Abstract 381: Human Cardiac Progenitor Cells Regenerate Cardiomyocytes and Coronary Vessels Repairing the Infarcted Myocardium
Human cardiac progenitor cells (hCPCs) were obtained from surgical samples. CPCs expressed the stem cell antigen c-kit and were self-renewing, clonogenic and multipotent. Coronary ligation was performed in immunodeficient mice and immunosuppressed rats and c-kit positive hCPCs were injected in the border zone. Foci of myocardial regeneration were identified at 2–3 weeks after infarction and consisted of myocytes, resistance arterioles and capillaries. Transcription factors indicative of myocyte and vascular cell lineages - GATA-4, MEF2C, GATA-6 and Ets1 - were detected in the new myocardium. The human origin of the developing myocardium was demonstrated by the recognition of human Alu DNA and human myosin light chain 2v (MLC2v) DNA. The surviving myocardium did not contain human Alu or human MLC2v DNA and showed rat MLC2v DNA. The expression of human mRNA for Nkx2.5, MLC2v, connexin 43, smooth muscle myosin heavy chain, and von Willebrand factor was detected by real time RT-PCR in rats treated with clonogenic c-kitPOS-hCSCs. The transcripts of these genes indicative of human myocyte and vessel formation increased with time. Human mRNA was restricted to the infarcted LV while it was undetectable in the surviving rat myocardium. The presence of connexin 43 and N-cadherin in the developing human myocytes strongly suggested that the human myocardium was functionally competent. The functional integration of EGFP-positive human myocytes with the surrounding myocardium was documented ex vivo by two-photon microscopy. At 2 weeks after coronary artery ligation and cell implantation, hearts were perfused with Tyrode solution containing the calcium indicator Rhod-2. Contraction and spontaneous activity were prevented by adding to the perfusate cytochalasin D and acetylcholine. After stimulation at 1 Hz, synchronous calcium transients were apparent in EGFP-positive human myocytes and EGFP-negative mouse myocytes. The synchronicity in calcium transients between human and mouse myocytes documented unequivocally that the human myocardium was functionally integrated with the mouse myocardium. In conclusion, HCPCs can be isolated and expanded in vitro for subsequent regeneration of dead myocardium in patients affected by heart failure of ischemic origin.