Basic fibroblast growth factor release by human coronary artery endothelial cells is enhanced by matrix proteins, 17 beta-estradiol, and a PKC signaling pathway

Endothelial cell function is regulated by interactions among cells, the ext racellular matrix (ECM), and soluble mediators. We investigated this intera ction by examining the effect of 17 beta-estradiol (E2) on release of basic fibroblast growth factor (FGF-2) by human coronary artery endothelial cell s (HCAEC) cultured on ECM proteins. After estrogen-depleted HCAEC were trea ted with E2 for 2 h, the conditioned media and cell layers were evaluated b y immunoblot or ELISA for FGF-2. Release of FGF-2 into conditioned media wa s enhanced 10-fold compared to that on plastic and a further 2.4-fold by E2 , As FGF-2 release from cells into the media increases, there is a correspo nding decrease in the cellular content of FGF-2. By ELISA, FGF-2 release in creased 406, 179, and 262%, on type TV collagen, laminin, or fibronectin, r espectively. HCAEC cultured on type I collagen did not show ES-enhanced FGF -2 release by ELISA or immunoblot analysis. No changes were noted in HCAEC release of lactate dehydrogenase, tested as a control protein for cellular integrity. The estrogen receptor antagonist ICI182,780 blocked E2-induced, but not basal, FGF-2 release. Increased FGF-2 release occurred via a cycloh eximide-insensitive pathway. Neither brefeldin-A nor genistein inhibited E2 enhancement of FGF-2 release by HCAEC cultured on fibronectin. However, th e protein kinase C inhibitor calphostin C inhibited the E2-augmented FGF-2 release. These data show that E2 enhances FGF-2 release by HCAEC cultured o n basement membrane proteins in the absence of wounding. This action requir es the estrogen receptor and PKC activity, but does not require new protein synthesis, endoplasmic reticulum-to-Golgi-mediated secretion, or protein t yrosine phosphorylation. E2-enhanced FGF-2 release could contribute to the cardioprotective effects of estrogen. (C) 1998 Academic Press.