Phospholamban as a crucial determinant of the inotropic response of human pluripotent stem cell-derived ventricular cardiomyocytes and engineered 3-dimensional tissue constructs

Gaopeng Chen; Sen Li; Ioannis Karakikes; Lihuan Ren; Maggie Zi-Ying Chow; Anant Chopra; Wendy Keung; Bin Yan; Camie W Y Chan; Kevin D Costa; Chi-Wing Kong; Roger J Hajjar; Christopher S Chen; Ronald A Li

doi:10.1161/CIRCEP.114.002049

Phospholamban as a crucial determinant of the inotropic response of human pluripotent stem cell-derived ventricular cardiomyocytes and engineered 3-dimensional tissue constructs

Circ Arrhythm Electrophysiol. 2015 Feb;8(1):193-202. doi: 10.1161/CIRCEP.114.002049. Epub 2014 Dec 10.

Authors

Affiliations

¹ From the Cardiovascular Research Center, Icahn School of Medicine at Mount Sinai, Manhattan, NY (G.C., I.K., K.D.C., R.J.H., R.A.L.); Department of Physiology (G.C., S.L., L.R., M.Z.-Y.C., W.K., C.-W.K., R.A.L.), Stem Cell and Regenerative Medicine Consortium (G.C., S.L., L.R., M.Z.-Y.C., W.K., B.Y., C.W.Y.C., C.-W.K., R.A.L.), Department of Anatomy (C.W.Y.C.), LKS Faculty of Medicine, University of Hong Kong, Pokfulam, Hong Kong; Department of Bioengineering, Boston University, MA (A.C., C.S.C.); Harvard Wyss Institute for Biologically Inspired Engineering, Boston, MA (A.C., C.S.C.); and Department of Biology, Hong Kong Baptist University, Hong Kong (B.Y.).
² From the Cardiovascular Research Center, Icahn School of Medicine at Mount Sinai, Manhattan, NY (G.C., I.K., K.D.C., R.J.H., R.A.L.); Department of Physiology (G.C., S.L., L.R., M.Z.-Y.C., W.K., C.-W.K., R.A.L.), Stem Cell and Regenerative Medicine Consortium (G.C., S.L., L.R., M.Z.-Y.C., W.K., B.Y., C.W.Y.C., C.-W.K., R.A.L.), Department of Anatomy (C.W.Y.C.), LKS Faculty of Medicine, University of Hong Kong, Pokfulam, Hong Kong; Department of Bioengineering, Boston University, MA (A.C., C.S.C.); Harvard Wyss Institute for Biologically Inspired Engineering, Boston, MA (A.C., C.S.C.); and Department of Biology, Hong Kong Baptist University, Hong Kong (B.Y.). ronaldli@hku.hk.

Abstract

Background: Human (h) embryonic stem cells (ESCs) and induced pluripotent stem cells (iPSCs) serve as a potential unlimited ex vivo source of cardiomyocytes (CMs). However, a well-accepted roadblock has been their immature phenotype. hESC/iPSC-derived ventricular (v) CMs and their engineered cardiac microtissues (hvCMTs) similarly displayed positive chronotropic but null inotropic responses to β-adrenergic stimulation. Given that phospholamban (PLB) is robustly present in adult but poorly expressed in hESC/iPSC-vCMs and its defined biological role in β-adrenergic signaling, we investigated the functional consequences of PLB expression in hESC/iPSC-vCMs and hvCMTs.

Methods and results: First, we confirmed that PLB protein was differentially expressed in hESC (HES2, H9)- and iPSC-derived and adult vCMs. We then transduced hES2-vCMs with the recombinant adenoviruses (Ad) Ad-PLB or Ad-S16E-PLB to overexpress wild-type PLB or the pseudophosphorylated point-mutated variant, respectively. As anticipated from the inhibitory effect of unphosphorylated PLB on sarco/endoplasmic reticulum Ca2+-ATPase, Ad-PLB transduction significantly attenuated electrically evoked Ca2+ transient amplitude and prolonged the 50% decay time. Importantly, Ad-PLB-transduced hES2-vCMs uniquely responded to isoproterenol. Ad-S16E-PLB-transduced hES2-vCMs displayed an intermediate phenotype. The same trends were observed with H9- and iPSC-vCMs. Directionally, similar results were also seen with Ad-PLB-transduced and Ad-S16E-transduced hvCMTs. However, Ad-PLB altered neither the global transcriptome nor ICa,L, implicating a PLB-specific effect.

Conclusions: Engineered upregulation of PLB expression in hESC/iPSC-vCMs restores a positive inotropic response to β-adrenergic stimulation. These results not only provide a better mechanistic understanding of the immaturity of hESC/iPSC-vCMs but will also lead to improved disease models and transplantable prototypes with adult-like physiological responses.

Keywords: adrenergic effects; phospholamban; pluripotent stem cells; tissues.

Publication types

Research Support, Non-U.S. Gov't

MeSH terms

Adrenergic beta-Agonists / pharmacology
Calcium Signaling
Calcium-Binding Proteins / genetics
Calcium-Binding Proteins / metabolism*
Cell Differentiation* / drug effects
Cells, Cultured
Coculture Techniques
Embryonic Stem Cells / drug effects
Embryonic Stem Cells / metabolism*
Humans
Induced Pluripotent Stem Cells / drug effects
Induced Pluripotent Stem Cells / metabolism*
Isoproterenol / pharmacology
Mutation
Myocardial Contraction* / drug effects
Myocytes, Cardiac / drug effects
Myocytes, Cardiac / metabolism*
Phenotype
Sarcoplasmic Reticulum / metabolism
Sarcoplasmic Reticulum Calcium-Transporting ATPases / metabolism
Signal Transduction
Tissue Engineering / methods*
Transduction, Genetic
Transfection
Up-Regulation

Substances

Adrenergic beta-Agonists
Calcium-Binding Proteins
phospholamban
Sarcoplasmic Reticulum Calcium-Transporting ATPases
Isoproterenol

Abstract

Publication types

MeSH terms

Substances

Grants and funding