Up-regulation of Bcl-2 in microvascular endothelial cells enhances intratumoral angiogenesis and accelerates tumor growth

Vascular endothelial growth factor (VEGF) has been shown to be a potent med iator of angiogenesis that functions as a survival factor for endothelial c ells by up-regulating Bcl-2 expression. We have recently reported that huma n dermal microvascular endothelial cells (HDMECs) seeded in biodegradable s ponges and implanted into severe combined immunodeficient (SCID) mice organ ize into functional human microvessels that transport mouse blood cells. In this study, we implanted sponges seeded with OSCC-3 (oral squamous cell ca rcinoma) or SLK (Kaposi's sarcoma) together with endothelial cells into SCI D mice to generate human tumors vascularized with human microvessels, This model system was used to examine the rule or both endothelial cell Bcl-2 an d the proangiogenic chemokine interleukin-8 (IL-8) on tumor growth and intr atumoral microvascular density. Coimplantation of HDMECs overexpressing Bcl -2 (HDMEC-Bcl-2) and tumor cells resulted in a 3-fold enhancement of tumor growth when compared with the coimplantation of control HDMECs and tumor ce lls, This was associated with increased intratumoral microvascular density and enhanced endothelial cell survival. To determine whether the enhanced n eovascularization mediated by Bcl-2 overexpression in endothelial cells was influenced by the synthesis of endogenous mediators of angiogenesis, we sc reened these cells for expression of VEGF, basic fibroblast growth factor ( hFGF), and IL-8 by ELISA. HDMEC-Bcl-2 cells and VEGF-treated HDMECs exhibit ed a 15-fold and 4-fold increase, respectively, in the expression of the pr oangiogenic chemokine IL-8 in vitro, whereas the expression of VEGF and bFG F remained unchanged. Transfection of antisense Bcl-2 into HDMECs blocked V EGF-mediated induction of IL-8. Conditioned media from HDMEC-Bcl-2 induced proliferation and sprouting of endothelial cells in vitro and neovasculariz ation in rat corneas. Anti-IL-8 antibody added to HDMEC-Bcl-2 conditioned m edia markedly reduced the potency of these responses. SCID mice bearing VEG F-producing tumor implants that were treated with anti-IL-8 antibody exhibi ted a 43% reduction in microvessel density and a 50% reduction in tumor wei ght compared with treatment with a nonspecific antibody. These results demo nstrate that the up-regulation of Bcl-2 expression in endothelial cells tha t constitute tumor microvessels enhances intratumoral microvascular surviva l and density and accelerates tumor growth. Furthermore, endothelial cells that overexpress Bcl-2 have more angiogenic potential than control cells, a nd IL-8-neutralizing antibodies attenuate their angiogenic activity in vitr o and in vivo.