α-Melanocyte-stimulating hormone inhibits angiogenesis through attenuation of VEGF/VEGFR2 signaling pathway.

ABSTRACT

BACKGROUND: Gene therapy of proopiomelanocortin, the precursor of α-melanocyte-stimulating hormone (α-MSH), suppresses the neovascularization in tumors. However, the roles of α-MSH in angiogenesis remain unclear. METHODS: The influence of α-MSH on angiogenesis was evaluated by ex vivo rat aorta and in vivo, including transgenic zebrafish and chicken chorioallantoic membrane (CAM) assays. The effect of α-MSH on proliferation, matrix metalloproteinase (MMP) secretion, migration and tube formation was examined using human umbilical vein endothelial cells (HUVECs). The expression of vascular endothelial growth factor (VEGF) and VEGF receptor 2 (VEGFR2) was investigated by quantitative RT-PCR, immunoblot and immunofluorescent analysis. Antibodies' neutralization was employed to dissect the receptor(s) transmitting α-MSH signaling. RESULTS: Application of α-MSH potently suppressed the microvessels sprouting in organotypic aortic rings. Besides, α-MSH perturbed the vessels development in zebrafish and chicken embryos. α-MSH (0.01-10nM) inhibited the MMP-2 secretion, migration and tube formation of HUVECs without affecting proliferation. Mechanistic studies unveiled α-MSH decreased the VEGF expression and release in HUVECs. Besides, α-MSH downregulated the VEGFR2 expression at transcriptional and translational levels. Importantly, α-MSH attenuated the Akt phosphorylation, but enhanced the expression of PTEN, endogenous antagonist of PI3K/Akt signaling. Expression analysis and antibody neutralization revealed that MC1-R and MC2-R participated in α-MSH-induced blockage of migration and VEGF/VEGFR2/Akt signaling. However, VEGF supply failed to reverse the anti-angiogenic function of α-MSH. CONCLUSIONS: α-MSH inhibits the physiological angiogenesis by attenuating VEGF/VEGFR2/Akt signaling in endothelial cells. GENERAL SIGNIFICANCE: α-MSH is a potent angiogenesis inhibitor targeting at endothelial VEGF/VEGFR2 signaling, which may have potential for therapeutic application.