Expression and actions of HIF prolyl-4-hydroxylase in the rat kidneys

Abstract

Hypoxia inducible factor (HIF) prolyl-4-hydroxylase domain-containing proteins (PHDs) promote the degradation of HIF-1␣. Because HIF-1␣ is highly expressed in the renal medulla and HIF-1␣-targeted genes such as nitric oxide synthase, cyclooxygenase, and heme oxygenase are important in the regulation of renal medullary function, we hypothesized that PHD regulates HIF-1␣ levels in the renal medulla and, thereby, participates in the control of renal Na ϩ excretion. Using real-time RT-PCR, Western blot, and immunohistochemical analyses, we have demonstrated that all three isoforms of PHD, PHD1, PHD2, and PHD3, are expressed in the kidneys and that PHD2 is the most abundant isoform. Regionally, all PHDs exhibited much higher levels in renal medulla than cortex. A furosemide-induced increase in renal medullary tissue PO2 significantly decreased PHD levels in renal medulla, whereas hypoxia significantly increased mRNA levels of PHDs in cultured renal medullary interstitial cells, indicating that O2 regulates PHDs. Functionally, the PHD inhibitor L-mimosine (L-Mim, 50 mg ⅐ kg Ϫ1 ⅐ day Ϫ1 ip for 2 wk) substantially upregulated HIF-1␣ expression in the kidneys, especially in the renal medulla, and remarkably enhanced (by Ͼ80%) the natriuretic response to renal perfusion pressure in Sprague-Dawley rats. Inhibition of HIF transcriptional activity by renal medullary transfection of HIF-1␣ decoy oligodeoxynucleotides attenuated L-Mim-induced enhancement of pressure natriuresis, which confirmed that HIF-1␣ mediated the effect of L-Mim. These results indicate that highly expressed PHDs in the renal medulla make an important contribution to the control of renal Na ϩ excretion through regulation of HIF-1␣ and its targeted genes. fluid homeostasis; anoxia; natriuretic factor; gene transcription; renal tubules; renal hemodynamics IT IS WELL KNOWN that PO 2 is much lower in the renal medulla than in the renal cortex because of relative "underperfusion," the countercurrent O 2 diffusion between descending and ascending vasa recta, and the high metabolic ion transport activity of the thick ascending limb of Henle in the renal medullary region (6, 10). To adapt to this low-PO 2 milieu, the renal medulla has naturally developed various mechanisms to protect the cells from ischemic and hypoxic injury (5,6,15) and to ensure that the cells in this kidney region function normally in a hypoxic environment (6, 10). Increase in tissue blood perfusion to facilitate the O 2 supply and inhibition of tubular metabolic activity to decrease O 2 demands (5,6,15,53) are the common outcomes of various adaptive actions induced by different renal medullary factors such as nitric oxide (NO), prostaglandins, and heme oxygenase (HO) products. Hypoxia inducible factor (HIF)-1␣, the master regulator of adaptation to hypoxia, has been shown to activate gene transcription of many O 2 -sensitive genes such as NO synthase (NOS), cyclooxygenase (COX), HO, and vascular endothelial growth factor (21,25,26,32,38,45,50,53). The participation of the products controlled...

Hypoxia-inducible Factors and the Prevention of Acute Organ Injury

“…However, the threshold and extent of HIF activation may depend on the hypoxic stimulus and cell type involved. To some extent, these cellular variations may refl ect diff erent expression of various PHD isoforms in diff erent tissues [5][6][7]. As HIF stimulation may potentiate hypoxia tolerance, studies were conducted to explore its clinical application.…”

“…Diverse characteristics and distribution patterns of diff erent PHDs [5][6][7] and particular actions of various PHD inhibitors [11,37] might enable selective manipulation of the HIF system in a more desired way, selectively favoring advantageous HIF-dependent responses in preferred tissues. Furthermore, it is believed that activation of adverse responses requires protracted HIF stimulation, whereas short-term and transient HIF activa tion might suffi ce to activate tissue-protective systems without continuing induction of harmful systems.…”

miR-29c is downregulated in renal interstitial fibrosis in humans and rats and restored by HIF-α activation

“…In the presence of oxygen, two ␣-prolyl residues undergo enzymatic hydroxylation, which is required for its proteasomal degradation. Pharmacological activation of HIF-␣ could be achieved in an oxygen-independent manner by using prolyl 4-hydroxylase (PHD) inhibitors such as L-mimosine (L-Mim) (23).…”

Morg1 heterozygous mice are protected from acute renal ischemia-reperfusion injury

“…The differential expression of PHD1-3 has been recently studied in rat kidneys (17,38). All three PHDs are expressed in tubules, where PHD2 is the most abundant isoform (17,38).…”