Contractility Drives Cardiomyocyte Maturation and the Response to Nanopatterns

YAP Promotes Microtubule Growth to Facilitate Sarcomere Disassembly in Adult Cardiomyocytes

It is shown that microtubule expression and network density decline with cardiac maturation, which identifies microtubule networks as an essential regulator modulating CM dedifferentiation and sarcomere reorganization, which is critical for CM cytokinesis and cardiac regenerative repair.

An integrated cardiac microtissue proteome map extends therapeutic remodelling by nanovesicles.

Human cardiac microtissues are a promising model to study cardiac biology and disease, but their application is constrained by therapeutic remodelling strategies and limited knowledge of their functional protein expression profiles. Here, we define the use of human cardiac microtissue (hCMT) model generated by assembling iPSC-derived endothelial cells, cardiac fibroblasts, and cardiomyocytes to model ischemia-reperfusion injury (IRI) through a model of hypoxia and reoxygenation and nanovesicle-mediated functional remodelling. Engineered nanovesicles (NVs), generated directly from human stem cells, have been shown to influence cardiac tissue and cell repair, and provide a platform for scalable and reproducible cell free-mediated therapy. We show the functional regulation of the hCMT model and define that administration of NVs (from human induced pluripotent stem cell origin) during reoxygenation significantly increase cardiomyocyte survival and preserve contractility function (contractile duration, relaxation time, relaxation:contraction velocity). We establish NV uptake and transfer with target cells from the hCMT model. Quantitative proteomics was applied to decipher the cell proteome dynamics and molecular mechanisms of IRI in our in vitro model following NV treatment, linked with networks associated with cell survival, energy production, and stress response regulation. Notably, cell type-specific enrichment analysis revealed that NVs drive distinct proteomic remodeling based on their cell origin, where CERA NVs selectively upregulate cytoprotective and structural networks (such as HSP70, MYH6, and XIRP1) within parenchymal cardiomyocytes, whereas CL2 NVs predominantly suppress non-myocyte activation and extracellular matrix remodeling factors within the endothelial and fibroblast compartments. Our findings provide an advanced human stem cell-based platform to understand underlying mechanisms of IRI and assess cell-free therapeutic cardioprotective strategies.

Methodologies for Manipulating Cardiomyocyte Physiology: In Vitro and In Vivo Perspectives

Cardiac tissue is primarily made up of cardiomyocytes, which are regulated by the autonomic nervous system. We have used and developed approaches such as patch clamping and electrical stimulation-combined calcium imaging, computer modeling, optogenetics and chemogenetics combining with video-based Short-Time Fourier transformation (STFT) method to study the physiological activities of cardiomyocytes. The action potential of cardiomyocytes was found to be synchronized with calcium signals, which can be grouped into two categories by STFT. A mathematical model was developed to simulate the changes in electrical activities within cardiomyocytes caused by energy depletion, especially for 2-deoxy-D-glucose (2DG) treatment. Optogenetic and chemogenetics tools, such as ChR2(H134R), OptoXR-β2AR and hM3Dq accelerated beating, while GR, ACR1 and hM4Di inhibited cardiomyocytes’ beating. A video-based STFT method was developed to visualize the beating frequency during these manipulations. An in vitro co-culture method was developed to study the relationship between sympathetic neuronal firing and calcium dynamics in cardiomyocytes. In vivo, electrocardiograph (ECG) measurements showed that Clozapine N-oxide (CNO) caused heart rates increasement in cTnT-hM3Dq virus injected mouse. However, it had no impact on cTnT-hM4Di virus injected mouse. This study provides comprehensive methodologies for studying cardiomyocyte physiology and manipulating heart rates in vitro and in vivo.

Mechanical unloading promotes adult cardiomyocyte proliferation through epicardial NRG1–ERBB4 signaling

Calr-mediated calcium buffering: a molecular barrier to in vivo cardiac reprogramming

In a recent publication in Cell Stem Cell , Cai et al. 1 employed an in vivo Perturb-seq strategy to systematically identify molecular barriers that limit direct cardiac reprogramming following myocardial infarction (MI). This study provides a conceptual and technological framework for improving fi broblast-to-cardiomyocyte conversion in vivo and identi fi es a key regulatory pathway that can be targeted to enhance cardiac regeneration. The limited regenerative capacity of the adult mammalian heart and the resulting loss of functional myocardium after MI under-score the urgent need for strategies that can restore cardiac muscle and improve heart function. Direct cardiac reprogramming has emerged as a promising approach for myocardial repair whereby fi broblasts can be directly converted into induced cardiomyocytes (iCMs). Pioneering studies have demonstrated that de fi ned transcription factors including Mef2c, Gata4, and Tbx5 (collectively referred to as MGT), and later extended combinations such as Myocd and Sall4 (collectively termed MGTMyoS) can induce transdifferentiation of fi broblasts to a cardiomyocyte-like state both in vitro and in vivo. 2 – 4 However, the ef fi ciency and fi delity of in vivo direct reprogramming remain low, representing a major obstacle for clinical translation. This limitation is thought to arise, at least in part, from the complex post-injury microenvironment, where in fl ammatory signaling, extracellular matrix remodeling, and cellular stress responses collectively constrain cell fate conversion. Despite growing recognition of these in fl uences, the molecular mechanisms that restrict cardiac reprogramming ef fi ciency in the injured heart have not been de fi ned. A comprehensive and quantitative understanding of the molecular barriers operating in this context is therefore

A biomimetic mesenchymal stem cell membrane-coated nanodelivery system attenuates cardiac fibrosis by restoring mitochondrial homeostasis.

Myocardial fibrosis is a critical pathological endpoint in heart failure, yet effective targeted therapies remain lacking. Dysregulation of mitochondrial dynamics, particularly Drp1-mediated excessive fission, drives cardiomyocyte dysfunction and pro-fibrotic signaling. Mdivi-1 is a selective Drp1 inhibitor, but suffers from poor solubility, lack of cardiac targeting, and potential systemic toxicity. To address these limitations, we developed a mesenchymal stem cell membrane-coated biomimetic nanodelivery system (MM@NPs/Mdivi-1) with high drug loading capacity and pH-responsive release properties. In vitro, MM@NPs/Mdivi-1 enhanced cellular uptake in injured cardiomyocytes, restored mitochondrial network integrity and membrane potential, and suppressed Drp1 phosphorylation. In an isoproterenol-induced murine cardiac fibrosis model, the system achieved cardiac-specific enrichment, improved cardiac function, reduced collagen deposition, and restored mitochondrial ultrastructure. Mechanistically, transcriptomics combined with functional rescue experiments revealed that its anti-fibrotic effects were dependent on FUNDC1-mediated mitophagy activation. Collectively, MM@NPs/Mdivi-1 effectively attenuates myocardial fibrosis by restoring mitochondrial homeostasis, offering a promising targeted nanotherapeutic strategy for heart failure.