Project description:Small molecule cardiac troponin activators could potentially enhance cardiac muscle contraction in the treatment of systolic heart failure. We designed a small molecule, RPI-194, to bind cardiac/slow skeletal muscle troponin (Cardiac muscle and slow skeletal muscle share a common isoform of the troponin C subunit.) Using solution NMR and stopped flow fluorescence spectroscopy, we determined that RPI-194 binds to cardiac troponin with a dissociation constant KD of 6-24 μM, stabilizing the activated complex between troponin C and the switch region of troponin I. The interaction between RPI-194 and troponin C is weak (KD 311 μM) in the absence of the switch region. RPI-194 acts as a calcium sensitizer, shifting the pCa50 of isometric contraction from 6.28 to 6.99 in mouse slow skeletal muscle fibers and from 5.68 to 5.96 in skinned cardiac trabeculae at 100 μM concentration. There is also some cross-reactivity with fast skeletal muscle fibers (pCa50 increases from 6.27 to 6.52). In the slack test performed on the same skinned skeletal muscle fibers, RPI-194 slowed the velocity of unloaded shortening at saturating calcium concentrations, suggesting that it slows the rate of actin-myosin cross-bridge cycling under these conditions. However, RPI-194 had no effect on the ATPase activity of purified actin-myosin. In isolated unloaded mouse cardiomyocytes, RPI-194 markedly decreased the velocity and amplitude of contractions. In contrast, cardiac function was preserved in mouse isolated perfused working hearts. In summary, the novel troponin activator RPI-194 acts as a calcium sensitizer in all striated muscle types. Surprisingly, it also slows the velocity of unloaded contraction, but the cause and significance of this is uncertain at this time. RPI-194 represents a new class of non-specific troponin activator that could potentially be used either to enhance cardiac muscle contractility in the setting of systolic heart failure or to enhance skeletal muscle contraction in neuromuscular disorders.

Project description:Citrus psorosis was reported for the first time in Florida in 1896 and was confirmed as a graft-transmissible disease in 1934. Citrus psorosis virus (CPsV) is the presumed causal agent of this disease. It is considered as a type species of the genus Ophiovirus, within the family Aspiviridae. CPsV genome is a negative single-stranded RNA (-ssRNA) with three segments. It has a coat protein (CP) of 48 kDa and its particles are non-enveloped with naked filamentous nucleocapsids existing as either circular open structures or collapsed pseudo-linear forms. Numerous rapid and sensitive immuno-enzymatic and molecular-based detection methods specific to CPsV are available. CPsV occurrence in key citrus growing regions across the world has been spurred the establishment of the earliest eradication and virus-free budwood programs. Despite these efforts, CPsV remains a common and serious challenge in several countries and causes a range of symptoms depending on the isolate, the cultivar, and the environment. CPsV can be transmitted mechanically to some herbaceous hosts and back to citrus. Although CPsV was confirmed to be seedborne, the seed transmission is not efficient. CPsV natural spread has been increasing based on both CPsV surveys detection and specific CPsV symptoms monitoring. However, trials to ensure its transmission by a soil-inhabiting fungus and one aphid species have been unsuccessful. Psorosis disease control is achieved using CPsV-free buds for new plantations, launching budwood certification and indexing programs, and establishing a quarantine system for the introduction of new varieties. The use of natural resistance to control CPsV is very challenging. Transgenic resistance to at least some CPsV isolates is now possible in at least some sweet orange varieties and constitutes a promising biotechnological alternative to control CPsV. This paper provides an overview of the most remarkable achievements in CPsV research that could improve the understanding of the disease and lead the development of better control strategies.

Project description:Coronavirus disease 2019 (COVID-19), an acute onset pneumonia caused by a novel Betacoronavirus, severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), emerged in the Wuhan City of China in December 2019 and evolved into a global pandemic. To date, there are no proven drugs or vaccines against this virus. Hence, the situation demands an urgent need to explore all potential therapeutic strategies that can be made available to prevent the disease progression and improve patient outcomes. In absence of clinically proven treatment guidelines, several repurposed drugs and investigational agents are currently being evaluated in clinical trials for their probable benefits in the treatment of COVID-19. These include antivirals (remdesivir, lopinavir/ritonavir, umifenovir, and favipiravir), interferon, antimalarials (chloroquine/hydroxychloroquine), antiparasitic drugs (ivermectin and nitazoxanide), biologics (monoclonal antibodies and interleukin receptor antagonist), cellular therapies (mesenchymal stem cells and natural killer cells), convalescent plasma, and cytokine adsorber. Though several observational studies have claimed many of these agents to be effective based on their in vitro activities and extrapolated evidence from SARS and Middle East respiratory syndrome (MERS) epidemics, the currently available data remains inconclusive because of ill-defined patient selection criteria, small sample size, lack of concurrent controls, and use of intermediary outcomes instead of patient-relevant outcomes. Moreover, there is a need to clearly define the patient populations who warrant therapy and also the timing of initiation of treatment. Understanding the disease pathology responsible for the clinical manifestations of COVID-19 is imperative to identify the potential targets for drug development. This review explains the pathophysiology of COVID-19 and summarizes the potential treatment candidates, which can provide guidance in developing effective therapeutic strategies.

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Poorly understood aspects of striated muscle contraction.

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Poorly understood aspects of striated muscle contraction.

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