Vasohibin as an endothelium-derived negative feedback regulator of angiogenesis
J. Clin. Invest. Kazuhide Watanabe, et al. 114:898 doi:10.1172/JCI21152 [Go to this article.]

Figure 1
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).