Lymphangiogenesis and its regulation in human neuroblastoma.

For the first time, we could detect lymph vessels in neuroblastoma (NB) by immunohistochemistry with the antibody D2_40. Furthermore, we demonstrate expression of the lymphangiogenic factors VEGF-C and VEGF-D and their receptors VEGFR-2 and VEGFR-3 in NB in vitro and in vivo by RT-PCR. However, addition of recombinant human VEGF-C or -D results in the absence of autocrine growth stimulus in NB cells. Treatment of NB cells with retinoic acid did not lead to a change in VEGF-C or VEGF-D mRNA expression. Incubation of the NB cells Lan-5 with 5-Aza-2'-deoxycytidine led to the up-regulation of VEGF-C mRNA expression, suggesting that the promotor of VEGF-C is methylated. Finally, VEGF-C mRNA expression could be effectively down-regulated by transfection with a specific siRNA in the NB cells Kelly. We conclude that lymphangiogenesis is involved in NB biology and that siRNA directed against VEGF-C may have a future role in anti-lymphangiogenic strategies in NB.