Abstract
Procurement and characterization of intact human cells are essential for studies in regenerative medicine and translational medical research. The selection of the currently available approaches to isolate intact cells depends on the age of the hearts. To isolate cardiomyocytes from the fetal or neonatal myocardium, the myocardium can be minced into small tissue blocks followed by enzyme incubation. However, the fetal and neonatal cardiomyocytes are very soft and the morphology changes from long rod or spindle shape to spheres after isolation. Because of the dense packing of the cardiomyocytes and the strong cell-cell connection in adult myocardium, it is difficult to isolate the cardiomyocytes from adult myocardium by enzyme incubation only. A perfusion method is necessary to deliver the enzyme solution to the deep layers of the myocardium. However, intact hearts, which are very rare, are required for the perfusion method. Therefore, lacking methods to efficiently isolate cardiomyocytes from myocardium of various ages builds a barrier between basic research and clinical studies. Here, we describe a method for the isolation of intact cardiomyocytes from fresh or frozen human myocardium or fresh mouse hearts and the quantification of multinucleation, cardiomyocyte size, cell cycle activity, and total cardiomyocyte count per heart. We generalize this fixation-digestion method by isolating cells from a variety of mouse organs, including the liver, lung, and thymus.
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Acknowledgments
This research was supported by the Richard King Mellon Foundation Institute for Pediatric Research (UPMC Children’s Hospital of Pittsburgh), by a Transatlantic Network of Excellence grant by Foundation Leducq (15CVD03), Children’s Cardiomyopathy Foundation, and NIH grant R01HL106302 (to B.K.). This project was supported, in part, by UPMC Children’s Hospital of Pittsburgh (to H.L.), Genomics Discovery Award (UPMC Children’s Hospital), UPP Physicians, Vascular Medicine Institute, Aging Institute, and NIH grant UL1TR001857 from the Clinical and Translational Sciences Institute (University of Pittsburgh, to B.K.).
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Liu, H., Bersell, K., Kühn, B. (2021). Isolation and Characterization of Intact Cardiomyocytes from Frozen and Fresh Human Myocardium and Mouse Hearts. In: Poss, K.D., Kühn, B. (eds) Cardiac Regeneration. Methods in Molecular Biology, vol 2158. Humana, New York, NY. https://doi.org/10.1007/978-1-0716-0668-1_15
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DOI: https://doi.org/10.1007/978-1-0716-0668-1_15
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