Bioprinting of pre-vascularized constructs for enhancedin vivoneo-vascularization.

ABSTRACT

Pre-vascularization has been receiving significant attention for developing implantable engineered 3D tissues. While various pre-vascularization techniques have been developed to improve graft vascularization, the effect of pre-vascularized patterns onin vivoneo-vessel formation has not been studied. In this study, we developed a functional pre-vascularized construct that significantly promotes graft vascularization and conductedin vivoevaluations of the micro-vascular patterns (µVPs) in various printed designs.µVP formation, composed of high-density capillaries, was induced by the co-printing of endothelial cells and adipose-derived stem cells (ADSC). We implanted the printed constructs with variousµVP designs into a murine femoral arteriovenous bundle model and evaluated graft vascularization via 3D visualization and immune-histological analysis of the neo-vessels. TheµVP-distal group (µVP located away from the host vessel) showed approximately two-fold improved neo-vascularization compared to theµVP-proximal group (µVP located near the host vessel). Additionally, we confirmed that theµVP-distal group can generate the angiogenic factor gradient spatial environment for graft vascularization via computational simulations. Based on these results, the ADSC mono pattern (AMP), which secretes four times higher angiogenic factors thanµVP, was added to theµVP + AMP group design. TheµVP + AMP group showed approximately 1.5- and 1.9-fold higher total sprouted neo-vessel volume than theµVP only and AMP only groups, respectively. In immunohistochemical staining analysis, theµVP + AMP group showed two-fold improved density and diameter of the matured neo-vessels. To summarize, these findings demonstrate graft vascularization accelerated due to design optimization of our pre-vascularized constructs. We believe that the developed pre-vascularization printing technique will facilitate new possibilities for the upscaling of implantable engineered tissues/organs.