Abstract 3977: Multilayered Composite Cell Sheets: A Further Step Towards Safe and Effective Myocardial Regeneration by Cardiac Progenitors Derived From Human Embryonic Stem Cells

The safety and efficacy of myocardial regeneration using embryonic stem cells (ESC) are limited by the risk of teratoma and the high rate of cell death. To try addressing these hurdles, we tested the following approach: 10 Rhesus monkeys underwent transient (90 minutes) coronary artery balloon occlusion and simultaneous autologous adipose tissue biopsy to derive stroma/stem cells (ADSC). Allogeneic Rhesus ESC expressing GFP under the control of α -actin were driven towards a cardiac lineage by exposure to BMP-2 and a FGF inhibitor and immunomagnetically sorted on the basis of a positive expression for CD15. Two weeks after infarction, the ADSC were cultured on temperature-responsive dishes to fabricate cell sheets (6 × 10 6 cells/sheet). Three sheets were stacked and the innest one (to be in direct contact with the heart) was seeded with 6 × 10 6 CD15 + progenitors. The 6–10 cm 2 composite sheets were then applied onto the infarcted areas through a thoracotomy. Ciclosporine immunosuppression was given for 2 months, at which time animals were sacrificed. Preimplantation studies showed that CD15 + progenitors expressed cardiac markers (including connexin 43) and had lost pluripotency (as assessed by RT-PCR, immunofluorescence, micro-RNA expression and epigenetic profiling). Postmortem analysis of >1,200 heart sections showed that these CD15 + progenitors could differentiate into cardiomyocytes with a robust repopulation of infarcted tissue by ADSC-derived CD90 + cells and an increased angiogenesis (mean group number of vessels per mm 2 : 820 (90%CI: 564–1076) vs. 588 (247–928) in controls). LV function, as assessed by echocardiography, was better preserved compared with controls, with a two-fold greater reduction in ESV compared with baseline (−0.65 mL vs. −0.33 mL). There was no teratoma or extra-cardiac tumor. Likewise, immunodeficient (RAG2−/−, γ c −/−, C5−/−) mice which were injected in parallel with CD15 + cardiac progenitors equally failed to develop teratoma. Put together, these data, collected in a clinically relevant large animal model, suggest that the combined epicardial delivery of CD15 + ESC-derived cardiac progenitors and ADSC providing trophic support may improve the safety/efficacy of regeneration of the chronically infarcted myocardium.