Abstract
The development of a new pharmacological strategy, the angiostatic therapy, to inhibit solid tumor progression has increased the need of powerful in vitro models to screen the angiostatic potential of new drug candidates. We produced an endothelialized reconstructed connective tissue (ERCT) that promotes the spontaneous formation of a human capillary-like network by coculture of human endothelial cells isolated from umbilical cord or from newborn foreskin, with dermal fibroblasts in a collagen sponge. Three inhibitors of angiogenesis, tamoxifen, ilomastat, and echistatin, were used to assess the efficiency of our ERCT to discriminate, in vitro, an angiostatic potential. The capillary-like structures were characterized by their immunoreactivity to human platelet-endothelial cellular adhesion molecule-1 antibodies and were quantified on histological cross-sections of biopsies taken after 10, 17, 24, and 31 days of culture. A dose-response significant inhibition of the capillary-like formation was detected when increasing concentrations of tamoxifen, ilomastat, or echistatin were added for 1 week to the culture medium of the ERCT. Tamoxifen was found to be angiogenic at 10 μM and to have a cytotoxic effect at 40 μM 1 week after drug removal. Echistatin induced a rapid, slight, and reversible inhibition of capillary-like formation, whereas ilomastat caused a very precocious, strong, and reversible inhibition of angiogenesis. In addition, a 16-h hypoxia promoted the formation of 10 times larger vessels (>300 μm2), compared with normoxic condition. These results suggest that our model could be efficiently used to study the long-term angiostatic potential of drugs in vitro in a very physiological environment.
Footnotes
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This study was supported by the Canadian Institutes of Health Research (CIHR) Grant MOP-14364, “Fondation de l'Hôpital du Saint-Sacrement”, “Fondation des Pompiers du Québec pour les Grands Brûlés”, and “France-Québec” exchange program. P.-L.T. was recipient of a scholarship from the Natural Sciences and Engineering Research Council of Canada (NSERC). F.B. was the recipient of a Fellowship from Fonds de Recherche en Santé du Québec, and L.G. was the recipient of a Canada Research Chair on stem cells and tissue engineering.
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Article, publication date, and citation information can be found at http://jpet.aspetjournals.org.
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doi:10.1124/jpet.105.089524.
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ABBREVIATIONS: VEGF, vascular endothelial growth factor; HUVEC, human umbilical vein endothelial cell(s); CLS, capillary-like structure(s); ERCT, endothelialized reconstructed connective tissue; HMVEC, human microvascular endothelial cell(s); PECAM-1, platelet-endothelial cellular adhesion molecule-1; MMP, matrix metallo proteinase.
- Received May 16, 2005.
- Accepted July 26, 2005.
- The American Society for Pharmacology and Experimental Therapeutics
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