INDUCTION OF ENDOTHELIAL-CELL DIFFERENTIATION IN-VITRO BY FIBROBLAST-DERIVED SOLUBLE FACTORS

Recent studies have suggested that fibroblasts, widely distributed mes enchymal cells, not only function to sustain various organs and tissue s as stroma cells but also act directly to regulate adjacent cell beha vior including migration, proliferation, and differentiation. Since fi broproliferative diseases and lesions (fibroplasia) are accompanied by new capillary growth (angiogenesis), we hypothesized that fibroblasts may have direct effects on endothelial cell behavior, independent of the elaboration of extracellular matrix, that are relevant to complex process of angiogenesis. To test this hypothesis, bovine aortic endoth elial cells were cocultured in collagen gels with human skin fibroblas ts. This coculture system caused the endothelial cells to become spind le shaped and to organize into a capillary-like structure within the c ollagen gels. We found that fibroblast-conditioned medium (FCM) also i nduced endothelial cells initially to elongate and subsequently to org anize into a capillary-like structure within collagen gels. While FCM had no significant effect on endothelial cell DNA synthesis, the solub le factor(s) in FCM increased endothelial cell motility in an in vitro wound assay and in a Boyden chamber assay. The chemoattractant(s) in FCM was alkaline (pH 9.0)-and acid (pH 3.0)-stable, relatively heat st able (stable at 60 degrees C for 30 min, unstable at 98 degrees C for 3 min), dithiothreitol (DTT)-sensitive, and bound to an anionic exchan ge resin (DEAE-cellulose). Another factor(s) stimulated endothelial ce ll reorganization into capillary-like structure both within a collagen gel and on a reconstituted basement membrane matrix, Matrigel. This f actor(s) was alkaline (pH 9.0)- and acid (pH 3.0)-stable, heat (98 deg rees C for 3 min)stable, and DTT-sensitive and bound an anionic exchan ge resin (DEAE-cellulose). These in vitro results suggest that fibrobl asts secrete soluble factors that can influence endothelial cell behav iors relevant to the angiogenesis process with possible implications f or vascularization in fibroproliferative conditions. (C) 1994 Academic Press, Inc.