Abstract
Many common diseases involve the injury, loss, or death of organ tissues. For these patients, organ transplantation is often the only viable solution. Nonetheless, organ transplantation is seriously limited by the relative scarcity of living and non-living donors, a situation that is worsening with aging of the world population. Tissue Engineering (TE) is a research discipline in regenerative medicine that aims to generate tissues in the laboratory that can replace diseased and damaged tissues in patients. Crucially, engineered tissues must have a vascular network that guarantees adequate nutrient supply, gas exchange, and elimination of waste products. Therefore, the search for clinically relevant sources of vasculogenic cells and the subsequent development of methods to achieve rapid vascularization is of utmost importance. We and others have previously shown that human blood-derived endothelial colony-forming cells (ECFCs) have the required vasculogenic capacity to form functional vascular networks in vivo. These studies demonstrated that, in the presence of an appropriate source of perivascular cells, ECFCs can self-assemble into microvascular networks and connect to the host vasculature, a process that takes approximately 7days in vivo. The prospect is to incorporate these vascular networks into future engineered tissues. However, engineered tissues must have a functional vasculature immediately after implantation in order to preserve viability and function. Thus, it is critical to further develop strategies for rapid formation of perfused vascular network in vivo. Here, we describe a methodology to deliver ECFCs and bone marrow-derived mesenchymal stem cells (MSCs) subcutaneously into immunodeficient mice in the presence of fibroblast growth factor-2 (FGF-2). This approach significantly reduces the time needed to achieve functional anastomoses between bioengineered human blood vessels and the host vasculature. This methodology includes (1) isolation, characterization and culture of ECFCs, (2) isolation, characterization and culture of MSCs, and (3) implantation of ECFCs and MSCs, in the presence of FGF-2, into immunodeficient mice to generate perfused vascular networks.