Hyperbaric oxygen preconditioning and the role of NADPH oxidase inhibition in postischemic acute kidney injury induced in spontaneously hypertensive rats

Abstract

Renal ischemia/reperfusion injury is a common cause of acute kidney injury (AKI) and hypertension might contribute to the increased incidence of AKI. The purpose of this study was to investigate the effects of single and combined hyperbaric oxygen (HBO) preconditioning and NADPH oxidase inhibition on oxidative stress, kidney function and structure in spontaneously hypertensive rats (SHR) after renal ischemia reperfusion injury. HBO preconditioning was performed by exposing to pure oxygen (2.026 bar) twice a day for two consecutive days for 60 minutes, and 24h before AKI induction. For AKI induction, the right kidney was removed and ischemia was performed by clamping the left renal artery for 45 minutes. NADPH oxidase inhibition was induced by apocynin (40 mg/kg b.m., intravenously) 5 minutes before reperfusion. AKI significantly increased renal vascular resistance and reduced renal blood flow, which were significantly improved after apocynin treatment. Also, HBO preconditioning, with or without apocynin treatment showed improvement on renal hemodynamics. AKI significantly increased plasma creatinine, urea, phosphate levels and lipid peroxidation in plasma. Remarkable improvement, with decrease in creatinine, urea and phosphate levels was observed in all treated groups. HBO preconditioning, solitary or with apocynin treatment decreased lipid peroxidation in plasma caused by AKI induction. Also, combined with apocynin, it increased catalase activity and solitary, glutathione reductase enzyme activity in erythrocytes. While AKI induction significantly increased plasma KIM- 1 levels, HBO preconditioning, solitary or with apocynin decreased its levels. Considering renal morphology, significant morphological alterations present after AKI induction were significantly improved in all treated groups with reduced tubular dilatation, tubular necrosis in the cortico-medullary zone and PAS positive cast formation. Our results reveal that NADPH oxidase inhibition and hyperbaric oxygen preconditioning, with or without NADPH oxidase inhibition may have beneficial effects, but their protective role should be evaluated in further studies.

Fig 1. Renal hemodynamic parameters.

RVR-renal vascular resistance, RBF-renal blood flow. ***p<0.001 vs. SHAM group, #p<0.05, ###p<0.001 vs. AKI group.

Fig 2. Plasma creatinine (PCr), urea (PU) and phosphate (PPhos) concentration 24 h after reperfusion.

***p<0.001 vs. SHAM group, #p<0.05, ##p<0.01, ###p<0.001 vs. AKI group.

Fig 3. Plasma TBARS levels 24 hours after reperfusion.

**p<0.01 vs. SHAM group, #p<0.05, ##p<0.01vs. AKI group.

Fig 4. Erythrocytes catalase (CAT), superoxide dismutase (SOD), glutathione peroxidase (GSH-Px) and glutathione reductase (GR) activity 24 hours after reperfusion.

*p<0.05,***p<0.001 vs. SHAM, #p<0.05, ##p<0.01 vs. AKI.

Fig 5. Plasma KIM—1 levels 24h after reperfusion.

*** p<0.001 vs. SHAM, # p<0.05 vs. AKI.

Fig 6. Histopathology of the representative kidney samples collected in different experimental groups (PAS staining, x 20 magnification): Normal morphology of renal tissue including glomerular and tubulointerstitial compartments in the sham-operated animals (A), proximal tubular dilatation (arrows), necrosis of tubular epithelial cells (*) and PAS-positive casts (**) in the AKI group (B), moderately intensive tubular necrosis, reduced tubular dilatation and less number of PAS-positive casts in AKI+APO (C), AKI+HBO (D), AKI+APO+HBO (E) groups and histopathological score as a sum of present morphological changes (F).

***p<0.001 vs. SHAM, ##p<0.01, ###p<0.001 vs. AKI.