Biology subjects

Tomek, J.

Publications and source records attributed to Tomek, J..

9 recordsLinked to original sources

bioRxiv2025

Targeting Troponin I Phosphorylation for Precision Treatment of Diastolic Dysfunction

BackgroundDefective cardiac relaxation (diastolic dysfunction) is prevalent in heart failure, particularly in the context of diabetes, obesity, hypertension, and ageing, and is associated with increased mortality. Yet there is no available treatment that directly targets this abnormal relaxation. Phosphorylation of troponin I by cAMP-dependent protein kinase A (PKA) promotes relaxation, but global PKA activation has widespread, undesirable effects. Here we investigated whether selectively enhancing troponin I phosphorylation could improve relaxation without engaging the broader PKA signalling network. MethodsTo resolve cAMP signalling at subcellular scales, we combined real-time cAMP measurements with FRET-based genetically encoded reporters targeted to defined nanodomains, alongside biochemical and genetic approaches. Protein-protein interaction surfaces were mapped using peptide-array technology, which guided the design of a disruptor peptide to displace specific interactions. The peptides effects on cardiomyocyte function were assessed in vitro using rodent and human cardiomyocytes with biochemical assays, real-time imaging, and work-loop analysis. In vivo effects were evaluated through echocardiography and haemodynamic measurements. ResultsWe identified a previously unrecognized regulatory mechanism in which the cAMP-hydrolyzing enzyme PDE4D9 interacts with troponin I and restricts local cAMP levels within a nanometre-scale domain, thereby selectively controlling troponin I phosphorylation. We found that PDE4D9-troponin I association is markedly increased in cardiac disease in both rodents and humans. A peptide that displaces PDE4D9 from troponin I in a mouse model of heart failure selectively enhances troponin I phosphorylation, accelerates cardiomyocyte relaxation, and prevents diastolic dysfunction, without compromising systolic contractility. ConclusionsThese findings reveal a highly specific regulatory nanodomain governing troponin I phosphorylation and points to a first-in-class, mechanism-based therapeutic strategy with the potential to address a major unmet contributor to heart failure. Clinical PerspectiveO_ST_ABSWhat Is New?C_ST_ABSO_LIWe identify a previously unrecognized, nanometre-scale regulatory mechanism in which PDE4D9 binds troponin I to locally suppress cAMP, limit troponin I phosphorylation and attenuate relaxation. C_LIO_LIWe demonstrate that this mechanism becomes maladaptive in disease: PDE4D9-troponin I interaction is markedly upregulated in failing hearts in both rodents and humans, which can mechanistically explain the impaired cardiac relaxation. C_LIO_LIWe show that this interaction is druggable: a rationally designed peptide can selectively displace PDE4D9, restore localized cAMP-PKA signalling, and normalize myocardial relaxation without global cAMP perturbation. C_LIO_LIDisplacement of PDE4D9 from TPNI improves relaxation with no impact on the calcium transient amplitude and without adversely affecting systolic output and cardiac reserve. C_LI What Are the Clinical Implications?O_LIDiastolic dysfunction, a major cause of heart failure with no targeted treatments, may be amenable to therapies that modulate cAMP signalling with subcellular precision rather than global pathway modulation. C_LIO_LISelective disruption of PDE4D9-troponin I binding offers a mechanism-based approach with potential to restore diastolic function while minimizing off-target effects typical of GPCR-directed therapies. C_LIO_LIBy enhancing relaxation while preserving systolic function and cardiac reserve, this approach may overcome limitations of myofilament-targeted therapies currently under evaluation for diastolic dysfunction that exhibit negative inotropic effects. C_LIO_LIMore broadly, this work establishes a platform for targeting localized signalling domains, a concept that may extend beyond cardiovascular disease to other pathological conditions in which dysregulated cAMP contributes to dysfunction. C_LI

cell biology↗

bioRxiv2025

T-World: A highly general computational model of a human ventricular myocyte

Cardiovascular disease is the leading cause of death, demanding new tools to improve mechanistic understanding and overcome limitations of stem cell and animal-based research. We introduce T-World, a highly general virtual model of human ventricular cardiomyocyte suitable for multiscale studies. T-World shows comprehensive agreement with human physiology, from electrical activation to contraction, and is the first to replicate all key cellular mechanisms driving life-threatening arrhythmias. Extensively validated on unseen data, it demonstrates strong predictivity across applications and scales. Using T-World we revealed a likely sex-specific arrhythmia risk in females related to restitution properties, identified arrhythmia drivers in type 2 diabetes, and describe unexpected pro-arrhythmic role of NaV1.8 in heart failure. T-World demonstrates strong performance in predicting drug-induced arrhythmia risk and opens new opportunities for predicting and explaining drug efficacy, demonstrated by unpicking effects of mexiletine in Long QT syndrome 2. T-World is available as open-source code and an online app.

cell biology↗

bioRxiv2025

Sex-specific human electromechanical multiscale in-silico models for virtual therapy evaluation

Background and AimWomen are significantly under-represented in cardiovascular research and in the evaluation of treatment safety and efficacy, leading to poorer patient outcomes. Quantification and investigation of sex differences in human electromechanical function and underlying mechanisms is crucial. To address this, we present sex-specific human cellular and biventricular electromechanical models for mechanistic investigations into sex-differences in therapy evaluation through simulations. MethodsProtein genomic expression data from healthy human myocytes were used to calibrate sex-specific models of human cellular electrophysiology, subsequently integrated in biventricular electromechanical models with male and female anatomies. A validation, verification and uncertainty evaluation were implemented at the cellular and biventricular level, including validation using sex-specific datasets from randomised controlled trials for Dofetilide and Verapamil, with known sex-differences. Ionic mechanisms underlying sex-differences in drug response were mechanistically investigated. ResultsSex-specific electromechanical models recapitulate sex-differences from ionic currents to ECG biomarkers including QTc interval (Male: 312ms; Female: 339ms; 9% difference), T-wave amplitude (6-9% difference) and ST-steepness through electrophysiological changes alone. Sex-specific simulations demonstrate both ECG biomarkers and mechanical biomarkers (Female LVEF: 68%, Male LVEF: 50%) within healthy ranges in clinical data for male and female population in the UK Biobank (n= 806, 46% Male). ECGs sex-differences are primarily explained by ionic currents, whereas mechanical sex-differences are driven by anatomical differences, and secondarily through more robust calcium function in females. Under Dofetilide, simulations show exacerbated QT prolongation in women compared to men (54-78% increase in effect), and a T-wave amplitude decrease in males (up to 0.25 mV), consistent with clinical data. This is explained in simulations by lower repolarisation reserve in women (due to low potassium and high calcium currents) than men. Verapamil shows no effect on simulated QT in either sex, and divergent T-wave modulation (increased amplitude in females, decreased in males) consistent with clinical trial data. Simulations identify enhanced contractile reservoir in female compared to male, with lesser decreases to ejection fraction with calcium current block. ConclusionSimulations using novel, sex-specific cellular and biventricular electromechanical models reveal the primary role of ionic currents sex-differences in ECG and drug response, whereas mechanical sex-differences are also underpinned by anatomical differences. Main ContributionsO_LIDevelopment, calibration and validation of sex-specific human ventricular electromechanical, multiscale models. C_LIO_LIAn analysis of clinical randomised trial data in the context of sex-specific effects of multi-channel blockers on the ECG by dosage, where previous analysis focused on pharmacokinetics. C_LIO_LIConsideration of both sex-specific electrophysiology and anatomy explains sex-specific differences on the impact of drugs on ECG and mechanics. C_LIO_LISimulations demonstrate that the reduced repolarisation reserve in females increases the susceptibility to QTc prolongation via potassium channel block compared to males, and more robust calcium dynamics protect against t-wave amplitude reduction and the more severe contractility loss through L-type calcium inhibition observed in males. C_LI

physiology↗

bioRxiv2022

Simultaneous widefield voltage and interferometric dye-free optical mapping quantifies electromechanical waves in human iPSC-cardiomyocytes.

Coupled electro-mechanical waves define hearts function in health and disease. Genetic abnormalities, drug-triggered or acquired pathologies can disrupt and uncouple these waves with potentially lethal consequences. Optical mapping of electrical waves using fluorescent dyes or genetically-encoded sensors in human induced pluripotent stem cell derived cardiomyocytes (iPSC-CMs) offers mechanistic insights into cardiac conduction abnormalities. Interferometric dye-free/label-free wave mapping (without specific sensors) presents an alternative, likely capturing the mechanical aspects of cardiac conduction. Because of its non-invasive nature and spectral flexibility (not restricted to a specific excitation wavelength), it is an attractive chronic imaging tool in iPSC-CMs, as part of all-optical high-throughput platforms. In this study, we developed simultaneous widefield voltage and interferometric dye-free optical imaging methodology that was used: 1) to validate dye-free optical mapping for quantification of cardiac wave properties in human iPSC-CMs; 2) to demonstrate low-cost optical mapping of electromechanical waves in hiPSC-CMs using recent near-infrared (NIR) voltage sensors and orders of magnitude cheaper miniature CMOS cameras; 3) to uncover previously underexplored frequency- and space-varying parameters of cardiac electromechanical waves in hiPSC-CMs. We find similarity in the frequency-dependent responses of electrical (NIR fluorescence imaged) and mechanical (dye-free imaged) waves, with the latter being more sensitive to faster rates and showing steeper restitution and earlier appearance of wave-front tortuosity. During regular pacing, the dye-free imaged conduction velocity and the electrical wave velocity are correlated; both modalities being sensitive to pharmacological uncoupling and both dependent on gap-junctional protein (connexins) determinants of wave propagation. We uncover strong frequency-dependence of the electromechanical delay (EMD) locally and globally in hiPSC-CMs on a rigid substrate. The presented framework and results offer new means to track the functional responses of hiPSC-CM inexpensively and non-invasively for counteracting heart disease and aiding cardiotoxicity testing and drug development.

bioengineering↗

bioRxiv2022

Ionic remodelling following myocardial infarction explains phenotypic variability in ECG and arrhythmic substrate but not in ejection fraction

AimsSudden death after myocardial infarction (MI) is associated with electrophysiological heterogeneities and ionic remodelling, which are reflected as variable phenotypes. Low ejection fraction (EF) is used in risk stratification, but its mechanistic links with the post-MI pro-arrhythmic heterogeneities are unknown. We aim to provide a mechanistic explanation of clinical phenotypes in acute and chronic MI, from ionic remodeling to ECG and EF, using human electromechanical modelling and simulation to augment experimental and clinical investigations. Methods and ResultsA human ventricular electromechanical modelling and simulation framework is constructed and validated with rich experimental and clinical datasets. Abnormalities caused by scar and border zone ionic remodeling are introduced in varying degrees as reported in experimental data obtained in acute and chronic infarction. Simulations enabled reproducing and explaining clinical phenotypes post-MI, from ionic remodelling to ECGs and pressure-volume loops. In acute MI, T-wave inversion and Brugada phenocopy were explained by up to 57 ms of local APD prolongation and activation failure due to the inhibition of potassium, sodium and calcium channels in the border zone. In chronic MI, upright tall T-waves highlight large repolarisation dispersion caused by uneven potassium channel expression in border and remote zones, which promoted ectopic propagation at fast pacing. Post-MI ionic remodelling reduced EF by up to 10% through inhibition of calcium transient amplitude due to weaker calcium currents or SERCA activity, but the EF at resting heart rate was not sensitive to the extent of repolarisation heterogeneity and the risk of repolarisation abnormalities at fast pacing. ConclusionsMulti-scale modelling and simulation coherently integrates experimental and clinical data at subcellular, tissue, and organ scales to unravel electromechanical disease mechanisms in MI. In acute post-MI, ionic remodelling and its effect on refractoriness and propagation failure in the BZ have a strong impact on phenotypic ECG variability, whereas in chronic post-MI, the repolarisation dispersion across the BZ is crucial. T-wave and QT abnormalities are better indicators of repolarisation heterogeneities than EF in post-MI.

bioengineering↗

bioRxiv2020

ToR-ORd-dynCl: an update of the ToR-ORd model of human ventricular cardiomyocyte with dynamic intracellular chloride

Recently, our group published a new model of human ventricular cardiomyocyte named ToR-ORd (Tomek et al., 2019). Its development, calibration, and validation, were performed using a broad range of human experimental data and brought general insights into modelling of ionic channels. Model calibration ensured the reproduction of key physiological cellular features, with independent multiscale validation demonstrating a correct response to channel blocking drugs and pathophysiological remodelling. However, for very long simulations (several hours rather than minutes), the ToR-ORd simulations display a drift in its behaviour, caused by modelling chloride concentrations as constant values. This may be a limitation for simulations considering extremely long protocols, or for studies on model stability. To remedy this, we present here an updated version, termed ToR-ORd-dynCl, with dynamic representation of intracellular chloride. This model behaves very similarly to the original ToR-ORd, but with stable properties over long simulations and only a small increase in model complexity.

physiology↗

bioRxiv2019

A ‘comb’ algorithm for accurate detection of calcium transients and action potentials in regularly activating cardiac preparations

Cardiac imaging and electrophysiological measurements yield vast amounts of data that typically need to be processed automatically. However, the detection and segmentation of calcium transients or action potentials is complicated by signal noise or signal drift, which may cause both false positive and negative segmentation. This article presents a simple but accurate comb algorithm for detection of calcium transients and action potentials in such data where the pattern of activation is regular and its frequency is known. This corresponds either to cases where the cardiac preparation is paced externally, or where the preparation is beating in a stable rhythm. The prior knowledge of the heart rate is leveraged to overcome a broad range of artefacts and complications which arise in experimental data, such as different types of noise, signal drift, or alternans. The algorithm is simple to implement and has only a single free parameter, which is furthermore simple to set. A Matlab/Octave implementation of the comb algorithm is provided.

bioinformatics↗

bioRxiv2019

Optical interrogation of sympathetic neuronal effects on macroscopic cardiac monolayer dynamics

Alterations in autonomic function are known to occur in cardiac conditions including sudden cardiac death. Cardiac stimulation via sympathetic neurons can potentially trigger arrhythmias. Dissecting direct neural-cardiac interactions at the cellular level is technically challenging and understudied due to the lack of experimental model systems and methodologies. Here we demonstrate the utility of optical interrogation of sympathetic neurons and their effects on macroscopic cardiac monolayer dynamics to address research targets such as the effects of adrenergic stimulation via the release of neurotransmitters, the effect of neuronal numbers on cardiac wave behaviour and the applicability of optogenetics in mechanistic in vitro studies. We combine photo-uncaging or optogenetic neural stimulation with imaging of cardiac monolayers to measure electrical activity in an automated fashion, illustrating the power and high throughput capability of such interrogations. The methods described highlight the challenges and benefits of co-cultures as experimental model systems.

biophysics↗

bioRxiv2017

Widespread impact of DNA replication on mutational mechanisms in cancer

DNA replication plays an important role in mutagenesis, yet little is known about how it interacts with other mutagenic processes. Here, we use somatic mutation signatures - each representing a mutagenic process - derived from 3056 patients spanning 19 cancer types to quantify the asymmetry of mutational signatures around replication origins and between early and late replicating regions. We observe that 22 out of 29 mutational signatures are significantly impacted by DNA replication. The distinct associations of different signatures with replication timing and direction around origins shed new light on several mutagenic processes, for example suggesting that oxidative damage to the nucleotide pool substantially contributes to the mutational landscape of esophageal adenocarcinoma. Together, our results indicate an involvement of DNA replication and associated damage repair in most mutagenic processes.

cancer biology↗