THE ROLE OF EXOGENOUS ENDOGENOUS BASIC FIBROBLAST GROWTH-FACTOR (FGF2) AND TRANSFORMING GROWTH-FACTOR-BETA (TGF-BETA-1) ON HUMAN CORNEAL ENDOTHELIAL-CELLS PROLIFERATION IN-VITRO

Adult human corneal endothelial cells (HCEC) have extremely low turnov er rates but undergo rapid division in vitro when stimulated with solu ble growth factors. We have investigated the role played by FGF2 and T GF beta-1 in the regulation of HCEC growth stimulation. HCEC from dono rs who were over 30 years old were cultured and experiments performed on cultures between the 2nd and the 6th passage in the presence of 5% NCS. Cell counts revealed a maximal stimulation of 2.1x for FGF2 and 1 .9x for TGF beta-1 compared to control cultures. When both factors wer e added, a synergistic effect was noticed with a maximal stimulation o f the proliferation rate of 4.5X over controls. In addition, endogenou s FGF2 produced by HCEC was quantitated in a sensitive EIA assay. Afte r 5 days in culture, 10(6) cells contained 150 ng FGF2 and 35 ng was e xtracted from trypsin-digested ECM. Two molar NaCl washes of ECM relea sed 15.6 ng FGF2, which induced a slight mitogenic activity (1.5x over control) in HCEC cultures, which was partially inhibited by an anti-F GF2 antibody. Northern blot analysis of HCEC extracts revealed the pre sence of FGF receptors R1 and R2 mRNA. The bioactive FGFRs were demons trated by the toxic effect of a mitotoxin FGF2-SAP. These results sugg est that FGF2 could participate in the autocrine regulation of HCEC pr oliferation and survival. The synergy between exogenously added FGF2 a nd TGF beta demonstrates that a combination of different growth factor s may be important to stimulate proliferation of these cells in vivo. (C) 1995 Academic Press, Inc.