Mesenchymal stromal cells enhance self-assembly of a HUVEC tubular network through uPA-uPAR/VEGFR2/integrin/NOTCH crosstalk

Endothelial cells (ECs) degrade the extracellular matrix of vessel walls and contact surrounding cells to facilitate migration during angiogenesis, leading to formation of an EC-tubular network (ETN). Mesenchymal stromal cells (MSC) support ETN formation when co-cultured with ECs, but the mechanism is incompletely understood. We examined the role of the urokinase-type plasminogen activator (uPA) system, i.e. the serine protease uPA, its inhibitor PAI-1, receptor uPAR/CD87, clearance by the low-density lipoprotein receptor-related protein (LRP1) and their molecular partners, in the formation of ETNs supported by adipose tissue-derived MSC. Co-culture of human umbilical vein ECs (HUVEC) with MSC increased mRNA expression levels of uPAR, MMP14, VEGFR2, TGFβ1, integrin β3 and Notch pathway components (Notch1 receptor and ligands: Dll1, Dll4, Jag1) in HUVECs and uPA, uPAR, TGFβ1, integrin β3, Jag1, Notch3 receptor in MSC. Inhibition at several steps in the activation process indicates that uPA, uPAR and LRP1 cross-talk with αv-integrins, VEGFR2 and Notch receptors/ligands to mediate ETN formation in HUVEC-MSC co-culture. The urokinase system mediates ETN formation through the coordinated action of uPAR, uPA's catalytic activity, its binding to uPAR and its nuclear translocation. These studies identify potential targets to help control aberrant angiogenesis with minimal impact on healthy vasculature. •MSC form fibronectin scaffold to support tube formation (ETN) by HUVEC in co-culture•uPA/uPAR cross-talk with avb3 integrins, VEGFR2 and Notch to mediate ETN formation•Urokinase mediates ETN formation through proteolitic activity and nuclear translocation.