Vasohibin as an endothelium-derived negative feedback regulator of angiogenesis
Kazuhide Watanabe, … , Hikaru Sonoda, Yasufumi Sato
Kazuhide Watanabe, … , Hikaru Sonoda, Yasufumi Sato
Published October 1, 2004
Citation Information: J Clin Invest. 2004;114(7):898-907. https://doi.org/10.1172/JCI21152.
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Article Angiogenesis

Vasohibin as an endothelium-derived negative feedback regulator of angiogenesis

Abstract

Negative feedback is a crucial physiological regulatory mechanism, but no such regulator of angiogenesis has been established. Here we report a novel angiogenesis inhibitor that is induced in endothelial cells (ECs) by angiogenic factors and inhibits angiogenesis in an autocrine manner. We have performed cDNA microarray analysis to survey VEGF-inducible genes in human ECs. We characterized one such gene, KIAA1036, whose function had been uncharacterized. The recombinant protein inhibited migration, proliferation, and network formation by ECs as well as angiogenesis in vivo. This inhibitory effect was selective to ECs, as the protein did not affect the migration of smooth muscle cells or fibroblasts. Specific elimination of the expression of KIAA1036 in ECs restored their responsiveness to a higher concentration of VEGF. The expression of KIAA1036 was selective to ECs, and hypoxia or TNF-α abrogated its inducible expression. As this molecule is preferentially expressed in ECs, we designated it “vasohibin.” Transfection of Lewis lung carcinoma cells with the vasohibin gene did not affect the proliferation of cancer cells in vitro, but did inhibit tumor growth and tumor angiogenesis in vivo. We propose vasohibin to be an endothelium-derived negative feedback regulator of angiogenesis.

Authors

Kazuhide Watanabe, Yasuhiro Hasegawa, Hiroshi Yamashita, Kazue Shimizu, Yuanying Ding, Mayumi Abe, Hideki Ohta, Keiichi Imagawa, Kanji Hojo, Hideo Maki, Hikaru Sonoda, Yasufumi Sato

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Figure 1

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KIAA1036 is an endothelium-derived VEGF-inducible secretory protein. (A)...
KIAA1036 is an endothelium-derived VEGF-inducible secretory protein. (A) The deduced amino acid sequences of the human and mouse KIAA1036 (KIAA) proteins are shown. Asterisks indicate identical amino acids between human and mouse. (B) A single KIAA1036 mRNA was induced by VEGF. HUVECs were stimulated with VEGF (1 nM) for the indicated periods and then Northern blotting was performed. (C) VEGF increased KIAA1036 mRNA in a concentration-dependent manner. HUVECs were stimulated with the indicated concentration of VEGF for 24 hours and then real-time RT-PCR was performed. (D) KIAA1036 protein was synthesized and secreted. GM7373 cells transfected with KIAA1036 gene were lysed. Equal amounts of protein were applied to lane 1 and lane 2, and transferred to the filter. The filter was then separated into 2 parts. Western blotting was performed with anti_KIAA1036 mAb (lane 1). Prior to Western blotting, anti_KIAA1036 mAb was absorbed with antigen peptide (lane 2). HUVECs were stimulated with VEGF (1 nM) for the following periods and were lysed for Western blotting: lane 3, 0 hours; lane 4, 12 hours; lane 5, 24 hours; lane 6, 48 hours. HUVECs were cultured for 3 days in the growth medium and then cells were lysed for Western blotting (lane 7). After this incubation, the medium was collected and concentrated. Five hundred microliters of concentrated medium was subjected to immunoprecipitation followed by Western blotting (lane 8). Asterisk indicates protein in the medium. (E) KIAA1036 protein does not colocalize with ER. HUVECs in the growth medium were used for the immunostaining of calnexin (red) and KIAA1036 protein (green).