Erythropoietin Synthesis in Renal Myofibroblasts Is Restored by Activation of Hypoxia Signaling

Abstract

Erythropoietin (Epo) is produced by renal Epo-producing cells (REPs) in a hypoxia-inducible manner. The conversion of REPs into myofibroblasts and coincident loss of Epo-producing ability are the major cause of renal fibrosis and anemia. However, the hypoxic response of these transformed myofibroblasts remains unclear. Here, we used complementary in vivo transgenic and live imaging approaches to better understand the importance of hypoxia signaling in Epo production. Live imaging of REPs in transgenic mice expressing green fluorescent protein from a modified Epo-gene locus revealed that healthy REPs tightly associated with endothelium by wrapping processes around capillaries. However, this association was hampered in states of renal injury-induced inflammation previously shown to correlate with the transition to myofibroblast-transformed renal Epo-producing cells (MF-REPs). Furthermore, activation of hypoxia-inducible factors (HIFs) by genetic inactivation of HIF-prolyl hydroxylases (PHD1, PHD2, and PHD3) selectively in Epo-producing cells reactivated Epo production in MF-REPs. Loss of PHD2 in REPs restored Epo-gene expression in injured kidneys but caused polycythemia. Notably, combined deletions of PHD1 and PHD3 prevented loss of Epo expression without provoking polycythemia. Mice with PHD-deficient REPs also showed resistance to LPS-induced Epo repression in kidneys, suggesting that augmented HIF signaling counterbalances inflammatory stimuli in regulation of Epo production. Thus, augmentation of HIF signaling may be an attractive therapeutic strategy for treating renal anemia by reactivating Epo synthesis in MF-REPs.

Distinct subpopulations of FOXD1 stroma-derived cells regulate renal erythropoietin

“…In contrast, a larger proportion of cortical and outer medullary EGFP + Epo -cells expressed the P0 lineage (14), it is plausible that non-P0 lineage interstitial cells with EPO-producing capacity are derived from FOXD1 stromal cells that existed prior to E13.5. PDGFRB is a well-recognized marker of both pericytes and REPCs, and recent imaging studies support the notion that renal pericytes are capable of synthesizing EPO (28). In our genetic model of Foxd1-Cre-mediated Phd2 inactivation we detected Pdgfrb transcripts in approximately 80% of Epo-expressing interstitial cells, suggesting that PDGFRB may not be an exclusive marker of REPCs.…”

“…This discrepancy is explained by the very small number of renal interstitial cells targeted with the use of the Epo-Cre transgene. The ability of PHD inhibition to protect the kidney's EPOproducing ability under injury conditions suggests that pharmacologic HIF activation increases EPO synthesis in fibrotic kidneys either by converting non-EPO-producing renal interstitial cells into REPCs, or, as proposed by Souma and colleagues, by maintaining Epo transcription in EPO-producing cells that are undergoing myofibroblast transformation (28). The possibility that even severely diseased kidneys do not completely lose their ability to synthesize EPO in response to HIF-2 activation has been suggested based on studies in CKD patients whose serum EPO levels increased upon ascent to high altitude or following treatment with HIF-2α-stabilizing compounds (30)(31)(32).…”

“…The reduced ability to mount an adequate EPO response appears to be reversible to a certain degree, as treatment with glucocorticoids or removal of fibrotic stimuli in experimental fibrosis models protected the EPO-producing ability of the injured kidney (14,29). In a recent genetic study in mice, Souma and colleagues showed that PHD inactivation in Epo-expressing cells was capable of reversing the decline of renal EPO production in a unilateral ureteral obstruction model (28). This study made use of a Cre-recombinase transgene that was controlled by Epo regulatory elements and thus was oxygenregulated, i.e., inducible by hypoxia.…”

Hypoxia and hypoxia-inducible factors in chronic kidney disease

“…In contrast, the expression of HIF-2α in the hypoxic kidney is limited to the endothelial and interstitial cells. Recent studies have revealed that renal erythropoietin (EPO) is produced in fibroblast-like cells in the interstitium in a HIF-2α-dependent manner [17][18][19].…”

The HIF System Response to ESA Therapy in CKD‐Anemia

“…Moreover, the pharmacological inhibition of the PHD in CKD patients stimulates endogenous EPO production further supporting a deranged oxygen sensing [27]. A recent study in mice by Souma et al [48] also strengthened this hypothesis, by showing that inflammatory cytokines and/or fibrosis factors suppress HIF activation through the over-activation of PHD even under pathologic hypoxic conditions, and that the inhibition of PHD restores EPO production.…”