Expression of the oxygen-regulated protein ORP150 accelerates wound healing by modulating intracellular VEGF transport
Kentaro Ozawa, … , Satoshi Ogawa, Tohru Ohshima
Kentaro Ozawa, … , Satoshi Ogawa, Tohru Ohshima
Published July 1, 2001
Citation Information: J Clin Invest. 2001;108(1):41-50. https://doi.org/10.1172/JCI11772.
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Article

Expression of the oxygen-regulated protein ORP150 accelerates wound healing by modulating intracellular VEGF transport

Abstract

Expression of angiogenic factors such as VEGF under conditions of hypoxia or other kinds of cell stress contributes to neovascularization during wound healing. The inducible endoplasmic reticulum chaperone oxygen-regulated protein 150 (ORP150) is expressed in human wounds along with VEGF. Colocalization of these two molecules was observed in macrophages in the neovasculature, suggesting a role of ORP150 in the promotion of angiogenesis. Local administration of ORP150 sense adenovirus to wounds of diabetic mice, a treatment that efficiently targeted this gene product to the macrophages of wound beds, increased VEGF antigen in wounds and accelerated repair and neovascularization. In cultured human macrophages, inhibition of ORP150 expression caused retention of VEGF antigen within the endoplasmic reticulum (ER), while overexpression of ORP150 promoted the secretion of VEGF into hypoxic culture supernatants. Taken together, these data suggest an important role for ORP150 in the setting of impaired wound repair and identify a key, inducible chaperone-like molecule in the ER. This novel facet of the angiogenic response may be amenable to therapeutic manipulation.

Authors

Kentaro Ozawa, Toshikazu Kondo, Osamu Hori, Yasuko Kitao, David M. Stern, Wolfgang Eisenmenger, Satoshi Ogawa, Tohru Ohshima

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Expression of VEGF and ORP150 in human macrophages exposed to hypoxia. (...
Expression of VEGF and ORP150 in human macrophages exposed to hypoxia. (a) Macrophages were either exposed to hypoxia (filled bars; 0–24 hours) or maintained under normoxic conditions (open bars). VEGF content in the culture supernatant was measured by ELISA as described in the text (n = 8; mean ± SD). *P < 0.01 by multiple contrast analysis. (b–d) Macrophages were exposed to hypoxia (0–24 hours). (b) RNA was prepared at the indicated times and subjected to RT-PCR analysis using primers for either VEGF (upper panel) or β-actin (lower panel). Protein extracts were prepared and subjected to Western blot analysis using anti-human ORP150 Ab (c). Cell lysates were incubated with either anti-human ORP150 Ab or anti-VEGF Ab. (d) The immunoprecipitant was separated in SDS-PAGE (8–12%; nonreducing condition for VEGF and reducing condition for ORP150) and subjected to Western blot analysis using either anti-VEGF Ab (left panel) or anti-ORP150 Ab (right panel). Migrations of molecular-weight markers are shown in the middle. The filled circles denote the signals derived from IgG used as primary Ab. (e–g) Human macrophages were double-stained with anti-ORP150 Ab (e) and anti-VEGF Ab (f). VEGF and ORP150 signals were digitally overlapped (×400) (g).