TY - JOUR
T1 - Maturation of hiPSC-derived cardiomyocytes promotes adult alternative splicing of SCN5A and reveals changes in sodium current associated with cardiac arrhythmia
AU - Campostrini, Giulia
AU - Kosmidis, Georgios
AU - Ward-van Oostwaard, Dorien
AU - Davis, Richard Paul
AU - Yiangou, Loukia
AU - Ottaviani, Daniele
AU - Veerman, Christiaan Cornelis
AU - Mei, Hailiang
AU - Orlova, Valeria Viktorovna
AU - Wilde, Arthur Arnold Maria
AU - Bezzina, Connie Rose
AU - Verkerk, Arie Otto
AU - Mummery, Christine Lindsay
AU - Bellin, Milena
N1 - Funding Information: The authors are grateful to C. Freund (the LUMC iPSC core facility, https://www.lumc.nl/research/facilities/hipsc-core-facility/). We thank R. van Helden for help with the ddPCR technique and V. Meraviglia and B. Gabbin for support with immunofluorescence analysis. Graphical abstract and schematics in Figure 4A and B and Supplementary material online, Figure S8 were created with biorender.com. This work was supported by the following grants: the Netherlands Organisation for Health Research and Development ZonMW (MKMD project no. 114022504); European Union’s Horizon 2020 Research and Innovation Programme under the Marie Sklodowska-Curie (MSCA-IF 838985 SiGNATURE); European Research Council (ERC-CoG 101001746 Mini-HEART; ERCAdG 323182 STEMCARDIOVASC; ERC-StG #638030 STEMCARDIORISK); the Rembrandt Institute of Cardiovascular Science; Netherlands Organ-on-Chip Initiative Gravitation project funded by Nederlandse Organisatie voor Wetenschappelijk Onderzoek (NWO 024.003.001); a VIDI and a VICI fellowship from the Netherlands Organisation for Scientific Research (NWO #91715303 and NWO #016.150.610, respectively); Transnational Research Project on Cardiovascular Diseases (JTC2016_FP-40-021 ACM-HF); Health̰Holland (TKI-LSH PPP-allowance, LSHM17013-H007); and Hartstichting, Netherlands CardioVascular Research Initiative (CVON) (PREDICT2). Publisher Copyright: © The Author(s) 2022. Published by Oxford University Press on behalf of the European Society of Cardiology.
PY - 2023/1/1
Y1 - 2023/1/1
N2 - Aims Human-induced pluripotent stem cell-cardiomyocytes (hiPSC-CMs) are widely used to study arrhythmia-associated mutations in ion channels. Among these, the cardiac sodium channel SCN5A undergoes foetal-to-adult isoform switching around birth. Conventional hiPSC-CM cultures, which are phenotypically foetal, have thus far been unable to capture mutations in adult gene isoforms. Here, we investigated whether tri-cellular cross-talk in a three-dimensional (3D) cardiac microtissue (MT) promoted post-natal SCN5A maturation in hiPSC-CMs.Methods We derived patient hiPSC-CMs carrying compound mutations in the adult SCN5A exon 6B and exon 4. and results Electrophysiological properties of patient hiPSC-CMs in monolayer were not altered by the exon 6B mutation compared with isogenic controls since it is not expressed; further, CRISPR/Cas9-mediated excision of the foetal exon 6A did not promote adult SCN5A expression. However, when hiPSC-CMs were matured in 3D cardiac MTs, SCN5A underwent isoform switch and the functional consequences of the mutation located in exon 6B were revealed. Up-regulation of the splicing factor muscleblind-like protein 1 (MBNL1) drove SCN5A post-natal maturation in microtissues since its overexpression in hiPSC-CMs was sufficient to promote exon 6B inclusion, whilst knocking-out MBNL1 failed to foster isoform switch.Conclusions Our study shows that (i) the tri-cellular cardiac microtissues promote post-natal SCN5A isoform switch in hiPSC-CMs, (ii) adult splicing of SCN5A is driven by MBNL1 in these tissues, and (iii) this model can be used for examining post-natal cardiac arrhythmias due to mutations in the exon 6B.Translational The cardiac sodium channel is essential for conducting the electrical impulse in the heart. Postnatal alternative splicing perspective regulation causes mutual exclusive inclusion of fetal or adult exons of the corresponding gene, SCN5A. Typically, immature hiPSCCMs fall short in studying the effect of mutations located in the adult exon. We describe here that an innovative tri-cellular three-dimensional cardiac microtissue culture promotes hiPSC-CMs maturation through upregulation of MBNL1, thus revealing the effect of a pathogenic genetic variant located in the SCN5A adult exon. These results help advancing the use of hiPSC-CMs in studying adult heart disease and for developing personalized medicine applications.
AB - Aims Human-induced pluripotent stem cell-cardiomyocytes (hiPSC-CMs) are widely used to study arrhythmia-associated mutations in ion channels. Among these, the cardiac sodium channel SCN5A undergoes foetal-to-adult isoform switching around birth. Conventional hiPSC-CM cultures, which are phenotypically foetal, have thus far been unable to capture mutations in adult gene isoforms. Here, we investigated whether tri-cellular cross-talk in a three-dimensional (3D) cardiac microtissue (MT) promoted post-natal SCN5A maturation in hiPSC-CMs.Methods We derived patient hiPSC-CMs carrying compound mutations in the adult SCN5A exon 6B and exon 4. and results Electrophysiological properties of patient hiPSC-CMs in monolayer were not altered by the exon 6B mutation compared with isogenic controls since it is not expressed; further, CRISPR/Cas9-mediated excision of the foetal exon 6A did not promote adult SCN5A expression. However, when hiPSC-CMs were matured in 3D cardiac MTs, SCN5A underwent isoform switch and the functional consequences of the mutation located in exon 6B were revealed. Up-regulation of the splicing factor muscleblind-like protein 1 (MBNL1) drove SCN5A post-natal maturation in microtissues since its overexpression in hiPSC-CMs was sufficient to promote exon 6B inclusion, whilst knocking-out MBNL1 failed to foster isoform switch.Conclusions Our study shows that (i) the tri-cellular cardiac microtissues promote post-natal SCN5A isoform switch in hiPSC-CMs, (ii) adult splicing of SCN5A is driven by MBNL1 in these tissues, and (iii) this model can be used for examining post-natal cardiac arrhythmias due to mutations in the exon 6B.Translational The cardiac sodium channel is essential for conducting the electrical impulse in the heart. Postnatal alternative splicing perspective regulation causes mutual exclusive inclusion of fetal or adult exons of the corresponding gene, SCN5A. Typically, immature hiPSCCMs fall short in studying the effect of mutations located in the adult exon. We describe here that an innovative tri-cellular three-dimensional cardiac microtissue culture promotes hiPSC-CMs maturation through upregulation of MBNL1, thus revealing the effect of a pathogenic genetic variant located in the SCN5A adult exon. These results help advancing the use of hiPSC-CMs in studying adult heart disease and for developing personalized medicine applications.
UR - http://www.scopus.com/inward/record.url?scp=85159143765&partnerID=8YFLogxK
U2 - https://doi.org/10.1093/cvr/cvac059
DO - https://doi.org/10.1093/cvr/cvac059
M3 - Article
C2 - 35394010
SN - 0008-6363
VL - 119
SP - 167
EP - 182
JO - Cardiovascular research
JF - Cardiovascular research
IS - 1
ER -