Melanoma cell-derived factor stimulation of fibroblast glycosaminoglycan synthesis - The role of platelet-derived growth factor

The hyaluronan-rich matrix surrounding many tumours may facilitate tumour g rowth, invasion and angiogenesis, with the majority of this hyaluronan appa rently being synthesised by normal fibroblasts, stimulated to do so by tumo ur cell-derived factors. Melanoma cell-conditioned medium (CIM) stimulates up to a 6-fold increase in fibroblast glycosaminoglycan (GAG) synthesis, wi th the active factors being present in tumour CIM ultrafiltration fractions > 30 kDa and < 1 kDa. These fractions are poorly active individually, but when recombined, the activity is substantially greater than the additive ef fect. The objective of this study was to identify the factors present in th e ultrafiltration fraction > 30 kDa that produce a greater than additive ef fect with the fraction < 1 kDa in stimulating the incorporation of H-3 gluc osamine into fibroblast GAGs. A number of factors including basic fibroblas t growth factor (bFGF), interleukin (IL)-1 beta, pleiotrophin, platelet-der ived growth factor (PDGF), transforming growth factor-beta (TGF-beta), tumo ur necrosis factor-alpha (TNF-alpha) and vascular endothelial growth factor (VEGF) failed to stimulate any significant increase in GAG synthesis, but when added to the < 1 kDa tumour CIM fraction, both PDGF and to a lesser ex tent, bFGF, exhibited potent stimulating activities. Neutralising antibodie s to PDGF and bFGF added to the melanoma CIM decreased the fibroblast GAG-s timulating activity by 29% and 40%, respectively, in C8161 melanoma CIM and by 47% and 45%, respectively, in Hs294T melanoma CIM. The activities of PD GF-AA and PDGF-BB isoforms were indistinguishable, suggesting the PDGF-alph a receptor plays a role in the GAG-stimulatory response. Western analysis f ollowing treatment with PDGF, bFGF or melanoma CM revealed banding patterns for PDGF and melanoma CIM that were similar. Immunoprecipitation of the PD GF-alpha receptor revealed it to be phosphorylated in fibroblasts treated w ith PDGF and melanoma CM, but not with control fibroblast CIM. These studie s suggest that PDGF plays an important role in the GAG-stimulating activity of the melanoma CIM, but requires the presence of an as yet unidentified n ovel low molecular weight factor for full activity. (C) 1999 Elsevier Scien ce Ltd. All rights reserved.