Adrenomedullin expression and regulation in human glioblastoma, cultured human glioblastoma cell lines and pilocytic astrocytoma.

Clinical and experimental studies suggest that angiogenesis is a prerequisite for solid tumour growth. Glioblastoma (GBM) and pilocytic astrocytoma (PA), both angiogenic tumours display strong contrast enhancement associated with peripheral oedema in GBM but not in PA indicating differences in vascular permeability in these two types of gliomas. Here we show that expression of adrenomedullin (AM) mRNA is induced in GBM whereas is barely detectable in PA. In situ analysis of tumour specimens undergoing neovascularisation shows that the production of AM is specifically induced in a subset of GBM cells distinguished by their immediate proximity to necrotic foci (presumably hypoxic regions), suggesting a hypoxic induction of AM expression in GBM. Vascular endothelial growth factor (VEGF) mRNA levels are increased in GBM and moderate in PA. Immunohistochemical study showed that cytoplasmic AM, VEGF and HIF-1α nuclear immunoreactivity were recorded in GBM located near large necrotic areas whereas they were not expressed by PA tumour cells. Interestingly, double fluorescence immunostaining demonstrated that 85% of AM immunoreactivity colocalised with VEGF. AM transduces its effects through calcitonin receptor-like receptor/receptor activity modifying protein-2 and -3 (CLR/RAMP2 and CLR/RAMP3). Real-time quantitative RT-PCR showed expression of RAMP2, RAMP3 and CLR in PA and GBM, suggesting that AM may function as an autocrine/paracrine growth factor for GBM cells. These observations strongly support the concept that tumour angiogenesis is regulated by paracrine mechanisms and identify beside VEGF, AM as a potential tumour angiogenesis factor in vivo which constitutes a potential interesting molecular target in GBM treatment.