Glutathione S-Transferase and Clusterin, New Players in the Ischemic Preconditioning Renal Protection in a Murine Model of Ischemia and Reperfusion

Abstract

BACKGROUND/AIMS: Renal ischemia and reperfusion injury (IRI) involves oxidative stress, disruption of microvasculature due to endothelial cell damage, loss of epithelial cell polarity secondary to cytoskeletal alterations, inflammation, and the subsequent transition into a mesenchymal phenotype. Ischemic preconditioning (IPC) has been proposed as a therapeutic strategy to avoid/ameliorate the IRI. Since previous results showed that IPC could have differential effects in kidney cortex vs. kidney medulla, in the present study we analyzed the effectiveness and molecular mechanisms implicated in IPC in both kidney regions. METHODS: We evaluated 3 experimental groups of BALB/c male mice: control (sham surgery); renal ischemia (30 min) by bilateral occlusion of the renal pedicle and reperfusion (48 hours) (I/R); and renal IPC (two cycles of 5 min of ischemia and 5 min of reperfusion) applied just before I/R. Acute kidney injury was evaluated by glomerular filtration rate (GFR), Neutrophil Gelatinase-Associated Lipocalin (NGAL) blood level, and histologic analysis. Oxidative stress was studied measurement the Glutathione S-Transferase (GST) activity, GSH/GSSG ratio, and lipoperoxidation levels. Inflammatory mediators (IL-1β, IL-6, Foxp3, and IL-10) were quantified by qRT-PCR. The endothelial (PECAM-1), epithelial (AQP-1), mesenchymal (Vimentin, Fascin, and Hsp47), iNOS, clusterin, and Hsp27 expression were evaluated (qRT-PCR and/or Western blot). RESULTS: The IPC protocol prevented the decrease of GFR, reduced the plasma NGAL, and ameliorated morphological damage in the kidney cortex after I/R. The IPC also prevented the downregulation of GST activity, lipoperoxidation and ameliorated the oxidized glutathione. In addition, IPC prevented the upregulation of vimentin, fascin, and Hsp47, which was associated with the prevention of the downregulation of AQP1 after I/R. The protective effect of IPC was associated with the upregulation of Hsp27, Foxp3, and IL-10 expression in the renal cortex. However, the upregulation of iNOS, IL-1β, IL-6, and clusterin by I/R were not modified by IPC. CONCLUSION: IPC conferred better protection in the kidney cortex as compared to the kidney medulla. The protective effect of IPC was associated with amelioration of oxidative stress, tubular damage, and the induction of markers of Treg lymphocytes activity in the cortical region. Further studies are needed to evaluate if lower tubular cell stress/damage after I/R may explain the preferential induction of Treg response in the kidney cortex induced by IPC.

Early loss of glutathione ‐s‐ transferase (GST) activity during diverse forms of acute renal tubular injury

“…Second, while it is universally recognized that GST mediates cellular protection via GSH conjugation to free radicals and xenobiotics, we did not confirm that free fatty acid‐induced GST inhibition directly contributes to tubular cell injury (given that this was not the goal of this study). However, it is notable that GST induction has been reported to confer renal protective effects (Pasten et al, 2021 ), and that provision of the GST substrate, GSH, also mitigates experimental AKI (e.g., ref (Paller, 1998 ; Sharma et al, 2004 ). Thus, the literature does imply that fatty acid‐induced GST inhibition could play an injury promoting role.…”

Advancing Renal Amyloidosis Care: The Role of Modern Diagnostic Techniques with the Potential of Enhancing Patient Outcomes

“…Glutathione-S transferase (GST) and phosphatidylethanolamine-binding protein 1 (PEB1) showed the highest upregulation in the proteome of early-stage amyloidosis. Both function as antioxidants, and PEB1 plays a key role in regulating the ferroptosis pathway [ 26 , 27 ]. Finally, the accumulation behavior of amyloidogenic proteins is independent of other proteins, suggesting that kidney toxicity in this condition cannot be solely attributed to amyloid levels.…”