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Original Article | Open Access

Osteocyte Egln1/Phd2 links oxygen sensing and biomineralization via FGF23

Megan L. Noonan1, Pu Ni1, Emmanuel Solis1, Yamil G. Marambio1, Rafiou Agoro1, Xiaona Chu1, Yue Wang1, Hongyu Gao1, Xiaoling Xuei1, Erica L. Clinkenbeard1, Guanglong Jiang1, Sheng Liu1, Steve Stegen2, Geert Carmeliet2, William R. Thompson3,4, Yunlong Liu1,5, Jun Wan1, Kenneth E. White1,6( )
Department of Medical and Molecular Genetics, Indiana University School of Medicine, Indianapolis, IN 46202, USA
Laboratory of Clinical and Experimental Endocrinology, Department of Chronic Diseases and Metabolism, KU Leuven, 3000 Leuven, Belgium
Department of Physical Therapy, Indiana University School of Medicine, Indianapolis, IN 46202, USA
Department of Anatomy, Cell Biology, and Physiology, Indiana University School of Medicine, Indianapolis, IN 46202, USA
Center for Computational Biology and Bioinformatics, Indiana University School of Medicine, Indianapolis, IN 46202, USA
Departments of Medicine/Division of Nephrology, Indiana University School of Medicine, Indianapolis, IN 46202, USA
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Abstract

Osteocytes act within a hypoxic environment to control key steps in bone formation. FGF23, a critical phosphate-regulating hormone, is stimulated by low oxygen/iron in acute and chronic diseases, however the molecular mechanisms directing this process remain unclear. Our goal was to identify the osteocyte factors responsible for FGF23 production driven by changes in oxygen/iron utilization. Hypoxia-inducible factor-prolyl hydroxylase inhibitors (HIF-PHI) which stabilize HIF transcription factors, increased Fgf23 in normal mice, as well as in osteocyte-like cells; in mice with conditional osteocyte Fgf23 deletion, circulating iFGF23 was suppressed. An inducible MSC cell line (‘MPC2’) underwent FG-4592 treatment and ATACseq/RNAseq, and demonstrated that differentiated osteocytes significantly increased HIF genomic accessibility versus progenitor cells. Integrative genomics also revealed increased prolyl hydroxylase Egln1 (Phd2) chromatin accessibility and expression, which was positively associated with osteocyte differentiation. In mice with chronic kidney disease (CKD), Phd1-3 enzymes were suppressed, consistent with FGF23 upregulation in this model. Conditional loss of Phd2 from osteocytes in vivo resulted in upregulated Fgf23, in line with our findings that the MPC2 cell line lacking Phd2 (CRISPR Phd2-KO cells) constitutively activated Fgf23 that was abolished by HIF1α blockade. In vitro, Phd2-KO cells lost iron-mediated suppression of Fgf23 and this activity was not compensated for by Phd1 or −3. In sum, osteocytes become adapted to oxygen/iron sensing during differentiation and are directly sensitive to bioavailable iron. Further, Phd2 is a critical mediator of osteocyte FGF23 production, thus our collective studies may provide new therapeutic targets for skeletal diseases involving disturbed oxygen/iron sensing.

References

1

Urakawa, I. et al. Klotho converts canonical FGF receptor into a specific receptor for FGF23. Nature 444, 770–774 (2006).
Crossref Google Scholar
2

Shimada, T. et al. FGF-23 is a potent regulator of vitamin D metabolism and phosphate homeostasis. J. Bone Min. Res 19, 429–435 (2004).
Crossref Google Scholar
3

Farrow, E. G. et al. Iron deficiency drives an autosomal dominant hypophosphatemic rickets (ADHR) phenotype in fibroblast growth factor-23 (Fgf23) knock-in mice. Proc. Natl. Acad. Sci. USA 108, E1146–E1155 (2011).
Crossref Google Scholar
4

David, V. et al. Inflammation and functional iron deficiency regulate fibroblast growth factor 23 production. Kidney Int. 89, 135–146 (2016).
Crossref Google Scholar
5

Spencer, J. A. et al. Direct measurement of local oxygen concentration in the bone marrow of live animals. Nature 508, 269–273 (2014).
Crossref Google Scholar
6

Harrison, J. S., Rameshwar, P., Chang, V. & Bandari, P. Oxygen saturation in the bone marrow of healthy volunteers. Blood 99, 394 (2002).
Crossref Google Scholar
7

Mohyeldin, A., Garzon-Muvdi, T. & Quinones-Hinojosa, A. Oxygen in stem cell biology: a critical component of the stem cell niche. Cell Stem Cell 7, 150–161 (2010).
Crossref Google Scholar
8

Rankin, E. B., Giaccia, A. J. & Schipani, E. A central role for hypoxic signaling in cartilage, bone, and hematopoiesis. Curr. Osteoporos. Rep. 9, 46–52 (2011).
Crossref Google Scholar
9

Hirao, M. et al. Oxygen tension is an important mediator of the transformation of osteoblasts to osteocytes. J. Bone Miner. Metab. 25, 266–276 (2007).
Crossref Google Scholar
10

Stegen, S. et al. Osteocytic oxygen sensing controls bone mass through epigenetic regulation of sclerostin. Nat. Commun. 9, 2557 (2018).
Crossref Google Scholar
11

Forsythe, J. A. et al. Activation of vascular endothelial growth factor gene transcription by hypoxia-inducible factor 1. Mol. Cell. Biol. 16, 4604–4613 (1996).
Crossref Google Scholar
12

Semenza, G. L. & Wang, G. L. A nuclear factor induced by hypoxia via de novo protein synthesis binds to the human erythropoietin gene enhancer at a site required for transcriptional activation. Mol. Cell. Biol. 12, 5447–5454 (1992).
Crossref Google Scholar
13

Imel, E. A. et al. Iron modifies plasma FGF23 differently in autosomal dominant hypophosphatemic rickets and healthy humans. J. Clin. Endocrinol. Metab. 96, 3541–3549 (2011).
Crossref Google Scholar
14

Econs, M. J. & McEnery, P. T. Autosomal dominant hypophosphatemic rickets/osteomalacia: clinical characterization of a novel renal phosphate-wasting disorder. J. Clin. Endocrinol. Metab. 82, 674–681 (1997).
Crossref Google Scholar
15

White, K. E. et al. Autosomal-dominant hypophosphatemic rickets (ADHR) mutations stabilize FGF-23. Kidney Int. 60, 2079–2086 (2001).
Crossref Google Scholar
16

Hum, J. M. et al. The metabolic bone disease associated with the Hyp mutation is independent of osteoblastic HIF1alpha expression. Bone Rep. 6, 38–43 (2017).
Crossref Google Scholar
17

Zhang, Q. et al. The hypoxia-inducible factor-1alpha activates ectopic production of fibroblast growth factor 23 in tumor-induced osteomalacia. Bone Res. 4, 16011 (2016).
Crossref Google Scholar
18

Imel, E. A., Hui, S. L. & Econs, M. J. FGF23 concentrations vary with disease status in autosomal dominant hypophosphatemic rickets. J. Bone Min. Res 22, 520–526 (2007).
Crossref Google Scholar
19

Imel, E. A. et al. Oral iron replacement normalizes fibroblast growth factor 23 in iron-deficient patients with autosomal dominant hypophosphatemic rickets. J. Bone Min. Res. 35, 231–238 (2020).
Crossref Google Scholar
20

Koh, N. et al. Severely reduced production of klotho in human chronic renal failure kidney. Biochem Biophys. Res Commun. 280, 1015–1020 (2001).
Crossref Google Scholar
21

Hu, M. C., Kuro-o, M. & Moe, O. W. Klotho and kidney disease. J. Nephrol. 23, S136–S144 (2010).
Google Scholar
22

Hu, M. C. et al. Klotho deficiency causes vascular calcification in chronic kidney disease. J. Am. Soc. Nephrol. 22, 124–136 (2011).
Crossref Google Scholar
23

Hu, M. C., Shiizaki, K., Kuro-o, M. & Moe, O. W. Fibroblast growth factor 23 and Klotho: physiology and pathophysiology of an endocrine network of mineral metabolism. Annu. Rev. Physiol. 75, 503–533 (2013).
Crossref Google Scholar
24

Babitt, J. L. & Lin, H. Y. Mechanisms of anemia in CKD. J. Am. Soc. Nephrol. 23, 1631–1634 (2012).
Crossref Google Scholar
25

Noonan, M. L. et al. Erythropoietin and a hypoxia-inducible factor prolyl hydroxylase inhibitor (HIF-PHDi) lowers FGF23 in a model of chronic kidney disease (CKD). Physiol. Rep. 8, e14434 (2020).
Crossref Google Scholar
26

Noonan, M. L. et al. The HIF-PHI BAY 85-3934 (Molidustat) improves anemia and is associated with reduced levels of circulating FGF23 in a CKD Mouse Model. J. Bone Min. Res. 36, 1117–1130 (2021).
Crossref Google Scholar
27

Faul, C. et al. FGF23 induces left ventricular hypertrophy. J. Clin. Invest. 121, 4393–4408 (2011).
Crossref Google Scholar
28

Leifheit-Nestler, M. et al. Induction of cardiac FGF23/FGFR4 expression is associated with left ventricular hypertrophy in patients with chronic kidney disease. Nephrol. Dial. Transpl. 31, 1088–1099 (2016).
Crossref Google Scholar
29

Mehta, R. et al. Iron status, fibroblast growth factor 23 and cardiovascular and kidney outcomes in chronic kidney disease. Kidney Int. 100, 1292–1302 (2021).
Crossref Google Scholar
30

Prideaux, M. et al. Generation of two multipotent mesenchymal progenitor cell lines capable of osteogenic, mature osteocyte, adipogenic, and chondrogenic differentiation. Sci. Rep. 11, 22593 (2021).
Crossref Google Scholar
31
E. L. Clinkenbeard et al., Increased FGF23 protects against detrimental cardio-renal consequences during elevated blood phosphate in CKD. JCI Insight 4 (2019).
Crossref
32

Onal, M. et al. A novel distal enhancer mediates inflammation-, PTH-, and early onset murine kidney disease-induced expression of the mouse Fgf23 Gene. JBMR 2, 32–47 (2018).
Crossref Google Scholar
33

Ronkainen, V. P. et al. Hypoxia-inducible factor 1-induced G protein-coupled receptor 35 expression is an early marker of progressive cardiac remodelling. Cardiovasc. Res 101, 69–77 (2014).
Crossref Google Scholar
34

Zhang, Y., Shi, T. & He, Y. GPR35 regulates osteogenesis via the Wnt/GSK3beta/beta-catenin signaling pathway. Biochem. Biophys. Res. Commun. 556, 171–178 (2021).
Crossref Google Scholar
35

Gess, B. et al. The cellular oxygen tension regulates expression of the endoplasmic oxidoreductase ERO1-Lalpha. Eur. J. Biochem. 270, 2228–2235 (2003).
Crossref Google Scholar
36

Chen, C., Li, H., Jiang, J., Zhang, Q. & Yan, F. Inhibiting PHD2 in bone marrow mesenchymal stem cells via lentiviral vector-mediated RNA interference facilitates the repair of periodontal tissue defects in SD rats. Oncotarget 8, 72676–72699 (2017).
Crossref Google Scholar
37

Imel, E. A. et al. Serum fibroblast growth factor 23, serum iron and bone mineral density in premenopausal women. Bone 86, 98–105 (2016).
Crossref Google Scholar
38

Wolf, M., Koch, T. A. & Bregman, D. B. Effects of iron deficiency anemia and its treatment on fibroblast growth factor 23 and phosphate homeostasis in women. J. Bone Min. Res. 28, 1793–1803 (2013).
Crossref Google Scholar
39

Haase, V. H. HIF-prolyl hydroxylases as therapeutic targets in erythropoiesis and iron metabolism. Hemodial. Int. 21, S110–S124 (2017).
Crossref Google Scholar
40

Yeh, T. L. et al. Molecular and cellular mechanisms of HIF prolyl hydroxylase inhibitors in clinical trials. Chem. Sci. 8, 7651–7668 (2017).
Crossref Google Scholar
41

Clinkenbeard, E. L. et al. Erythropoietin stimulates murine and human fibroblast growth factor-23, revealing novel roles for bone and bone marrow. Haematologica 102, e427–e430 (2017).
Crossref Google Scholar
42

Hanudel, M. R. et al. Effects of erythropoietin on fibroblast growth factor 23 in mice and humans. Nephrol. Dial. Transpl. 34, 2057–2065 (2018).
Crossref Google Scholar
43

Daryadel, A. et al. Erythropoietin stimulates fibroblast growth factor 23 (FGF23) in mice and men. Pflug. Arch. 470, 1569–1582 (2018).
Crossref Google Scholar
44

Kwon, S. Y. et al. Hypoxia enhances cell properties of human mesenchymal stem cells. Tissue Eng. Regen. Med 14, 595–604 (2017).
Crossref Google Scholar
45

Antebi, B. et al. Short-term physiological hypoxia potentiates the therapeutic function of mesenchymal stem cells. Stem Cell. Res. Ther. 9, 265 (2018).
Crossref Google Scholar
46

Wang, Y. et al. The hypoxia-inducible factor alpha pathway couples angiogenesis to osteogenesis during skeletal development. J. Clin. Invest. 117, 1616–1626 (2007).
Crossref Google Scholar
47

Hung, S. P., Ho, J. H., Shih, Y. R., Lo, T. & Lee, O. K. Hypoxia promotes proliferation and osteogenic differentiation potentials of human mesenchymal stem cells. J. Orthop. Res. 30, 260–266 (2012).
Crossref Google Scholar
48

Wu, C. et al. Oxygen-sensing PHDs regulate bone homeostasis through the modulation of osteoprotegerin. Genes Dev. 29, 817–831 (2015).
Crossref Google Scholar
49

Cheng, S., Xing, W., Pourteymoor, S. & Mohan, S. Conditional disruption of the prolyl hydroxylase domain-containing protein 2 (Phd2) gene defines its key role in skeletal development. J. Bone Min. Res 29, 2276–2286 (2014).
Crossref Google Scholar
50

Stiers, P. J. et al. Inhibition of the oxygen sensor PHD2 enhances tissue-engineered endochondral bone formation. J. Bone Min. Res. 34, 333–348 (2019).
Crossref Google Scholar
51

Roszko, K. L. et al. C-Terminal, but not intact, FGF23 and EPO are strongly correlatively elevated in patients with gain-of-function mutations in HIF2A: Clinical evidence for EPO regulating FGF23. J. Bone Min. Res. 36, 315–321 (2021).
Crossref Google Scholar
52

Block, G. A. et al. A pilot randomized trial of ferric citrate coordination complex for the treatment of advanced CKD. J. Am. Soc. Nephrol. 30, 1495–1504 (2019).
Crossref Google Scholar
53

Wolf, M. et al. Effects of Iron Isomaltoside vs. Ferric Carboxymaltose on Hypophosphatemia in iron-deficiency anemia: two randomized clinical trials. JAMA 323, 432–443 (2020).
Crossref Google Scholar
54

Macdougall, I. C. et al. Iron management in chronic kidney disease: conclusions from a “Kidney Disease: Improving Global Outcomes” (KDIGO) Controversies Conference. Kidney Int 89, 28–39 (2016).
Crossref Google Scholar
55

Stevens, L. A., Djurdjev, O., Cardew, S., Cameron, E. C. & Levin, A. Calcium, phosphate, and parathyroid hormone levels in combination and as a function of dialysis duration predict mortality: evidence for the complexity of the association between mineral metabolism and outcomes. J. Am. Soc. Nephrol. 15, 770–779 (2004).
Crossref Google Scholar
56

Slinin, Y., Foley, R. N. & Collins, A. J. Calcium, phosphorus, parathyroid hormone, and cardiovascular disease in hemodialysis patients: the USRDS waves 1, 3, and 4 study. J. Am. Soc. Nephrol. 16, 1788–1793 (2005).
Crossref Google Scholar
57

Clinkenbeard, E. L. et al. Conditional deletion of Murine Fgf23: Interruption of the Normal skeletal responses to phosphate challenge and rescue of genetic hypophosphatemia. J. Bone Min. Res. 31, 1247–1257 (2016).
Crossref Google Scholar
58

Mazzone, M. et al. Heterozygous deficiency of PHD2 restores tumor oxygenation and inhibits metastasis via endothelial normalization. Cell 136, 839–851 (2009).
Crossref Google Scholar
59

Lu, Y. et al. DMP1-targeted Cre expression in odontoblasts and osteocytes. J. Dent. Res. 86, 320–325 (2007).
Crossref Google Scholar
60

Diehl, K. H. et al. A good practice guide to the administration of substances and removal of blood, including routes and volumes. J. Appl. Toxicol. 21, 15–23 (2001).
Crossref Google Scholar
61

Diwan, V., Small, D., Kauter, K., Gobe, G. C. & Brown, L. Gender differences in adenine-induced chronic kidney disease and cardiovascular complications in rats. Am. J. Physiol. Ren. Physiol. 307, F1169–F1178 (2014).
Crossref Google Scholar
62

Woo, S. M., Rosser, J., Dusevich, V., Kalajzic, I. & Bonewald, L. F. Cell line IDG-SW3 replicates osteoblast-to-late-osteocyte differentiation in vitro and accelerates bone formation in vivo. J. Bone Min. Res. 26, 2634–2646 (2011).
Crossref Google Scholar
63

Zhou, M. et al. The pro-angiogenic role of hypoxia inducible factor stabilizer FG-4592 and its application in an in vivo tissue engineering chamber model. Sci. Rep. 9, 6035 (2019).
Crossref Google Scholar
64

Byrnes, C. et al. Iron dose-dependent differentiation and enucleation of human erythroblasts in serum-free medium. J. Tissue Eng. Regen. Med. 10, E84–E89 (2016).
Crossref Google Scholar
65

Dobin, A. et al. STAR: ultrafast universal RNA-seq aligner. Bioinformatics 29, 15–21 (2013).
Crossref Google Scholar
66

Liao, Y., Smyth, G. K. & Shi, W. featureCounts: an efficient general purpose program for assigning sequence reads to genomic features. Bioinformatics 30, 923–930 (2014).
Crossref Google Scholar
67

Robinson, M. D., McCarthy, D. J. & Smyth, G. K. edgeR: a Bioconductor package for differential expression analysis of digital gene expression data. Bioinformatics 26, 139–140 (2010).
Crossref Google Scholar
68

Dennis, G. Jr. et al. DAVID: Database for annotation, visualization, and integrated discovery. Genome Biol. 4, P3 (2003).
Crossref Google Scholar
69

Kramer, A., Green, J., Pollard, J. Jr. & Tugendreich, S. Causal analysis approaches in Ingenuity Pathway Analysis. Bioinformatics 30, 523–530 (2014).
Crossref Google Scholar
70

Buenrostro, J. D., Giresi, P. G., Zaba, L. C., Chang, H. Y. & Greenleaf, W. J. Transposition of native chromatin for fast and sensitive epigenomic profiling of open chromatin, DNA-binding proteins and nucleosome position. Nat. Methods 10, 1213–1218 (2013).
Crossref Google Scholar
71

Langmead, B. & Salzberg, S. L. Fast gapped-read alignment with Bowtie 2. Nat. Methods 9, 357–359 (2012).
Crossref Google Scholar
72

Zhang, Y. et al. Model-based analysis of ChIP-Seq (MACS). Genome Biol. 9, R137 (2008).
Crossref Google Scholar
73

E. P. Consortium. An integrated encyclopedia of DNA elements in the human genome. Nature 489, 57–74 (2012).
Crossref Google Scholar
74

Amemiya, H. M., Kundaje, A. & Boyle, A. P. The ENCODE blacklist: identification of problematic regions of the genome. Sci. Rep. 9, 9354 (2019).
Crossref Google Scholar
75

Piper, J. et al. Wellington: a novel method for the accurate identification of digital genomic footprints from DNase-seq data. Nucleic Acids Res. 41, e201 (2013).
Crossref Google Scholar
76

McCarthy, D. J., Chen, Y. & Smyth, G. K. Differential expression analysis of multifactor RNA-Seq experiments with respect to biological variation. Nucleic Acids Res. 40, 4288–4297 (2012).
Crossref Google Scholar
77

Heinz, S. et al. Simple combinations of lineage-determining transcription factors prime cis-regulatory elements required for macrophage and B cell identities. Mol. Cell 38, 576–589 (2010).
Crossref Google Scholar
78

Livak, K. J. & Schmittgen, T. D. Analysis of relative gene expression data using real-time quantitative PCR and the 2(-Delta Delta C(T)) Method. Methods 25, 402–408 (2001).
Crossref Google Scholar
Bone Research
Volume 11,
December 2023
Article number: 7
DOI: 10.1038/s41413-022-00241-w
Cite this article:
Noonan ML, Ni P, Solis E, et al. Osteocyte Egln1/Phd2 links oxygen sensing and biomineralization via FGF23. Bone Research, 2023, 11: 7. https://doi.org/10.1038/s41413-022-00241-w

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Received: 07 April 2022
Revised: 29 September 2022
Accepted: 03 November 2022
Published: 18 January 2023
© The Author(s) 2023

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