Cell-based delivery of dATP via gap junctions enhances cardiac contractility.
The transplantation of human pluripotent stem cell-derived cardiomyocytes (hPSC-CMs) is a promising strategy to treat myocardial infarction and reverse heart failure, but to date the contractile benefit in most studies remains modest. We have previously shown that the nucleotide 2-deoxyadenosine triphosphate (dATP) can substitute for ATP as the energy substrate for cardiac myosin, and increasing cellular dATP content by globally overexpressing ribonucleotide reductase (R1R2) can dramatically enhance cardiac contractility. Because dATP is a small molecule, we hypothesized that it would diffuse readily between cells via gap junctions and enhance the contractility of neighboring coupled wild type cells. To test this hypothesis, we performed studies with the goals of (1) validating gap junction-mediated dATP transfer in vitro and (2) investigating the use of R1R2-overexpressing hPSC-CMs in vivo as a novel strategy to increase cardiac function. We first performed intracellular dye transfer studies using dATP conjugated to fluorescein and demonstrated rapid gap junction-mediated transfer between cardiomyocytes. We then cocultured wild type cardiomyocytes with either cardiomyocytes or fibroblasts overexpressing R1R2 and saw more than a twofold increase in the extent and rate of contraction of wild type cardiomyocytes. Finally, we transplanted hPSC-CMs overexpressing R1R2 into healthy uninjured rat hearts and noted an increase in fractional shortening from 41±4% to 53±5% just five days after cell transplantation. These findings demonstrate that dATP is an inotropic factor that spreads between cells via gap junctions. Our data suggest that transplantation of dATP-producing hPSC-CMs could significantly increase the effectiveness of cardiac cell therapy.
Department of Bioengineering, University of Washington, Seattle, WA 98195, USA; Institute for Stem Cell and Regenerative Medicine, University of Washington, Seattle, WA 98195, USA.
Scott D
SD
Lundy
Department of Bioengineering, University of Washington, Seattle, WA 98195, USA.
Sean A
SA
Murphy
Institute for Stem Cell and Regenerative Medicine, University of Washington, Seattle, WA 98195, USA; Center for Cardiovascular Biology, University of Washington, Seattle, WA 98195, USA.
Sarah K
SK
Dupras
Department of Bioengineering, University of Washington, Seattle, WA 98195, USA.
Jin
J
Dai
Department of Bioengineering, University of Washington, Seattle, WA 98195, USA; Institute for Stem Cell and Regenerative Medicine, University of Washington, Seattle, WA 98195, USA; Center for Cardiovascular Biology, University of Washington, Seattle, WA 98195, USA; Department of Pathology, University of Washington, Seattle, WA 98195, USA; Department of Medicine/Cardiology, University of Washington, Seattle, WA 98195, USA.
Charles E
CE
Murry
Department of Bioengineering, University of Washington, Seattle, WA 98195, USA; Institute for Stem Cell and Regenerative Medicine, University of Washington, Seattle, WA 98195, USA; Center for Cardiovascular Biology, University of Washington, Seattle, WA 98195, USA; Department of Pathology, University of Washington, Seattle, WA 98195, USA.
Michael A
MA
Laflamme
Department of Bioengineering, University of Washington, Seattle, WA 98195, USA; Institute for Stem Cell and Regenerative Medicine, University of Washington, Seattle, WA 98195, USA; Center for Cardiovascular Biology, University of Washington, Seattle, WA 98195, USA. Electronic address: mregnier@u.washington.edu.
Michael
M
Regnier
D000818:Animals; D000831:Animals, Newborn; D001692:Biological Transport; D002454:Cell Differentiation; D064987:Cell- and Tissue-Based Therapy / Q000379:methods*; D018920:Coculture Techniques; D003838:Deoxyadenine Nucleotides / Q000494:pharmacology*; D005347:Fibroblasts / Q000166:cytology / Q000378:metabolism; D017629:Gap Junctions / Q000187:drug effects* / Q000378:metabolism; D015870:Gene Expression; D006321:Heart / Q000502:physiology; D006352:Heart Ventricles / Q000166:cytology / Q000378:metabolism; D006801:Humans; D008297:Male; D009200:Myocardial Contraction / Q000502:physiology*; D032383:Myocytes, Cardiac / Q000166:cytology* / Q000378:metabolism / Q000637:transplantation*; D039904:Pluripotent Stem Cells / Q000166:cytology / Q000378:metabolism; D061251:Primary Cell Culture; D051381:Rats; D011923:Rats, Nude; D012264:Ribonucleotide Reductases / Q000235:genetics / Q000378:metabolism; D014183:Transplantation, Heterologous
D003838:Deoxyadenine Nucleotides; D012264:Ribonucleotide Reductases; C026600:2'-deoxyadenosine triphosphate
Cardiac cell therapy; Cardiac regeneration; Gap junction intercellular communication; Ribonucleotide reductase; Stem cells; dATP