Department of Pathology, College of Korean Medicine, Wonkwang University, Iksan, Jeonbuk 54538, Republic of Korea.
Department of Microbiology and Immunology, School of Medicine, Wonkwang University, Iksan, Jeonbuk 54538, Republic of Korea.
Int J Mol Med. 2017 Dec;40(6):1957-1964. doi: 10.3892/ijmm.2017.3181. Epub 2017 Oct 10.
Cisplatin is one of the most widely used chemotherapeutic agents for the treatment of human cancers. However, the nephrotoxicity of cisplatin limits its use as a therapeutic agent. It has been suggested that oxidative stress and p53 activation play important roles in cisplatin-induced nephrotoxicity. It has been demonstrated that the eukaryotic translation initiation factor 2α (eIF2α) may protect HK-2 human renal proximal tubular cells against cisplatin-induced apoptosis through inhibition of reactive oxygen species (ROS)‑mediated p53 activation. The aim of the present study was to investigate the effects of siRNA‑mediated knockdown of the PKR-like endoplasmic reticulum kinase (PERK) gene, which induces the phosphorylation of eIF2α, or Sal003, a selective inhibitor of eIF2α dephosphorylation, on cisplatin‑induced apoptosis in HK-2 cells. Cisplatin induced eIF2α phosphorylation as well as p53 activation. In particular, inhibition of p53 by pifithrin‑α, and upregulation of eIF2α phosphorylation by Sal003, reduced cisplatin-induced apoptosis. Of note, Sal003‑mediated upregulation of eIF2α phosphorylation suppressed cisplatin‑induced p53 activation. Furthermore, reduction of eIF2α phosphorylation by PERK knockdown enhanced cisplatin-induced p53 activation and apoptosis. In addition, the ROS scavenger N-acetyl-L-cysteine inhibited eIF2α phosphorylation as well as p53 activation in HK-2 cells treated with cisplatin, suggesting that oxidative stress induced by cisplatin may lead to apoptosis through p53 activation; furthermore, this stress may confer resistance to apoptosis via eIF2α phosphorylation, which was further supported by the finding that cisplatin‑induced ROS generation was attenuated by Sal003, whereas it was enhanced by PERK knockdown. Furthermore, cisplatin induced the expression of activating transcription factor 4 (ATF4) and heme oxygenase-1 (HO-1) that were enhanced by Sal003 and reduced by PERK knockdown. Taken together, these results suggest that phosphorylation of eIF2α suppresses cisplatin‑induced p53 activation and apoptosis by attenuating oxidative stress via ATF4-mediated HO-1 expression in HK-2 cells, as ATF4 expression is usually dependent on the phosphorylation of eIF2α and may also transcriptionally induce the expression of HO-1 in response to oxidative stress. Therefore, regulation of eIF2α phosphorylation may play an important role in alleviating cisplatin-induced nephrotoxicity.
顺铂是治疗人类癌症最广泛使用的化疗药物之一。然而，顺铂的肾毒性限制了其作为治疗剂的应用。已经表明，氧化应激和 p53 激活在顺铂诱导的肾毒性中发挥重要作用。已经证明，真核翻译起始因子 2α（eIF2α）可通过抑制活性氧（ROS）介导的 p53 激活来保护 HK-2 人肾近端肾小管细胞免受顺铂诱导的凋亡。本研究旨在探讨 siRNA 介导的蛋白激酶 R 样内质网激酶（PERK）基因敲低，其诱导 eIF2α 的磷酸化，或选择性抑制 eIF2α 去磷酸化的 Sal003，对 HK-2 细胞中顺铂诱导的细胞凋亡的影响。顺铂诱导 eIF2α 磷酸化以及 p53 激活。特别是，pifithrin-α抑制 p53，以及 Sal003 上调 eIF2α 磷酸化，减少顺铂诱导的凋亡。值得注意的是，Sal003 介导的 eIF2α 磷酸化上调抑制了顺铂诱导的 p53 激活。此外，PERK 敲低减少 eIF2α 磷酸化增强了顺铂诱导的 p53 激活和凋亡。此外，ROS 清除剂 N-乙酰-L-半胱氨酸抑制了顺铂处理的 HK-2 细胞中 eIF2α 的磷酸化以及 p53 的激活，表明顺铂诱导的氧化应激可能通过 p53 激活导致细胞凋亡；此外，这种应激可能通过 eIF2α 磷酸化赋予细胞凋亡抗性，这进一步得到了以下发现的支持：Sal003 减弱了顺铂诱导的 ROS 生成，而 PERK 敲低增强了 ROS 生成。此外，顺铂诱导激活转录因子 4（ATF4）和血红素加氧酶 1（HO-1）的表达，Sal003 增强了它们的表达，PERK 敲低则降低了它们的表达。总之，这些结果表明，eIF2α 的磷酸化通过 ATF4 介导的 HO-1 表达来减轻氧化应激，从而抑制顺铂诱导的 p53 激活和凋亡，因为 ATF4 的表达通常依赖于 eIF2α 的磷酸化，并且也可能响应氧化应激转录诱导 HO-1 的表达。因此，调节 eIF2α 磷酸化可能在减轻顺铂诱导的肾毒性中发挥重要作用。
