Distinct subpopulations of FOXD1 stroma-derived cells regulate renal erythropoietin
Hanako Kobayashi, Qingdu Liu, Thomas C. Binns, Andres A. Urrutia, Olena Davidoff, Pinelopi P. Kapitsinou, Andrew S. Pfaff, Hannes Olauson, Annika Wernerson, Agnes B. Fogo, Guo-Hua Fong, Kenneth W. Gross, Volker H. Haase
Hanako Kobayashi, Qingdu Liu, Thomas C. Binns, Andres A. Urrutia, Olena Davidoff, Pinelopi P. Kapitsinou, Andrew S. Pfaff, Hannes Olauson, Annika Wernerson, Agnes B. Fogo, Guo-Hua Fong, Kenneth W. Gross, Volker H. Haase
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Research Article Nephrology

Distinct subpopulations of FOXD1 stroma-derived cells regulate renal erythropoietin

Abstract

Renal peritubular interstitial fibroblast-like cells are critical for adult erythropoiesis, as they are the main source of erythropoietin (EPO). Hypoxia-inducible factor 2 (HIF-2) controls EPO synthesis in the kidney and liver and is regulated by prolyl-4-hydroxylase domain (PHD) dioxygenases PHD1, PHD2, and PHD3, which function as cellular oxygen sensors. Renal interstitial cells with EPO-producing capacity are poorly characterized, and the role of the PHD/HIF-2 axis in renal EPO-producing cell (REPC) plasticity is unclear. Here we targeted the PHD/HIF-2/EPO axis in FOXD1 stroma-derived renal interstitial cells and examined the role of individual PHDs in REPC pool size regulation and renal EPO output. Renal interstitial cells with EPO-producing capacity were entirely derived from FOXD1-expressing stroma, and Phd2 inactivation alone induced renal Epo in a limited number of renal interstitial cells. EPO induction was submaximal, as hypoxia or pharmacologic PHD inhibition further increased the REPC fraction among Phd2–/– renal interstitial cells. Moreover, Phd1 and Phd3 were differentially expressed in renal interstitium, and heterozygous deficiency for Phd1 and Phd3 increased REPC numbers in Phd2–/– mice. We propose that FOXD1 lineage renal interstitial cells consist of distinct subpopulations that differ in their responsiveness to Phd2 inactivation and thus regulation of HIF-2 activity and EPO production under hypoxia or conditions of pharmacologic or genetic PHD inactivation.

Authors

Hanako Kobayashi, Qingdu Liu, Thomas C. Binns, Andres A. Urrutia, Olena Davidoff, Pinelopi P. Kapitsinou, Andrew S. Pfaff, Hannes Olauson, Annika Wernerson, Agnes B. Fogo, Guo-Hua Fong, Kenneth W. Gross, Volker H. Haase

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Figure 4

Epo induction in Foxd1-Phd2–/– mutants is submaximal.

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Epo induction in Foxd1-Phd2–/– mutants is submaximal. Shown are the res...
Shown are the results of multiplex ISH for Epo and EGFP RNA using formalin-fixed, paraffin-embedded kidney tissue sections from Foxd1-mT/mG (Co) and Foxd1-mT/mG-Phd2–/– (Phd2–/–) mutant mice at baseline and after phlebotomy (n = 5 for each group). (A) Representative images of kidney tissue sections containing peritubular interstitial cells expressing Epo (red signal) and/or EGFP (green signal). Nuclei are stained with DAPI (blue signal). Scale bars: 100 μm (baseline and phlebotomy), 10 μm (high-magnification images). White arrows identify Epo-expressing cells; asterisks depict glomeruli. (B) Top left panel: Quantification of Epo-expressing cells. Shown is the absolute number of Epo+ cells per square millimeter of kidney tissue. Bottom left panel: Epo mRNA levels relative to 18S in total kidney tissue homogenates as determined by real-time PCR (n = 3–5). Top right panel: Quantification of EGFP-expressing cells. Shown is the absolute number of EGFP+ cells per square millimeter. Bottom right panel: Fraction of Epo-expressing cells among EGFP+ cells. Data are represented as mean ± SEM; 2-way ANOVA with post hoc Tukey’s test; *P < 0.05, **P < 0.01, ***P < 0.001.