The Interplay between Mechanoregulation and ROS in Heart Physiology, Disease, and Regeneration.

Cardiovascular diseases are currently the most common cause of death in developed countries. Due to lifestyle and environmental factors, this problem is only expected to increase in the future. Reactive oxygen species (ROS) are a key player in the onset of cardiovascular diseases but also have important functions in healthy cardiac tissue. Here, the interplay between ROS generation and cardiac mechanical forces is shown, and the state of the art and a perspective on future directions are discussed. To this end, an overview of what is currently known regarding ROS and mechanosignaling at a subcellular level is first given. There the role of ROS in mechanosignaling as well as the interplay between both factors in specific organelles is emphasized. The consequences at a larger scale across the population of heart cells are then discussed. Subsequently, the roles of ROS in embryogenesis, pathogenesis, and aging are further discussed, exemplifying some aspects of mechanoregulation. Finally, different models that are currently in use are discussed to study the topics above.

A time-efficient workflow to purify a library of alanine mutants of surface-exposed residues of the domain III of the envelope protein of dengue virus serotype 1.

Dengue complex is formed by four viral serotypes that cause the disease of the same name. Dengue is the arthropod-borne disease with the highest incidence worldwide. The envelope glycoprotein comprises three structural domains. The domain III (DIII) induces neutralizing antibodies and is involved in the interactions with soluble plasma factors from human host. Recombinant DIII proteins have been used as analytical tools for the characterization of virus-host interactions and have been evaluated as sub-unit vaccines. Here, we report a purification procedure of recombinant DIII protein and seventy-four alanine mutants refolding by size exclusion chromatography that allows obtaining highly homogeneous protein preparations and suitable for efficient purification and folding check. Four positions are identified that significantly affect either the protein expression or folding of recombinant DIIIE1, K310, G304, D330 and P332.

Quantum Sensing for Real-Time Monitoring of Drug Efficacy in Synovial Fluid from Arthritis Patients.

Diamond-based T1 relaxometry is a new technique that allows nanoscale magnetic resonance measurements. Here we present its first application in patient samples. More specifically, we demonstrate that relaxometry can determine the free radical load in samples from arthritis patients. We found that we can clearly differentiate between osteoarthritis and rheumatoid arthritis patients in both the synovial fluid itself and cells derived from it. Furthermore, we tested how synovial fluid and its cells respond to piroxicam, a common nonsteroidal anti-inflammatory drug (NSAID). It is known that this drug leads to a reduction in reactive oxygen species production in fibroblast-like synoviocytes (FLS). Here, we investigated the formation of free radicals specifically. While FLS from osteoarthritis patients showed a drastic decrease in the free radical load, cells from rheumatoid arthritis retained a similar radical load after treatment. This offers a possible explanation for why piroxicam is more beneficial for patients with osteoarthritis than those with rheumatoid arthritis.

Intracellular Quantum Sensing of Free-Radical Generation Induced by Acetaminophen (APAP) in the Cytosol, in Mitochondria and the Nucleus of Macrophages.

Acetaminophen overdoses cause cell injury in the liver. It is widely accepted that liver toxicity is initiated by the reactive N-acetyl-para-aminophenol (APAP) metabolite N-acetyl-p-benzoquinone imine (NAPQI), which first depletes glutathione and then irreversibly binds to mitochondrial proteins and nuclear DNA. As a consequence, mitochondrial respiration is inhibited, and DNA strands break. NAPQI also promotes the oxidative stress since glutathione is one of the main free-radical scavengers in the cell. However, so far it is unknown where exactly free radicals are generated. In this study, we used relaxometry, a novel technique that allows nanoscale magnetic resonance imaging detection of free radicals. The method is based on fluorescent nanodiamonds, which change their optical properties based on their magnetic surrounding. To achieve subcellular resolution, these nanodiamonds were targeted to cellular locations, that is, the cytoplasm, mitochondria, and the nucleus. Since relaxometry is sensitive to spin noise from radicals, we were able to measure the radical load in these different organelles. For the first time, we measured APAP-induced free-radical production in an organelle-specific manner, which helps predict and better understand cellular toxicity.