Mechanotransduction by proprioceptive sensory organs is poorly understood. Evidence was recently shown that muscle spindle and hair follicle primary afferents (lanceolates) constantly release glutamate from synaptic-like vesicles (SLVs) within the terminals. The secreted glutamate activates a highly unusual metabotropic glutamate receptor (mGluR) to modulate the firing rate (spindles) and SLV recycling (lanceolates). This receptor has yet to be isolated and sequenced. To further investigate this receptor's pharmacology, ligands selective for classical mGluRs have been recently characterised for their ability to alter stretch-evoked spindle firing and SLV endocytosis in these different endings. Here, it is described how the results of these screens facilitated the development of novel compounds to be used in the process of isolating and sequencing of this non-canonical mGluR. This study shows how the compounds were tested for their ability to alter stretch-evoked afferent firing in muscle spindles and SLV endocytosis in the lanceolate endings of hair follicles to ensure they maintained their ability to bind to the receptor. For the development of novel compounds, kainate was chosen as the parent ligand due to its potency and ease of chemical modification. Novel kainate derivatives were then synthesised and tested to find potent analogues suitable for 'click-chemistry', an established technique for relatively quick, cheap, stereospecific and high-yield chemical modifications (Angewandte Chemie (International ed. in English), 40, 2001, pp2004). Of the novel kainate analogues developed, unfortunately ZCZ49 and ZCZ50 lost the ability to produce a significant change in spindle stretch-evoked firing. However, ZCZ90 was as potent as kainate, increasing firing by a similar margin at 1 μm (n = 8; P < 0.001). The addition of either a biotin or a fluorescein side group to ZCZ90, using the click-chemistry technique, did not affect the potency and hence these compounds will be used in further studies of the receptor. As well as the development of these compounds, the study found not only many similarities, but also some key differences between the two types of primary mechanosensory endings investigated. These differences must be taken into account in further study. However, they also present an intriguing opportunity for these receptors to be targeted selectively to modulate ending sensitivity as treatments for muscle spasm in multiple sclerosis and spinal cord injury, and possibly even baroreceptor firing to treat hypertension.
Senescence is a highly organized and well-regulated process. As much as 75% of total cellular nitrogen may be located in mesophyll chloroplasts of C(3)-plants. Proteolysis of chloroplast proteins begins in an early phase of senescence and the liberated amino acids can be exported to growing parts of the plant (e.g. maturing fruits). Rubisco and other stromal enzymes can be degraded in isolated chloroplasts, implying the involvement of plastidial peptide hydrolases. Whether or not ATP is required and if stromal proteins are modified (e.g. by reactive oxygen species) prior to their degradation are questions still under debate. Several proteins, in particular cysteine proteases, have been demonstrated to be specifically expressed during senescence. Their contribution to the general degradation of chloroplast proteins is unclear. The accumulation in intact cells of peptide fragments and inhibitor studies suggest that multiple degradation pathways may exist for stromal proteins and that vacuolar endopeptidases might also be involved under certain conditions. The breakdown of chlorophyll-binding proteins associated with the thylakoid membrane is less well investigated. The degradation of these proteins requires the simultaneous catabolism of chlorophylls. The breakdown of chlorophylls has been elucidated during the last decade. Interestingly, nitrogen present in chlorophyll is not exported from senescencing leaves, but remains within the cells in the form of linear tetrapyrrolic catabolites that accumulate in the vacuole. The degradation pathways for chlorophylls and chloroplast proteins are partially interconnected.
Background . A new endemic disease has spread across Wuhan City, China, in December 2019. Within few weeks, the World Health Organization (WHO) announced a novel coronavirus designated as coronavirus disease 2019 (COVID‐19). In late January 2020, WHO declared the outbreak of a “public‐health emergency of international concern” due to the rapid and increasing spread of the disease worldwide. Currently, there is no vaccine or approved treatment for this emerging infection; thus, the objective of this study is to design a multiepitope peptide vaccine against COVID‐19 using an immunoinformatics approach. Method . Several techniques facilitating the combination of the immunoinformatics approach and comparative genomic approach were used in order to determine the potential peptides for designing the T‐cell epitope‐based peptide vaccine using the envelope protein of 2019‐nCoV as a target. Results . Extensive mutations, insertion, and deletion were discovered with comparative sequencing in the COVID‐19 strain. Additionally, ten peptides binding to MHC class I and MHC class II were found to be promising candidates for vaccine design with adequate world population coverage of 88.5% and 99.99%, respectively. Conclusion . The T‐cell epitope‐based peptide vaccine was designed for COVID‐19 using the envelope protein as an immunogenic target. Nevertheless, the proposed vaccine rapidly needs to be validated clinically in order to ensure its safety and immunogenic profile to help stop this epidemic before it leads to devastating global outbreaks.
Summary: Global quantification of total RNA is used to investigate steady state levels of gene expression. However, being able to differentiate pre-existing RNA (that has been synthesized prior to a defined point in time) and newly transcribed RNA can provide invaluable information e.g. to estimate RNA half-lives or identify fast and complex regulatory processes. Recently, new techniques based on metabolic labeling and RNA-seq have emerged that allow to quantify new and old RNA: Nucleoside analogs are incorporated into newly transcribed RNA and are made detectable as point mutations in mapped reads. However, relatively infrequent incorporation events and significant sequencing error rates make the differentiation between old and new RNA a highly challenging task. We developed a statistical approach termed GRAND-SLAM that, for the first time, allows to estimate the proportion of old and new RNA in such an experiment. Uncertainty in the estimates is quantified in a Bayesian framework. Simulation experiments show our approach to be unbiased and highly accurate. Furthermore, we analyze how uncertainty in the proportion translates into uncertainty in estimating RNA half-lives and give guidelines for planning experiments. Finally, we demonstrate that our estimates of RNA half-lives compare favorably to other experimental approaches and that biological processes affecting RNA half-lives can be investigated with greater power than offered by any other method. GRAND-SLAM is freely available for non-commercial use at http://software.erhard-lab.de; R scripts to generate all figures are available at zenodo (doi: 10.5281/zenodo.1162340).
complex that upon activation by calcium binding/thrombin cleavage covalently cross-links preformed fibrin clots protecting them from premature fibrinolysis. Our study characterizes the recently disclosed three calcium binding sites of FXIII-A concerning evolution, mutual crosstalk, thermodynamic activation profile, substrate binding, and interaction with other similarly charged ions. We demonstrate unique structural aspects within FXIII-A calcium binding sites that give rise to functional differences making FXIII unique from other transglutaminases. The first calcium binding site showed an antagonistic relationship towards the other two. The thermodynamic profile of calcium/thrombin-induced FXIII-A activation explains the role of bulk solvent in transitioning its zymogenic dimeric form to an activated monomeric form. We also explain the indirect effect of solvent ion concentration on FXIII-A activation. Our study suggests FXIII-A calcium binding sites could be putative pharmacologically targetable regions.
This trial was conducted to determine the dose-limiting toxicities (DLTs) and maximum tolerated dose (MTD) of the first in class NEDD8-activating enzyme (NAE) inhibitor, pevonedistat, and to investigate pevonedistat pharmacokinetics and pharmacodynamics in patients with acute myeloid leukaemia (AML) and myelodysplastic syndromes (MDS). Pevonedistat was administered via a 60-min intravenous infusion on days 1, 3 and 5 (schedule A, n = 27), or days 1, 4, 8 and 11 (schedule B, n = 26) every 21-days. Dose escalation proceeded using a standard '3 + 3' design. Responses were assessed according to published guidelines. The MTD for schedules A and B were 59 and 83 mg/m(2) , respectively. On schedule A, hepatotoxicity was dose limiting. Multi-organ failure (MOF) was dose limiting on schedule B. The overall complete (CR) and partial (PR) response rate in patients treated at or below the MTD was 17% (4/23, 2 CRs, 2 PRs) for schedule A and 10% (2/19, 2 PRs) for schedule B. Pevonedistat plasma concentrations peaked after infusion followed by elimination in a biphasic pattern. Pharmacodynamic studies of biological correlates of NAE inhibition demonstrated target-specific activity of pevonedistat. In conclusion, administration of the first-in-class agent, pevonedistat, was feasible in patients with MDS and AML and modest clinical activity was observed.
Extracellular vesicles (EVs) are released by many cell types and distributed within various biofluids. EVs have a lipid membrane-confined structure that allows for carrying unique molecular information originating from their parent cells. The species and quantity of EV cargo molecules, including nucleic acids, proteins, lipids, and metabolites, may vary largely owing to their parent cell types and the pathophysiologic status. Such heterogeneity in EV populations provides immense challenges to researchers, yet allows for the possibility to prognosticate the pathogenesis of a particular tissue from unique molecular signatures of dispersing EVs within biofluids. However, the inherent nature of EV's small size requires advanced methods for EV purification and evaluation from the complex biofluid. Recently, the interdisciplinary significance of EV research has attracted growing interests, and the EV analytical platforms for their diagnostic prospect have markedly progressed. This review summarizes the recent advances in these EV detection techniques and methods with the intention of translating an EV-based liquid biopsy into clinical practice. This article aims to present an overview of current EV assessment techniques, with a focus on their progress and limitations, as well as an outlook on the clinical translation of an EV-based liquid biopsy that may augment current paradigms for the diagnosis, prognosis, and monitoring the response to therapy in a variety of disease settings.
The phenothiazinium salt methylene blue [3,7-bis(dimethylamino)phenothiazinium chloride] is the oldest known synthetic antimalarial drug, its clinical efficacy having been reported in 1891. The role of methylene blue in the evolution of the modern antimalarial armoury is often unappreciated, yet it can be linked directly to standard drugs such as chloroquine and its congeners. Also, in the face of increasing plasmodial resistance to modern antimalarials, phenothiazinium derivatives have again featured as lead compounds in drug research. The precise mode of action of methylene blue and its commercial analogues against Plasmodium spp. remains a cause for conjecture, having been variously described as nucleic acid intercalation, food vacuole basification, parasite redox cycle interference and haem polymerization inhibition. That the activity of the series may be due to more than one route - i.e. a multifactorial activity - underlines the utility of these compounds in antimalarial research either as single drugs or as adjuvants (partners in a drug combination), particularly in the face of resistant parasitic strains.
Pharmacologically active compounds with preferential cytotoxic activity for senescent cells, known as senolytics, can ameliorate or even revert pathological manifestations of senescence in numerous preclinical mouse disease models, including cancer models. However, translation of senolytic therapies to human disease is hampered by their suboptimal specificity for senescent cells and important toxicities that narrow their therapeutic windows. We have previously shown that the high levels of senescence-associated lysosomal β-galactosidase (SA-β-gal) found within senescent cells can be exploited to specifically release tracers and cytotoxic cargoes from galactose-encapsulated nanoparticles within these cells. Here, we show that galacto-conjugation of the BCL-2 family inhibitor Navitoclax results in a potent senolytic prodrug (Nav-Gal), that can be preferentially activated by SA-β-gal activity in a wide range of cell types. Nav-Gal selectively induces senescent cell apoptosis and has a higher senolytic index than Navitoclax (through reduced activation in nonsenescent cells). Nav-Gal enhances the cytotoxicity of standard senescence-inducing chemotherapy (cisplatin) in human A549 lung cancer cells. Concomitant treatment with cisplatin and Nav-Gal in vivo results in the eradication of senescent lung cancer cells and significantly reduces tumour growth. Importantly, galacto-conjugation reduces Navitoclax-induced platelet apoptosis in human and murine blood samples treated ex vivo, and thrombocytopenia at therapeutically effective concentrations in murine lung cancer models. Taken together, we provide a potentially versatile strategy for generating effective senolytic prodrugs with reduced toxicities.
Next-generation sequencing (NGS) is the method of choice when large numbers of sequences have to be obtained. While the technique is widely applied, varying error rates have been observed. We analysed millions of reads obtained after sequencing of one single sequence on an Illumina sequencer. According to our analysis, the index-PCR for sample preparation has no effect on the observed error rate, even though PCR is traditionally seen as one of the major contributors to enhanced error rates in NGS. In addition, we observed very persistent pre-phasing effects although the base calling software corrects for these. Removal of shortened sequences abolished these effects and allowed analysis of the actual mutations. The average error rate determined was 0.24 ± 0.06% per base and the percentage of mutated sequences was found to be 6.4 ± 1.24%. Constant regions at the 5'- and 3'-end, e.g., primer binding sites used in in vitro selection procedures seem to have no effect on mutation rates and re-sequencing of samples obtains very reproducible results. As phasing effects and other sequencing problems vary between equipment and individual setups, we recommend evaluation of error rates and types to all NGS-users to improve the quality and analysis of NGS data.
Hydrogen sulfide (H₂S) has traditionally been thought of as a phytotoxin, having deleterious effects on the plant growth and survival. It is now recognized that plants have enzymes which generate H₂S, cysteine desulfhydrase, and remove it, O-acetylserine lyase. Therefore, it has been suggested that H₂S is considered as a signalling molecule, alongside small reactive compounds such as hydrogen peroxide (H₂O₂) and nitric oxide (NO). Exposure of plants to low of H₂S, for example from H₂S donors, is revealing that many physiological effects are seen. H₂S seems to have effects on stomatal apertures. Intracellular effects include increases in glutathione levels, alterations of enzyme activities and influences on NO and H₂O₂ metabolism. Work in animals has shown that H₂S may have direct effects on thiol modifications of cysteine groups, work that will no doubt inform future studies in plants. It appears therefore, that instead of thinking of H₂S as a phytotoxin, it needs to be considered as a signalling molecule that interacts with reactive oxygen species and NO metabolism, as well as having direct effects on the activity of proteins. The future may see H₂S being used to modulate plant physiology in the field or to protect crops from postharvest spoilage.
Many cellular signaling processes are initiated by dimerization or oligomerization of membrane proteins. However, since the spatial scale of these interactions is below the diffraction limit of the light microscope, the dynamics of these interactions have been difficult to study on living cells. We have developed a novel high-speed hyperspectral microscope (HSM) to perform single particle tracking of up to 8 spectrally distinct species of quantum dots (QDs) at 27 frames per second. The distinct emission spectra of the QDs allows localization with ∼10 nm precision even when the probes are clustered at spatial scales below the diffraction limit. The capabilities of the HSM are demonstrated here by application of multi-color single particle tracking to observe membrane protein behavior, including: 1) dynamic formation and dissociation of Epidermal Growth Factor Receptor dimers; 2) resolving antigen induced aggregation of the high affinity IgE receptor, FcεR1; 3) four color QD tracking while simultaneously visualizing GFP-actin; and 4) high-density tracking for fast diffusion mapping.
H-NMR). The colloidal stability of GNR when exposed to skin, and their preferential accumulation into excised human skin layers were investigated. Confocal laser scanning microscopy, transmission electron microscope (TEM) and inductively coupled plasma-optical emission spectroscopy (ICP-OES) were utilized to track the penetration of GNR into different skin layers. The results demonstrated that cholesterol-PEG coated GNR were preferentially loaded up in the upper layers of skin (stratum corneum), while phospholipid-PEG coated counterparts were drastically deposited in skin dermis. Neutral methoxy-PEG-coated GNR were distributed in both SC and dermis skin layers, while charged GNR (anionic-carboxylic acid-PEG-GNR and cationic-amine-PEG-GNR) revealed a minimal accumulation into skin. DSPE-PEG-GNR and Chol-PEG-GNR demonstrated antibacterial activities against Staphylococcus aureus (S aureus) at MIC values of 0.011 nM and 0.75 nM, respectively. Photothermal treatment for S. aureus at sub-MIC concentrations resulted in a significant bactericidal effect when using Chol-PEG-GNR but not DSPE-PEG-GNR. Gold-based nanoscale systems have great value as a promising platform for skin diseases therapy.
Scopus is among the largest curated abstract and citation databases, with a wide global and regional coverage of scientific journals, conference proceedings, and books, while ensuring only the highest quality data are indexed through rigorous content selection and re-evaluation by an independent Content Selection and Advisory Board. Additionally, extensive quality assurance processes continuously monitor and improve all data elements in Scopus. Besides enriched metadata records of scientific articles, Scopus offers comprehensive author and institution profiles, obtained from advanced profiling algorithms and manual curation, ensuring high precision and recall. The trustworthiness of Scopus has led to its use as bibliometric data source for large-scale analyses in research assessments, research landscape studies, science policy evaluations, and university rankings. Scopus data have been offered for free for selected studies by the academic research community, such as through application programming interfaces, which have led to many publications employing Scopus data to investigate topics such as researcher mobility, network visualizations, and spatial bibliometrics. In June 2019, the International Center for the Study of Research was launched, with an advisory board consisting of bibliometricians, aiming to work with the scientometric research community and offering a virtual laboratory where researchers will be able to utilize Scopus data.
Proteolysis-targeting chimeras are a new drug modality that exploits the endogenous ubiquitin proteasome system to degrade a protein of interest for therapeutic benefit. As the first-generation of proteolysis-targeting chimeras have now entered clinical trials for oncology indications, it is timely to consider the theoretical safety risks inherent with this modality which include off-target degradation, intracellular accumulation of natural substrates for the E3 ligases used in the ubiquitin proteasome system, proteasome saturation by ubiquitinated proteins, and liabilities associated with the "hook effect" of proteolysis-targeting chimeras This review describes in vitro and non-clinical in vivo data that provide mechanistic insight of these safety risks and approaches being used to mitigate these risks in the next generation of proteolysis-targeting chimera molecules to extend therapeutic applications beyond life-threatening diseases.
Reactive oxygen species (ROS) play a dual role in the initiation, development, suppression, and treatment of cancer. Excess ROS can induce nuclear DNA, leading to cancer initiation. Not only that, but ROS also inhibit T cells and natural killer cells and promote the recruitment and M2 polarization of macrophages; consequently, cancer cells escape immune surveillance and immune defense. Furthermore, ROS promote tumor invasion and metastasis by triggering epithelial-mesenchymal transition in tumor cells. Interestingly, massive accumulation of ROS inhibits tumor growth in two ways: (1) by blocking cancer cell proliferation by suppressing the proliferation signaling pathway, cell cycle, and the biosynthesis of nucleotides and ATP and (2) by inducing cancer cell death via activating endoplasmic reticulum stress-, mitochondrial-, and P53- apoptotic pathways and the ferroptosis pathway. Unfortunately, cancer cells can adapt to ROS via a self-adaption system. This review highlighted the bidirectional regulation of ROS in cancer. The study further discussed the application of massively accumulated ROS in cancer treatment. Of note, the dual role of ROS in cancer and the self-adaptive ability of cancer cells should be taken into consideration for cancer prevention.
Small molecule, dietary antioxidants exert a remarkably broad range of bioactivities, and many of these can be explained by the influence of antioxidants on the redox homeostasis. Such compounds help to modulate the levels of harmful reactive oxygen/nitrogen species, and therefore participate in the regulation of various redox signaling pathways. However, upon ingestion, antioxidants usually undergo extensive metabolism that can generate a wide range of bioactive metabolites. This makes it difficult, but otherwise a need, to identify the ones responsible for the different activities of antioxidants. By better understanding their ways of action, the use of antioxidants in therapy can be improved. This review provides a summary on the role of the in vivo metabolic changes and the oxidized metabolites on the mechanisms behind the bioactivity of antioxidants. A special attention is given to metabolites described as products of biomimetic oxidative chemical reactions, which can be considered as models of free radical scavenging. During such reactions a wide variety of metabolites are formed, and they can exert completely different specific bioactivities as compared to their parent antioxidants. This implies that exploring the free radical scavenging-related metabolite fingerprint of each antioxidant molecule, collectively defined here as the scavengome, will lead to a deeper understanding of the bioactivity of these compounds. Furthermore, this paper aims to be a working tool for systematic studies on oxidized metabolic fingerprints of antioxidants, which will certainly reveal an often-neglected segment of chemical space that is a treasury of bioactive compounds.
Plants respond to insect herbivory with the production of volatiles that attract carnivorous enemies of the herbivores, a phenomenon called indirect defence or 'plants crying for help'. Plants are under selection to maximize Darwinian fitness, and this can be done by making the right 'decisions' (i.e. by responding to environmental stress in ways that maximize seed production). Plant decisions related to the response to herbivory in terms of the emission of herbivore-induced volatiles include 'to respond or not to respond', 'how fast to respond', 'how to respond' and 'when to stop responding'. In this review, the state-of-the-art of the research field is presented in the context of these decisions that plants face. New questions and directions for future research are identified. To understand the consequences of plant responses in a community context, it is important to expand research from individual interactions to multispecies interactions in a community context. To achieve this, detailed information on underlying mechanisms is essential and first steps on this road have been made. This selective review addresses the ecology of herbivore-induced plant volatiles (HIPVs) by integrating information on mechanisms and ecological functions. New questions are identified as well as challenges for extending current information to community ecology.
In a striking glimpse into extreme physics, scientists have captured the split-second chaos that unfolds when powerful laser flashes blast matter into a superheated plasma。 By combining two cutting-edge lasers, researchers were able to track how copper atoms lose and regain electrons in trillionths of a second, creating and dissolving highly charge
Liquid biopsy, characterized by minimally invasive detection through biofluids such as blood, saliva, and urine, has emerged as a revolutionary strategy for cancer diagnosis and prognosis prediction. Exosomes are a subset of extracellular vesicles (EVs) that shuttle molecular cargoes from donor cells to recipient cells and play a crucial role in mediating intercellular communication. Increasing studies suggest that exosomes have a great promise to serve as novel biomarkers in liquid biopsy, since large quantities of exosomes are enriched in body fluids and are involved in numerous physiological and pathological processes. However, the further clinical application of exosomes has been greatly restrained by the lack of high-quality separation and component analysis methods. This review aims to provide a comprehensive overview on the conventional and novel technologies for exosome isolation, characterization and content detection. Additionally, the roles of exosomes serving as potential biomarkers in liquid biopsy for the diagnosis, treatment monitoring, and prognosis prediction of cancer are summarized. Finally, the prospects and challenges of applying exosome-based liquid biopsy to precision medicine are evaluated.