Activation of the Hypoxia-Inducible Factor Pathway Inhibits Epithelial Sodium Channel–Mediated Sodium Transport in Collecting Duct Principal Cells

Abstract

Background Active sodium reabsorption is the major factor influencing renal oxygen consumption and production of reactive oxygen species (ROS). Increased sodium reabsorption uses more oxygen, which may worsen medullary hypoxia and produce more ROS via enhanced mitochondrial ATP synthesis. Both mechanisms may activate the hypoxiainducible factor (HIF) pathway. Because the collecting duct is exposed to low oxygen pressure and variations of active sodium transport, we assessed whether the HIF pathway controls epithelial sodium channel (ENaC)-dependent sodium transport. Methods We investigated HIF's effect on ENaC expression in mpkCCDcl4 cells (a model of collecting duct principal cells) using real-time PCR and Western blot and ENaC activity by measuring amiloride-sensitive current. We also assessed the effect of hypoxia and sodium intake on abundance of kidney sodium transporters in wild-type and inducible kidney tubule-specific Hif1α knockout mice. Results In cultured cells, activation of the HIF pathway by dimethyloxalylglycine or hypoxia inhibited sodium transport and decreased expression of βENaC and γENaC, as well as of Na,K-ATPase. HIF1α silencing increased βENaC and γENaC expression and stimulated sodium transport. A constitutively active mutant of HIF1α produced the opposite effect. Aldosterone and inhibition of the mitochondrial respiratory chain slowly activated the HIF pathway, suggesting that ROS may also activate HIF. Decreased γENaC abundance induced by hypoxia in normal mice was abolished in Hif1α knockout mice. Similarly, Hif1α knockout led to increased γENaC abundance under high sodium intake. Conclusions This study reveals that γENaC expression and activity are physiologically controlled by the HIF pathway, which may represent a negative feedback mechanism to preserve oxygenation and/or prevent excessive ROS generation under increased sodium transport.

Analysis of the Hypoxic Response in a Mouse Cortical Collecting Duct-Derived Cell Line Suggests That Esrra Is Partially Involved in Hif1α-Mediated Hypoxia-Inducible Gene Expression in mCCDcl1 Cells

“…RT-qPCR results confirmed that these genes were indeed significantly regulated by hypoxia (Figure 2). Furthermore, the mRNA expression levels of Scnn1a (αENaC) and Scnn1b (βENaC) were significantly reduced following hypoxia, consistent with the RNAseq data, confirming the observations of Dizin and colleagues [17].…”

“…In this study, we provided evidence that mCCDcl1 cells respond to hypoxia in a Hif-1-dependent manner. Along with the study of Dizin and colleagues, who showed that activation of the Hif pathway leads to a reduction in ENaC and of Na + ,K + -ATPase expression in mpkCCDcl4 cells [17], this raises the possibility that cells of the aldosterone-sensitive distal nephron are responsive to hypoxia overall. It is well described that the glomeruli, proximal tubules, and thick ascending limb of Henle's loop are the most sensitive to hypoxia given their high energy consumption rates [21]; however, it is intriguing that aldosterone levels decrease following acute hypoxia, while most studies focusing on renin secretion have reported a rapid rise in plasmatic and renal renin levels, leading to a temporary uncoupling between aldosterone and renin [22,23].…”

“…It is highly likely that the decrease in aldosterone, together with increased natriuresis upon hypoxia, is an energy-sparing mechanism to reduce Na + ,K + -ATPase-mediated ATP consumption within the cortical collecting duct, in order to protect the more sensitive parts of the nephron from insufficient energy supply [17]. In line with this hypothesis, the levels of the Na + ,K + -ATPase were also reduced in the transcriptome of mCCD cl1 cells following hypoxia (Table S1).…”

“…Interestingly, for decades, numerous studies have consistently reported a decrease in aldosterone secretion following hypoxia [11][12][13][14][15][16]. A recent study by Dizin and colleagues [17] that was conducted in the cortical collecting duct mpkCCD cl4 cell line demonstrated that the activation of the HIF pathway following hypoxia or dimethyloxalylglycine treatment leads to decreased sodium transport and ENaC expression. Similar results were obtained following the induction of reactive oxygen species (ROS).…”

“…The effects of HIF-1α on ENaC and Na + , K + -ATPase are likely indirect however, since no putative HIF binding site was found in the proximal regulatory regions of their genes. The authors suggested that the inhibition of ENaCmediated sodium transport may prevent increased ATP-and thus oxygen-consumption via the basolateral Na + , K + -ATPase under hypoxic conditions [17].…”