Struggling to Prepare an Injectable Self-Assembling Human Cardiac Matrix and Facing Unexpected Failure

Restoring myocardium integrity is the most ambitious and demanding undertaking of cardiovascular regenerative medicine. Several attempts to recover myocardium integrity and contractile function by either direct injection of stem cells in the heart wall or functional three-dimensional tissue constructs of scaffolds and cells, were made in the past decade. Obviously, an injectable preparation of decellularized extracellular matrix (d-ECM), avoiding open chest cardiac surgery would be a tremendous innovation in cardiac regenerative medicine. Aiming at preparing human cardiac d-ECM solution to be used as stem cell delivery medium and embedding scaffold, we decellularized ECM incubating cardiac samples first with 1% SDS for 12 h, then with 1% Triton X-100 for 30 min. To obtain an injectable form, we lyophilized and dissolved d-ECM in pepsin solution as previously reported. Although we scrupulously followed the described methodology, d-ECM did not dissolve. Adjustments were made until, with a treatment of 24 h at 37°C on tube rotator, d-ECM dissolved. It is reported that d-ECM-pepsin solution spontaneously gels at 37°C in about 1h. Again, strictly following previously reported procedures, our results differed significantly. We struggled with several adjustments of temperature, time, d-ECM batch and concentration in pepsin solution, but we never had the d-ECM gelling. Since in previous reports d-ECM was obtained from animal models or human pericardium, we infer that gelling depends on collagen content and, hence, they do not make a good substitute for ECM of human cardiac myocardium.