title: Inhibition of TGFβ signaling increases direct conversion of fibroblasts to induced cardiomyocytes.
abstract: Recent studies have been successful at utilizing ectopic expression of transcription factors to generate induced cardiomyocytes (iCMs) from fibroblasts, albeit at a low frequency in vitro. This work investigates the influence of small molecules that have been previously reported to improve differentiation to cardiomyocytes as well as reprogramming to iPSCs in conjunction with ectopic expression of the transcription factors Hand2, Nkx2.5, Gata4, Mef2C, and Tbx5 on the conversion to functional iCMs. We utilized a reporter system in which the calcium indicator GCaMP is driven by the cardiac Troponin T promoter to quantify iCM yield. The TGFβ inhibitor, SB431542 (SB), was identified as a small molecule capable of increasing the conversion of both mouse embryonic fibroblasts and adult cardiac fibroblasts to iCMs up to ∼5 fold. Further characterization revealed that inhibition of TGFβ by SB early in the reprogramming process led to the greatest increase in conversion of fibroblasts to iCMs in a dose-responsive manner. Global transcriptional analysis at Day 3 post-induction of the transcription factors revealed an increased expression of genes associated with the development of cardiac muscle in the presence of SB compared to the vehicle control. Incorporation of SB in the reprogramming process increases the efficiency of iCM generation, one of the major goals necessary to enable the use of iCMs for discovery-based applications and for the clinic.
authors.affiliation: Department of Cell and Developmental Biology, Institute for Regenerative Medicine, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, United States of America.
authors.forename: Jamie L
authors.initials: JL
authors.lastname: Ifkovits
authors.affiliation: Department of Cell and Developmental Biology, Institute for Regenerative Medicine, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, United States of America.
authors.forename: Russell C
authors.initials: RC
authors.lastname: Addis
authors.affiliation: Department of Cell and Developmental Biology, Institute for Regenerative Medicine, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, United States of America.
authors.forename: Jonathan A
authors.initials: JA
authors.lastname: Epstein
authors.affiliation: Department of Cell and Developmental Biology, Institute for Regenerative Medicine, Perelman School of Medicine, University of Pennsylvania, Philadelphia, Pennsylvania, United States of America.
authors.forename: John D
authors.initials: JD
authors.lastname: Gearhart
mesh_terms: D000818:Animals; D001549:Benzamides / Q000494:pharmacology; D002454:Cell Differentiation; D004149:Dioxoles / Q000494:pharmacology; D005347:Fibroblasts / Q000187:drug effects / Q000502:physiology*; D051379:Mice; D032383:Myocytes, Cardiac / Q000502:physiology*; D018125:Receptors, Transforming Growth Factor beta / Q000037:antagonists & inhibitors*; D015398:Signal Transduction / Q000187:drug effects; D014157:Transcription Factors / Q000378:metabolism; D016212:Transforming Growth Factor beta / Q000037:antagonists & inhibitors*
chemical_list: C459179:4-(5-benzo(1,3)dioxol-5-yl-4-pyridin-2-yl-1H-imidazol-2-yl)benzamide; D001549:Benzamides; D004149:Dioxoles; D018125:Receptors, Transforming Growth Factor beta; D014157:Transcription Factors; D016212:Transforming Growth Factor beta