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Histone deacetylases (HDACs) are a class of enzymes that control chromatin state and influence cell fate. We evaluated the chromatin accessibility and transcriptome dynamics of zinc-containing HDACs during cell differentiation in vitro coupled with chemical perturbation to identify the role of HDACs in mesendoderm cell fate specification. Single-cell RNA sequencing analyses of HDAC expression during human pluripotent stem cell (hPSC) differentiation in vitro and mouse gastrulation in vivo reveal a unique association of HDAC1 and -3 with mesendoderm gene programs during exit from pluripotency. Functional perturbation with small molecules reveals that inhibition of HDAC1 and -3, but not HDAC2, induces mesoderm while impeding endoderm and early cardiac progenitor specification. These data identify unique biological functions of the structurally homologous enzymes HDAC1–3 in influencing hPSC differentiation from pluripotency toward mesendodermal and cardiac progenitor populations. [Display omitted] • Single-cell transcriptomics find HDAC1–3 to be regulators of cardiac differentiation • Inhibition of HDAC1–3 activity results in dynamic chromatin accessibility changes • Loss of HDAC1 and -3, but not HDAC2, impairs cardiac progenitor specification • HDAC1 and -3 are required during early stages of mesendoderm specification Sinniah et al. identify HDAC1 and -3 as required for early mesendoderm and cardiac progenitor specification during human pluripotent stem cell differentiation. Changes to chromatin accessibility and transcriptome dynamics upon perturbation of HDAC activity reveal distinct roles of HDAC1–3 in germ-layer and cardiac-lineage commitment following exit from pluripotency. [ABSTRACT FROM AUTHOR]

Engineered cardiac tissues hold tremendous promise for in vitro drug discovery, studies of heart development and disease, and therapeutic applications. Here, we describe a versatile "frame-hydrogel" methodology to generate engineered cardiac tissues with highly mature functional properties. This methodology has been successfully utilized with a variety of cell sources (neonatal rat ventricular myocytes, human and mouse pluripotent stem cell-derived cardiomyocytes) to generate tissues with diverse 3D geometries (patch, bundle, network) and levels of structural and functional anisotropy. Maturation of such engineered cardiac tissues is rapidly achieved without the need for exogenous electrical or mechanical stimulation or use of complex bioreactors, with tissues routinely reaching conduction velocities and specific forces of 25 cm/s and 20 mN/mm 2 , respectively, and forces per input cardiomyocyte of up to 12 nN. This method is reproducible and readily scalable to generate small tissues ideal for in vitro testing as well as tissues with large, clinically relevant dimensions.