title: Same-Single-Cell Analysis of Pacemaker-Specific Markers in Human Induced Pluripotent Stem Cell-Derived Cardiomyocyte Subtypes Classified by Electrophysiology.
abstract: Insights into the expression of pacemaker-specific markers in human induced pluripotent stem cell (hiPSC)-derived cardiomyocyte subtypes can facilitate the enrichment and track differentiation and maturation of hiPSC-derived pacemaker-like cardiomyocytes. To date, no study has directly assessed gene expression in each pacemaker-, atria-, and ventricular-like cardiomyocyte subtype derived from hiPSCs since currently the subtypes of these immature cardiomyocytes can only be identified by action potential profiles. Traditional acquisition of action potentials using patch-clamp recordings renders the cells unviable for subsequent analysis. We circumvented these issues by acquiring the action potential profile of a single cell optically followed by assessment of protein expression through immunostaining in that same cell. Our same-single-cell analysis for the first time revealed expression of proposed pacemaker-specific markers-hyperpolarization-activated cyclic nucleotide-modulated (HCN)4 channel and Islet (Isl)1-at the protein level in all three hiPSC-derived cardiomyocyte subtypes. HCN4 expression was found to be higher in pacemaker-like hiPSC-derived cardiomyocytes than atrial- and ventricular-like subtypes but its downregulation over time in all subtypes diminished the differences. Isl1 expression in pacemaker-like hiPSC-derived cardiomyocytes was initially not statistically different than the contractile subtypes but did become statistically higher than ventricular-like cells with time. Our observations suggest that although HCN4 and Isl1 are differentially expressed in hiPSC-derived pacemaker-like relative to ventricular-like cardiomyocytes, these markers alone are insufficient in identifying hiPSC-derived pacemaker-like cardiomyocytes. Stem Cells 2016;34:2670-2680.
authors.affiliation: Department of Internal Medicine, Division of Cardiovascular Medicine, University of California, Davis, California, USA.
authors.forename: Sergey
authors.initials: S
authors.lastname: Yechikov
authors.affiliation: Bridges to Stem Cell Research Program, California State University, Sacramento, California, USA.
authors.forename: Raul
authors.initials: R
authors.lastname: Copaciu
authors.affiliation: Department of Internal Medicine, Division of Cardiovascular Medicine, University of California, Davis, California, USA.
authors.forename: Jessica M
authors.initials: JM
authors.lastname: Gluck
authors.affiliation: Department of Biochemistry and Molecular Medicine, University of California, Davis, California, USA.
authors.forename: Wenbin
authors.initials: W
authors.lastname: Deng
authors.affiliation: Department of Internal Medicine, Division of Cardiovascular Medicine, University of California, Davis, California, USA.|Department of Veterans Affairs, Northern California Health Care System, Mather, California, USA.
authors.forename: Nipavan
authors.initials: N
authors.lastname: Chiamvimonvat
authors.affiliation: Center for Biophotonics, University of California, Davis, Sacramento, California, USA.|Department of Pathology and Laboratory Medicine, University of California, Davis, Sacramento, California, USA.
authors.forename: James W
authors.initials: JW
authors.lastname: Chan
authors.affiliation: Department of Internal Medicine, Division of Cardiovascular Medicine, University of California, Davis, California, USA.
authors.forename: Deborah K
authors.initials: DK
authors.lastname: Lieu
mesh_terms: D000200:Action Potentials / Q000502:physiology*; D015415:Biomarkers / Q000378:metabolism; D002454:Cell Differentiation; D002460:Cell Line; D019070:Cell Lineage / Q000235:genetics; D004594:Electrophysiology; D015870:Gene Expression; D006325:Heart Atria / Q000166:cytology / Q000378:metabolism*; D006329:Heart Conduction System / Q000166:cytology / Q000378:metabolism*; D006352:Heart Ventricles / Q000166:cytology / Q000378:metabolism*; D006801:Humans; D064428:Hyperpolarization-Activated Cyclic Nucleotide-Gated Channels / Q000235:genetics / Q000378:metabolism; D007150:Immunohistochemistry; D057026:Induced Pluripotent Stem Cells / Q000166:cytology / Q000378:metabolism*; D060850:LIM-Homeodomain Proteins / Q000235:genetics / Q000378:metabolism; D009124:Muscle Proteins / Q000235:genetics / Q000378:metabolism; D032383:Myocytes, Cardiac / Q000166:cytology / Q000378:metabolism*; D009928:Organ Specificity; D015221:Potassium Channels / Q000235:genetics / Q000378:metabolism; D059010:Single-Cell Analysis / Q000379:methods; D014157:Transcription Factors / Q000235:genetics / Q000378:metabolism
chemical_list: D015415:Biomarkers; C119587:HCN4 protein, human; D064428:Hyperpolarization-Activated Cyclic Nucleotide-Gated Channels; D060850:LIM-Homeodomain Proteins; D009124:Muscle Proteins; D015221:Potassium Channels; D014157:Transcription Factors; C063843:insulin gene enhancer binding protein Isl-1
keywords: Cardiomyocyte subtype; Electrophysiology; Human induced pluripotent stem cells; Pacemaker