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  • Title: Paracrine interactions between fibroblasts and endothelial cells in a serum-free coculture model. Modulation of angiogenesis and collagen gel contraction.
    Author: Villaschi S, Nicosia RF.
    Journal: Lab Invest; 1994 Aug; 71(2):291-9. PubMed ID: 7521446.
    Abstract:
    BACKGROUND: Previous data suggest that paracrine interactions between fibroblasts and endothelial cells modulate the formation of granulation tissue during wound healing. The study of these interactions in vivo is hindered by the interference of serum and other cell types. To overcome this limitation, we developed a serum-free in vitro model in which microvessels were cocultured with fibroblasts in a 3-dimensional collagen gel. EXPERIMENTAL DESIGN: Microvascular networks were obtained by culturing rat aortic endothelial cells between two layers of collagen. Microvessels were cultured in serum-free medium with or without fibroblasts embedded in the collagen. The cultures were studied by phase contrast microscopy, conventional electron microscopy, and by light and electron immunohistochemistry. The role of endothelial cells and the endothelial-derived peptide endothelin-1 in the transformation of fibroblasts into myofibroblasts was studied by evaluating fibroblast alpha-smooth muscle actin expression and collagen contraction. The angiogenic properties of fibroblasts were investigated with the rat aorta assay. RESULTS: Microvessels cultured without fibroblasts degenerated within 3 to 4 days. Fibroblasts stabilized the microvessels greatly prolonging their life span. This effect was associated with an increased deposition of subendothelial extracellular matrix. Both fibroblasts and fibroblast-conditioned medium stimulated angiogenesis in the rat aorta assay. Endothelial cells and endothelin-1 enhanced the expression of alpha-smooth muscle actin in fibroblasts and stimulated fibroblast-mediated collagen contraction. CONCLUSIONS: This study demonstrates that endothelial-fibroblast interactions can be studied in vitro under serum-free conditions. Our results suggest that paracrine mechanisms operating between endothelial cells and fibroblasts play a key role in the formation and contraction of granulation tissue during wound healing. We propose that fibroblasts stimulate angiogenesis and stabilize the neovascular endothelium which in turn promotes the morphological and functional transformation of fibroblasts into myofibroblasts.
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