In vivo engraftment of vascular structures in 3D-engineered artificial tissues vascularized by cord blood- or peripheral blood-derived endothelial colony-forming cells

Abstract

Constructing human vascularized artificial tissues (HVATs) that form stable vascular structures after transplantation remains challenging in regenerative medicine. Using the cryopreserved cell accumulation method (CP-CAM), a scaffold-free approach based on extracellular matrix nanofilm-coated cells, we generated HVATs from human adipose tissue-derived mesenchymal stromal cells (hASCs) and cord blood- or peripheral blood-derived endothelial colony-forming cells (hCB-ECFCs and hPB-ECFCs). Both ECFCs formed vascular networks within four days after seeding, with no significant differences in two-dimensional vascular length density or area fraction compared with HUVEC-based constructs. hCB-ECFC-derived vascular structures showed incomplete endothelial closure and reduced perivascular differentiation of hASCs in vitro, whereas hPB-ECFC-derived structures exhibited more continuous vascular morphology. Angiogenesis-related factor profiling of HVAT supernatants suggested distinct patterns among endothelial cell sources. After subcutaneous transplantation into nude mice, hCB-ECFC-derived vessels exhibited continuous endothelial lumens and surrounding pericyte- or smooth muscle-like cells, with further structural maturation over four weeks. HVATs constructed with hPB-ECFCs also engrafted and formed human-derived vascular structures containing host blood cells and surrounded by perivascular cells. These findings demonstrate that both hCB-ECFCs and hPB-ECFCs can be used to construct transplantable HVATs by CP-CAM and support the potential of blood-derived ECFCs as endothelial cell sources for vascularized tissue engineering and therapeutic applications.