Nuclear factor erythroid 2-related factor 2 (Nrf2) and hypoxia-inducible factor-1a (HIF1a) transcription factors protect against ischemic acute kidney injury (AKI) by upregulating metabolic and cytoprotective gene expression. In this study, we tested the hypothesis that Nrf2 is required for HIF1a-mediated hypoxic responses using Nrf2-sufficient (wild-type) and Nrf2-deficient (Nrf2-/-) primary murine renal/kidney tubular epithelial cells (RTECs) and human immortalized tubular epithelial cells (HK2 cells) with HIF1 inhibition and activation. The HIF1 pathway inhibitor digoxin blocked hypoxia-stimulated HIF1a activation and heme oxygenase (HMOX1) expression in HK2 cells. Hypoxia-mimicking cobalt (II) chloride-stimulated HMOX1 expression was significantly lower in Nrf2-/- RTECs than in wild-type counterparts. Similarly, hypoxia-stimulated HIF1a-dependent metabolic gene expression was markedly impaired in Nrf2-/- RTECs. Nrf2 deficiency impaired hypoxia-induced HIF1a stabilization independent of increased prolyl 4-hydroxylase gene expression. We found decreased HIF1a mRNA levels in Nrf2-/- RTECs under both normoxia and hypoxia-reoxygenation conditions. In silico analysis and chromatin immunoprecipitation assays demonstrated Nrf2 binding to the HIF1a promoter in normoxia, but its binding decreased in hypoxia-exposed HK2 cells. However, Nrf2 binding at the HIF1a promoter was enriched following reoxygenation, demonstrating that Nrf2 maintains constitutive HIF1a expression. Consistent with this result, we found decreased levels of Nrf2 in hypoxia and that were restored following reoxygenation. Inhibition of mitochondrial complex I prevented hypoxia-induced Nrf2 downregulation and also increased basal Nrf2 levels. These results demonstrate a crucial role for Nrf2 in optimal HIF1a activation in hypoxia and that mitochondrial signaling downregulates Nrf2 levels in hypoxia, whereas reoxygenation restores it. Nrf2 and HIF1a interact to provide optimal metabolic and cytoprotective responses in ischemic AKI.
- Acute kidney injury
- Hypoxia-inducible factor-1a
- Nuclear factor erythroid 2-related factor 2
ASJC Scopus subject areas