Bioprinting of 3D pre-vascularized stem cell delivery platform for the treatment of ischemic cardiac diseases

摘要

Introduction: Stem cell therapy emerges as a new therapeutic method for the treatment of ischemic heart diseases. Towards the enhancement of therapeutic efficiency, a functionalized patch type cell delivery platform can be a potential alternative to achieve high retention, survival, engraftment, and differentiation of cells. Although there have been many successful results, fabricating an organized vascular plexus within the 3D structure remains a key barrier to enhancement of cell survival and function. 3D bioprinting is considered as a promising approach to generate 3D engineered tissues by emulating the cellular organization of the native tissue with high repeatability. Here, we developed a 3D pre-vascularized patch type stem cell delivery platform by patterning vascular cells and cardiac progenitor cells. Materials and Methods: Two 3D patches were fabricated using heart tissue-derived decellularized extracellular matrix (hdECM) and fibrin bioink. The fabricated bioink patches (hdECM and fibrin bioink) were implanted into the rat myocardiac infarction (MI) model for verifying the therapeutic efficacy of bioink. Human c-kit+ cardiac tissue derived cardiac progenitor cells (CPCs) and human turbinate tissue derived mesenchymal stem cells (MSCs) were prepared by following protocol described in elsewhere. To print the patch for stem cell delivery, we used three different bioinks' formulation as follow: 1) CPCs-laden hdECM bioink (Bioink Ⅰ), 2) MSCs-laden hdECM bioink with 10 μg/ml VEGF (Bioink Ⅱ) for vessel formation and 3) the mixture of CPCs and MSCs-laden hdECM bioink with 10 μg/ml VEGF (Bioink Ⅲ). We printed the cell deliverable patch spatially alternating Bioink Ⅰ and Bioink Ⅱ (patterned patch) using lab-built multi-head bioprinter. The mixed patch was fabricated using Bioink Ⅲ. The printed patches were implanted either in the mice subcutaneous site to investigate the vascularization capability or transplanted the printed patch into rat MI model to verify the therapeutic effect of the patch. Results and Discussion: hdECM bioink enhanced angiogenesis and promoted fetal reprogramming through the endogenous paracrine signaling of epicardium compared to fibrin bioink. In addition, the patterned patch shortened the time for inosculation with the host circulatory system and promoted strong vasculogenesis which is distinct from the capillary formation made by the mixed patch. After 8 weeks of transplantation of the patch to the infarct tissue, the major cardiac functions were significantly enhanced in the group of patterned patch (ejection fraction 8.12±1.1 %, fractional shortening 6.24±2.46 % compared to baseline). It is because the patterned patch may provide a suitable environment which contributes to prolonged therapeutic efficacy and viability of delivered cells. Moreover, partial re-muscularization was progressed in both the implanted patch and the infarct region. The larger number of cells were migrated from the patterned patch to the infarct and differentiated into the endothelium compared to the mixed patch. Conclusion: A major advantage of the developed platform technology is the spatial patterning of each cell to promote rapid vascularization in the 3D structure and prolonged survival of delivered cells in vivo. This 3D pre-vascularized stem cell delivery platform may open new avenues for delivering cells with high retention capability and regenerating ischemic tissue area.