EXPRESSION OF FIBROBLAST GROWTH-FACTOR-I (FGF-1), FGF-2 AND FGF RECEPTOR-1 IN A HUMAN SALIVARY-GLAND ADENOCARCINOMA CELL-LINE - EVIDENCE OFAUTOCRINE GROWTH

Fibroblast growth factor-1 (FGF-1) and FGF-2 are heparin-binding polyp eptides which express potent mitogenic properties in neoplastic cells. In the present study, we have examined the contribution of endogenous FGF-1 and FGF-2 to the autocrine growth of HSY human salivary-gland a denocarcinoma cells in vitro. Using specific monoclonal antibodies aga inst FGF-1 and FGF-2, immunohistochemical analysis of HSY cells reveal ed strong expression of both FGF-1 and FGF-2 in the cytoplasm and nucl eus. Consistent with these data, 2 molecular mass species of FGF-1 (16 and 18 kDa) and 3 FGF-2 (18, 24 and 27 kDa) were identified in HSY ce lls by Western-blot analysis. Scatchard analysis of FGF binding sites on HSY cells indicated the presence of 23,000 [I-125]FGF-1 binding sit es/cells with a dissociation constant (K-D) of 178 pM and 13,000 [I-12 5]FCF-2 binding sites/cell with a K-D of 102 pM. In addition, HSY cell s were shown to express the mRNA for FGF receptor-1 (FGFR-1) by revers e transcription-polymerase chain reaction (RT-PCR), confirming the exi stence of high-affinity FGF binding sites. The influence of endogenous FGF-1 and FGF-2 on HSY cell growth was evaluated by suppressing the e xpression and activity of FGF by using anti-sense oligonucleotides and neutralizing antibodies. The addition of 50 mu M FGF-1-specific anti- sense oligonucleotides to HSY cells resulted in a 61% inhibition of ce ll growth, while 50 mu M FGF-2-specific anti-sense oligonucleotides re sulted in a 76% inhibition. These effects were dose-dependent and spec ific, since sense oligonucleotides were ineffective in inhibiting HSY cell growth at the same concentration. Furthermore, HSY cell growth wa s suppressed in the presence of anti-FGF-1 or anti-FGF-2 neutralizing antibody, resulting in a 58% inhibition at 8 mu mg/ml. Our observation s suggest that FGF-1 and FGF-2 may act as autocrine regulators by inte racting with FGF receptors on HSY cells. (C) 1996 Wiley-Liss, Inc.