Self-assembling scaffold of fibrin and decellularized cardiac matrix for the delivery of cardiac progenitor cells for myocardial regeneration

Cardiac tissue engineering (CTE) is a promising approach to replace and regenerate the injured and not functional cardiac tissue. Injectable self-assembling scaffolds consisting of biomaterials mimicking the native extracellular environment are an attractive option to promote specific homing and cell retention with minimal invasiveness. Among biological injectable materials, fibrin has been tested in several pathological conditions, either in combination with autologous stem cells or not, to stimulate wound healing through tissue regeneration, as it offers the unrivalled advantages of being autologous, enriched in growth factors and capable of gelling at 37 °C. In this study we combined fibrin with human cardiac decellularized extracellular matrix (d-ECM) and human Cardiac Progenitor Cells (hCPCs) to prepare an injectable hybrid three-dimensional scaffold serving as a natural cardiac platform for the delivery and engraftment of hCPCs. To this aim, samples of adult human hearts of recipients of heart transplantation were either decellularized to obtain the native cardiac d-ECM or disaggregated mechanically and by enzymatic digestion to obtain hCPCs. d-ECM was then lyophilized and solubilized to be incorporated along with the hCPCs into fibrin gels that were prepared by combining fibrinogen and thrombin. We tested three ratios of fibrin:d-ECM to determine the ideal combination. Specifically, we prepared 1:1, 1:2 and 2:1 fibrin:d-ECM gels and we allowed them to gel at 37°C. Gelling occurred in all preparations, but differences in the time needed for gelling and organization of gels occurred. The 2:1 preparation gelled after one hour, but solubilized d-ECM and fibrin did not combine well as the fibrin gel floated into the solubilized d-ECM. The 1:1 and 2:1 preparations, instead, gelled in a time ranging between eight and twenty hours, respectively, and produced homogenous gels. Gels were processed for histological analysis after three days of culture and hematoxylin-Eosin staining showed the presence and distribution of hCPCs throughout the entire thickness of the gels with 1:1 ratio, while for 1:2 and 2:1 ratios hCPCs coated the surface but were not visible inside the gels. Furthermore, the presence of collagen and ECM glycoproteins was detected by Sirius Red, Periodic Acid Schiff and Masson's Trichrome stainings exclusively in the gel with 1:1 ratio. Our results suggest that the combination of fibrin and d-ECM at 1:1 ratio yields a well-structured self-assembling scaffold that might be used as a minimally invasive method to deliver hCPCs and cardiac d-ECM in the injured myocardium to boost cardiac regeneration.