Abstract
Infantile hemangiomas are localized and rapidly growing regions of disorganized angiogenesis. We show that expression of vascular endothelial growth factor receptor-1 (VEGFR1) in hemangioma endothelial cells (hemECs) and hemangioma tissue is markedly reduced compared to controls. Low VEGFR1 expression in hemECs results in VEGF-dependent activation of VEGFR2 and downstream signaling pathways. In hemECs, transcription of the gene encoding VEGFR1 (FLT1) is dependent on nuclear factor of activated T cells (NFAT). Low VEGFR1 expression in hemECs is caused by reduced activity of a pathway involving β1 integrin, the integrin-like receptor tumor endothelial marker-8 (TEM8), VEGFR2 and NFAT. In a subset of individuals with hemangioma, we found missense mutations in the genes encoding VEGFR2 (KDR) and TEM8 (ANTXR1). These mutations result in increased interactions among VEGFR2, TEM8 and β1 integrin proteins and in inhibition of integrin activity. Normalization of the constitutive VEGFR2 signaling in hemECs with soluble VEGFR1 or antibodies that neutralize VEGF or stimulate β1 integrin suggests that local administration of these or similar agents may be effective in hemangioma treatment.
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Acknowledgements
This work was supported by the John B. Mulliken Foundation and grants AR36820 and AR48564 from the US National Institutes of Health (to B.R.O.). We are indebted to J.B. Mulliken (Children's Hospital, Boston) and L. Boon (Universite Catholique de Louvain) for providing essential surgical material (control and hemangioma tissues and blood samples) for these studies. We thank N.A. Clipstone (Feinberg School of Medicine, Northwestern University) for providing the constitutively active form of NFATc1, M. Kurabayashi (Gunma University Graduate School of Medicine) for providing the promoter construct for FLT1, S. Dias (Instituto Portugues de Oncologia Francisco Gentil) for providing expression vectors for FLT1, L. Claesson-Welsh (Uppsala University) for providing expression vectors for KDR and mutant FLT1, S. Liu (US National Institute of Allergy and Infectious Diseases) for providing expression vector for ANTXR1 and T. Kitamura (University of Tokyo,) for the pMXs vector. We thank R. Ruijtenbeek and R. Houtman (both from PamGene Corporation) for providing reagents and equipment for kinase arrays, F. Naji and M. Dankers for kinase array bioinformatics assistance, T. Rector for assistance with protein multiplexing, J. Eastcott and J. Wylie-Sears for flow cytometry and technical assistance, S. Feske (New York University School of Medicine) for antibody to NFATc2 (clone 67.1) and advice and W. Kuo for technical advice. We thank Y. Pittel, S. Plotkina, N. Liu, A. Heilmann, Y. Ishida and Y. Yamamura for technical assistance and D. Glotzer for comments and advice on the manuscript.
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M.J., cell signaling studies, immunoblotting and immunoprecipitation analyses, transcription and promoter studies, integrin studies; D.M., multiplex ELISA and kinase assays, proliferation and migration analyses; L.P., sequencing of candidate genes; N.L., allele-specific DNA sequencing; Y.L., initial signaling studies; E. Boscolo, hemangioma tissue characterization; J.B., isolation of hemEC lines, control cells and tissues; M.V., collection of control and hemangioma tissues and DNAs; E. Boye, planning, NFAT and Ca2+ studies; B.R.O., planning and directing the study, manuscript writing.
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Jinnin, M., Medici, D., Park, L. et al. Suppressed NFAT-dependent VEGFR1 expression and constitutive VEGFR2 signaling in infantile hemangioma. Nat Med 14, 1236–1246 (2008). https://doi.org/10.1038/nm.1877
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DOI: https://doi.org/10.1038/nm.1877
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