Elucidating the molecular mechanisms of VEGF signaling in vasculogenesis.
During embryonic development, pluripotent stem cells (PSCs) differentiate to mesoderm and then in a process called vasculogenesis, mesodermal precursors differentiate into ECs and form a primary vascular plexus. It is well documented that VEGF (Vascular Endothelial Growth Factor) plays a crucial role in vasculogenesis (Carmeliet et al., Nature 1996, 380(6573):435-9),(Ferrara N et al., Nature 1996, 380(6573):439-42). However, there is a scientific gap at this specific point with almost no information concerning the cascades of VEGF that induce differentiation of human mesodermal progenitor cells to vascular progenitor cells. Given the undoubtable necessity of VEGF in embryonic development of the vasculature and previous work from our group on VEGF signal transduction in primary endothelial cells and in angiogenesis (Angiogenesis section), we are very keen to elucidate VEGF pathways in vasculogenesis. For that reason, we developed a differentiation protocol of PSCs to vascular progenitor cell (VPCs), where the commitment to endothelial lineage solely depends on VEGF (Vascular progenitor cells (VPCs) derived from hESCs, hiPSCs in feeder free and serum free conditions). A detailed characterization of PSC-VPCs was attained by performing proteomics using high resolution mass spectrometry combined with multivariate analysis. In order to explore the molecular mechanism by which VEGF drives differentiation of mesodermal progenitor cells to endothelial cells, it is also necessary to investigate the signaling events which take place just after the administration of VEGF. Bioinformatic analysis of the VEGF-induced phosphorylated proteins and the concomitant gene expression pattern will provide information on VEGF-dependent signaling cascades and transcription factors that are involved in the differentiation to VPCs. The above complementary approaches, using “omics” methods both shortly (during the first 2 hours) and at a later time point (48 hours) following VEGF administration combined with additional gain-of-function and loss-of-function-experiments, will allow us to gain a thorough knowledge of VEGF signaling during vasculogenesis. These projects are ongoing in the lab and can be summarized as follows:
- Understanding the overall organization of VPCs differentiation to ECs by VEGF
- Determining the critical signalling pathway(s) responsible for this differentiation
- Dissecting the interconnection between differentiation and signalling/trafficking of VEGF and its receptors
- Eventually, exploring the translational potential of the identified regulatory circuits in anti-angiogenic or pro-angiogenic applications
|Model of the transition of hESCs into differentiated cells. hESCs differentiate into mesodermal intermediates and then into CD34+ VPCs and CD34− cells, under feeder-free conditions, using defined media. CD34+ synthesize known vascular cell markers, as well as proteins related to vasculogenesis and smooth muscle differentiation, suggesting a bipotent phenotype. Indeed, CD34+ can differentiate further to both directions depending on the growth conditions. CD34− cells consist of a mixed population over-synthesizing secreted factors that can promote angiogenesis, suggesting a paracrine effect on CD34+ cells and also synthesizing proteins that can lead to differentiation to other lineages (muscle, bone, cartilage).|