Tumor vascularization is a hallmark of cancer central to disease progression and metastasis. Current anti-angiogenic therapies have limited success prompting the need to better understand the cellular origin of tumor vessels. Using fate-mapping analysis of endothelial cell populations in melanoma, we report the very early infiltration of endovascular progenitors (EVP) in growing tumors. These cells harbored self-renewal and reactivated the expression of SOX18 transcription factor, initiating a vasculogenic process as single cells, progressing towards a transit amplifying stage and ultimately differentiating into more mature endothelial phenotypes that comprised arterial, venous and lymphatic subtypes within the core of the tumor. Molecular profiling by RNA sequencing of purified endothelial fractions characterized EVPs as quiescent progenitors remodeling the extracellular matrix with significant paracrine activity promoting growth. Functionally, EVPs did not rely on VEGF-A signaling whereas endothelial-specific loss of Rbpj depleted the population and strongly inhibited metastasis. The understanding of endothelial heterogeneity opens new avenues for more effective anti-vascular therapies in cancer.

Endothelial to mesenchymal transition (EndMT) is a leading cause of fibrosis and disease, however its mechanism has yet to be elucidated. The endothelium possesses a profound regenerative capacity to adapt and reorganize that is attributed to a population of vessel-resident endovascular progenitors (EVP) governing an endothelial hierarchy. Here, using fate analysis, we show that two transcription factors SOX9 and RBPJ specifically affect the murine EVP numbers and regulate lineage specification. Conditional knock-out of Sox9 from the vasculature (Sox9fl/fl/Cdh5-CreERRosaYFP) depletes EVP while enhancing Rbpj expression and canonical Notch signalling. Additionally, skin wound analysis from Sox9 conditional knock-out mice demonstrates a significant reduction in pathological EndMT resulting in reduced scar area. The converse is observed with Rbpj conditionally knocked-out from the murine vasculature (Rbpjfl/fl/Cdh5-CreER RosaYFP) or inhibition of Notch signaling in human endothelial colony forming cells, resulting in enhanced Sox9 and EndMT related gene (Snail, Slug, Twist1, Twist2, TGF-β) expression. Similarly, increased endothelial hedgehog signaling (Ptch1fl/fl/Cdh5-CreER RosaYFP), that upregulates the expression of Sox9 in cells undergoing pathological EndMT, also results in excess fibrosis. Endothelial cells transitioning to a mesenchymal fate express increased Sox9, reduced Rbpj and enhanced EndMT. Importantly, using topical administration of siRNA against Sox9 on skin wounds can substantially reduce scar area by blocking pathological EndMT. Overall, here we report distinct fates of EVPs according to the relative expression of Rbpj or Notch signalling and Sox9, highlighting their potential plasticity and opening exciting avenues for more effective therapies in fibrotic diseases.

The cardiovascular system is composed around the central function of the endothelium that lines the inner surfaces of its vessels. In recent years, the existence of a progenitor population within the endothelium has been validated through the study of endothelial colony-forming cells (ECFCs) in human peripheral blood and certain vascular beds. However, our knowledge on endothelial populations in vivo that can give rise to ECFCs in culture has been limited. In this review we report and analyse recent attempts at describing progenitor populations in vivo from murine studies that reflect the self-renewal and stemness capacity observed in ECFCs. We pinpoint seminal discoveries within the field, which have phenotypically defined, and functionally scrutinised these endothelial progenitors. Furthermore, we review recent publications utilising single-cell sequencing technologies to better understand the endothelium in homeostasis and pathology.

The aggressiveness of solid cancers, such as melanoma, relies on their metastatic potential. It has become evident that this key cause of mortality is largely conferred by the tumour-associated stromal cells, especially endothelial cells. In addition to their essential role in the formation of the tumour vasculature, endothelial cells significantly contribute to the establishment of the tumour microenvironment, thus enabling the dissemination of cancer cells. Melanoma tumour vascularization occurs through diverse biological processes. Vasculogenesis is the formation of de novo blood vessels from endothelial progenitor cells (EPCs), and recent research has shown the role of EPCs in melanoma tumour vascularization. A more detailed understanding of the complex role of EPCs and how they contribute to the abnormal vessel structures in tumours is of importance. Moreover, anti-angiogenic drugs have a limited effect on melanoma tumour vascularization, and the role of these drugs on EPCs remains to be clarified. Overall, targeting cancer vasculature remains a challenge, and the role of anti-angiogenic drugs and combination therapies in melanoma, a focus of this review, is an area of extensive exploration.