DIFFERENT EFFECTS OF MUCOSAL, BOVINE LUNG AND CHEMICALLY-MODIFIED HEPARIN ON SELECTED BIOLOGICAL PROPERTIES OF BASIC FIBROBLAST GROWTH-FACTOR

Heparins from bovine mucosa and lung, and chemically modified heparins were assayed for their capacity to: (i) protect human recombinant bas ic fibroblast growth factor (bFGF) from tryptic cleavage; (ii) prevent I-125-bFGF binding to heparan sulphate proteoglycans present in the e xtracellular matrix and on the eel surface of fetal bovine aortic endo thelial GM 7373 cell cultures, (iii) affect I-125-bFGF binding to high -affinity tyrosine kinase FGF receptors present on the cell membrane o f GM 7373 cells; (iv) inhibit the mitogenic activity exerted by bFGF i n the same cells. The results demonstrate that the potency shown by mu cosal heparins in the different assays is a direct function of size, v ery-low-molecular-mass heparin (2.0 kDa) being significantly less effe ctive on a molar basis than unfractionated heparin (13.6 kDa). Increas ed flexibility of the backbone structure, as observed in reduced/oxidi zed heparins of different size, does not affect the capacity of the po lysaccharide to interact with bFGF. In contrast, selective 2-O-desulph ation, but not 6-O-desulphation, drastically reduced the capacity of h eparin to protect bFGF from proteolytic cleavage, to affect its intera ction with low- and high-affinity sites, and to inhibit its mitogenic activity. Two preparations of bovine lung heparin, differing in molecu lar mass, were as effective as mucosal heparin in the bFGF-tryptic-dig estion assay and the endothelial-cell proteoglycan-binding assay, but they were highly inefficient at inhibiting the capacity of bFGF to int eract with its tyrosine kinase receptors. Bovine lung heparins were al so less effective than mucosal heparin as bFGF antagonists in GM 7373- cell-proliferation assays. N-Desulphated/N-acetylated bovine lung hepa rin retained only a significant capacity to protect bFGF from tryptic cleavage. The results demonstrate that different chemical features of the heparin molecule, including decrease in molecular mass, selective desulphation, disaccharide composition and clustering, affect differen tly the capacity of the glycosaminoglycan to interact with bFGF and to influence its biological behaviour in different assays in vitro and i n endothelial cell cultures. Our findings should aid the design of syn thetic oligosaccharides aimed at improving the,bioavailability of bFGF when administered in vivo as a therapeutic agent.