询问分子机制。我们发现，在GSH充足的条件下，CysNO激发（100-500 μM）导致蛋白质S-亚硝基化在不存在明显S-氧化的情况下显著但可逆的增加。相反，在GSH缺乏的条件下，CysNO诱导升高和持续水平的S-亚硝基化和S-氧化。在各种癌细胞系中的实验显示，单独施用CysNO或BSO通常诱导最小的细胞毒性，而BSO/CysNO组合疗法导致广泛的细胞死亡。在HeLa癌细胞中的研究表明，用BSO/CysNO处理导致GSH和Trx系统的双重抑制，从而放大氧化还原应激并引起细胞功能障碍。特别是，BSO/CysNO诱导肌动蛋白细胞骨架网络的快速氧化和崩溃，随后线粒体功能丧失，导致ATP水平的深刻和不可逆的下降。进一步的观察表明，BSO/CysNO诱导的细胞死亡发生通过半胱天冬酶独立的机制，涉及多个应激诱导的途径。目前的研究结果提供了新的见解细胞亚硝基化/氧化，巯基抗氧化剂防御和细胞死亡之间的关系。这些结果可能有助于未来开发基于NO/氧化还原的抗癌方法。
Nitric oxide (NO)-dependent signaling and cytotoxic effects are mediated in part via protein S-nitrosylation. The magnitude and duration of S-nitrosylation are governed by the two main thiol reducing systems, the glutathione (GSH) and thioredoxin (Trx) antioxidant systems. In recent years, approaches have been developed to harness the cytotoxic potential of NO/nitrosylation to inhibit tumor cell growth. However, progress in this area has been hindered by insufficient understanding of the balance and interplay between cellular nitrosylation, other oxidative processes and the GSH/Trx systems. In addition, the mechanistic relationship between thiol redox imbalance and cancer cell death is not fully understood. Herein, we explored the redox and cellular effects induced by the S-nitrosylating agent, S-nitrosocysteine (CysNO), in GSH-sufficient and -deficient human tumor cells. We used L-buthionine-sulfoximine (BSO) to induce GSH deficiency, and employed redox, biochemical and cellular assays to interrogate molecular mechanisms. We found that, under GSH-sufficient conditions, a CysNO challenge (100-500 μM) results in a marked yet reversible increase in protein S-nitrosylation in the absence of appreciable S-oxidation. In contrast, under GSH-deficient conditions, CysNO induces elevated and sustained levels of both S-nitrosylation and S-oxidation. Experiments in various cancer cell lines showed that administration of CysNO or BSO alone commonly induce minimal cytotoxicity whereas BSO/CysNO combination therapy leads to extensive cell death. Studies in HeLa cancer cells revealed that treatment with BSO/CysNO results in dual inhibition of the GSH and Trx systems, thereby amplifying redox stress and causing cellular dysfunction. In particular, BSO/CysNO induced rapid oxidation and collapse of the actin cytoskeletal network, followed by loss of mitochondrial function, leading to profound and irreversible decrease in ATP levels. Further observations indicated that BSO/CysNO-induced cell death occurs via a caspase-independent mechanism that involves multiple stress-induced pathways. The present findings provide new insights into the relationship between cellular nitrosylation/oxidation, thiol antioxidant defenses and cell death. These results may aid future efforts to develop NO/redox-based anticancer approaches.
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影响因子：
7.4
作者：
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通讯作者：
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