Hypoxia-induced angiogenesis and vascular endothelial growth factor secretion in human melanoma

Abstract

Summary Tumour cells exposed to hypoxia in vitro can show increased expression of several selected genes, including the gene encoding the vascular endothelial growth factor (VEGF), suggesting that hypoxia followed by reoxygenation might promote the malignant progression of tumours. An in vitro/in vivo study was conducted to investigate whether hypoxia can increase the angiogenic potential of tumour cells through increased VEGF secretion. Four human melanoma cell lines (A-07, D-12, R-18, U-25) were included in the study. Cell cultures were exposed to hypoxia (oxygen concentration <10 p.p.m.) in vitro using the steel chamber method. Rate of VEGF secretion was measured in vitro in aerobic and hypoxic cell cultures by ELISA. Angiogenesis was assessed in vivo using the intradermal angiogenesis assay. Aliquots of cells harvested from aerobic cultures or cultures exposed to hypoxia for 24 h were inoculated intradermally in the flanks of adult female BALB/cnu/nu mice. Tumours developed and angiogenesis was quantified by scoring the number of capillaries in the dermis oriented towards the tumours. The number of tumour-oriented capillaries did not differ significantly between tumours from hypoxic and aerobic cultures for A-07 and U-25, whereas tumours from hypoxic cultures showed a larger number of tumour-oriented capillaries than tumours from aerobic cultures for D-1 2 and R-1 8. The VEGF secretion under aerobic conditions and the absolute increase in VEGF secretion induced by hypoxia were lower for D-12 and R-18 than for A-07 and U-25, whereas the relative increase in VEGF secretion induced by hypoxia was higher for D-12 and R-18 than for A-07 and U-25. VEGF is not a limiting factor in the angiogenesis of some tumours under normoxic conditions. Hypoxia can increase the angiogenic potential of tumour cells by increasing the secretion of VEGF, but only of tumour cells showing low VEGF secretion under normoxia. Transient hypoxia might promote the malignant progression of tumours by temporarily increasing the angiogenic potential of tumour cells showing low VEGF expression under normoxic conditions. Keywords: angiogenesis; hypoxia; melanoma; vascularization; VEGF Many malignant tumours develop regions of hypoxic cells during growth (Coleman, 1988;Vaupel et al, 1989;Brown and Giaccia, 1994). Two types of hypoxia have been recognized: chronic hypoxia, arising from limitations in oxygen diffusion, and acute hypoxia, resulting from transient stoppages in microregional blood flow (Stone et al, 1993;Horsman, 1995). Reoxygenation of hypoxic cells occurs during unperturbed tumour growth as a result of reopening of temporarily closed vessels and during therapy as a result of therapy-induced tumour cell inactivation (Kallman, 1972;Brown, 1979;Chaplin et al, 1987). Hypoxia followed by reoxygenation might promote the malignant progression of tumours (Hill, 1990). Thus, tumour cells exposed to hypoxia in vitro can show increased expression of several selected genes, including genes encoding cell cycle-regulatory proteins, haematopoietic...

The tumor microenvironment and metastatic disease

“…Its receptors, VEGFR1 and-2, are also induced under hypoxic conditions [124]. The role of this protein in metastatic disease has been extensively examined, with some studies suggesting a link between VEGF-A expression and metastatic disease [23,125,126], and others not [127,128], consistent with the concept that different cell types may demonstrate tumor specific effects. However, in driving the development of neo-vasculature VEGF-A may provide a mechanism of transport for the tumor cells as well as possibly enhancing the intravasation and extravasation stages of the metastatic process due to increased vascular permeability.…”

Vascular Endothelial Growth Factor (VEGF) as a Target of Bevacizumab in Cancer: From the Biology to the Clinic

“…Both VEGF-C and VEGF-D are more selective growth factors for the endothelium of lymphatic vessels and also bind to the VEGF-3 receptor [19,20].…”

True

“…Although different VEGF levels in tumours can originate as a result of hypoxia, VEGF secretion and mRNA levels do also differ significantly between tumour cells of the same histological origin grown under normoxic conditions in vitro (Westphal et al, 1997;Rofstad and Danielsen, 1998). Thus, it seems likely that genetic changes during tumour progression modulate the VEGF expression of the tumour cells.…”