Cardiac stereotactic body radiotherapy to treat malignant ventricular arrhythmias directly affects the cardiomyocyte electrophysiology.

Background: Promising as a treatment option for life-threatening ventricular arrhythmias, cardiac stereotactic body radiotherapy (cSBRT) has demonstrated early antiarrhythmic effects within days of treatment. The mechanisms underlying the immediate and short-term antiarrhythmic effects are poorly understood.

Objective: We hypothesize that cSBRT has a direct antiarrhythmic effect on cellular electrophysiology through reprogramming of ion channel and gap junction protein expression.

BIN1, Myotubularin, and Dynamin-2 Coordinate T-Tubule Growth in Cardiomyocytes.

Background: Transverse tubules (t-tubules) form gradually in the developing heart, critically enabling maturation of cardiomyocyte Ca homeostasis. The membrane bending and scaffolding protein BIN1 (bridging integrator 1) has been implicated in this process. However, it is unclear which of the various reported BIN1 isoforms are involved, and whether BIN1 function is regulated by its putative binding partners MTM1 (myotubularin), a phosphoinositide 3'-phosphatase, and DNM2 (dynamin-2), a GTPase believed to mediate membrane fission.

Membrane remodelling triggers maturation of excitation-contraction coupling in 3D-shaped human-induced pluripotent stem cell-derived cardiomyocytes.

The prospective use of human-induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CM) for cardiac regenerative medicine strongly depends on the electro-mechanical properties of these cells, especially regarding the Ca-dependent excitation-contraction (EC) coupling mechanism. Currently, the immature structural and functional features of hiPSC-CM limit the progression towards clinical applications. Here, we show that a specific microarchitecture is essential for functional maturation of hiPSC-CM.

Distress-Mediated Remodeling of Cardiac Connexin-43 in a Novel Cell Model for Arrhythmogenic Heart Diseases.

Gap junctions and their expression pattern are essential to robust function of intercellular communication and electrical propagation in cardiomyocytes. In healthy myocytes, the main cardiac gap junction protein connexin-43 (Cx43) is located at the intercalated disc providing a clear direction of signal spreading across the cardiac tissue. Dislocation of Cx43 to lateral membranes has been detected in numerous cardiac diseases leading to slowed conduction and high propensity for the development of arrhythmias.

Improved Generation of Human Induced Pluripotent Stem Cell-Derived Cardiac Pacemaker Cells Using Novel Differentiation Protocols.

Current protocols for the differentiation of human-induced pluripotent stem cells (hiPSC) into cardiomyocytes only generate a small amount of cardiac pacemaker cells. In previous work, we reported the generation of high amounts of cardiac pacemaker cells by co-culturing hiPSC with mouse visceral endoderm-like (END2) cells. However, potential medical applications of cardiac pacemaker cells generated according to this protocol, comprise an incalculable xenogeneic risk.

The Structural and the Functional Aspects of Intercellular Communication in iPSC-Cardiomyocytes.

Recent advances in the technology of producing novel cardiomyocytes from induced pluripotent stem cells (iPSC-cardiomyocytes) fuel new hope for future clinical applications. The use of iPSC-cardiomyocytes is particularly promising for the therapy of cardiac diseases such as myocardial infarction, where these cells could replace scar tissue and restore the functionality of the heart. Despite successful cardiogenic differentiation, medical applications of iPSC-cardiomyocytes are currently limited by their pronounced immature structural and functional phenotype.

Substrate Stiffness Influences Structural and Functional Remodeling in Induced Pluripotent Stem Cell-Derived Cardiomyocytes.

Novel treatment strategies for cardiac tissue regeneration are heading for the use of engineered cardiac tissue made from induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs). Despite the proven cardiogenic phenotype of these cells, a significant lack of structural and functional properties of mature myocytes prevents safe integration into the diseased heart. To date, maturation processes of cardiomyocytes remain largely unknown but may comprise biophysical cues from the immediate cell environment.

AAV-mediated expression of NFAT decoy oligonucleotides protects from cardiac hypertrophy and heart failure.

Previous studies have underlined the substantial role of nuclear factor of activated T cells (NFAT) in hypertension-induced myocardial hypertrophy ultimately leading to heart failure. Here, we aimed at neutralizing four members of the NFAT family of transcription factors as a therapeutic strategy for myocardial hypertrophy transiting to heart failure through AAV-mediated cardiac expression of a RNA-based decoy oligonucleotide (dON) targeting NFATc1-c4. AAV-mediated dON expression markedly decreased endothelin-1 induced cardiomyocyte hypertrophy in vitro and resulted in efficient expression of these dONs in the heart of adult mice as evidenced by fluorescent in situ hybridization.

Trigger-Specific Remodeling of K2 Potassium Channels in Models of Atrial Fibrillation.

Aim: Effective antiarrhythmic treatment of atrial fibrillation (AF) constitutes a major challenge, in particular, when concomitant heart failure (HF) is present. HF-associated atrial arrhythmogenesis is distinctly characterized by prolonged atrial refractoriness. Small-conductance, calcium-activated K (K, SK, ) channels contribute to cardiac action potential repolarization and are implicated in AF susceptibility and therapy.

Inhibition of cardiac K4.3 (I) channel isoforms by class I antiarrhythmic drugs lidocaine and mexiletine.

Transient outward K current, I, contributes to cardiac action potential generation and is primarily carried by K4.3 (KCND3) channels. Two K4.

Shaping the heart: Structural and functional maturation of iPSC-cardiomyocytes in 3D-micro-scaffolds.

Cardiomyocytes derived from induced pluripotent stem cells (iPSC-CMs) represent the best cell source for cardiac regenerative purposes but retain an immature phenotype after differentiation with significant limitations compared to adult cardiomyocytes. Apart from an incomplete cardiomyocyte-specific structure and microarchitecture, cells show at the level of Ca signaling only slow Ca release and reuptake properties. Here, we investigated the effect of restructuring single iPSC-CMs in specially designed 3D-micro-scaffolds on cell morphology and Ca handling.

Endothelial cell modulation of cardiomyocyte gene expression.

The anatomic arrangement of microvascular endothelial cells and cardiomyocytes in vivo enables close interactions among these cells. In our in vitro co-culture system, ANP and BNP expression in the mouse atrial cardiomyocyte cell line HL-1 and subsequent ANP release were significantly upregulated when co-cultured with mouse cardiac microvascular endothelial cells or exposed to endothelial cell-conditioned medium. Endothelin-1 (ET-1) activation of endothelial cells remarkably enhanced their paracrine effect on cardiomyocyte gene expression, suggesting that ET-1 stimulation of endothelial cells affects expression of fetal genes such as ANP and BNP in adult cardiomyocytes through paracrine signalling.

Somatic mutations and promotor methylation of the ryanodine receptor 2 is a common event in the pathogenesis of head and neck cancer.

Genomic sequencing projects unraveled the mutational landscape of head and neck squamous cell carcinoma (HNSCC) and provided a comprehensive catalog of somatic mutations. However, the limited number of significant cancer-related genes obtained so far only partially explains the biological complexity of HNSCC and hampers the development of novel diagnostic biomarkers and therapeutic targets. We pursued a multiscale omics approach based on whole-exome sequencing, global DNA methylation and gene expression profiling data derived from tumor samples of the HIPO-HNC cohort (n = 87), and confirmed new findings with datasets from The Cancer Genome Atlas (TCGA).

The VAMP-associated protein VAPB is required for cardiac and neuronal pacemaker channel function.

Hyperpolarization-activated cyclic nucleotide-gated (HCN) channels encode neuronal and cardiac pacemaker currents. The composition of pacemaker channel complexes in different tissues is poorly understood, and the presence of additional HCN modulating subunits was speculated. Here we show that vesicle-associated membrane protein-associated protein B (VAPB), previously associated with a familial form of amyotrophic lateral sclerosis 8, is an essential HCN1 and HCN2 modulator.

Improving electrical properties of iPSC-cardiomyocytes by enhancing Cx43 expression.

The therapeutic potential of induced pluripotent stem cell-derived cardiomyocytes (iPSC-CMs) is limited by immature functional features including low impulse propagation and reduced cell excitability. Key players regulating electrical activity are voltage-gated Na channels (Na1.5) and gap junctions built from connexin-43 (Cx43).

Subtype-specific differentiation of cardiac pacemaker cell clusters from human induced pluripotent stem cells.

Background: Human induced pluripotent stem cells (hiPSC) harbor the potential to differentiate into diverse cardiac cell types. Previous experimental efforts were primarily directed at the generation of hiPSC-derived cells with ventricular cardiomyocyte characteristics. Aiming at a straightforward approach for pacemaker cell modeling and replacement, we sought to selectively differentiate cells with nodal-type properties.

Bacopa monnieri extract increases rat coronary flow and protects against myocardial ischemia/reperfusion injury.

Background: This study explored Bacopa monnieri, a medicinal Ayurvedic herb, as a cardioprotectant against ischemia/reperfusion injury using cardiac function and coronary flow as end-points.

Methods: In normal isolated rat hearts, coronary flow, left ventricular developed pressure, heart rate, and functional recovery were measured using the Langendorff preparation. Hearts were perfused with either (i) Krebs-Henseleit (normal) solution, (control), or with 30, 100 μg/ml B.

Functional characterization of orbicularis oculi and extraocular muscles.

The orbicularis oculi are the sphincter muscles of the eyelids and are involved in modulating facial expression. They differ from both limb and extraocular muscles (EOMs) in their histology and biochemistry. Weakness of the orbicularis oculi muscles is a feature of neuromuscular disorders affecting the neuromuscular junction, and weakness of facial muscles and ptosis have also been described in patients with mutations in the ryanodine receptor gene.

Slow conduction in mixed cultured strands of primary ventricular cells and stem cell-derived cardiomyocytes.

Modern concepts for the treatment of myocardial diseases focus on novel cell therapeutic strategies involving stem cell-derived cardiomyocytes (SCMs). However, functional integration of SCMs requires similar electrophysiological properties as primary cardiomyocytes (PCMs) and the ability to establish intercellular connections with host myocytes in order to contribute to the electrical and mechanical activity of the heart. The aim of this project was to investigate the properties of cardiac conduction in a co-culture approach using SCMs and PCMs in cultured cell strands.

Functional Characterization and Comparison of Intercellular Communication in Stem Cell-Derived Cardiomyocytes.

One novel treatment strategy for the diseased heart focuses on the use of pluripotent stem cell-derived cardiomyocytes (SC-CMs) to overcome the heart's innate deficiency for self-repair. However, targeted application of SC-CMs requires in-depth characterization of their true cardiogenic potential in terms of excitability and intercellular coupling at cellular level and in multicellular preparations. In this study, we elucidated the electrical characteristics of single SC-CMs and intercellular coupling quality of cell pairs, and concomitantly compared them with well-characterized murine native neonatal and immortalized HL-1 cardiomyocytes.

Development and Characterization of a Scaffold-Free 3D Spheroid Model of Induced Pluripotent Stem Cell-Derived Human Cardiomyocytes.

Cardiomyocytes (CMs) are terminally differentiated cells in the adult heart, and ischemia and cardiotoxic compounds can lead to cell death and irreversible decline of cardiac function. As testing platforms, isolated organs and primary cells from rodents have been the standard in research and toxicology, but there is a need for better models that more faithfully recapitulate native human biology. Hence, a new in vitro model comprising the advantages of 3D cell culture and the availability of induced pluripotent stem cells (iPSCs) of human origin was developed and characterized.

Characterisation of connexin expression and electrophysiological properties in stable clones of the HL-1 myocyte cell line.

The HL-1 atrial line contains cells blocked at various developmental stages. To obtain homogeneous sub-clones and correlate changes in gene expression with functional alterations, individual clones were obtained and characterised for parameters involved in conduction and excitation-contraction coupling. Northern blots for mRNAs coding for connexins 40, 43 and 45 and calcium handling proteins (sodium/calcium exchanger, L- and T-type calcium channels, ryanodine receptor 2 and sarco-endoplasmic reticulum calcium ATPase 2) were performed.

Dynamic patterns of ventricular remodeling and apoptosis in hearts unloaded by heterotopic transplantation.

Background: Mechanical unloading of failing hearts can trigger functional recovery but results in progressive atrophy and possibly detrimental adaptation. In an unbiased approach, we examined the dynamic effects of unloading duration on molecular markers indicative of myocardial damage, hypothesizing that potential recovery may be improved by optimized unloading time.

Methods: Heterotopically transplanted normal rat hearts were harvested at 3, 8, 15, 30, and 60 days.

Establishment of a human skeletal muscle-derived cell line: biochemical, cellular and electrophysiological characterization.

Excitation-contraction coupling is the physiological mechanism occurring in muscle cells whereby an electrical signal sensed by the dihydropyridine receptor located on the transverse tubules is transformed into a chemical gradient (Ca2+ increase) by activation of the ryanodine receptor located on the sarcoplasmic reticulum membrane. In the present study, we characterized for the first time the excitation-contraction coupling machinery of an immortalized human skeletal muscle cell line. Intracellular Ca2+ measurements showed a normal response to pharmacological activation of the ryanodine receptor, whereas 3D-SIM (super-resolution structured illumination microscopy) revealed a low level of structural organization of ryanodine receptors and dihydropyridine receptors.