Abstract 16288: A Practical Freezing Method of Human-Induced Pluripotent Stem Cells-Derived Cardiomyocytes Toward a Clinical Application
Introduction: Cardiomyocytes (CMs) derived from human iPS cells (hiPSCs) implantation have the potential to lead to new heart failure therapy, while additional technology, such as an efficient cryopreservation method, should be developed to achieve clinical application. In this study, we hypothesized that the freezing protocol for hiPSCs-derived CMs can preserve their functional and physiological potentials.
Method: (1) Dissociated CMs were suspended in STEMCELLBANKER(R) and stocked using a controlled rate freezer at -1°C/min from 4°C to -80°C. We then evaluated the viability, functionality, and purity of cryopreserved CMs by flow cytometry analysis of cardiac troponin T before and after thawing. (2) To test their electrophysiological ability, the thawed CMs were reseeded onto thermo-responsive culture dishes to make cell sheets and analyzed by a microelectrode array (MEA). (3) To check the impacts on cardiac performance in vivo, we transplanted the cell sheets of non-cryopreserved CMs (non-cryo: n=6) and cryopreserved CMs (cryo: n=7) into nude rats with myocardial infarction and compared with a sham-operated group (S group: n=6).
Results: (1) We found that the cell recovery rate reached 72±4% in freezing conditions (purity range: 21-92%, average: 47%, n=7). The purity of CMs did not change from before freezing to after thawing; in addition, cardiac cell purity had no significant contribution to the cell viability. (2) Cell sheets could be made from CMs preserved by slow freezing with STEMCELLBANKER(R); the MEA revealed that the CMs sheet showed a spontaneous synchronized beating with the expression of cardiac troponin T and sarcomeric α-actinin. (3) Four weeks after transplantation, the ejection fraction (EF) was significantly higher in the non-cryo group (51±3%) and cryo group (51±5%) compared to the S group (39±1%), measured by echocardiography (p <0.01). There was no significant difference in the EF between the non-cryo and cryo groups, indicating that the efficiency of cryopreserved CMs in vivo was comparable to that of freshly prepared CMs.
Conclusion: hiPSCs-derived CMs can be efficiently preserved by the use of STEMCELLBANKER(R) with a slow freezing method, suggesting that this protocol may contribute to the clinical application of these cells.
Author Disclosures: F. Ohashi: Employment; Significant; Terumo Corporation. S. Miyagawa: None. S. Fukushima: None. S. Yoshida: None. A. Saito: None. S. Masuda: None. H. Iseoka: Employment; Significant; TERUMO COPORATION. E. Ito: None. T. Ishikawa: None. T. Sameshima: Employment; Significant; Terumo Coporation. Y. Sato: None. Y. Sawa: None.
- © 2016 by American Heart Association, Inc.