Diuresis Efficacy in Ambulatory Congested Heart Failure Patients: Intra-patient Comparison of Three Diuretic Regimens (DEA-HF).

BACKGROUND: Limited evidence exists regarding efficacy and safety of diuretic regimens in ambulatory, congestion-refractory, chronic heart failure (CHF) patients. OBJECTIVE: To compare the potency and safety of commonly used diuretic regimens in CHF patients. METHODS: A prospective, randomized, open-label, crossover study conducted in NYHA class II-IV CHF patients, treated in an ambulatory day-care unit. Each patient received 3 different diuretic regimens: intravenous (IV) furosemide 250mg; IV furosemide 250mg plus oral metolazone 5mg; and IV furosemide 250mg plus IV acetazolamide 500mg. Treatments were administered once a week, in one of six randomized sequences. The primary endpoint was total sodium excretion, and the secondary was total urinary volume excreted, both measured for 6 hours post-treatment initiation. RESULTS: A total of 42 patients were recruited. Administration of furosemide plus metolazone resulted in the highest weight of sodium excreted, 4691 mg (95% CI: 4153-5229) compared to furosemide alone 3835 mg (95% CI: 3279-4392), P=0.015 and to furosemide plus acetazolamide 3584 mg (95% CI: 3020-4148), P=0.001. Furosemide plus metolazone resulted in 1.84 liters of urine (95% CI: 1.63-2.05), compared to 1.58 liters (95% CI: 1.37-1.8) P=0.039 collected following administration of furosemide plus acetazolamide and 1.71 liters (95% CI 1.49-1.93) following furosemide alone. The incidence of worsening renal function (WRF) was significantly higher when adding metolazone (41%) to furosemide compared to furosemide alone (17%) and to furosemide plus acetazolamide (2.6%), P<0.001. CONCLUSIONS: In ambulatory CHF patients, furosemide plus metolazone resulted in a significantly higher natriuresis compared to IV furosemide alone or furosemide plus acetazolamide.

Synthetic cells with self-activating optogenetic proteins communicate with natural cells.

Development of regulated cellular processes and signaling methods in synthetic cells is essential for their integration with living materials. Light is an attractive tool to achieve this, but the limited penetration depth into tissue of visible light restricts its usability for in-vivo applications. Here, we describe the design and implementation of bioluminescent intercellular and intracellular signaling mechanisms in synthetic cells, dismissing the need for an external light source. First, we engineer light generating SCs with an optimized lipid membrane and internal composition, to maximize luciferase expression levels and enable high-intensity emission. Next, we show these cells' capacity to trigger bioprocesses in natural cells by initiating asexual sporulation of dark-grown mycelial cells of the fungus Trichoderma atroviride. Finally, we demonstrate regulated transcription and membrane recruitment in synthetic cells using bioluminescent intracellular signaling with self-activating fusion proteins. These functionalities pave the way for deploying synthetic cells as embeddable microscale light sources that are capable of controlling engineered processes inside tissues.

Optogenetic Control of Human Induced Pluripotent Stem Cell-Derived Cardiac Tissue Models.

Background Optogenetics, using light-sensitive proteins, emerged as a unique experimental paradigm to modulate cardiac excitability. We aimed to develop high-resolution optogenetic approaches to modulate electrical activity in 2- and 3-dimensional cardiac tissue models derived from human induced pluripotent stem cell (hiPSC)-derived cardiomyocytes. Methods and Results To establish light-controllable cardiac tissue models, opsin-carrying HEK293 cells, expressing the light-sensitive cationic-channel CoChR, were mixed with hiPSC-cardiomyocytes to generate 2-dimensional hiPSC-derived cardiac cell-sheets or 3-dimensional engineered heart tissues. Complex illumination patterns were designed with a high-resolution digital micro-mirror device. Optical mapping and force measurements were used to evaluate the tissues' electromechanical properties. The ability to optogenetically pace and shape the tissue's conduction properties was demonstrated by using single or multiple illumination stimulation sites, complex illumination patterns, or diffuse illumination. This allowed to establish in vitro models for optogenetic-based cardiac resynchronization therapy, where the electrical activation could be synchronized (hiPSC-derived cardiac cell-sheets and engineered heart tissue models) and contractile properties improved (engineered heart tissues). Next, reentrant activity (rotors) was induced in the hiPSC-derived cardiac cell-sheets and engineered heart tissue models through optogenetics programmed- or cross-field stimulations. Diffuse illumination protocols were then used to terminate arrhythmias, demonstrating the potential to study optogenetics cardioversion mechanisms and to identify optimal illumination parameters for arrhythmia termination. Conclusions By combining optogenetics and hiPSC technologies, light-controllable human cardiac tissue models could be established, in which tissue excitability can be modulated in a functional, reversible, and localized manner. This approach may bring a unique value for physiological/pathophysiological studies, for disease modeling, and for developing optogenetic-based cardiac pacing, resynchronization, and defibrillation approaches.

Optogenetic modulation of cardiac action potential properties may prevent arrhythmogenesis in short and long QT syndromes.

Abnormal action potential (AP) properties, as occurs in long or short QT syndromes (LQTS and SQTS, respectively), can cause life-threatening arrhythmias. Optogenetics strategies, utilizing light-sensitive proteins, have emerged as experimental platforms for cardiac pacing, resynchronization, and defibrillation. We tested the hypothesis that similar optogenetic tools can modulate the cardiomyocyte's AP properties, as a potentially novel antiarrhythmic strategy. Healthy control and LQTS/SQTS patient-specific human induced pluripotent stem cell-derived cardiomyocytes (hiPSC-CMs) were transduced to express the light-sensitive cationic channel channelrhodopsin-2 (ChR2) or the anionic-selective opsin, ACR2. Detailed patch-clamp, confocal-microscopy, and optical mapping studies evaluated the ability of spatiotemporally defined optogenetic protocols to modulate AP properties and prevent arrhythmogenesis in the hiPSC-CMs cell/tissue models. Depending on illumination timing, light-induced ChR2 activation induced robust prolongation or mild shortening of AP duration (APD), while ACR2 activation allowed effective APD shortening. Fine-tuning these approaches allowed for the normalization of pathological AP properties and suppression of arrhythmogenicity in the LQTS/SQTS hiPSC-CM cellular models. We next established a SQTS-hiPSC-CMs-based tissue model of reentrant-arrhythmias using optogenetic cross-field stimulation. An APD-modulating optogenetic protocol was then designed to dynamically prolong APD of the propagating wavefront, completely preventing arrhythmogenesis in this model. This work highlights the potential of optogenetics in studying repolarization abnormalities and in developing novel antiarrhythmic therapies.

Literature Review and Knowledge Distribution During an Outbreak: A Methodology for Managing Infodemics.

PROBLEM: The COVID-19 pandemic has challenged health care systems in an unprecedented way by imposing new demands on health care resources and scientific knowledge. There has also been an exceedingly fast accumulation of new information on this novel virus. As the traditional peer-review process takes time, there is currently a significant gap between the ability to generate new data and the ability to critically evaluate them. This problem of an excess of mixed-quality data, or infodemic, is echoing throughout the scientific community. APPROACH: The authors aimed to help their colleagues at the Rambam Medical Center, Haifa, Israel, manage the COVID-19 infodemic with a methodologic solution: establishing an in-house mechanism for continuous literature review and knowledge distribution (March-April 2020). Their methodology included the following building blocks: a dedicated literature review team, artificial intelligence-based research algorithms, brief written updates in a graphical format, large-scale webinars and online meetings, and a feedback loop. OUTCOMES: During the first month (April 2020), the project produced 21 graphical updates. After consideration of feedback from colleagues and final editing, 13 graphical updates were uploaded to the center's website; of these, 31% addressed the clinical presentation of the disease and 38% referred to specific treatments. This methodology as well as the graphical updates it generated were adopted by the Israeli Ministry of Health and distributed in a hospital preparation kit. NEXT STEPS: The authors believe they have established a novel methodology that can assist in the battle against COVID-19 by making high-quality scientific data more accessible to clinicians. In the future, they expect this methodology to create a favorable uniform standard for evidence-guided health care during infodemics. Further evolution of the methodology may include evaluation of its long-term sustainability and impact on the day-to-day clinical practice and self-confidence of clinicians who treat COVID-19 patients.

Generating ring-shaped engineered heart tissues from ventricular and atrial human pluripotent stem cell-derived cardiomyocytes.

The functions of the heart are achieved through coordination of different cardiac cell subtypes (e.g., ventricular, atrial, conduction-tissue cardiomyocytes). Human pluripotent stem cell-derived cardiomyocytes (hPSC-CMs) offer unique opportunities for cardiac research. Traditional studies using these cells focused on single-cells and utilized mixed cell populations. Our goal was to develop clinically-relevant engineered heart tissues (EHTs) comprised of chamber-specific hPSC-CMs. Here we show that such EHTs can be generated by directing hPSCs to differentiate into ventricular or atrial cardiomyocytes, and then embedding these cardiomyocytes in a collagen-hydrogel to create chamber-specific, ring-shaped, EHTs. The chamber-specific EHTs display distinct atrial versus ventricular phenotypes as revealed by immunostaining, gene-expression, optical assessment of action-potentials and conduction velocity, pharmacology, and mechanical force measurements. We also establish an atrial EHT-based arrhythmia model and confirm its usefulness by applying relevant pharmacological interventions. Thus, our chamber-specific EHT models can be used for cardiac disease modeling, pathophysiological studies and drug testing.

Human Induced Pluripotent Stem Cell-Derived Cardiac Cell Sheets Expressing Genetically Encoded Voltage Indicator for Pharmacological and Arrhythmia Studies.

Fulfilling the potential of human induced pluripotent stem cell (hiPSC)-derived cardiomyocytes for studying conduction and arrhythmogenesis requires development of multicellular models and methods for long-term repeated tissue phenotyping. We generated confluent hiPSC-derived cardiac cell sheets (hiPSC-CCSs), expressing the genetically encoded voltage indicator ArcLight. ArcLight-based optical mapping allowed generation of activation and action-potential duration (APD) maps, which were validated by mapping the same hiPSC-CCSs with the voltage-sensitive dye, Di-4-ANBDQBS. ArcLight mapping allowed long-term assessment of electrical remodeling in the hiPSC-CCSs and evaluation of drug-induced conduction slowing (carbenoxolone, lidocaine, and quinidine) and APD prolongation (quinidine and dofetilide). The latter studies also enabled step-by-step depiction of drug-induced arrhythmogenesis ("torsades de pointes in the culture dish") and its prevention by MgSO4 and rapid pacing. Phase-mapping analysis allowed biophysical characterization of spiral waves induced in the hiPSC-CCSs and their termination by electrical cardioversion and overdrive pacing. In conclusion, ArcLight mapping of hiPSC-CCSs provides a powerful tool for drug testing and arrhythmia investigation.

Cardiac optogenetics: the next frontier.

The emerging technology of optogenetics uses optical and genetic means to monitor and modulate the electrophysiological properties of excitable tissues. While transforming the field of neuroscience, the technology has recently gained popularity also in the cardiac arena. Here, we describe the basic principles of optogenetics, the available and evolving optogenetic tools, and the unique potential of this technology for basic and translational cardiac electrophysiology. Specifically, we discuss the ability to control (augment or suppress) the cardiac tissue's excitable properties using optogenetic actuators (microbial opsins), which are light-gated ion channels and pumps that can cause light-triggered membrane depolarization or hyperpolarization. We then focus on the potential clinical implications of this technology for the treatment of cardiac arrhythmias by describing recent efforts for developing optogenetic-based cardiac pacing, resynchronization, and defibrillation experimental strategies. Finally, the significant obstacles and challenges that need to be overcome before any future clinical translation can be expected are discussed.