Microfibrous Scaffolds Enhance Endothelial Differentiation and Organization of Induced Pluripotent Stem Cells.

INTRODUCTION: Human induced pluripotent stem cells (iPSCs) are a promising source of endothelial cells (iPSC-ECs) for engineering three-dimensional (3D) vascularized cardiac tissues. To mimic cardiac microvasculature, in which capillaries are oriented in parallel, we hypothesized that endothelial differentiation of iPSCs within topographically aligned 3D scaffolds would be a facile one-step approach to generate iPSC-ECs as well as induce aligned vascular organization. METHODS: Human iPSCs underwent endothelial differentiation within electrospun 3D polycaprolactone (PCL) scaffolds having either randomly oriented or parallel-aligned microfibers. Using transcriptional, protein, and endothelial functional assays, endothelial differentiation was compared between conventional two-dimensional (2D) films and 3D scaffolds having either randomly oriented or aligned microfibers. Furthermore, the role of parallel-aligned microfiber patterning on the organization of vessel-like networks was assessed. RESULTS: The cells in both the randomly oriented and aligned 3D scaffolds demonstrated an 11-fold upregulation in gene expression of the endothelial phenotypic marker, CD31, compared to cells on 2D films. This upregulation corresponded to >3-fold increase in CD31 protein expression in 3D scaffolds, compared to 2D films. Concomitantly, other endothelial phenotypic markers including CD144 and endothelial nitric oxide synthase also showed significant transcriptional upregulation in 3D scaffolds by >7-fold, compared to 2D films. Nitric oxide production, which is characteristic of endothelial function, was produced 4-fold more abundantly in 3D scaffolds, compared to on 2D PCL films. Within aligned scaffolds, the iPSC-ECs displayed parallel-aligned vascular-like networks with 70% longer branch length, compared to cells in randomly oriented scaffolds, suggesting that fiber topography modulates vascular network-like formation and patterning. CONCLUSION: Together, these results demonstrate that 3D scaffold structure promotes endothelial differentiation, compared to 2D substrates, and that aligned topographical patterning induces anisotropic vascular network organization.

Bilayered vascular graft derived from human induced pluripotent stem cells with biomimetic structure and function.

BACKGROUND: We developed an aligned bi-layered vascular graft derived from human induced pluripotent stem cells (iPSCs) that recapitulates the cellular composition, orientation, and anti-inflammatory function of blood vessels. MATERIALS & METHODS: The luminal layer consisted of longitudinal-aligned nanofibrillar collagen containing primary endothelial cells (ECs) or iPSC-derived ECs (iPSC-ECs). The outer layer contained circumferentially oriented nanofibrillar collagen with primary smooth muscle cells (SMCs) or iPSC-derived SMCs(iPSC-SMCs). RESULTS: On the aligned scaffolds, cells organized F-actin assembly within 8º from the direction of nanofibrils. When compared to randomly-oriented scaffolds, EC-seeded aligned scaffolds had significant reduced inflammatory response, based on adhesivity to monocytes. CONCLUSION: This study highlights the importance of anisotropic scaffolds in directing cell form and function, and has therapeutic significance as physiologically relevant blood vessels.