A cell-based screen identifies HDAC inhibitors as activators of RIG-I signaling Enhancing the immune microenvironment in cancer by targeting the nucleic acid sensors is becoming a potent therapeutic strategy. Among the nucleic acid sensors, activation of the RNA sensor Retinoic Acid-inducible Gene (RIG-I) using small hairpin RNAs has been shown to elicit powerful innate and adaptive immune responses. Given the challenges inherent in pharmacokinetics and delivery of RNA based agonists, we set out to discover small molecule agonists of RIG-I using a cell-based assay. To this end, we established and validated a robust high throughput screening assay based on a commercially available HEK293 reporter cell line with a luciferase reporter downstream of tandem interferon stimulated gene 54 (ISG54) promoter elements. We first confirmed that the luminescence in this cell line is dependent on RIG-I and the interferon receptor using a hairpin RNA RIG-I agonist. We established a 96-well and a 384-well format HTS based on this cell line and performed a proof-of-concept screen using an FDA approved drug library of 1200 compounds. Surprisingly, we found two HDAC inhibitors Entinostat, Mocetinostat and the PLK1 inhibitor Volasertib significantly enhanced ISG-luciferase activity. This luminescence was substantially diminished in the null reporter cell line indicating the increase in signaling was dependent on RIG-I expression. Combination treatment of tumor cell lines with Entinostat increased RIG-I induced cell death in a mammary carcinoma cell line that is resistant to either Entinostat or RIG-I agonist alone. Taken together, our data indicates an unexpected role for HDAC1,-3 inhibitors in enhancing RIG-I signaling and highlight potential opportunities for therapeutic combinations.
MYB insufficiency disrupts proteostasis in hematopoietic stem cells leading to age-related neoplasia The Myb transcription factor plays critical roles in normal and malignant hematopoiesis. Acquired genetic dysregulation of Myb, which plays a central role in hematopoietic stem cell (HSC) gene regulation, is involved in the etiology of a number of leukemias. Also, inherited non-coding variants of the Myb gene are a factor in susceptibility to many hematological conditions, including myeloproliferative neoplasms (MPN), but the mechanisms by which variations in Myb levels predispose to disease, including age-dependency in disease occurrence, are completely unknown. Here, we address these key points by showing that Myb insufficiency in mice leads in later life to MPN, myelodysplasia, and leukemia, mirroring the age profile of equivalent human diseases. This age-dependence is intrinsic to HSC, involving progressive accumulation of subtle changes. Interestingly, and linking to previous studies showing the importance of proteostasis to the maintenance of normal HSC, we observed altered proteosomal activity in young Myb-insufficient mice and later elevated ribosome activity. We propose that these alterations collectively cause an imbalance in proteostasis, potentially creating a cellular milieu favoring disease initiation by driver mutations.
A Hierarchy of Biomolecular Proportional-Integral-Derivative Feedback Controllers for Robust Perfect Adaptation and Dynamic Performance Proportional-Integral-Derivative (PID) feedback controllers have been the most widely used controllers in industry for almost a century due to their good performance, simplicity, and ease of tuning. Motivated by their success in various engineering disciplines, PID controllers recently found their way into synthetic biology, where the design of feedback molecular control systems has been identified as an important goal. In this paper, we consider the mathematical realization of PID controllers via biomolecular interactions. We propose an array of topologies that offer a compromise between simplicity and high performance. We first demonstrate that different Proportional-Integral (PI) controllers exhibit different capabilities for enhancing the dynamics and reducing variance (cell-to-cell variability). Next, we introduce several derivative controllers that are realized based on incoherent feedforward loops acting in a feedback configuration. Alternatively, we show that differentiators can be realized by placing molecular integrators in a negative feedback loop--an arrangement that can then be augmented by PI components to yield PID feedback controllers. We demonstrate that the derivative component can be exploited for enhancing system stability, dramatically increasing the molecular control systems dynamic performance, and for reducing the noise effect on the output of interest. The PID controller features are established through various deterministic and stochastic analyses as well as numerical simulations. Finally, we provide an experimental demonstration using a recently developed hybrid setup, the cyberloop, where the controller is implemented in silico to control a biological genetic circuit in vivo. The large array of novel biomolecular PID controllers introduced here forms a basis for the design and construction of advanced high-performance biomolecular control systems that robustly regulate the dynamics of living systems. An NLR Integrated Domain toolkit to identify plant pathogen effector targets Plant resistance genes (or NLR "Nod-like Receptors") are known to contain atypical domains procuring them with a decoy capacity. Some of these integrated domains (or ID) allow the plant to lure the virulence determinants ("effectors") of pathogens and triggering a specific NLR immune reaction.

In this work, our goal was to generate a library of known IDs that could be screened with plant pathogen effectors in order to identify putative new effector virulence targets and NLR-effector pairs.

We curated the IDs contained in NLRs from seven model and crop plant species. We cloned 52 IDs representing 31 distinct Pfam domains. This library was screened for interaction by yeast-two-hybrid with a set of 31 conserved Ralstonia solanacearum type III effectors. This screening and the further in planta interaction assay allowed us to identify three interactions, involving different IDs (kinase, DUF3542, WRKY) and two type III effectors (RipAE and PopP2).

PopP2 was found to physically interact with ID#85, an atypical WRKY domain integrated in the GmNLR-ID85 NLR protein from Soybean. Using a imaging method in living plant cells, we showed that PopP2 associates with ID#85 in the nucleus. But unlike the known WRKY-containing Arabidopsis RRS1-R NLR receptor, this newly identified soybean WRKY domain could not be acetylated by PopP2 and its atypical sequence (WRKYGKR) also probably renders it inefficient in plant immunity triggering.

This ID toolkit is available for screening with other plant pathogen effectors and should prove useful to discover new effectors targets and potentially engineer new plant resistance genes. Cell Rearrangement Generates Pattern Emergence as a Function of Temporal Morphogen Exposure. As tissues elongate, cell rearrangement alters positional information in manner that must be accounted for to generate gene expression pattern. How this is achieved during paraxial mesoderm elongation is unknown. By reverse-engineering gene regulatory networks that predict single cell expression trajectories across the tissue, we find a network capable of recapitulating the full range of dynamic differentiation profiles observed both in vivo and in vitro. Simulating gene expression profiles on in toto cell tracking data sets reveal that temporal exposure to Wnt and FGF is generated by cell movement. The absence of reversal in gene expression towards a more premature gene expression state predicts the generation of aberrant tbx6 expression in the posterior progenitor zone that we then confirm by quantitative single cell imaging. Taken together, these results demonstrate cell rearrangement tunes the dynamics of mesoderm progenitor differentiation to generate pattern emergence as a function of temporal Wnt and FGF exposure. Drug screening at single-organoid resolution via bioprinting and interferometry High-throughput drug screening is an established approach to investigate tumor biology and identify therapeutic leads. Traditional platforms for high-throughput screening use two-dimensional cultures of immortalized cell lines which do not accurately reflect the biology of human tumors. More clinically relevant model systems, such as three-dimensional tumor organoids, can be difficult to screen and scale. For example, manually seeded organoids coupled to destructive endpoint assays allow for the characterization of response to treatment, but do not capture the transitory changes and intra-sample heterogeneity underlying clinically observed resistance to therapy. We therefore developed a pipeline to generate bioprinted tumor organoids linked to label-free, real-time imaging via high-speed live cell interferometry (HSLCI) and machine learning-based quantitation of individual organoids. Bioprinting cells gives rise to 3D organoid structures that preserve tumor histology and gene expression. HSLCI imaging in tandem with machine learning-based image segmentation and organoid classification tools enables accurate, label-free parallel mass measurements for thousands of bioprinted organoids. We demonstrate that our method quantitatively identifies individual organoids as insensitive, transiently sensitive, or persistently sensitive to specific treatments. This opens new avenues for rapid, actionable therapeutic selection using automated tumor organoid screening. Avian haemosporidians (Plasmodium and Haemoproteus) status in selected bird groups (Old world Flycatchers, Warblers, Babblers, and Thrushes) of India and their phylogenetic relationships with other lineages of the world The avian haemosporidians (Plasmodium and Haemoproteus) are widely distributed and may affect the host populations from body damage at individual level to the extinction of a population. The knowledge about their status may help in future avifauna conservation plans. Hence, we examined the avian haemosporidians status, in selected bird groups (Old world Flycatchers, Warblers, Babblers, and Thrushes) of India, and their phylogenetic relationships with other known lineages of the world. We used the common genetic marker (Cytochrome b gene fragment of 479 bp) with information on the geographic distribution of parasite and host species available at MalAvi database. The prevalence of avian haemosporidians, from northern, eastern, and southern parts of India and phylogenetic genetic analysis of lineages was carried out to know the genetic relatedness among them at local and world level. The MCC tree revealed six Haemosporidian lineages in which one was common (H_MSP01) and five were unique (H_CYOPOL01, H_CHD01, H_CYORUB01, H_EUMTHA01, and P_GEOCIT01). The avian host richness Index was 2.0852. 9.9%, prevalence of Haemosporidian infection was found in 111 DNA samples belonging to 6 host species. The Haemoproteus prevalence was found to be 9.0 % across five host species (Phylloscopus trochiloides, Cyornis poliogenys, C. hainanus dialilaemus, C. rubeculoides, Eumiyas thalassinus) and Plasmodium prevalence was 0.9% in Geokichla citrina. Spatial phylogeny at global level showed H_MSP01 lineage, found in different host species in India, was genetically related to H. pallidus lineages (COLL2 and PFC1) in parts of Africa, Europe, North America, Malaysia, and Philippines. The Plasmodium lineage (P_GEOCIT01) was related to PADOM16 in Egypt with poor sequence similarity (93.89%). The statistical analysis suggested that the haemosporidians host species distribution range was directly and significantly associated with the altitude, minimum temperature, and relative humidity. H_MSP01 distribution was in accordance with H. pallidus having a wide geographic and host range. Structural predictions of the SNX-RGS proteins suggest they belong to a new class of lipid transfer proteins Recent advances in protein structure prediction using machine learning such as AlphaFold2 and RosettaFold presage a revolution in structural biology. Genome-wide predictions of protein structures are providing unprecedented insights into their architecture and intradomain interactions, and applications have already progressed towards assessing protein complex formation. Here we present detailed analyses of the sorting nexin proteins that contain regulator of G-protein signalling domains (SNX-RGS proteins), providing a key example of the ability of AlphaFold2 to reveal novel structures with previously unsuspected biological functions. These large proteins are conserved in most eukaryotes and are known to associate with lipid droplets (LDs) and sites of LD-membrane contacts, with key roles in regulating lipid metabolism. They possess five domains, including an N-terminal transmembrane domain that anchors them to the endoplasmic reticulum, an RGS domain, a lipid interacting phox homology (PX) domain and two additional domains named the PXA and PXC domains of unknown structure and function. Here we report the crystal structure of the RGS domain of sorting nexin 25 (SNX25) and show that the AlphaFold2 prediction closely matches the experimental structure. Analysing the full-length SNX-RGS proteins across multiple homologues and species we find that the distant PXA and PXC domains in fact fold into a single unique structure that notably features a large and conserved hydrophobic pocket. The nature of this pocket strongly suggests a role in lipid or fatty acid binding, and we propose that these molecules represent a new class of conserved lipid transfer proteins. Analysis of single-cell RNA sequencing data basedon autoencoders BackgroundSingle-cell RNA sequencing (scRNA-Seq) experiments are gaining ground to study the molecular processes that drive normal development as well as the onset of different pathologies. Finding an effective and efficient low-dimensional representation of the data is one of the most important steps in the downstream analysis of scRNA-Seq data, as it could provide a better identification of known or putatively novel cell-types. Another step that still poses a challenge is the integration of different scRNA-Seq datasets. Though standard computational pipelines to gain knowledge from scRNA-Seq data exist, a further improvement could be achieved by means of machine learning approaches.

ResultsAutoencoders (AEs) have been effectively used to capture the non-linearities among gene interactions of scRNA-Seq data, so that the deployment of AE-based tools might represent the way forward in this context. We introduce here scAEspy, a unifying tool that embodies: (1) four of the most advanced AEs, (2) two novel AEs that we developed on purpose, (3) different loss functions. We show that scAEspy can be coupled with various batch-effect removal tools to integrate data by different scRNA-Seq platforms, in order to better identify the cell-types. We benchmarked scAEspy against the most used batch-effect removal tools, showing that our AE-based strategies outperform the existing solutions.

ConclusionsscAEspy is a user-friendly tool that enables using the most recent and promising AEs to analyse scRNA-Seq data by only setting up two user-defined parameters. Thanks to its modularity, scAEspy can be easily extended to accommodate new AEs to further improve the downstream analysis of scRNA-Seq data. Considering the relevant results we achieved, scAEspy can be considered as a starting point to build a more comprehensive toolkit designed to integrate multi single-cell omics. Nuclear Progestin Receptor Mediated Linkage of Blood Coagulation and Ovulation Ovulation is a dramatic remodeling process that includes rupture of blood capillaries and clotting, but coagulation is not thought to directly regulate this process. Herein, we report remarkable increases of coagulation factors V (f5, [~]3145-fold) and tissue factor (f3a, [~]120-fold) in zebrafish ovarian follicle cells during ovulation. This increase was mediated through the nuclear progestin receptor (Pgr), which is essential for ovulation in zebrafish, and was totally abolished in ovarian follicular cells from pgr-/- mutants. In addition, promoter activities of f5 and f3a were significantly enhanced by progestin (DHP) via Pgr. Similar regulation of human F5 promoter activity was induced via human PGRB, suggesting a conserved mechanism. Site-directed mutagenesis of the zebrafish f5 promoter further demonstrated a direct regulation of coagulation factors via progestin response elements. Moreover, a stark increase of erythrocytes occurred in capillaries meshed in wildtype preovulatory follicles but was absent in pgr-/- mutants. Interestingly, anticoagulants significantly inhibited ovulation both in vitro and in vivo, respectively. Furthermore, reduced fecundity was observed in f5+/- female zebrafish. Taken together, our study provides plausible evidence for steroid regulation of coagulation factors, and a new hypothesis for blood clotting triggered ovulation in vertebrates. Collagen polarization promotes epithelial elongation by stimulating locoregional cell proliferation. Epithelial networks are commonly generated by processes where multicellular aggregates elongate and branch. Here we focus on understanding cellular mechanisms for elongation, using an organotypic culture system as a model of mammary epithelial anlage. Isotropic cell aggregates broke symmetry and slowly elongated when transplanted into collagen 1 gels. The elongating regions of aggregates displayed enhanced cell proliferation that was necessary for elongation to occur. Strikingly, this loco-regional increase in cell proliferation occurred where collagen 1 fibrils reorganized into bundles which were polarized with the elongating aggregates. Applying external stretch as a cell-independent way to reorganize the ECM, we found that collagen polarization stimulated regional cell proliferation to precipitate symmetry-breaking and elongation. This required {beta}1-integrin and ERK signaling. We propose that collagen polarization supports epithelial anlagen elongation by stimulating loco-regional cell proliferation. This could provide a long-lasting structural memory of the initial axis that is generated when anlage break symmetry. Protein scaffold-based multimerization of soluble ACE2 efficiently blocks SARS-CoV-2 infection in vitro and in vivo Soluble ACE2 (sACE2) decoy receptors are promising agents to inhibit SARS-CoV-2, as their efficiency is less likely to be affected by common escape mutations in viral proteins. However, their success may be limited by their relatively poor potency. To address this challenge, we developed a large decoy library of sACE2 fusion proteins, generated with several protease inhibitors or multimerization tags. Among these decoys, multimeric sACE2 consisting of SunTag or MoonTag systems, which were originally utilized for signal amplification or gene activation systems, were extremely effective in neutralizing SARS-CoV-2 in pseudoviral systems and in clinical isolates. These novel sACE2 fusion proteins exhibited greater than 100-fold SARS-CoV-2 neutralization efficiency, compared to monomeric sACE2. SunTag or MoonTag in combination with a more potent version of sACE2, which has multiple point mutations for greater binding (v1), achieved near complete neutralization at a sub-nanomolar range, comparable with clinical monoclonal antibodies. Pseudoviruses bearing mutant versions of Spike, alpha, beta, gamma or delta variants, were also neutralized efficiently with SunTag or MoonTag fused sACE2(v1). Finally, therapeutic treatment of sACE2(v1)-MoonTag provided protection against SARS-CoV-2 infection in an in vivo mouse model. Overall, we suggest that the superior activity of the sACE2-SunTag or sACE2-MoonTag fusions is due to the greater occupancy of the multimeric sACE2 receptors on Spike protein as compared to monomeric sACE2. Therefore, these highly potent multimeric sACE2 decoy receptors may offer a promising treatment approach against SARS-CoV-2 infections.

One Sentence SummaryMultimerization of sACE2 markedly enhanced the neutralization of SARS-CoV-2 by blocking multiple viral spike proteins simultaneously. Design of anti-SARS-CoV-2 molecules from hydroxychloroquine (HCQ)for better binding SARS-CoV-2 N protein Great efforts have led to successfully developing the spike-based vaccines but challenges still exist to completely terminate the SARS-CoV-2 pandemic. SARS-CoV-2 nucleocapsid (N) protein plays the essential roles in almost all key steps of the viral life cycle, thus representing a top drug target. Almost all key functions of N protein including liquid-liquid phase separation (LLPS) depend on its capacity in interacting with nucleic acids. Therefore, only the variants with their N proteins functional in binding nucleic acids might survive and spread in evolution and indeed, the residues critical for binding nucleic acids are highly conserved. Very recently, hydroxychloroquine (HCQ) was shown to prevent the transmission in a large-scale clinical study in Singapore but so far, no specific SARS-CoV-2 protein was experimentally identified to be targeted by HCQ. Here by NMR, we unambiguously decode that HCQ specifically binds NTD and CTD of SARS-CoV-2 N protein with Kd of 112.1 and 57.1 M respectively to inhibit their interaction with nucleic acid, as well as to disrupt LLPS essential for the viral life cycle. Most importantly, HCQ-binding residues are identical in SARS-CoV-2 variants and therefore HCQ is likely effective to them all. The results not only provide a structural basis for the anti-SARS-CoV-2 activity of HCQ, but also renders HCQ to be the first known drug capable of targeting LLPS. Furthermore, the unique structure of the HCQ-CTD complex decodes a promising strategy for further design of better anti-SARS-CoV-2 drugs from HCQ. Therefore, HCQ is a promising candidate to help terminate the pandemic. Artificial seawater based long-term culture of colonial ascidians Tunicates are highly diverse marine invertebrate filter-feeders that are vertebrates closest relatives. These organisms, despite a drastically different body plan during their adulthood, have a tissue complexity related to that of vertebrates. Ascidians, which compose most of the Tunicata, are benthic sessile hermaphrodites that reproduce sexually through a motile tadpole larval stage. Over half of the known ascidians species are able to reproduce asexually by budding, typically leading to the formation of colonies where animals, called zooids, are interconnected through an external vascular system. In addition, colonial ascidians are established models for important biological processes including allorecognition, immunobiology, aging, angiogenesis and whole-body regeneration. However, the current paucity in breeding infrastructures limits the study of these animals to coastal regions.

To promote a wider scientific spreading and popularity of colonial ascidians, we have developed a flexible recirculating husbandry setup for their long-term in-lab culture. Our system is inspired both by the flow-through aquariums used by coastal ascidian labs, as well as by the recirculating in-lab systems used for zebrafish research. Our hybrid system thus combines colony breeding, water filtering and food culturing in a semi-automated system where specimens develop on hanging microscopy glass slides. Temperature, light/dark cycles, flow speed and feeding rates can be controlled independently in four different breeding environments to provide room for species-specific optimization as well as for running experiments. This setup is complemented with a quarantine for the acclimatization of wild isolates.

Herein we present our success in breeding Botrylloides diegensis, a species of colonial ascidians, for more than 3 years in recirculating artificial seawater over 600 km away from their natural habitat. We show that colonies adapt well to in-lab culturing provided that a suitable marine microbiome is present, and that a specific strain can be isolated, propagated and efficiently used for research over prolonged periods of time. The flexible and modular structure of our system can be scaled and adapted to the needs of specific species, such as Botryllus schlosseri, as well as of particular laboratory spaces. Overall, we show that Botrylloides diegensis can be proficiently bred in-land and suggest that our results can be extended to other species of colonial ascidians to promote research on these fascinating animals.

HighlightsO_LIFirst artificial seawater based recirculating aquaculture for colonial ascidians
C_LIO_LIOver 3 years of continuous breeding
C_LIO_LISemi-automated setup with minimized maintenance
C_LIO_LIGood biomass production for strain propagation
C_LIO_LI4 different culture conditions for optimized breeding for species of interest
C_LI EMBED: Essential Microbiome Dynamics, a dimensionality reduction approach for longitudinal microbiome studies Dimensionality reduction can offer unique insights into high dimensional microbiome dynamics by leveraging collective abundance fluctuations of multiple bacteria driven by similar ecological perturbations. However, methods providing lower-dimensional representations of microbiome dynamics both at the community and individual taxa level are not currently available. To that end, we present EMBED: Essential MicroBiomE Dynamics, a probabilistic non-linear tensor factorization approach. Similar to normal mode analysis in structural biophysics, EMBED infers ecological normal modes (ECNs), which represent the unique orthogonal modes capturing the collective behavior of microbial communities. A very small number of ECNs can accurately approximate microbiome dynamics across multiple data sets. Inferred ECNs reflect specific ecological behaviors, providing natural templates along which the dynamics of individual bacteria may be partitioned. Moreover, the multi-subject treatment in EMBED systematically identifies subject-specific and universal abundance dynamics that are not detected by traditional approaches. Collectively, these results highlight the utility of EMBED as a versatile dimensionality reduction tool for studies of microbiome dynamics. Hot and cold spots of pest-induced US urban tree death, 2020-2050 O_LIUrban trees are important nature-based solutions for future wellbeing and livability but are at high risk of mortality from insect pests. In the United States (US), 82% of the population live in urban settings and this number is growing, making urban tree mortality a matter of concern for most of its population. Until now, the magnitudes and spatial distributions of risks were unknown.
C_LIO_LIHere, we combine new models of street tree populations in [~]30,000 US communities, species-specific spread predictions for 57 invasive insect species, and estimates of tree death due to insect exposure for 48 host tree genera.
C_LIO_LIWe estimate that 1.4 million street trees will be killed by invasive insects from 2020 through 2050, costing an annualized average of US$ 30M. However, these estimates hide substantial variation: 23% of urban centers will experience 95% of all insect-induced mortality. Further, 90% of all mortality will be due to emerald ash borer (Agrilus planipennis, EAB), which is expected to kill virtually all ash trees (Fraxinus spp.) in >6000 communities.
C_LIO_LIWe define an EAB high-impact zone spanning 902,500km2, largely within the southern and central US, within which we predict the death of 98.8% of all ash trees. "Mortality hotspot cities" include Milwaukee, WI; Chicago, IL; and New York, NY.
C_LIO_LIWe identify Asian wood borers of maple and oak trees as the highest risk future invaders, where a new establishment could cost US$ 4.9B over 30 years.
C_LIO_LIPolicy implications: To plan effective mitigation, managers must know which tree species in which communities will be at the greatest risk, as well as the highest-risk insects. We provide the first country-wide, spatial forecast of urban tree mortality due to invasive insect pests. This framework identifies dominant pest insects and spatial impact hotspots, which can provide the basis for spatial prioritization of spread control efforts such as quarantines and biological control release sites. Our results highlight the need for EAB early-detection efforts as far from current infestations as Seattle, WA. Further, these findings produce a list of biotic and spatiotemporal risk factors for future high-impact US urban forest insect pests.
C_LI Diverse stem-chondrichthyan oral structures and evidence for an independently acquired acanthodid dentition The teeth of sharks famously form a series of transversely-organised files with conveyor-belt replacement that are borne directly on the jaw cartilages, in contrast to the dermal plate-borne dentition of bony fishes that undergoes site-specific replacement. A major obstacle in understanding how this system evolved is the poorly understood relationships of the earliest chondrichthyans and the profusion of morphologically and terminologically diverse bones, cartilages, splints and whorls that they possess. Here we use tomographic methods to investigate mandibular structures in several early branching  acanthodian-grade stem-chondrichthyans. We show that the dentigerous jaw bones of disparate genera of ischnacanthids are united by a common construction, being growing bones with non-shedding dentition. Mandibular splints, which support the ventro-lateral edge of the Meckels cartilage in some taxa, are formed from dermal bone and may be an acanthodid synapomorphy. We demonstrate that the teeth of Acanthodopsis are borne directly on the mandibular cartilage and that this taxon is deeply nested within an edentulous radiation, representing an unexpected independent origin of teeth. Many or even all of the range of unusual oral structures may be apomorphic, but they should nonetheless be considered when building hypotheses of tooth and jaw evolution, both in chondrichthyans and more broadly. Bioinspired Dual-Functional Solid Lipid Nanoformulations for Targeted Drug Delivery and Sustained Release for Enhancement of Potency of Albendazole, an Antihelminthic Drug Targeted delivery has not been achieved for anthelmintic treatment, resulting in the requirement of excess drugs dose leading to side effects and therapeutic resistance. Gastrointestinal helminths ingest lipid droplets from digestive fluid for energy production, development, and defense. Worms habit inspired us to develop biocompatible, oral administrable, bee-wax derived solid lipid nanoparticles (SLN) with excellent drug (albendazole) loading efficiency of 83.3 {+/-} 6.5 mg/g and sustained-release properties (86.4 {+/-} 3.9 % of drug released within 24 h). Rhodamine B-loaded SLN showed time-dependent release and distribution of dye in vivo in Haemonchous contortus. The intestinal sustained-release property was shown by the particles that caused enhancement of albendazole potency for up to 50 folds. Therefore, this formulation has immense potential as an anthelminthic drug delivery vehicle that will not only be able to reduce the dose but will also reduce the drug-induced side effects by enhancing the bioavailability of the drug.

HighlightsO_LIAlbendazole-loaded Solid Lipid Nanoparticles (SLN-A) were formulated using Beeswax as stating material showing high drug loading capacity of 83 mg/g with sustained-release properties and 84 {+/-} 3 % of drug release within 24 h.
C_LIO_LISLN-A particles showed 50 fold enhancement of Albendazole activity against Haemonchous contortus worm.
C_LIO_LIRhodamine B-loaded SLN particles showed the specific uptake and in-vivo sustained release of dye in the worm.
C_LI Motor planning brings human primary somatosensory cortex into action-specific preparatory states Motor planning plays a critical role in producing fast and accurate movement. Yet, the neural processes that occur in human primary motor and somatosensory cortex during planning, and how they relate to those during movement execution, remain poorly understood. Here we used 7T functional magnetic resonance imaging (fMRI) and a delayed movement paradigm to study single finger movement planning and execution. The inclusion of no-go trials and variable delays allowed us to separate what are typically overlapping planning and execution brain responses. Although our univariate results show widespread deactivation during finger planning, multivariate pattern analysis revealed finger-specific activity patterns in contralateral primary somatosensory cortex (S1), which predicted the planned finger action. Surprisingly, these activity patterns were as informative as those found in contralateral primary motor cortex (M1). Control analyses ruled out the possibility that the detected information was an artifact of subthreshold movements during the preparatory delay. Furthermore, we observed that finger-specific activity patterns during planning were highly correlated to those during execution. These findings reveal that motor planning activates the specific S1 and M1 circuits that are engaged during the execution of a finger press, while activity in both regions is overall suppressed. We propose that preparatory states in S1 may improve movement control through changes in sensory processing or via direct influence of spinal motor neurons.

Significance statementMotor planning is important for good behavioral performance, yet it is unclear which neural processes underlie the preparation of the nervous system for an upcoming movement. Using high-resolution functional neuroimaging, we investigated how motor planning for finger presses changes the activity state in primary motor and primary somatosensory cortex, and how brain responses during planning and execution relate to each other. We show that planning leads to finger-specific activation in both M1 and S1, which is highly similar to the finger-specific activity patterns elicited during execution. Our findings suggest that S1 is being specifically prepared for an upcoming action, either to actively contribute to the outflowing motor command or to enable action-specific sensory gating. If Research is Not the Evidence, What is it? The Lack of Research Citations in Health Vlogs Doctor-influencers have the knowledge needed to understand scientific research, and they have the social media popularity to share it with large audiences. In this article, I explore the treatment of evidence in vlogs by Egyptian doctor-influencers: how they present it and what they use instead of it. I answer the following questions: 1) How do doctor-influencers present evidence on social media? 2) If evidence is not the research, then what is it? The data was collected through in-depth interviews with 12 Egyptian doctor-influencers from different specializations; two focus groups; and a critical discourse analysis of 48 of the influencers most popular and most engaging videos. I found that the doctor-influencers cited academic research only in the videos about medical controversies, new treatments, misinformation, or common medical mistakes. In most of their videos, the Egyptian doctor-influencers scarcely referred to medical research. This is because they believe that their audiences would not understand evidence due to their low educational attainment, weak research skills, and the language barrier (as most medical research is published in English rather than Arabic). These findings highlight the challenges of communicating science in societies with a low level of education. In these settings, communicators may need to move beyond mediating research studies on social media and instead focus on simplifying, translating, and connecting evidence-based information to the cultural context. A novel nongenetic murine model of hyperglycemia and hyperlipidemia-associated aggravated atherosclerosis ObjectiveAtherosclerosis, the main pathology underlying cardiovascular diseases is accelerated in diabetic patients. Genetic mouse models require breeding efforts which are time-consuming and costly. Our aim was to establish a new nongenetic model of inducible metabolic risk factors that mimics hyperlipidemia, hyperglycemia, or both and allows the detection of phenotypic differences dependent on the metabolic stressor(s).

Methods and ResultsWild-type mice were injected with gain-of-function PCSK9D377Y (proprotein convertase subtilisin/kexin type 9) mutant adeno-associated viral particles (AAV) and streptozotocin and fed either a high-fat diet (HFD) for 12 or 20 weeks or a high-cholesterol/high-fat diet (Paigen diet, PD) for 8 weeks. To evaluate atherosclerosis, two different vascular sites (aortic sinus and the truncus of the brachiocephalic artery) were examined in the mice. Combined hyperlipidemic and hyperglycemic (HGHCi) mice fed a HFD or PD displayed characteristic features of aggravated atherosclerosis when compared to hyperlipidemia (HCi HFD or PD) mice alone. Atherosclerotic plaques of HGHCi HFD animals were larger, showed a less stable phenotype (measured by the increased necrotic core area, reduced fibrous cap thickness, and less -SMA-positive area) and had more inflammation (increased plasma IL-1{beta} level, aortic pro-inflammatory gene expression, and MOMA-2-positive cells in the BCA) after 20 weeks of HFD. Differences between the HGHCi and HCi HFD models were confirmed using RNA-seq analysis of aortic tissue, revealing that significantly more genes were dysregulated in mice with combined hyperlipidemia and hyperglycemia than in the hyperlipidemia-only group. The HGHCi-associated genes were related to pathways regulating inflammation (increased Cd68, iNos, and Tnfa expression) and extracellular matrix degradation (Adamts4 and Mmp14). When comparing HFD with PD, the PD aggravated atherosclerosis to a greater extent in mice and showed plaque formation after 8 weeks. Hyperlipidemic and hyperglycemic mice fed a PD (HGHCi PD) showed less collagen (Sirius red) and increased inflammation (CD68-positive cells) within aortic plaques than hyperlipidemic mice (HCi PD). HGHCi-PD mice represent a directly inducible hyperglycemic atherosclerosis model compared with HFD-fed mice, in which atherosclerosis is severe by 8 weeks.

ConclusionWe established a nongenetically inducible mouse model allowing comparative analyses of atherosclerosis in HCi and HGHCi conditions and its modification by diet, allowing analyses of multiple metabolic hits in mice. Game of theories: an agent-based model of how variation in contest cost-benefits determines the evolution of contest resolution rules Game-theory based models are used to understand rules that animals use to settle contests over indivisible resources. However, the empirical literature of contests indicates controversial support to models, with some species supporting different models and other species showing no support to any model. Since strategies used to resolve contests may have different associated costs, it is possible that different conditions have determined the evolution of distinct assessment strategies used by animals. We used an agent-based model to explore the importance of the following conditions: resource availability, probability of reproduction with resource, and damage costs on evolution of assessment strategies. We used self- and mutual-assessment models as a heurist framework to build agents with different assessment strategies. In our model, agents competed for resources in scenarios with different combinations of resource availability, probability of reproduction with resource, and damage costs. We found that agents following the self-assessment with damage strategy were prevalent in scenarios with no probability of reproduction without the resource, independently of other variables. We also found that agents following the non-aggressive strategy occurred in all scenarios. However, agents using the non-aggressive strategy were prevalent only in scenarios with probability of reproduction with the resource. Finally, we observed that agents using mutual-assessment occurred only in a scenario with high risk of damage, low availability of resources, and with probability of reproduction without the resource. These results indicate that agents following the self-assessment with damage and non-aggressive strategies may be able to stay at most scenarios. Mutagenic distinction between the receptor-binding and fusion subunits of the SARS-CoV-2 spike glycoprotein and its upshot We observe that a residue R of the spike glycoprotein of SARS-CoV-2 which has mutated in one or more of the current Variants of Concern or Interest or under Monitoring rarely participates in a backbone hydrogen bond if R lies in the S1 subunit and usually participates in one if R lies in the S2 subunit. A partial explanation for this based upon free energy is explored as a potentially general principle in the mutagenesis of viral glycoproteins. This observation could help target future vaccine cargos for the evolving coronavirus as well as more generally. A study of the Delta and Omicron variants suggests that Delta was an energetically necessary intermediary in the evolution from Wuhan-Hu-1 to Omicron. Mitochondrial genomes of two parasitic Cuscuta species lack clear evidence of horizontal gene transfer and retain unusually fragmented ccmFC genes BackgroundThe intimate association between parasitic plants and their hosts favours the exchange of genetic material, potentially leading to horizontal gene transfer (HGT) between plants. With the recent publication of several parasitic plant nuclear genomes, there has been considerable focus on such non-sexual exchange of genes. To enhance the picture on HGT events in a widely distributed parasitic genus, Cuscuta (dodders), we assembled and analyzed the organellar genomes of two recently sequenced species, C. australis and C. campestris, making this the first account of complete mitochondrial genomes (mitogenomes) for this genus.

ResultsThe mitogenomes are 265,696 and 275,898 bp in length and contain a typical set of mitochondrial genes, with ten missing or pseudogenized genes often lost from angiosperm mitogenomes. Each mitogenome also possesses a structurally unusual ccmFC gene, which exhibits splitting of one exon and a shift to trans-splicing of its intron. Based on phylogenetic analysis of mitochondrial genes from across angiosperms and similarity-based searches, there is little to no indication of HGT into the Cuscuta mitogenomes. A few candidate regions for plastome-to-mitogenome transfer were identified, with one suggestive of possible HGT.

ConclusionsThe lack of HGT is surprising given examples from the nuclear genomes, and may be due in part to the relatively small size of the Cuscuta mitogenomes, limiting the capacity to integrate foreign sequences. Local fitness and epistatic effects lead to distinct patterns of linkagedisequilibrium in protein-coding genes Selected mutations interfere and interact with evolutionary processes at nearby loci, distorting allele frequency trajectories and creating correlations between pairs of mutations. A number of recent studies have used patterns of linkage disequilibrium (LD) between selected variants to test for selective interference and epistatic interactions, with some disagreement over interpreting observations from data. Interpretation is hindered by a lack of analytic or even numerical expectations for patterns of variation between pairs of loci under the combined effects of selection, dominance, epistasis, and demography. Here, I develop a numerical approach to compute the expected two-locus sampling distribution under diploid selection with arbitrary epistasis and dominance, recombination, and variable population size. I use this to explore how epistasis and dominance affect expected signed LD, including for non-steadystate demography relevant to human populations. Using whole-genome sequencing data from humans, I explore genome-wide patterns of LD within protein-coding genes. I show that positive LD between missense mutations within genes is driven by strong positive allele-frequency correlations between pairs of mutations that fall within the same annotated conserved domain, pointing to compensatory mutations or antagonistic epistasis as the prevailing mode of interaction within conserved genic elements. LD between missense mutations is reduced outside of conserved domains, as would expected under Hill-Robertson interference. This variation in both mutational fitness effects and selective interactions within proteincoding genes calls for more refined inferences of the joint distribution of fitness and interactive effects, and the methods presented here should prove useful in that pursuit. Glucose inhibits haemostasis and accelerates diet-induced hyperlipidaemia in zebrafish larvae Hyperglycaemia damages the microvasculature in part through the reduced recruitment of immune cells and interference with platelet signalling, leading to poor wound healing and accelerated lipid deposition in mammals. We investigated the utility of zebrafish larvae to model the effect of glucose on neutrophil and macrophage recruitment to a tail wound, wound-induced haemostasis, and chicken egg yolk feed challenge-induced hyperlipidaemia by supplementing larvae with exogenous glucose by immersion or injection. Neither method of glucose supplementation affected the recruitment of neutrophils and macrophages following tail transection. Glucose injection reduced thrombocyte retention and fibrin plug formation while only thrombocyte retention was reduced by glucose immersion following tail transection. We observed accelerated lipid accumulation in glucose-injected larvae challenged with high fat chicken egg yolk feeding. Our study identifies conserved and divergent effects of high glucose on inflammation, haemostasis, and hyperlipidaemia in zebrafish larvae compared to mammals.