Abstract
Autophagy is an important renal-protective mechanism in septic acute kidney injury (AKI). Receptor interacting protein kinase 3 (RIP3) has been implicated in the renal tubular injury and renal dysfunction during septic AKI. Here we investigated the role and mechanism of RIP3 on autophagy in septic AKI. We showed an activation of RIP3, accompanied by an accumulation of the autophagosome marker LC3II and the autophagic substrate p62, in the kidneys of lipopolysaccharide (LPS)-induced septic AKI mice and LPS-treated cultured renal proximal tubular epithelial cells (PTECs). The lysosome inhibitor did not further increase the levels of LCII or p62 in LPS-treated PTECs. Moreover, inhibition of RIP3 attenuated the aberrant accumulation of LC3II and p62 under LPS treatment in vivo and in vitro. By utilizing mCherry-GFP-LC3 autophagy reporter mice in vivo and PTECs overexpression mRFP-GFP-LC3 in vitro, we observed that inhibition of RIP3 restored the formation of autolysosomes and eliminated the accumulated autophagosomes under LPS treatment. These results indicated that RIP3 impaired autophagic degradation, contributing to the accumulation of autophagosomes. Mechanistically, the nuclear translocation of transcription factor EB (TFEB), a master regulator of the lysosome and autophagy pathway, was inhibited in LPS-induced mice and LPS-treated PTECs. Inhibition of RIP3 restored the nuclear translocation of TFEB in vivo and in vitro. Co-immunoprecipitation further showed an interaction of RIP3 and TFEB in LPS-treated PTECs. Also, the expression of LAMP1 and cathepsin B, two potential target genes of TFEB involved in lysosome function, were decreased under LPS treatment in vivo and in vitro, and this decrease was rescued by inhibiting RIP3. Finally, overexpression of TFEB restored the autophagic degradation in LPS-treated PTECs. Together, the present study has identified a pivotal role of RIP3 in suppressing autophagic degradation through impeding the TFEB-lysosome pathway in septic AKI, providing potential therapeutic targets for the prevention and treatment of septic AKI.
Highlights
Sepsis is the leading cause of acute kidney injury (AKI)in critically ill patients, characterized by a rapid decrease of renal function accompanied by multiorgan dysfunctionPathologically, one of the major features of septic AKI is sublethal and lethal injury of renal tubular cells[6]
Receptor interacting protein kinase 3 (RIP3) has recently been identified to play a pivotal role in tubular injury and renal dysfunction in septic AKI8
The present study has demonstrated that RIP3 was activated to suppress autophagic degradation in septic AKI mice and in LPS-treated cultured proximal tubular epithelial cells (PTECs)
Summary
Sepsis is the leading cause of acute kidney injury (AKI)in critically ill patients, characterized by a rapid decrease of renal function accompanied by multiorgan dysfunctionPathologically, one of the major features of septic AKI is sublethal and lethal injury of renal tubular cells[6]. Our recent study showed that a RIP3-selective inhibitor attenuated renal tubular apoptosis and renal dysfunction in LPS-induced septic AKI mice[9]. Sureshbabu et al.[8], using proximal tubule-specific RIP3-knockout mice, showed an alleviation of tubular injury and restoration of renal function in septic AKI mice, further demonstrating the pathogenetic role of RIP3 in septic AKI. It is still unknown how RIP3 mediates nephrotoxicity and how RIP3 inhibitors protect the kidneys during septic AKI. Our previous study suggested RIP1 inhibited autophagy in LPS-induced cultured renal tubular cells[10]. We aimed to investigate the role of RIP3 in regulating autophagy during septic AKI
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