Multiple myeloma related angiogenesis: role of notch signaling pathway

The progression of multiple myeloma (MM) is associated to a angiogenic switch. MM shows a dysregulation of Notch pathway due to overexpressed Jag1 and Jag2 ligands. The presence of Jag1/2 promotes interactions between MM cells and BM resident cells carrying Notch receptors, including BM stromal cells (BMSCs), while the outcome on endothelial cells (ECs) still needs to be clarified. This study aims to investigate the role of Notch signaling in MM-associated angiogenesis evaluating the contribution of MM cells and BMSCs. We analyzed the interplay between MM cell lines, RPMI8226 and U266, the ECs (HPAECs) and the BMSC line HS5. Jag1 and 2 were silenced in MM cell lines (MMshJAG1/2) by specific siRNAs and we investigated the outcome on the ability of MM cells to alter ECs adhesion on fibronectin substrate, ECs ability to form tube-like structures (matrigel assay) and ECs motility (wound healing assay). We analyzed the effect of a direct interaction MM cells-ECs or MM-derived conditioned media (CM). Secondly, we analyzed the outcome of MM cell ability to activate the Notch signaling in BMSCs and induce VEGF secretion. At this purpose we used CM from MM-BMSC co-cultures. Our results indicate that MM cells improve angiogenesis, adhesion and migration of ECs through both direct contact and CM indicating also the involvement of soluble factors. These effects are significantly reduced in the absence of Jag ligands on MM cells, indicating the requirement of Notch signaling activation in ECs. MM cells are able to induce BMSCs to increase VEGF production, while MMshJAG1/2 cells are not, indicating again the necessity of Notch signaling activation These findings are a first indication that MM cells may promote tumor-associated angiogenesis by shaping the nearby microenvironment via a Notch-directed cell-cell communication and the release of soluble factors.