Optogenetics in striated muscle: defibrillation of the heart and direct stimulation of skeletal muscles with light

Optogenetic depolarization of cells using the non-selective cation channel Channelrhodopsin-2 (ChR2) enables precise control over the membrane potential of cells within a specific area of intact organs. Furthermore, the selective overexpression of light-gated proteins allows cell type-specific and pain-free stimulation which could be of great benefit for future scientific and therapeutic approaches. In my thesis, I explored two potential applications of optogenetic methods in striated muscle: optogenetic defibrillation to terminate ventricular arrhythmia in intact mouse hearts and direct optogenetic stimulation of skeletal muscles. These new approaches could lead in the future to the development of optogenetic defibrillators and laryngeal pacemakers. Most experiments were performed with explanted hearts, isolated skeletal fibers and muscles or larynges from transgenic ChR2 expressing mice. To add translational perspectives, we also explored optogenetic defibrillation and intralaryngeal muscles stimulation after ChR2 gene transfer to wild type mice using adeno-associated virus (AAV). Optogenetic defibrillation by epicardial illumination was highly efficient in terminating ventricular arrhythmia in transgenic hearts and the success rate of optogenetic defibrillation was depending on the pulse duration, the size of illumination and the light intensity. Importantly, we were also able to terminate ventricular arrhythmia in non-transgenic hearts even one year after AAV mediated gene transfer. The potential applicability of optogenetic defibrillation in the human heart was assessed in experimentally-calibrated computer simulations of a patient’s heart with infarct-related ventricular tachycardia. Because optogenetic stimulation would be in principle pain-free in patients, the proof for its feasibility could lay the foundation for the development of a new treatment option for patients at high risk for ventricular arrhythmia. Direct optogenetic stimulation of skeletal muscle was first proven in isolated Flexor digitorum brevis fibers and in intact soleus muscles, which could both be stimulated using brief light pulses. The force of light-induced single twitches could be precisely controlled by varying the pulse duration and light intensity. Optogenetic stimulation was most efficient with 10 ms long pulses at a repetition rate of 40 Hz reaching ~84% of the maximum force generated by electrical stimulation with 100 Hz. Recurrent nerve paralysis is a severe complication of neck surgery, malignant processes or central neurological diseases and results in a fixed paramedian position of the vocal cords as well as life-threatening dyspnea in the case of bilateral paralysis. Current treatment options consist only of destructive surgery. Unfortunately the use of local electrical stimulation to restore laryngeal function faces severe technical limitations. Therefore I sought to explore direct optogenetic stimulation of intralaryngeal muscles in explanted larynges from ChR2 transgenic mice. Specific illumination of the individual intralaryngeal muscle groups led to an opening or closing of the vocal cords depending on the site of illumination. This proves the sufficient spatial resolution of light for selective stimulation of the intralaryngeal muscles groups. In addition, we were able to induce opening of the vocal cords in wild type mice after AAV-based gene transfer of ChR2 with light. Thus optogenetic stimulation could become a new treatment option for patients suffering from bilateral laryngeal paralysis. In conclusion, optogenetic stimulation can overcome the severe limitations of electrical stimulation of the heart and skeletal muscles. The new technologies, I have developed and characterized in this thesis, allow for the design of completely new stimulation patterns to address open questions in muscle physiology. Furthermore, optogenetic stimulation of striated muscles could become a new treatment option for patients enabling selective and pain-free stimulation with few side effects

Effets de divers stimuli sur les caractéristiques des cardiomyocytes en culture dans le but de définir les conditions optimisées pour la fabrication de tissu cardiaque de remplacement

Encore en 2015, un grand nombre d’individus décèdent de pathologies du rythme cardiaque non contrôlées ou d’un manque de disponibilité de donneurs d’organes compatibles. Le génie tissulaire en créant, réparant ou améliorant la fonction des tissus est une option prometteuse afin de diminuer la mortalité associée à ces pathologies. L’objectif global de mon projet de recherche était de développer des outils et d’étudier l’impact fonctionnel des différents stimuli (mécanique et électrique) de l’environnement cardiaque dans le but de définir des conditions optimisées de culture pour la fabrication de tissu de remplacement par génie tissulaire. Cette thèse présente le développement d’un bioréacteur; un système qui optimise les conditions pour la culture cellulaire. L’efficacité du bioréacteur est validée par des expériences de culture cellulaire qui se concentrent sur la prolifération cellulaire, l’organisation cellulaire, l’expression génique et protéique de même que sur l’activité contractile spontanée. En premier lieu, nos résultats montrent, bien que la fréquence de contraction moyenne mesurée reste inchangée, une augmentation significative du nombre de cas de réentrées pour les cultures sur verre comparativement aux cultures sur Polydimethylsiloxane. Une augmentation de l’instabilité spatiotemporelle a été démontrée lorsque les cardiomyocytes étaient déposés sur un support de Polydimethylsiloxane et cette dernière corrèle avec une diminution non-significative de l’ARNm de la connexine-43 et une augmentation significative de l’ARNm pour CaV3.1 et HCN2. La culture sur Polydimethylsiloxane est également associée avec une plus forte réponse à l’isoprotérénol (β-adrénergique) et à l’acétylcholine (parasympathique). En second lieu, nous présentons les résultats du développement de notre bioréacteur en mettant l’emphase sur les caractéristiques (composantes accessibles, étirement uniaxial, électrode de carbone, stimulation biphasique) tout en validant notre approche pour optimiser les conditions de culture et améliorer la rentabilité des étapes de production du tissu de remplacement. Pour finir, nous partageons une nouvelle approche d’évaluation des caractéristiques contractiles de cellules cardiaques en culture. Nous avons développé des algorithmes qui utilisent les données de vidéomicroscopie pour valider l’impact de stimuli, évaluer l’hétérogénéité du signal enregistré et détecter des conditions favorables au développement d’arythmies.In 2015, there are still a large number of people who die due to diseases of uncontrolled heart rhythm or due to lack of availability of compatible donor organs. Tissue engineering aim to create, repair or improve the function by different techniques. Tissue engineering is a viable option to reduce the mortality associated with many heart conditions. The overall goal of my PhD research was to study the functional impact of different stimuli in cardiac environment (mechanical and electrical stimulation) on cardiac cell cultures. This, in order to define optimized culture conditions for the production of replacement tissue using tissue engineering. This thesis presents the stages of creation and development of a bioreactor; a system that permits the culture of cardiac cells by integrating various stimuli. The optimization of culture conditions by using the bioreactor was confirmed by cell culture experiments that focus on cell proliferation, cell organization, gene and protein expression as well as on spontaneous activity. In the first place, our results show that although mean frequency of spontaneous activity remained unaltered, incidence of reentrant activity was significantly higher in samples cultured on glass compared to PDMS substrates. Higher spatial and temporal instability of the spontaneous rate activation was found when cardiomyocytes were cultured on PDMS, and correlated with decreased connexin-43 (unsignificant) and a significant increased CaV3.1 and HCN2 mRNA levels. Compared to cultures on glass, cultures on PDMS were associated with the strongest response to isoproterenol (β-adrenergic) and acetylcholine (parasympathetic). Secondly, we present the design of our bioreactor with an emphasis on its characteristics and by putting in perspective the relevance of our approach to optimize culture conditions and to improve profitability culture experiences and production stages of replacement heart tissue. Finally, a new approach is proposed to evaluate the characteristics of the contractile cells in culture which allows to validate the functional impact of stimuli, evaluate the heterogeneity in the beating behavior of the cells and to detect localized abnormal activity that could favour arrhythmia

Applications of nanogenerators for biomedical engineering and healthcare systems

The dream of human beings for long living has stimulated the rapid development of biomedical and healthcare equipment. However, conventional biomedical and healthcare devices have shortcomings such as short service life, large equipment size, and high potential safety hazards. Indeed, the power supply for conventional implantable device remains predominantly batteries. The emerging nanogenerators, which harvest micro/nanomechanical energy and thermal energy from human beings and convert into electrical energy, provide an ideal solution for self‐powering of biomedical devices. The combination of nanogenerators and biomedicine has been accelerating the development of self‐powered biomedical equipment. This article first introduces the operating principle of nanogenerators and then reviews the progress of nanogenerators in biomedical applications, including power supply, smart sensing, and effective treatment. Besides, the microbial disinfection and biodegradation performances of nanogenerators have been updated. Next, the protection devices have been discussed such as face mask with air filtering function together with real‐time monitoring of human health from the respiration and heat emission. Besides, the nanogenerator devices have been categorized by the types of mechanical energy from human beings, such as the body movement, tissue and organ activities, energy from chemical reactions, and gravitational potential energy. Eventually, the challenges and future opportunities in the applications of nanogenerators are delivered in the conclusive remarks. The combination of nanogenerator and biomedicine have been accelerating the development of self‐powered biomedical devices, which show a bright future in biomedicine and healthcare such as smart sensing, and therapy

Diagnostic Challenges in Sports Cardiology

The foundations of sports cardiology include promoting physical activity and providing a safe environment for training and competition for all athletes at all levels, from professional to recreational. To combine these two aims, reliable tools to perform preparticipation screenings are needed. Moreover, those at high risk of potentially life-threatening events should be advised to limit their training load, while others should be reassured that there is no exercise-related cardiovascular risk. We are currently witnessing the advent of new portable devices for remote and mobile heart monitoring and several new and promising biochemical markers, which can support athletes’ diagnostic processes. In this Special Issue of the Diagnostics journal entitled “Diagnostic Challenges in Sports Cardiology”, we present a series of 13 manuscripts, including eight original works, three reviews, and two case reports, which give a glimpse into the current research topics in the area of sports cardiology

Pacing with restoration of respiratory sinus arrhythmia improved cardiac contractility and the left ventricular output: a translational study

Introduction: Respiratory sinus arrhythmia (RSA) is a prognostic value for patients with heart failure and is defined as a beat-to-beat variation of the timing between the heart beats. Patients with heart failure or patients with permanent cardiac pacing might benefit from restoration of RSA. The aim of this translational, proof-of-principle study was to evaluate the effect of pacing with or without restored RSAon parameters of LV cardiac contractility and the cardiac output

Applications of nanogenerators for biomedical engineering and healthcare systems

The dream of human beings for long living has stimulated the rapid development of biomedical and healthcare equipment. However, conventional biomedical and healthcare devices have shortcomings such as short service life, large equipment size, and high potential safety hazards. Indeed, the power supply for conventional implantable device remains predominantly batteries. The emerging nanogenerators, which harvest micro/nanomechanical energy and thermal energy from human beings and convert into electrical energy, provide an ideal solution for self-powering of biomedical devices. The combination of nanogenerators and biomedicine has been accelerating the development of self-powered biomedical equipment. This article first introduces the operating principle of nanogenerators and then reviews the progress of nanogenerators in biomedical applications, including power supply, smart sensing, and effective treatment. Besides, the microbial disinfection and biodegradation performances of nanogenerators have been updated. Next, the protection devices have been discussed such as face mask with air filtering function together with real-time monitoring of human health from the respiration and heat emission. Besides, the nanogenerator devices have been categorized by the types of mechanical energy from human beings, such as the body movement, tissue and organ activities, energy from chemical reactions, and gravitational potential energy. Eventually, the challenges and future opportunities in the applications of nanogenerators are delivered in the conclusive remarks.Web of Science4

Clinical and structural risk factors predicting atrial fibrillation

Atrial fibrillation (AF) is associated with a high morbidity and mortality. Early identification of patients with AF may reduce morbidity and mortality. Current models predicting AF have limitations and focus on mainly clinical variables which are not always apparent in AF patients. Models focusing on pathophysiological mechanisms such as blood based biomarkers and ECG markers may be more accurate in identifying patients with AF. This study is based on the Birmingham and Black Country Atrial Fibrillation Registry (BBC-AF Registry) which recruited a cohort of 800 patients with and without AF. Blood based biomarkers and ECG markers were compared between the two groups of patients. The blood based biomarker analysis using a novel proteomics chip technique demonstrated that BNP and a novel biomarker, fibroblast growth factor 23 (FGF-23) were increased in AF patients and were also independently predictive of AF. In the ECG analysis, QT interval was increased in AF patients and independently predicted AF. A combined model using blood based biomarkers, ECG markers and clinical variables demonstrated that a simple model consisting of simple clinical variables, QT interval, BNP and FGF-23 had a good ability to predict AF and performed better than contemporary AF prediction models in the current literature