title: Generation of Functional Cardiomyocytes from Efficiently Generated Human iPSCs and a Novel Method of Measuring Contractility.
abstract: Human induced pluripotent stem cells (iPSCs) derived cardiomyocytes (iCMCs) would provide an unlimited cell source for regenerative medicine and drug discoveries. The objective of our study is to generate functional cardiomyocytes from human iPSCs and to develop a novel method of measuring contractility of CMCs. In a series of experiments, adult human skin fibroblasts (HSF) and human umbilical vein endothelial cells (HUVECs) were treated with a combination of pluripotent gene DNA and mRNA under specific conditions. The iPSC colonies were identified and differentiated into various cell lineages, including CMCs. The contractile activity of CMCs was measured by a novel method of frame-by-frame cross correlation (particle image velocimetry-PIV) analysis. Our treatment regimen transformed 4% of HSFs into iPSC colonies at passage 0, a significantly improved efficiency compared with use of either DNA or mRNA alone. The iPSCs were capable of differentiating both in vitro and in vivo into endodermal, ectodermal and mesodermal cells, including CMCs with >88% of cells being positive for troponin T (CTT) and Gata4 by flow cytometry. We report a highly efficient combination of DNA and mRNA to generate iPSCs and functional iCMCs from adult human cells. We also report a novel approach to measure contractility of iCMCs.
authors.affiliation: Cardiovascular Research Institute, Division of Cardiovascular Diseases, Department of Internal Medicine, University of Kansas Medical Center, Kansas City, Kansas, United States of America.
authors.forename: Sheeja
authors.initials: S
authors.lastname: Rajasingh
authors.affiliation: Cardiovascular Research Institute, Division of Cardiovascular Diseases, Department of Internal Medicine, University of Kansas Medical Center, Kansas City, Kansas, United States of America.
authors.forename: Jayakumar
authors.initials: J
authors.lastname: Thangavel
authors.affiliation: Department of Anatomy and Cell Biology, University of Kansas Medical Center, Kansas City, Kansas, United States of America.
authors.forename: Andras
authors.initials: A
authors.lastname: Czirok
authors.affiliation: Cardiovascular Research Institute, Division of Cardiovascular Diseases, Department of Internal Medicine, University of Kansas Medical Center, Kansas City, Kansas, United States of America.
authors.forename: Saheli
authors.initials: S
authors.lastname: Samanta
authors.affiliation: Department of Anatomy and Cell Biology, University of Kansas Medical Center, Kansas City, Kansas, United States of America.
authors.forename: Katherine F
authors.initials: KF
authors.lastname: Roby
authors.affiliation: Cardiovascular Research Institute, Division of Cardiovascular Diseases, Department of Internal Medicine, University of Kansas Medical Center, Kansas City, Kansas, United States of America.
authors.forename: Buddhadeb
authors.initials: B
authors.lastname: Dawn
authors.affiliation: Cardiovascular Research Institute, Division of Cardiovascular Diseases, Department of Internal Medicine, University of Kansas Medical Center, Kansas City, Kansas, United States of America; Department of Biochemistry and Molecular Biology, University of Kansas Medical Center, Kansas City, Kansas, United States of America.
authors.forename: Johnson
authors.initials: J
authors.lastname: Rajasingh
mesh_terms: D000328:Adult; D000818:Animals; D018929:Cell Culture Techniques / Q000379:methods; D002454:Cell Differentiation; D002460:Cell Line; D005347:Fibroblasts / Q000166:cytology; D061307:Human Umbilical Vein Endothelial Cells / Q000166:cytology; D006801:Humans; D057026:Induced Pluripotent Stem Cells / Q000166:cytology*; D008297:Male; D016513:Mice, SCID; D009200:Myocardial Contraction; D032383:Myocytes, Cardiac / Q000166:cytology*