3D Culture Facilitates VEGF-Stimulated Endothelial Differentiation of Adipose-Derived Stem Cells

De novo vascularization of implantable tissue and whole organ constructs has been a significant challenge in the field of tissue engineering; the use of endothelial cell populations for this task is constrained by the cell population's limited regeneration capacity and potential for loss of function. Thus, there is a need for a stem-cell population that may be induced into an endothelial cell phenotype reliably. Adipose derived stem cells (ADSCs) are multipotent cells that can be readily isolated from donor fat and may have the potential to be readily induced into endothelial cells. The ability to stimulate endothelial differentiation of these cells has been limited in standard 2D culture. We hypothesized that 3D culture would yield better differentiation. To study the influence of cell density and culture conditions on the potential of ADSCs to differentiate into an endothelial-like state, we seeded these cells types within a 3D cell-adhesive, proteolytically degradable, peptide-modified poly(ethylene-glycol) (PEG) hydrogel. ADSCs were either cultured in basal media or pro-angiogenic media supplemented with 20 ng/mL of VEGF in 2D and then encapsulated at low or high densities within the PEG-based hydrogel. These encapsulated cells were maintained in either basal media or pro-angiogenic media. Cells were then isolated from the hydrogels and cultured in Matrigel to assess the potential for tubule formation. Our work shows that maintenance of ADSCs in a pro-angiogenic medium in 2D monoculture alone does not result in any CD31 expression. Furthermore, the level of CD31 expression was affected by the density of the cells encapsulated within the PEG-based hydrogel. Upon isolation of these cells, we found that these induced ADSCs were able to form tubules within Matrigel, indicative of endothelial function, while ADSCs cultured in basal medium could not. This finding points to the potential for this stem-cell population to serve as a safe and reliable source of endothelial cells for tissue engineering and regenerative medicine purposes.