Abstract 200: Engineering Evolution: Tetraploidization of Human Cardiac Stem Cells to Enhance Functional Activity

Abstract

Introduction: Reparative and regenerative capacity is consistently observed in lower vertebrate species, but translational implementation has failed to yield comparable results in the clinical setting. Cardiac stem cells (CSC) isolated from rodent or swine heart possess capacity for tetraploidization resulting in 4n chromatin content associated with override of cell cycle arrest and senescence, but comparable tetraploid CSC are not present in human samples. Furthermore, diploid:tetraploid CSC ratio in rodent myocardium increases following infarction injury suggesting ploidy is a dynamic cellular condition influencing response to pathologic damage. Since polyploidization correlates with enhanced regenerative throughout evolution, ploidy differences raise provocative questions regarding translational applicability of experimental studies of myocardial regeneration. Hypothesis: Mononuclear chromatin duplication in human stem cells improves functional capabilities by inhibiting cellular senescence and increasing stress resistance. Methods and Results: Tetraploidy was artificially induced in human CSCs, ESCs and MSCs from multiple patient samples by transient exposure to demecolcine that blocks mitotic spindle formation and chromosome segregation. Mononuclear tetraploid content confirmed by karyotype analysis was consistently induced in hCSCs and hEPCs, unlike hMSCs which underwent apoptosis. Mononuclear tetraploid content in hCSCs and hEPCs was stable for continued multiple passages demonstrated by flow cytometry with proliferation rates equivalent to the parent diploid line. Stress induction through hydrogen peroxide demonstrate tetraploid hCSCs respond equivalent or better than parental lines measured through apoptosis and necrosis. Double strand DNA breaks in tetraploid hCSCs were reduced by half compared to the parental line, measured by gamma H2AX per total DNA. Conclusion: Tetraploid content induction in human CSCs and EPCs enhances functional activity by increased resistance to stress and inhibition of senescence without promoting oncogenic transformation. Ongoing studies focus upon potential of tetraploid hCSCs to exert enhanced capabilities for mediating myocardial repair and regeneration.