Melanoma cell migration in vitro can be blocked in a modified Boyden chamber assay using the anti-angiogenic endogenous sister isoform VEGF165b.


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Daemon Dewing1, Maxine Emmett1, Paul McArthur1, Rowan Pritchard Jones1
1University of Liverpool, Merseyside, United Kingdom

Background

Background: Melanoma is the most lethal skin cancer with 2000 deaths annually in the UK. Angiogenesis, the growth of new blood vessels, is an absolute pre-requisite for tumour growth and metastasis. Central to this process is Vascular Endothelial Growth Factor (VEGF)[1]. The alternative splicing of the VEGF gene results in pro- (eg VEGF165) or anti-angiogenic isoforms (eg VEGF165b) [2]. This latter isoform binds at VEGF-R2 as an equipotent inhibitor of pro-angiogenic VEGF.

VEGF stimulates endothelial cell migration, sprouting of blood vessels and generates new vessel formation within tumours. Significantly, we have demonstrated that VEGF also stimulates melanoma cell migration.

Method

Methods: Using a modified Boyden chamber, migration of metastatic A375 melanoma cells was assessed in response to pro-angiogenic VEGF165 and anti-angiogenic VEGF165b, through a collagen coated porous membrane.

Results

Results: We demonstrated mean ±SEM dose dependent migration of 1.49±0.15 and 10.37±0.94 fold increases over the control, when using VEGF165 at concentrations of 20ng/ml and 160ng/ml respectively (ANOVA p<0.001). Furthermore, this VEGF165 induced migration can be completely blocked using equipotent anti-angiogenic VEGF165b in a dose dependent fashion.

Conclusion

Conclusion: Our data demonstrates, for the first time, not only that tumour cells migrate in a dose dependent manner, and hence potentially metastasise in response to pro-angiogenic VEGF, but that migration is inhibited by endogenous anti-angiogenic VEGF165b in a similarly dose dependent manner. This further supports our endeavor to consider the therapeutic possibilities of this molecule.