Angiogenic balance in human melanoma: Expression of VEGF, bFGF, IL-8, PDGFand angiostatin in relation to vascular density of xenografts in vivo

Tumor angiogenesis, a major requirement for tumor outgrowth and metastasis formation, is regulated by pro- and anti-angiogenic factors. We have studie d the expression of a panel of angiogenic factors, and of the angiogenesis inhibitor angiostatin, in a panel of human melanoma cell lines giving rise to xenografts with different vascular densities. Angiogenic-factor expressi on was analyzed in vitro (cell lines) and in vivo (xenografts), both at mRN A (RT-PCR and Northern blot) and at protein level (ELISA and Western blot). In vitro angiostatin generation was assessed by Western-blot analysis. Exp ression of bFGF and VEGF was clearly correlated with a high degree of vascu larization, confirming the importance of these factors for tumor angiogenes is. In addition, there was exclusive or elevated in vitro expression of ang iogenic factors IL-8, PDGF-AB, and, to a lesser extent, midkine in cell lin es that formed highly vascularized tumors. A similar angiogenic-factor-expr ession pattern was found in the corresponding xenografts, with the exceptio n of VEGF. in most cell lines, this factor had low expression in vitro whic h was strongly enhanced in vivo. Although all 8 melanoma cell lines were ab le to excise the angiostatin fragment from the plasminogen parent molecule in vitro, cell lines BLM and M14 showed the most potent angiostatin generat ion. In vitro angiostatin generation by cell lysates prepared from melanoma xenografts was comparable in all xenograft types. Thus, in our model syste m we found no correlation between angiostatin generation and vascular densi ty. Our study has limited the number of pro-angiogenic factors that may be involved in melanoma angiogenesis, and provides evidence for the notion tha t regulation of tumor angiogenesis is dependent on multiple factors. Inhibi tion of angiogenesis for therapeutic purposes, therefore, should preferably not concentrate on a single factor. (C) 2000 Wiley-Liss, Inc.