Slit2N and Robo4 regulate lymphangiogenesis through the VEGF-C/VEGFR-3 pathway

BACKGROUND: Signaling through vascular endothelial growth factor C (VEGF–C) and VEGF receptor 3 (VEGFR-3) plays a central role in lymphangiogenesis and the metastasis of several cancers via the lymphatics. Recently, the Slit2/Robo4 pathway has been recognized as a modulator of vascular permeability and integrity. Signaling via the Robo receptor inhibits VEGF-mediated effects; however, its effects on lymphatic endothelial cell function have not been well characterized. RESULTS: We found that pretreatment with Slit2N, an active fragment of Slit2, inhibited VEGF-C-mediated lung-derived lymphatic endothelial cell (L-LEC) proliferation, migration, and in vitro tube formation. Slit2N induced the internalization of VEGFR-3, which blocked its activation, and inhibited the activation of the PI3K/Akt pathway by VEGF-C in L-LECs. Moreover, we found that inhibition of VEGF-C-induced effects by Slit2N was Robo4-dependent. CONCLUSION: These results indicate that Slit2N/Robo4 modulates several key cellular functions, which contribute to lymphangiogenesis, and identify this ligand-receptor pair as a potential therapeutic target to inhibit lymphatic metastasis of VEGF-C-overexpressing cancers and manage lymphatic dysfunctions characterized by VEGF-C/VEGFR-3 activation.