Involvement of phosphatase and tensin homolog-induced putative kinase 1–Parkin-mediated mitophagy in septic acute kidney injury / 中华医学杂志(英文版)

ABSTRACT

Background@#Studies have reported mitophagy activation in renal tubular epithelial cells (RTECs) in acute kidney injury (AKI). Phosphatase and tensin homolog-induced putative kinase 1 (PINK1) and E3 ubiquitin-protein ligase Parkin are involved in mitophagy regulation; however, little is known about the role of PINK1-Parkin mitophagy in septic AKI. Here we investigated whether the PINK1-Parkin mitophagy pathway is involved in septic AKI and its effects on cell apoptosis in vitro and on renal functions in vivo.@*Methods@#Mitophagy-related gene expression was determined using Western blot assay in human RTEC cell line HK-2 stimulated with bacterial lipopolysaccharide (LPS) and in RTECs from septic AKI rats induced by cecal ligation and perforation (CLP). Autophagy-related ultrastructural features in rat RTECs were observed using electron microscopy. Gain- and loss-of-function approaches were performed to investigate the role of the PINK1-Parkin pathway in HK-2 cell mitophagy. Autophagy activators and inhibitors were used to assess the effects of mitophagy modulation on cell apoptosis in vitro and on renal functions in vivo.@*Results@#LPS stimulation could significantly induce LC3-II and BECN-1 protein expression (LC3-II 1.72 ± 0.05 vs. 1.00 ± 0.05, P < 0.05; BECN-1 5.33 ± 0.57 vs. 1.00 ± 0.14, P < 0.05) at 4 h in vitro. Similarly, LC3-II, and BECN-1 protein levels were significantly increased and peaked at 2 h after CLP (LC3-II 3.33 ± 0.12 vs. 1.03 ± 0.15, P < 0.05; BECN-1 1.57 ± 0.26 vs. 1.02 ± 0.11, P < 0.05) in vivo compared with those after sham operation. Mitochondrial deformation and mitolysosome-mediated mitochondria clearance were observed in RTECs from septic rats. PINK1 knockdown significantly attenuated LC3-II protein expression (1.35 ± 0.21 vs. 2.38 ± 0.22, P < 0.05), whereas PINK1 overexpression markedly enhanced LC3-II protein expression (2.07 ± 0.21 vs. 1.29 ± 0.19, P < 0.05) compared with LPS-stimulated HK-2 cells. LPS-induced proapoptotic protein expression remained unchanged in autophagy activator-treated HK-2 cells and was significantly attenuated in PINK1-overexpressing cells, but was remarkably upregulated in autophagy inhibitor-treated and in PINK1-depleted cells. Consistent results were observed in flow cytometric apoptosis assay and in renal function indicators in rats.@*Conclusion@#PINK1-Parkin-mediated mitophagy might play a protective role in septic AKI, serving as a potential therapeutic target for septic AKI.