Cell-Surface Marker Signature for Enrichment of Ventricular Cardiomyocytes Derived from Human Embryonic Stem Cells

Abstract

SummaryTo facilitate understanding of human cardiomyocyte (CM) subtype specification, and the study of ventricular CM biology in particular, we developed a broadly applicable strategy for enrichment of ventricular cardiomyocytes (VCMs) derived from human embryonic stem cells (hESCs). A bacterial artificial chromosome transgenic H9 hESC line in which GFP expression was driven by the human ventricular-specific myosin light chain 2 (MYL2) promoter was generated, and screened to identify cell-surface markers specific for MYL2-GFP-expressing VCMs. A CD77+/CD200− cell-surface signature facilitated isolation of >97% cardiac troponin I-positive cells from H9 hESC differentiation cultures, with 65% expressing MYL2-GFP. This study provides a tool for VCM enrichment when using some, but not all, human pluripotent stem cell lines. Tools generated in this study can be utilized toward understanding CM subtype specification, and enriching for VCMs for therapeutic applications.

Transcriptionally active HERV-H retrotransposons demarcate topologically associating domains in human pluripotent stem cells

“…To characterize the dynamics of genome organization during differentiation, we utilized a transgenic human embryonic stem cell (hESC) H9 MYL2:H2B-GFP ventricular cardiomyocyte reporter line, which can be differentiated into ventricular cardiomyocytes in a highly synchronized fashion 27 ( Supplementary Fig. 1a ).…”

Single-Cell RNA-Sequencing and Optical Electrophysiology of Human Induced Pluripotent Stem Cell-Derived Cardiomyocytes Reveal Discordance Between Cardiac Subtype-Associated Gene Expression Patterns and Electrophysiological Phenotypes

“…Together, this led us to profile individual cells via ArcLight and scRNA-seq at early (D12) and later (D40) stages of differentiation, and thus evaluate the cells both before and after distinct cardiomyocyte subtypes might presumably be detected. Electrophysiological classification of cardiomyocyte subtypes has frequently been performed during this time frame, adding additional relevance to our study design [11,[24][25][26][27][28][29].…”

“…Having established the utility of the ArcLight system and determined a classification system for APs, electrophysiological changes were mapped over the course of long-term culture of hiPSC-CMs (*6 weeks of differentiation), a time frame during which a mixed population of cardiomyocyte subtypes has often been described [11,[24][25][26][27][28][29]. Cardiomyocytes at D16 or earlier had a lower AP amplitude ( Fig.…”

Genome-wide association and multi-omic analyses reveal ACTN2 as a gene linked to heart failure

“…Cardiomyocyte differentiation model. To further probe the effects of the ACTN2 locus on cardiomyocyte function, we performed ATAC-seq, ChIP-seq of H3K4me3 and H3K27ac, RNA-seq and HiC experiments 39 , in an engineered H9 hESC (WiCell Research Institute) modified into H9 hESC MLC2v:H2B-GFP reporter transgenic line, which expresses H2B-GFP in differentiated ventricular cardiomyocytes 40 . This cell line was differentiated into cardiomyocytes using a wellestablished Wnt-based differentiation protocol 41 .…”