Background: Ischemia-reperfusion (IR) injury is the main cause of delayed graft function in solid organ transplantation. Hypoxia-inducible factors (HIFs) control the expression of genes related to preconditioning against IR injury. During normoxia, HIF-α subunits are marked for degradation by the egg-laying defective nine homolog (EGLN) family of prolyl-4-hydroxylases. The inhibition of EGLN stabilizes HIFs and protects against IR injury. The aim of this study was to determine whether the EGLN inhibitors sodium (S)-2-hydroxyglutarate [(S)-2HG] and succinic acid (SA) have a nephroprotective effect against renal IR injury in Wistar rats.
Methods: (S)-2HG was synthesized in a 22.96% yield from commercially available L-glutamic acid in a two-step methodology (diazotization/alkaline hydrolysis), and its structure was confirmed by nuclear magnetic resonance and polarimetry. SA was acquired commercially. (S)-2HG and SA were independently evaluated in male and female Wistar rats respectively after renal IR injury. Rats were divided into the following groups: sham (SH), nontoxicity [(S)-2HG: 12.5 or 25 mg/kg; SA: 12.5, 25, or 50 mg/kg], IR, and compound+IR [(S)-2HG: 12.5 or 25 mg/kg; SA: 12.5, 25, or 50 mg/kg]; independent SH and IR groups were used for each assessed compound. Markers of kidney injury (BUN, creatinine, glucose, and uric acid) and liver function (ALT, AST, ALP, LDH, serum proteins, and albumin), proinflammatory cytokines (IL-1β, IL-6, and TNF-α), oxidative stress biomarkers (malondialdehyde and superoxide dismutase), and histological parameters (tubular necrosis, acidophilic casts, and vascular congestion) were assessed. Tissue HIF-1α was measured by ELISA and Western blot, and the expression of Hmox1 was assessed by RT-qPCR.
Results: (S)-2HG had a dose-dependent nephroprotective effect, as evidenced by a significant reduction in the changes in the BUN, creatinine, ALP, AST, and LDH levels compared with the IR group. Tissue HIF-1α was only increased in the IR group compared to SH; however, (S)-2HG caused a significant increase in the expression of Hmox1, suggesting an early accumulation of HIF-1α in the (S)-2HG-treated groups. There were no significant effects on the other biomarkers. SA did not show a nephroprotective effect; the only changes were a decrease in creatinine level at 12.5 mg/kg and increased IR injury at 50 mg/kg. There were no effects on the other biochemical, proinflammatory, or oxidative stress biomarkers.
Conclusion: None of the compounds were hepatotoxic at the tested doses. (S)-2HG showed a dose-dependent nephroprotective effect at the evaluated doses, which involved an increase in the expression of Hmox1, suggesting stabilization of HIF-1α. SA did not show a nephroprotective effect but tended to increase IR injury when given at high doses.
Bibliographical noteFunding Information:
This study was financially supported by resources of the Liver Unit and the Department of Analytical Chemistry and did not receive any specific grant from funding agencies in the public, commercial, or not-for-profit sectors. Eduardo Cienfuegos-Pecina received a scholarship from the National Council of Science and Technology (CONACYT) through the project 2017-01-5652. There was no additional external funding received for this study. The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript.
Copyright 2020 Cienfuegos-Pecina et al.
All Science Journal Classification (ASJC) codes
- Biochemistry, Genetics and Molecular Biology(all)
- Agricultural and Biological Sciences(all)