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VEGF modulates erythropoiesis through regulation of adult hepatic erythropoietin synthesis

Nature Medicine volume 12, pages 793–800 (2006)Cite this article

A Corrigendum to this article was published on 01 April 2009

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Abstract

Vascular endothelial growth factor (VEGF) exerts crucial functions during pathological angiogenesis and normal physiology. We observed increased hematocrit (60–75%) after high-grade inhibition of VEGF by diverse methods, including adenoviral expression of soluble VEGF receptor (VEGFR) ectodomains, recombinant VEGF Trap protein and the VEGFR2-selective antibody DC101. Increased production of red blood cells (erythrocytosis) occurred in both mouse and primate models, and was associated with near-complete neutralization of VEGF corneal micropocket angiogenesis. High-grade inhibition of VEGF induced hepatic synthesis of erythropoietin (Epo, encoded by Epo) >40-fold through a HIF-1α–independent mechanism, in parallel with suppression of renal Epo mRNA. Studies using hepatocyte-specific deletion of the Vegfa gene and hepatocyte–endothelial cell cocultures indicated that blockade of VEGF induced hepatic Epo by interfering with homeostatic VEGFR2-dependent paracrine signaling involving interactions between hepatocytes and endothelial cells. These data indicate that VEGF is a previously unsuspected negative regulator of hepatic Epo synthesis and erythropoiesis and suggest that levels of Epo and erythrocytosis could represent noninvasive surrogate markers for stringent blockade of VEGF in vivo.

NOTE: In the version of this article initially published, the name of one of the authors, Nihar R. Nayak, was misspelled as Nihar R. Niyak. The error has been corrected in the HTML and PDF versions of the article.

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Figure 1: Adenoviruses encoding sVEGFRs increase hematocrit.
Figure 2: High-grade inhibition of VEGF induces production of Epo.
Figure 3: Soluble VEGFRs stimulate hepatic expression of Epo.
Figure 4: Elevated hematocrit and hepatic Epo mRNA in Ad-Cre–treated VegfaloxP/loxP mice.

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  • 06 April 2009

    In the version of this article initially published, the name of one of the authors, Nihar R. Nayak, was misspelled as Nihar R. Niyak. The error has been corrected in the HTML and PDF versions of the article.

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Acknowledgements

We thank members of the Kuo laboratory for discussion, J. Folkman for support of this project, C. Davis for histopathologic analysis of liver sections, C.-P. Chang for HA-Flt1-His cDNA, A. Patterson for determination of arterial oxygen tension, A. Wagers for demonstrating parabiotic surgeries, M. Socolovsky for murine Epo, J. Zehnder for advice with real-time PCR, C. Koch for EF5 and EF5-specific antibodies, G. Molineux for permission to cite unpublished data and E. Yu for initial assistance. We are indebted to H.-P. Gerber and N. Ferrara for their gift of VegfaloxP/loxP mice, Y.-M. Lin and T. Wandless for synthesis of ZD4190 and the UCSF Liver Center (supported by NIH P30 DK26743) for providing core services. B.Y.Y.T. is a Fonds de la Recherche en Santé du Québec fellow. K.W. is supported by a Medical Scientist Training Program Grant to Stanford University. This work was supported by grants from the US National Institutes of Health (1 R01 CA95654-01 and 1 R01 HL074267-01) and the Department of Defense (DAMD17-02-1-0143) to C.J.K. and funds from Amgen Corporation (to R.J.K.). C.J.K. is a Burroughs Wellcome Foundation Scholar in the Pharmacological Sciences and a Kimmel Foundation Scholar.

Author information

Author notes

  1. Betty Y Y Tam

    Present address: TargeGen Inc., 9380 Judicial Drive, San Diego, California, 92121, USA

  2. Sang-ki Park

    Present address: Department of Pharmaceutical Sciences, St. John's University, 8000 Utopia Parkway, Queens, New York, 11438, USA

  3. Betty Y Y Tam and Kevin Wei: These authors contributed equally to this work.

Authors and Affiliations

  1. Division of Hematology, Stanford University School of Medicine, 269 Campus Drive, CCSR 1155, Stanford, California, 94305, USA

    Betty Y Y Tam, Kevin Wei, Jana Hoffman, Jenny Yuan, Cecile Chartier, Lisa Ma, Grace Wu, Stanley L Schrier & Calvin J Kuo

  2. Regeneron Pharmaceuticals, Incorporated, 777 Old Saw Mill River Road, Tarrytown, 10591, New York, USA

    John S Rudge, Joceyln Holash, Shelly Jiang & George D Yancopoulos

  3. Division of Biological Sciences, University of California, San Diego, Molecular Biology Section, 9500 Gilman Drive, La Jolla, 92093, California, USA

    Sang-ki Park & Randall S Johnson

  4. Liver Center and Department of Medicine, S-357 University of California, San Francisco, 94143, California, USA

    Colleen Hefner & Jacquelyn J Maher

  5. Department of Genetics, Division of Molecular Medicine, Harvard Medical School, Children's Hospital, 300 Longwood Avenue, Boston, 02115, Massachusetts, USA

    Jeng-Shin Lee & Richard C Mulligan

  6. Division of Gynecology and Obstetrics, Stanford University School of Medicine, 300 Pasteur Drive, Stanford, 94305, California, USA

    Nihar R Nayak

  7. Children's Hospital Oakland Research Institute, 5700 Martin Luther King Jr. Way, Oakland, 94609, California, USA

    Frans A Kuypers

  8. Division of Pathology, Stanford University School of Medicine, 300 Pasteur Drive, Stanford, 94305, California, USA

    Uma Sundram

  9. Division of Cardiology MC8573, University of Texas Southwestern Medical Center at Dallas, 5323 Harry Hines Boulevard, Dallas, 75390, Texas, USA

    Joseph A Garcia

  10. TargeGen Inc., 9380 Judicial Drive, San Diego, California, 92121, USA

    Sang-ki Park

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Contributions

B.Y.Y.T. and K.W. designed and performed experiments and wrote the manuscript. J.S.R. performed animal studies with VEGF Trap protein and edited the manuscript. J. Hoffman performed Epo RT-QPCR. J.Y. and C.C. generated Ad VEGF Trap and Ad KDR-Fc. G.W. assisted on hematocrit and flow cytometry studies. L.M. and S.L.S. analyzed RBC precursors. U.S. performed pathologic analysis. J. Holash and S.J. performed animal studies with VEGF Trap protein. S.K.P. performed studies with VegfaloxP/loxP mice and RT-QPCR of hypoxic genes. R.S.J. and J.A.G. provided expertise in liver physiology. F.A.K. designed and performed RBC mass analysis. G.D.Y. supervised VEGF Trap studies. R.C.M. supervised adenoviral construction and production. C.J.K. was principal investigator, designed and conceptualized the study, analyzed and interpreted all data, and drafted and edited the manuscript.

Corresponding author

Correspondence to Calvin J Kuo.

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The authors declare no competing financial interests.

Supplementary information

Supplementary Fig. 1

Elevation of erythroid parameters in sVEGFR-treated mice (PDF 139 kb)

Supplementary Fig. 2

Stimulation of Epo mRNA production in hepatocytes of Ad-Flt1treated mice as determined by in situ hybridization. (PDF 519 kb)

Supplementary Fig. 3

Soluble Flt1 ectodomain induces hepatic Epo expression in the absence of direct liver infection in parabiotic mice. (PDF 117 kb)

Supplementary Fig. 4

Transient increase of Epo mRNA in hypoxic kidneys or livers. (PDF 60 kb)

Supplementary Fig. 5

Ad-Cremediated excision of the loxP-flanked stop cassette in ROSA26-LacZ mice allows expression of the lacZ gene. (PDF 2327 kb)

Supplementary Fig. 6

EF5 detection of hypoxia in tumors and Ad-Flt1treated liver. (PDF 206 kb)

Supplementary Fig. 7

Ad-Flt1 and Ad-Flk1-Fctreated livers do not show evidence of hemorrhage, necrosis or ischemic tissue damage. (PDF 818 kb)

Supplementary Fig. 8

Epo mRNA is selectively elevated in hepatocyte coculture with LSECs but not other endothelial cell types or fibroblasts. (PDF 117 kb)

Supplementary Fig. 9

Comparative inhibition of VEGF-stimulated corneal-micropocket angiogenesis by Ad-Flt1, ZD4190 and rVEGF Trap. (PDF 490 kb)

Supplementary Table 1

Elevation of Ter119+CD45 erythroid precursors by VEGF blockade. (PDF 58 kb)

Supplementary Table 2

Assessment of hydration and oxygenation status following VEGF inhibition. (PDF 117 kb)

Supplementary Methods (PDF 142 kb)

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Tam, B., Wei, K., Rudge, J. et al. VEGF modulates erythropoiesis through regulation of adult hepatic erythropoietin synthesis. Nat Med 12, 793–800 (2006). https://doi.org/10.1038/nm1428

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