Department of Nephrology, Henan Provincial Key Laboratory of Kidney Disease and Immunology, Henan Provincial Clinical Research Center for Kidney Disease, Henan Provincial People's Hospital and People's Hospital of Zhengzhou University, 7 Weiwu Road, Henan 450053, China; Academy of Medical Sciences, Zhengzhou University, Zhengzhou, Henan 450052, China.
Department of Nephrology, Henan Provincial Key Laboratory of Kidney Disease and Immunology, Henan Provincial Clinical Research Center for Kidney Disease, Henan Provincial People's Hospital and People's Hospital of Zhengzhou University, 7 Weiwu Road, Henan 450053, China.
Ecotoxicol Environ Saf. 2022 Sep 1;242:113881. doi: 10.1016/j.ecoenv.2022.113881. Epub 2022 Jul 18.
Cobalt is a transition element that abundantly exists in the environment. Besides direct hypoxia stress, cobalt ions indirectly induce hypoxia-reoxygenation injury (HRI), the main cause of acute kidney injury (AKI), a life-threatening clinical syndrome characterized by the necrosis of the proximal tubular epithelial cells (PTECs) and inflammation. Pyroptosis, a type of inflammatory programmed cell death, might play an essential role in HRI-AKI. However, whether pyroptosis is involved in cobalt chloride (CoCl)-induced HRI-AKI remains unknown. Autophagy is a cellular biological process maintaining cell homeostasis that is involved in cell damage in AKI, yet the underlying regulatory mechanism of autophagy on pyroptosis has not been fully understood. In this study, the in vitro and in vivo models of CoCl-induced HRI-AKI were established with HK-2 cell line and C57BL/6J mouse. Pyroptosis-related markers were detected with western blotting and immunofluorescence assays, and results showed that gasdermin E (GSDME)-mediated pyroptosis was involved in the cell damage in HRI-AKI. Specific chemical inhibitors of caspase 3, caspase 8, and caspase 9 significantly inhibited GSDME-mediated pyroptosis, verifying that GSDME-mediated pyroptosis was induced via the activation of caspase 3/8/9. The western blotting and immunofluorescence assays were adopted to detect the accumulation of the autophagosomes, and results suggested that HRI increased the autophagic level. The effects of autophagy on apoptosis and pyroptosis were evaluated using lentivirus transfection assays to knock down autophagy-specific genes atg5 and fip200, and results demonstrated that autophagy induced GSDME-mediated pyroptosis via apoptotic pathways in HRI-AKI. Our results revealed the involvement of GSDME-mediated pyroptosis in CoCl-induced HRI-AKI and promoted the understanding of the regulatory mechanism of GSDME cleavage. Our study might provide a potential therapeutic target for HRI-AKI, and will be helpful for the risk evaluation of cobalt exposure.
钴是一种环境中大量存在的过渡元素。除了直接缺氧应激外，钴离子还通过缺氧复氧损伤（HRI）间接诱导急性肾损伤（AKI），这是一种以近端肾小管上皮细胞（PTEC）坏死和炎症为特征的危及生命的临床综合征。细胞焦亡是一种炎症程序性细胞死亡，可能在 HRI-AKI 中发挥重要作用。然而，钴氯化物（CoCl）诱导的 HRI-AKI 是否涉及细胞焦亡尚不清楚。自噬是一种维持细胞内稳态的细胞生物学过程，它参与 AKI 中的细胞损伤，但自噬对细胞焦亡的潜在调节机制尚未完全了解。在这项研究中，我们建立了 CoCl 诱导的 HRI-AKI 的体外和体内模型，该模型采用 HK-2 细胞系和 C57BL/6J 小鼠。采用 Western blot 和免疫荧光法检测细胞焦亡相关标志物，结果表明，Gasdermin E（GSDME）介导的细胞焦亡参与了 HRI-AKI 中的细胞损伤。细胞凋亡特异性化学抑制剂 caspase 3、caspase 8 和 caspase 9 显著抑制 GSDME 介导的细胞焦亡，证实 GSDME 介导的细胞焦亡是通过 caspase 3/8/9 的激活诱导的。采用 Western blot 和免疫荧光法检测自噬小体的积累，结果表明 HRI 增加了自噬水平。通过慢病毒转染实验评价自噬对细胞凋亡和细胞焦亡的影响，敲低自噬特异性基因 atg5 和 fip200，结果表明自噬通过凋亡途径诱导 GSDME 介导的细胞焦亡在 HRI-AKI 中。我们的结果揭示了 GSDME 介导的细胞焦亡在 CoCl 诱导的 HRI-AKI 中的作用，并促进了对 GSDME 切割调节机制的理解。我们的研究可能为 HRI-AKI 提供了一个潜在的治疗靶点，并将有助于钴暴露风险评估。
