Indirect angiogenic agents do not release fibroblast growth factors from extracellular matrix

Vascular growth factors are categorized as either primary or secondary angiogenic factors. Primary angiogenic agents such as fibroblast growth factors, not only induce the complete angiogenic response, but also stimulate the individual components of vascular growth. Secondary angiogenic agents can induce vascular growth, but they do not act through the direct stimulation of endothelial proliferation, migration, and protease production. Since fibroblast growth factors are known to bind to components of the extracellular matrix, we assessed whether secondary agents act through liberating growth factors from matrix storage sites. The study utilized L6 skeletal myoblasts in culture, which we demonstrated were capable of synthesizing extracellular matrix containing heparin binding endothelial mitogens. The heparin-binding mitogenic activity accumulated in a time-dependent fashion, and matrix extracts contained a protein with immunologic identity to acidic fibroblast growth factor. The ability of secondary angiogenic agents and related compounds including adenosine, inosine, hypoxanthine, nicotinamide, lactic acid, phorbol esters, prostaglandin E2, and copper (at concentrations of 1 microM and 1 mM) to release heparin binding mitogenic activity from the matrix was evaluated. The results demonstrate that although heparin is capable of releasing heparin-binding growth factors from extracellular matrix storage sites in a dose dependent fashion, none of the known secondary angiogenesis factors are capable of functioning in a similar fashion. Thus these secondary angiogenic factors do not appear to exert their effect through increasing the bioavailability of preformed heparin-binding growth factors sequestered in the extracellular matrix. The mechanism(s) whereby these agents induce vascular growth remains to be elucidated.