397 results found
Schneider M, Golforoush P, Narasimhan P, et al., Selective protection of human cardiomyocytes from anthracycline cardiotoxicity by small molecule inhibitors of MAP4K4, Scientific Reports
Sparks H, Dvinskikh L, Firth J, et al., 2020, Development a flexible light-sheet fluorescence microscope for high-speed 3D imaging of calcium dynamics and 3D imaging of cellular microstructure, Journal of Biophotonics, Vol: 13, ISSN: 1864-063X
We report a flexible light‐sheet fluorescence microscope (LSFM) designed for studying dynamic events in cardiac tissue at high speed in 3D and the correlation of these events to cell microstructure. The system employs two illumination‐detection modes: the first uses angle‐dithering of a Gaussian light sheet combined with remote refocusing of the detection plane for video‐rate volumetric imaging; the second combines digitally‐scanned light‐sheet illumination with an axially‐swept light‐sheet waist and stage‐scanned acquisition for improved axial resolution compared to the first mode. We present a characterisation of the spatial resolution of the system in both modes. The first illumination‐detection mode achieves dual spectral‐channel imaging at 25 volumes per second with 1024 × 200 × 50 voxel volumes and is demonstrated by time‐lapse imaging of calcium dynamics in a live cardiomyocyte. The second illumination‐detection mode is demonstrated through the acquisition of a higher spatial resolution structural map of the t‐tubule network in a fixed cardiomyocyte cell.
Constantinou C, Miranda Almeida A, Chaves Guerrero P, et al., Human pluripotent stem cell-derived cardiomyocytes as a targetplatform for paracrine protection by cardiac mesenchymal stromalcells, Scientific Reports, ISSN: 2045-2322
Pitoulis FG, Hasan W, Papadaki M, et al., 2020, Intact myocardial preparations reveal intrinsic transmural heterogeneity in cardiac mechanics, Journal of Molecular and Cellular Cardiology, Vol: 141, Pages: 11-16, ISSN: 0022-2828
Determining transmural mechanical properties in the heart provides a foundation to understand physiological and pathophysiological cardiac mechanics. Although work on mechanical characterisation has begun in isolated cells and permeabilised samples, the mechanical profile of living individual cardiac layers has not been examined. Myocardial slices are 300 μm-thin sections of heart tissue with preserved cellular stoichiometry, extracellular matrix, and structural architecture. This allows for cardiac mechanics assays in the context of an intact in vitro organotypic preparation. In slices obtained from the subendocardium, midmyocardium and subepicardium of rats, a distinct pattern in transmural contractility is found that is different from that observed in other models. Slices from the epicardium and midmyocardium had a higher active tension and passive tension than the endocardium upon stretch. Differences in total myocyte area coverage, and aspect ratio between layers underlined the functional readouts, while no differences were found in total sarcomeric protein and phosphoprotein between layers. Such intrinsic heterogeneity may orchestrate the normal pumping of the heart in the presence of transmural strain and sarcomere length gradients in the in vivo heart.
Schobesberger S, Wright PT, Poulet C, et al., 2020, beta(3)-Adrenoceptor redistribution impairs NO/cGMP/PDE2 signalling in failing cardiomyocytes, eLife, Vol: 9, Pages: 1-15, ISSN: 2050-084X
Cardiomyocyte β3-adrenoceptors (β3-ARs) coupled to soluble guanylyl cyclase (sGC)-dependent production of the second messenger 3’,5’-cyclic guanosine monophosphate (cGMP) have been shown to protect from heart failure. However, the exact localization of these receptors to fine membrane structures and subcellular compartmentation of β3-AR/cGMP signals underpinning this protection in health and disease remain elusive. Here, we used a Förster Resonance Energy Transfer (FRET)-based cGMP biosensor combined with scanning ion conductance microscopy (SICM) to show that functional β3-ARs are mostly confined to the T-tubules of healthy rat cardiomyocytes. Heart failure, induced via myocardial infarction, causes a decrease of the cGMP levels generated by these receptors and a change of subcellular cGMP compartmentation. Furthermore, attenuated cGMP signals led to impaired phosphodiesterase two dependent negative cGMP-to-cAMP cross-talk. In conclusion, topographic and functional reorganization of the β3-AR/cGMP signalosome happens in heart failure and should be considered when designing new therapies acting via this receptor.
Fiedler LR, Chapman K, Xie M, et al., 2020, MAP4K4 Inhibition Promotes Survival of Human Stem Cell-Derived Cardiomyocytes and Reduces Infarct Size In Vivo (vol 24, 579.e1,2019), CELL STEM CELL, Vol: 26, Pages: 458-458, ISSN: 1934-5909
Pitoulis FG, Nunez-Toldra R, Kit-Anan WS, et al., 2020, Exploring Mechanical Load-Induced Cardiac Remodelling Using a Novel Organotypic Myocardial Slice Model, 64th Annual Meeting of the Biophysical-Society, Publisher: CELL PRESS, Pages: 425A-425A, ISSN: 0006-3495
Lyon A, Babalis D, Morley-Smith AC, et al., Investigation of the safety and feasibility of AAV1/SERCA2a gene transfer in patients with chronic heart failure supported with a left ventricular assist device – the SERCA-LVAD TRIAL, Gene Therapy, ISSN: 0969-7128
The SERCA-LVAD trial was a phase 2a trial assessing the safety and feasibility of delivering an adeno-associated vector 1 carrying the cardiac isoform of the sarcoplasmic reticulum calcium ATPase (AAV1/SERCA2a) to adult chronic heart failure patients implanted with a left ventricular assist device. Enrolled subjects were randomised to receive a single intracoronary infusion of 1x1013 DNase-resistant AAV1/SERCA2a particles or a placebo solution in a double-blinded design, stratified by presence of neutralising antibodies to AAV. Elective endomyocardial biopsy was performed at 6 months unless the subject had undergone cardiac transplantation, with myocardial samples assessed for the presence of exogenous viral DNA from the treatment vector. Safety assessments including ELISPOT were serially performed. Although designed as a 24 subject trial, recruitment was stopped after five subjects had been randomised and received infusion due to the neutral result from the CUPID 2 trial. Here we describe the results from the 5 patients, which confirmed that viral DNA was delivered to the failing human heart in 2 patients receiving gene therapy with vector detectable at follow up endomyocardial biopsy or cardiac transplantation. Absolute levels of detectable transgene DNA were low, and no functional benefit was observed. There were no safety concerns in this small cohort. This trial identified some of the challenges of performing gene therapy trials in this LVAD patient cohort, which may help guide future trial design.
Jabbour RJ, Owen TJ, Pandey P, et al., 2019, Future potential of engineered heart tissue patches for repairing the damage caused by heart attacks, EXPERT REVIEW OF MEDICAL DEVICES, Vol: 17, Pages: 1-3, ISSN: 1743-4440
Shun-Shin MJ, Leong KMW, Ng FS, et al., 2019, Ventricular conduction stability test: a method to identify and quantify changes in whole heart activation patterns during physiological stress, EP-Europace, Vol: 21, Pages: 1422-1431, ISSN: 1099-5129
AIMS: Abnormal rate adaptation of the action potential is proarrhythmic but is difficult to measure with current electro-anatomical mapping techniques. We developed a method to rapidly quantify spatial discordance in whole heart activation in response to rate cycle length changes. We test the hypothesis that patients with underlying channelopathies or history of aborted sudden cardiac death (SCD) have a reduced capacity to maintain uniform activation following exercise. METHODS AND RESULTS: Electrocardiographical imaging (ECGI) reconstructs >1200 electrograms (EGMs) over the ventricles from a single beat, providing epicardial whole heart activation maps. Thirty-one individuals [11 SCD survivors; 10 Brugada syndrome (BrS) without SCD; and 10 controls] with structurally normal hearts underwent ECGI vest recordings following exercise treadmill. For each patient, we calculated the relative change in EGM local activation times (LATs) between a baseline and post-exertion phase using custom written software. A ventricular conduction stability (V-CoS) score calculated to indicate the percentage of ventricle that showed no significant change in relative LAT (<10 ms). A lower score reflected greater conduction heterogeneity. Mean variability (standard deviation) of V-CoS score over 10 consecutive beats was small (0.9 ± 0.5%), with good inter-operator reproducibility of V-CoS scores. Sudden cardiac death survivors, compared to BrS and controls, had the lowest V-CoS scores post-exertion (P = 0.011) but were no different at baseline (P = 0.50). CONCLUSION: We present a method to rapidly quantify changes in global activation which provides a measure of conduction heterogeneity and proof of concept by demonstrating SCD survivors have a reduced capacity to maintain uniform activation following exercise.
Sattler S, Baxan N, Chowdhury R, et al., 2019, Characterization of acute TLR-7 agonist-induced hemorrhagic myocarditis in mice by multi-parametric quantitative cardiac MRI, Disease Models & Mechanisms, Vol: 12, Pages: 1-10, ISSN: 1754-8403
Hemorrhagic myocarditis is a potentially fatal complication of excessive levels of systemic inflammation. It has been reported in viral infection, but is also possible in systemic autoimmunity. Epicutaneous treatment of mice with the TLR-7 agonist Resiquimod induces auto-antibodies and systemic tissue damage including in the heart, and is used as an inducible mouse model of Systemic Lupus Erythematosus (SLE).Here, we show that over-activation of the TLR-7 pathway of viral recognition by Resiquimod-treatment of CFN mice induces severe thrombocytopenia and internal bleeding which manifests most prominently as hemorrhagic myocarditis. We optimized a cardiac magnetic resonance (CMR) tissue mapping approach for the in vivo detection of diffuse infiltration, fibrosis and hemorrhages using a combination of T1, T2 and T2* relaxation times, and compared results to ex vivo histopathology of cardiac sections corresponding to CMR tissue maps. This allowed a detailed correlation between in vivo CMR parameters and ex vivo histopathology, and confirmed the need to include T2* measurements to detect tissue iron for accurate interpretation of pathology associated with CMR parameter changes.In summary, we provide detailed histological and in vivo imaging-based characterization of acute hemorrhagic myocarditis as acute cardiac complication in the mouse model of Resiquimod-induced SLE, and a refined CMR protocol to allow non-invasive longitudinal in vivo studies of heart involvement in acute inflammation. We propose that adding T2* mapping to CMR protocols for myocarditis diagnosis will improve interpretation of disease mechanisms and diagnostic sensitivity.
Owen TJ, Harding SE, 2019, Multi-cellularity in cardiac tissue engineering, how close are we to native heart tissue?, Journal of Muscle Research and Cell Motility, Vol: 40, Pages: 151-157, ISSN: 0142-4319
Tissue engineering is a complex field where the elements of biology and engineering are combined in an attempt to recapitulate the native environment of the body. Tissue engineering has shown one thing categorically; that the human body is extremely complex and it is truly a difficult task to generate this in the lab. There have been varied attempts at trying to generate a model for the heart with numerous cell types and different scaffolds or materials. The common underlying theme in these approaches is to combine together matrix material and different cell types to make something similar to heart tissue. Multi-cellularity is an essential aspect of the heart and therefore critical to any approach which would try to mimic such a complex tissue. The heart is made up of many cell types that combine to form complex structures like: deformable chambers, a tri-layered heart muscle, and vessels. Thus, in this review we will summarise how tissue engineering has progressed in modelling the heart and what gaps still exist in this dynamic field.
Watson S, Duff J, Bardi I, et al., 2019, Biomimetic electromechanical stimulation to maintain adult myocardial slices in vitro, Nature Communications, Vol: 10, ISSN: 2041-1723
Adult cardiac tissue undergoes a rapid process of dedifferentiation when cultured outside the body. The in vivo environment, particularly constant electromechanical stimulation, is fundamental to the regulation of cardiac structure and function. We investigated the role of electromechanical stimulation in preventing culture-induced dedifferentiation of adult cardiac tissue using rat, rabbit and human heart failure myocardial slices. Here we report that the application of a preload equivalent to sarcomere length (SL) = 2.2 μm is optimal for the maintenance of rat myocardial slice structural, functional and transcriptional properties at 24 h. Gene sets associated with the preservation of structure and function are activated, while gene sets involved in dedifferentiation are suppressed. The maximum contractility of human heart failure myocardial slices at 24 h is also optimally maintained at SL = 2.2 μm. Rabbit myocardial slices cultured at SL = 2.2 μm remain stable for 5 days. This approach substantially prolongs the culture of adult cardiac tissue in vitro.
Hellen N, Pinto Ricardo C, Vauchez K, et al., 2019, Proteomic analysis reveals temporal changes in protein expression in human induced pluripotent stem cell-derived cardiomyocytes in vitro, Stem Cells and Development, Vol: 28, ISSN: 1547-3287
Human induced pluripotent stem cell-derived cardiomyocytes hold great promise for regenerative medicine and in vitro screening. Despite displaying key cardiomyocyte phenotypic characteristics, they more closely resemble foetal/neonatal cardiomyocytes and further characterisation is necessary. Combining the use of tandem mass tags to label cell lysates, followed by multiplexing, we have determined the effects of short term (30 day) in vitro culture on human induced pluripotent stem cell derived cardiomyocyte protein expression. We found that human induced pluripotent stem cell derived cardiomyocytes exhibit temporal changes in global protein expression; alterations in protein expression were pronounced during the first 2 weeks following thaw and dominated by reductions in proteins associated with protein synthesis and ubiquitination. Between 2 and 4 weeks proceeding thaw alterations in protein expression were dominated by metabolic pathways, indicating a potential temporal metabolic shift from glycolysis towards oxidative phosphorylation. Time-dependent changes in proteins associated with cardiomyocyte contraction, excitation-contraction coupling and metabolism were detected. While some were associated with expected functional outcomes in terms of morphology or electrophysiology, others such as metabolism did not produce the anticipated maturation of human induced pluripotent stem cell derived cardiomyocytes. In several cases, a predicted outcome was not clear because of the concerted changes in both stimulatory and inhibitory pathways. Nevertheless, clear development of human induced pluripotent stem cell derived cardiomyocytes over this time period was evident.
Schneider M, Fiedler L, Chapman K, et al., 2019, MAP4K4 inhibition promotes survival of human stem cell derived cardiomyocyte and reduces infarct size in vivo, Cell Stem Cell, Vol: 24, Pages: 579-591.e12, ISSN: 1875-9777
Heart disease is a paramount cause of global death and disability. Although cardiomyocyte death plays a causal role and its suppression would be logical, no clinical counter-measures target the responsible intracellular pathways. Therapeutic progress has been hampered by lack of preclinical human validation. Mitogen-activated protein kinase kinase kinase kinase-4 (MAP4K4) is activated in failing human hearts and relevant rodent models. Using human induced-pluripotent-stem-cell-derived cardiomyocytes (hiPSC-CMs) and MAP4K4 gene silencing, we demonstrate that death induced by oxidative stress requires MAP4K4. Consequently, we devised a small-molecule inhibitor, DMX-5804, that rescues cell survival, mitochondrial function, and calcium cycling in hiPSC-CMs. As proof of principle that drug discovery in hiPSC-CMs may predict efficacy in vivo, DMX-5804 reduces ischemia-reperfusion injury in mice by more than 50%. We implicate MAP4K4 as a well-posed target toward suppressing human cardiac cell death and highlight the utility of hiPSC-CMs in drug discovery to enhance cardiomyocyte survival.
Wang BX, Kit-Anan W, Whittaker T, et al., 2019, Human Cardiac Fibroblast-Secreted Exosomes Improve Efficiency of Human Cardiomyocyte Calcium Cycling, Publisher: SPRINGER, Pages: 269-269, ISSN: 0920-3206
Brito L, Mylonaki I, Moroz E, et al., 2019, Epicardial cell transfection with cationic polymeric nanocomplexes, British-Society-for-Gene-and-Cell-Therapy Autumn Conference, Publisher: MARY ANN LIEBERT, INC, Pages: A9-A9, ISSN: 1043-0342
Owen TJ, Smith JS, Bhagwan JR, et al., 2019, Isogenic pairs of patient-derived hiPSC-CMs with an apical hypertrophic cardiomyopathy/left ventricular non-compaction-associated ACTC1 E99K mutation unveil differential functional deficits, British-Society-for-Gene-and-Cell-Therapy Autumn Conference, Publisher: MARY ANN LIEBERT, INC, Pages: A7-A8, ISSN: 1043-0342
© 2019 Elsevier B.V. All rights reserved. Stem cells have emerged as a justifiable and promising medical therapy for patients with heart failure. Many cell types have been contemplated during the past years, and there is promising preclinical and clinical data on their efficacy and safety in the setting of acute myocardial infarction and chronic heart failure. However, there are still many hurdles to be overcome for the routine clinical application of these cells. In current preclinical models, small molecules, epigenetic modifications, tissue engineering and gene modifications are being used to treat a diverse array of diseases. A better understanding of the biology of stem cell-derived cardiovascular cells and their application will help in the design of future strategies for functional cardiac regeneration.
Dubey P, Humphrey E, Majid Q, et al., 2019, Polyhydroxyalkanoates, ideal materials for cardiac regeneration, ISSN: 1526-7547
© 2019 Omnipress - All rights reserved. Statement of purpose: Polyhydroxyalkanoates (PHAs), a family of biodegradable and biocompatible polymers with a range of material properties and degradation rates are known to be particularly cardio-regenerative in nature1,2,3. Myocardial infarction results in the generation of scar tissue with limited or no regeneration due to the modest nature of intrinsic myocardial regenerative capability. The concept of a cardiac patch is tailored to meet the unmet medical need of cardiac regeneration where a biomaterial-based patch with/without cells would be used to induce efficient cardiac regeneration. Medium chain length PHAs (MCL-PHAs) with monomer chain length between C6-C16 are highly elastomeric in nature1,2 and have been shown to be excellent substrates for the growth and function of neonatal cardiomyocytes3. This work describes an in-depth study of the potential of MCL-PHAs for the development of functional cardiac patches laden with human pluripotent stem cell derived cardiomyocytes (hiPSC-CMs), providing mechanical support and cell based therapy.
Sattler S, Couch LS, Harding SE, 2018, Takotsubo Syndrome: Latest addition to the expanding family of immune-mediated diseases?, JACC: Basic to Translational Science, Vol: 3, Pages: 779-781, ISSN: 2452-302X
Smith JGW, Owen T, Bhagwan JR, et al., 2018, Isogenic pairs of hiPSC-CMs with hypertrophic cardiomyopathy/LVNC-associated ACTC1 E99K mutation unveil differential functional deficits, Stem Cell Reports, Vol: 11, Pages: 1226-1243, ISSN: 2213-6711
Hypertrophic cardiomyopathy (HCM) is a primary disorder of contractility in heart muscle. To gain mechanistic insight and guide pharmacological rescue, this study models HCM using isogenic pairs of human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) carrying the E99K-ACTC1 cardiac actin mutation. In both 3D engineered heart tissues and 2D monolayers, arrhythmogenesis was evident in all E99K-ACTC1 hiPSC-CMs. Aberrant phenotypes were most common in hiPSC-CMs produced from the heterozygote father. Unexpectedly, pathological phenotypes were less evident in E99K-expressing hiPSC-CMs from the two sons. Mechanistic insight from Ca2+ handling expression studies prompted pharmacological rescue experiments, wherein dual dantroline/ranolazine treatment was most effective. Our data are consistent with E99K mutant protein being a central cause of HCM but the three-way interaction between the primary genetic lesion, background (epi)genetics, and donor patient age may influence the pathogenic phenotype. This illustrates the value of isogenic hiPSC-CMs in genotype-phenotype correlations.
Noseda M, Harding SE, 2018, Understanding dynamic tissue organization by studying the human body one cell at a time: the human cell atlas (HCA) project, CARDIOVASCULAR RESEARCH, Vol: 114, Pages: E93-E95, ISSN: 0008-6363
Panahi M, Papanikolaou A, Torabi A, et al., 2018, Immunomodulatory interventions in myocardial infarction and heart failure: a systematic review of clinical trials and meta-analysis of IL-1 inhibition, Cardiovascular Research, Vol: 114, Pages: 1445-1461, ISSN: 1755-3245
Following a myocardial infarction (MI), the immune system helps to repair ischaemic damage and restore tissue integrity, but excessive inflammation has been implicated in adverse cardiac remodelling and development towards heart failure (HF). Pre-clinical studies suggest that timely resolution of inflammation may help prevent HF development and progression. Therapeutic attempts to prevent excessive post-MI inflammation in patients have included pharmacological interventions ranging from broad immunosuppression to immunomodulatory approaches targeting specific cell types or factors with the aim to maintain beneficial aspects of the early post-MI immune response. These include the blockade of early initiators of inflammation including reactive oxygen species and complement, inhibition of mast cell degranulation and leucocyte infiltration, blockade of inflammatory cytokines, and inhibition of adaptive B and T-lymphocytes. Herein, we provide a systematic review on post-MI immunomodulation trials and a meta-analysis of studies targeting the inflammatory cytokine Interleukin-1. Despite an enormous effort into a significant number of clinical trials on a variety of targets, a striking heterogeneity in study population, timing and type of treatment, and highly variable endpoints limits the possibility for meaningful meta-analyses. To conclude, we highlight critical considerations for future studies including (i) the therapeutic window of opportunity, (ii) immunological effects of routine post-MI medication, (iii) stratification of the highly diverse post-MI patient population, (iv) the potential benefits of combining immunomodulatory with regenerative therapies, and at last (v) the potential side effects of immunotherapies.
Watson SA, Duff JD, Bardi IB, et al., 2018, A novel platform to maintain adult cardiac tissue in vitro: myocardial slices and electromechanical stimulation with physiological preload, European-Society-of-Cardiology Congress, Publisher: OXFORD UNIV PRESS, Pages: 1090-1090, ISSN: 0195-668X
Kapnisi M, Mansfield C, Marijon C, et al., 2018, Auxetic cardiac patches with tunable mechanical and conductive properties toward treating myocardial infarction, Advanced Functional Materials, Vol: 28, ISSN: 1616-301X
An auxetic conductive cardiac patch (AuxCP) for the treatment of myocardial infarction (MI) is introduced. The auxetic design gives the patch a negative Poisson's ratio, providing it with the ability to conform to the demanding mechanics of the heart. The conductivity allows the patch to interface with electroresponsive tissues such as the heart. Excimer laser microablation is used to micropattern a re‐entrant honeycomb (bow‐tie) design into a chitosan‐polyaniline composite. It is shown that the bow‐tie design can produce patches with a wide range in mechanical strength and anisotropy, which can be tuned to match native heart tissue. Further, the auxetic patches are conductive and cytocompatible with murine neonatal cardiomyocytes in vitro. Ex vivo studies demonstrate that the auxetic patches have no detrimental effect on the electrophysiology of both healthy and MI rat hearts and conform better to native heart movements than unpatterned patches of the same material. Finally, the AuxCP applied in a rat MI model results in no detrimental effect on cardiac function and negligible fibrotic response after two weeks in vivo. This approach represents a versatile and robust platform for cardiac biomaterial design and could therefore lead to a promising treatment for MI.
Couch LS, Harding SE, 2018, Takotsubo Syndrome: Stress or NO Stress?, JACC: Basic to Translational Science, Vol: 3, Pages: 227-229, ISSN: 2452-302X
Husveth-Toth M, Gara E, Nemes A, et al., 2018, Human pluripotent stem cell-derived endothelial cells are vasoactive in vitro and capable of engineering 3D vascular grafts, 5th Congress of the ESC-Council-on-Basic-Cardiovascular-Science on Frontiers in Cardio Vascular Biology, Publisher: Oxford University Press (OUP), Pages: S111-S111, ISSN: 1755-3245
Foldes G, Lawlor K, Harding SE, et al., 2018, STAT3 mediates differentiation and maintenance of human pluripotent stem-derived endothelial cells, 5th Congress of the ESC-Council-on-Basic-Cardiovascular-Science on Frontiers in Cardio Vascular Biology, Publisher: OXFORD UNIV PRESS, Pages: S39-S39, ISSN: 0008-6363
Background: High plasticity derivatives of human pluripotent stem cells (hPSC) such as embryonic stem cells (hESC) are being intensively developed for their use in endothelial replacement.Methods/Results: In this study, we found that transient addition of Activin A, followed by culture with VEGF165, BMP4 and FGF2 was an effective mechanism to induce differentiation of hESC toward the endothelial lineage. Indeed, human GeneChip microarray analysis revealed that endothelial gene regulatory networks were gradually increased during 12 days of differentiation. Isolated hESC-derived endothelial cells (hESC-EC) expressed mature endothelial-associated genes, including CD31, NRP1, VE-cadherin, Tie2, VWF and ICAM2, reaching levels comparable with human umbilical cord vascular endothelial cells by day 19. We found that a network of CD31+ tubes comprising endothelial precursor cells had formed in culture from 10 days after start of differentiation of hESC. As assessed by automated high content microscopy, the alignment and tube formation of the newly formed CD31+ vascular network were markedly decreased in response to C188-9, a novel small molecule inhibitor of STAT3 transcription factor (tube length: 57%, connected tube area: 22% of those in control, both p<0.001). Human ESC-EC were capable of transdifferentiating into mesenchymal cells in long-term cultures. Endothelial-mesenchymal transition was characterised by gradual loss of endothelial marker expression and increased mesenchymal marker FSP1 expression. We found that inhibition of STAT3 tyr705 phosphorylation by C188-9 resulted in a decreased proliferation of FSP1+ mesenchymal cells (2-fold decrease in Ki67%-positive population, p<0.001), and subsequently reduced number of FSP+ cells (36% reduction, p<0.05). At the same time, C188-9 increased the number of CD31+ hESC-EC by 30% (p=0.05, n=6). Viability remained unchanged in C188-9-treated cells (Topro3 necrosis marker, p=0.32, n=3).Conclusions: These results sugge
Wright PT, Bhogal N, Diakonov I, et al., 2018, Cardiomyocyte membrane structure and cAMP compartmentation produce anatomical variation in β2AR-cAMP responsiveness in murine hearts, Cell Reports, Vol: 23, Pages: 459-469, ISSN: 2211-1247
Cardiomyocytes from the apex but not the base of the heart increase their contractility in response to β2-adrenoceptor (β2AR) stimulation, which may underlie the development of Takotsubo cardiomyopathy. However, both cell types produce comparable cytosolic amounts of the second messenger cAMP. We investigated this discrepancy using nanoscale imaging techniques and found that, structurally, basal cardiomyocytes have more organized membranes (higher T-tubular and caveolar densities). Local membrane microdomain responses measured in isolated basal cardiomyocytes or in whole hearts revealed significantly smaller and more short-lived β2AR/cAMP signals. Inhibition of PDE4, caveolar disruption by removing cholesterol or genetic deletion of Cav3 eliminated differences in local cAMP production and equilibrated the contractile response to β2AR. We conclude that basal cells possess tighter control of cAMP because of a higher degree of signaling microdomain organization. This provides varying levels of nanostructural control for cAMP-mediated functional effects that orchestrate macroscopic, regional physiological differences within the heart.
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