Vascular differentiation of bone marrow stem cells is directed by a tunable three-dimensional matrix

Microenvironmental cues are critical in regulating cell behavior and fate. The roles that matrix mechanical signals play in regulating cell behavior have recently been elucidated. An artificial matrix that can maintain the appropriate characteristics for transplanted stem cells is therefore needed to achieve a desired cell phenotype. The objective of this study was to develop a three-dimensional (3-D) matrix with tunable physical and mechanical properties and investigate their effects on mesenchymal stem cell (MSC) differentiation towards vascular cell types. In this study we developed an extracellular microenvironment by modifying fibrinogen with various polyethylene glycol (PEG) derivatives. We hypothesized that adjusting the type of PEG derivative to modify the resultant physical and mechanical characteristics of fibrin would allow us to create a tunable system for use in culture or in vivo in conjunction with a regenerative medicine strategy. Human MSC (hMSC) were entrapped into PEGylated fibrin matrices at a density of 50,000 cells ml(-1). Cell phenotypes were confirmed by immunofluorescent staining as well as the use of oligonucleotide arrays. Vascular phenotypes were correlated with measured mechanical properties and fiber diameters of the PEGylated fibrin matrices. Blocking studies were performed to identify mechanistic factors controlling MSC differentiation through selected blocking of matrix degradation or cell contraction. Cell-matrix interactions were also examined in vivo. Our results demonstrate that transdifferentiation of MSC towards an endothelial cell phenotype is profoundly affected by the 3-D matrix microenvironment. Our work provides a predictive road map for the creation of fibrin-based matrices that support robust endothelial cell gene expression and tubulogenesis.