Miracle-dz/UniFAR-FTUs huggingface dataset:
The readily-releasable pool dynamically regulates multivesicular release

The number of neurotransmitter-filled vesicles released into the synaptic cleft with each action potential dictates the reliability of synaptic transmission. Variability of this fundamental property provides diversity of synaptic function across brain regions, but the source of this variability is unclear. The prevailing view is that release of a single (univesicular release, UVR) or multiple vesicles (multivesicular release, MVR) reflects variability in vesicle release probability, a notion that is well-supported by the calcium-dependence of release mode. However, using mouse brain slices, we now demonstrate that the number of vesicles released is regulated by the size of the readily-releasable pool, upstream of vesicle release probability. Our results point to a model wherein protein kinase A and its vesicle-associated target, synapsin, dynamically control release site occupancy to dictate the number of vesicles released without altering release probability. Together these findings define molecular mechanisms that control MVR and functional diversity of synaptic signaling.
Generation and protective efficacy of a cold-adapted attenuated genotype 2b porcine epidemic diarrhea virus

The recent emergence and re-emergence of porcine epidemic diarrhea virus (PEDV) underscore the urgent need for the development of novel, safe, and effective vaccines against the prevailing strain. In this study, we generated a cold-adapted live attenuated vaccine candidate (Aram-P29-CA) by short-term passage of a virulent PEDV isolate at successively lower temperatures in Vero cells. Whole genome sequencing identified 12 amino acid changes in the cold-adapted strain with no insertions and deletions throughout the genome. Animal inoculation experiments confirmed the attenuated phenotype of Aram-P29-CA virus in the natural host. Pregnant sows were orally administered P29-CA live vaccines two doses at 2-week intervals prior to parturition, and the newborn piglets were challenged with the parental virus. The oral homologous prime-boost vaccination of P29-CA significantly improved the survival rate of the piglets and notably mitigated the severity of diarrhea and PEDV fecal shedding after the challenge. Furthermore, strong antibody responses to PEDV were detected in the sera and colostrum of immunized sows and in the sera of their offspring. These results demonstrated that the cold-adapted attenuated virus can be used as a live vaccine in maternal vaccination strategies to provide durable lactogenic immunity and confer passive protection to litters against PEDV.
Toward Microfluidic Label‐Free Isolation and Enumeration of Circulating Tumor Cells from Blood Samples

The isolation, analysis, and enumeration of circulating tumor cells (CTCs) from cancer patient blood samples are a paradigm shift for cancer patient diagnosis, prognosis, and treatment monitoring. Most methods used to isolate and enumerate these target cells rely on the expression of cell surface markers, which varies between patients, cancer types, tumors, and stages. Here, we propose a label‐free high‐throughput platform to isolate, enumerate, and size CTCs on two coupled microfluidic devices. Cancer cells were purified through a Vortex chip and subsequently flowed in‐line to an impedance chip, where a pair of electrodes measured fluctuations of an applied electric field generated by cells passing through. A proof‐of‐concept of the coupling of those two devices was demonstrated with beads and cells. First, the impedance chip was tested as a stand‐alone device: (1) with beads (mean counting error of 1.0%, sizing information clearly separated three clusters for 8, 15, and 20 um beads, respectively) as well as (2) with cancer cells (mean counting error of 3.5%). Second, the combined setup was tested with beads, then with cells in phosphate‐buffered saline, and finally with cancer cells spiked in healthy blood. Experiments demonstrated that the Vortex HT chip enriched the cancer cells, which then could be counted and differentiated from smaller blood cells by the impedance chip based on size information. Further discrimination was shown with dual high‐frequency measurements using electric opacity, highlighting the potential application of this combined setup for a fully integrated label‐free isolation and enumeration of CTCs from cancer patient samples. © 2019 International Society for Advancement of Cytometry
Gene-Wide Analysis of Aquaporin Gene Family in Malus domestica and Heterologous Expression of the Gene MpPIP2;1 Confers Drought and Salinity Tolerance in Arabidposis thaliana

The aquaporins (AQPs) are a family of integral membrane proteins involved in the transcellular membrane transport of water and other small molecules. A scan of the apple (Malus domestica) genome revealed the presence of 42 genes encoding putative AQPs. Based on a phylogenetic analysis of the deduced peptide sequences of the AQPs generated by Arabidopsis thaliana, poplar (Populus trichocarpa), and rubber (Hevea brasiliensis), the apple AQPs were each assigned membership of the five established AQP subfamilies, namely the PIPs (eleven members), the TIPs (thirteen members), the NIPs (eleven members), the SIPs (five members), and the XIPs (two members). The apple AQPs included asparagine-proline-alanine (NPA) motifs, an aromatic/arginine (ar/R) selectivity filter, and the Froger’s positions. The heterologous expression of MpPIP2;1 in A. thaliana was shown to enhance the level of tolerance exhibited against both drought and salinity.
Revisiting the classification of adhesion GPCRs

G protein–coupled receptors (GPCRs) are encoded by over 800 genes in the human genome. Motivated by different scientific rationales, the two classification systems that are mainly in use, the ABC and GRAFS systems, organize GPCRs according to their pharmacological features and phylogenetic relations, respectively. Within those systems, adhesion GPCRs (aGPCRs) constitute a group of over 30 mammalian homologs, most of which are still orphans with undefined activating signals and signal transduction properties. Previous efforts have further subdivided mammalian aGPCRs into nine subfamilies to indicate phylogenetic relationships. However, this subclassification scheme has shortcomings and inconsistencies that require attention. Here, we have reassessed the phylogenetic relationships of aGPCRs from vertebrate and invertebrate species. Our findings confirm that secretin receptor–like GPCRs most probably emerged from ancestral aGPCRs. We show that reassignment of several aGPCRs to families essentially requires input from functional data. Our analyses establish the need for introducing novel aGPCR subfamilies due to aGPCR sequences from invertebrate species that are not readily assignable to any existing subfamily. We conclude that the current classification systems ought to be updated to consider an unambiguous taxonomy of a hierarchically organized classification and pharmacological properties, and to accommodate phylogenetic affiliations between aGPCR genes within mammals and across the animal kingdom.
Nanoscale mapping of non-uniform heterogeneous nucleation kinetics mediated by surface chemistry.

Nucleation underlies the formation of many liquid-phase synthetic and natural materials with applications in materials chemistry, geochemistry, biophysics, and structural biology. Most liquid-phase nucleation processes are heterogeneous, occurring at specific nucleation sites at a solid-liquid interface; however, the chemical and topographical identity of these nucleation sites and how nucleation kinetics vary from site-to-site remains mysterious. Here we utilize in situ liquid cell electron microscopy to unveil counterintuitive nanoscale non-uniformities in heterogeneous nucleation kinetics on a macroscopically uniform solid-liquid interface. Time-resolved in situ electron microscopy imaging of silver nanoparticle nucleation at a water-silicon nitride interface showed apparently randomly located nucleation events at the interface. However, nanometric maps of local nucleation kinetics uncovered nanoscale interfacial domains with either slow or rapid nucleation. Interestingly, the interfacial domains vanished at high supersaturation ratio, giving way to rapid spatially uniform nucleation kinetics. Atomic force microscopy and nanoparticle labeling experiments revealed a topographically flat, chemically heterogeneous interface with nanoscale interfacial domains of functional groups similar in size to those observed in the nanometric nucleation maps. These results, along with a semi-quantitative nucleation model, indicate that a chemically non-uniform interface presenting different free energy barriers to heterogeneous nucleation underlies our observations of non-uniform nucleation kinetics. Overall, our results introduce a new imaging modality, nanometric nucleation mapping, and provide important new insights into the impact of surface chemistry on nanoscopic spatial variations in heterogeneous nucleation kinetics that have not been previously observed.
Arctium lappa Extract Suppresses Inflammation and Inhibits Melanoma Progression

Background: Arctium lappa has been used as popular medicinal herb and health supplement in Chinese societies. Bioactive components from A. lappa have attracted the attention of researchers due to their promising therapeutic effects. In this study, we investigated the effects of A. lappa hydroalcoholic extract (Alhe) during different models of inflammation, in vivo. Methods: The anti-inflammatory activity was evaluated through the air pouch model. For this, mice received an inflammatory stimulus with lipopolysaccharide (LPS) and were later injected with Alhe. To assess anti-tumoral activity, the animals were inoculated with B16F10 cells and injected with Alhe every 5 days, along the course of 30 days. Controls were submitted to the same conditions and injected with the vehicle. Peritoneal or air pouch fluids were collected to evaluate leukocyte counting or cellular activation via quantification of cytokines and nitric oxide. Results: Alhe injection reduced the neutrophil influx and production of inflammatory mediators in inflammatory foci after LPS or tumor challenges. Furthermore, Alhe injection reduced tumor growth and enhanced mice survival. Conclusions: Collectively, these data suggest that Alhe regulates immune cell migration and activation, which correlates with favorable outcome in mouse models of acute inflammation and melanoma progression.
Aristoteline, an Indole-Alkaloid, Induces Relaxation by Activating Potassium Channels and Blocking Calcium Channels in Isolated Rat Aorta

Alkaloids derived from plants have shown great medicinal benefits, and are often reported for their use in cardiovascular disease management. Aristotelia chilensis (Molina) Stuntz (Maqui) has shown important medicinal properties in traditional useage. In this study, we evaluated the effect of the indole-alkaloid aristoteline (ARI), isolated from leaves of Maqui, on vascular reactivity of isolated aortic rings from normotensive rats. ARI induced relaxation (100%) in a concentration-dependent manner in intact or denuded-endothelium aortic rings pre-contracted with phenylephrine (PE; 1 μM). However, a specific soluble guanylyl cyclase inhibitor (ODQ; 1 μM) significantly reduced the relaxation to ARI in aortic rings pre-contracted with PE. In the presence of ARI, the contraction induced by KCl or PE was significantly (p < 0.05) decreased. Interestingly, the potassium channel blockade with 10 μM BaCl2 (Kir), 10 μM glibenclamide (KATP), 1 mM tetraethylammonium (TEA; KCa1.1), or 1 mM 4-aminopyridine (4-AP; Kv) significantly (p < 0.05) reduced the ARI-induced relaxation. ARI significantly (p < 0.05) reduced the contractile response to agonist of CaV1.2 channels (Bay K8644; 10 nM), likely reducing the influx of extracellular calcium through plasma membrane. The mechanisms associated with this process suggest an activation of the potassium channels, a calcium-induced antagonism and endothelium independent vasodilation that possibly involves the nitric oxide-independent soluble guanylate cyclase pathway.
Influence of Smoking Status on Risk of Incident Heart Failure: A Systematic Review and Meta-Analysis of Prospective Cohort Studies

Smoking is a well-known risk factor for atherosclerotic cardiovascular disease. However, there are insufficient data regarding the predictive influence of smoking status on the risk of incident heart failure (HF). This study involved a systematic review and meta-analysis of prospective cohort studies to identify the association of smoking status with incident risk of HF. Peer-reviewed articles published in PubMed, Embase, Web of Science, Cochrane, and CINAHL up to May 2019 were identified. Seven studies, based on 42,759 participants and 4826 HF cases, were included. Pooled hazard ratios (HRs) and their 95% confidence intervals (CI) were estimated using the fixed effects model. Subgroup analyses were conducted to define possible sources of heterogeneity. Current smokers aged 18 years and over had a greater risk of HF incidence compared with non-smokers (never or former smokers) (HR = 1.609, 95% CI, 1.470–1.761). Additionally, former smokers had a greater risk of HF incidence compared with never smokers (HR = 1.209, 95% CI, 1.084–1.348). The present study highlighted that never smokers have more obvious cardiovascular benefits than current or former smokers. Therefore, health professionals should support cessation at the earliest among current smokers and encourage young people and non-smokers not to start smoking.
Bacterial dynamin‐like protein DynA mediates lipid and content mixing

Many bacterial species contain dynamin‐like proteins (DLPs). However, so far the functional mechanisms of bacterial DLPs are poorly understood. DynA in Bacillus subtilis is a 2‐headed DLP, mediating nucleotide‐independent membrane tethering in vitro and contributing to the innate immunity of bacteria against membrane stress and phage infection. Here, we employed content mixing and lipid mixing assays in reconstituted systems to study if DynA induces membrane full fusion, characterize its subunits in membrane fusion, and further test the possibility that GTP hydrolysis of DynA may act on the fusion‐through‐hemifusion pathway. Our results based on fluorescence resonance energy transfer indicated that DynA could induce aqueous content mixing even in the absence of GTP. Moreover, DynA‐induced membrane fusion in vitro is a thermo‐promoted slow process, and it has phospholipid and membrane curvature preferences. The D1 part of DynA is crucial for membrane binding and fusion, whereas D2 subunit plays a role in facilitating membrane fusion. Surprisingly, digestion of DynA mediated an instant rise of content exchange, supporting the assumption that disassembly of DynA is a driving force for fusion‐through‐hemifusion. DynA is a rare example of a membrane fusion catalyst that lacks a transmembrane domain and hence sets this system apart from well‐characterized fusion systems such as the soluble N‐ethyl maleimide sensitive factor attachment protein receptor complexes. —Guo, L. , Bramkamp, M. Bacterial dynamin‐like protein DynA mediates lipid and content mixing. FASEB J. 33, 11746–11757 (2019). www.fasebj. org
Schedulability analysis and low-criticality execution support for mixed-criticality real-time systems on uniprocessors

An increasing trend in safety-critical real-time systems is towards open computing environments, where applications with different degrees of importance are developed independently and integrated together on a single computing platform. In these systems, the correct behavior of some application (e.g., flight control) is more important (“critical”) to the overall safety of the system than that of another (e.g., in-flight cooling). Such a system that consolidates multiple functionalities of different importance or criticality is called a mixed-criticality system. Classical real-time scheduling theory can be used to guarantee the correctness of mixed-criticality systems, such that applications at different levels of criticality are verified with the corresponding degree of assurance. However, conservative resource reservations are often used to achieve this correctness at high assurance levels, and this results in severe under-utilization of system resources. To address this problem, real-time scheduling theory for mixed-criticality systems has been developed. The primary objective of this theory is to improve resource utilization, while still guaranteeing the safe execution of critical applications. Focusing on mixed-criticality scheduling theory, this dissertation addresses two important problems in the design of scheduling algorithms for mixed-criticality systems on uniprocessor platforms. First, we focus on the design of scheduling algorithms and analysis techniques for mixed-criticality systems that can successfully schedule a large proportion of such systems. Since mixed-criticality scheduling has been shown to be a NP-Hard problem, it is important to develop techniques that improve schedulability while still having efficient runtimes. To address this problem, we propose a demand bound function (DBF) based schedulability analysis technique for earliest deadline first (EDF) scheduling of mixed-criticality systems, and also develop a corresponding scheduling strategy. This analysis considers the different execution modes in a mixed-criticality system collectively, and is thus able to overcome some of the pessimistic assumptions in existing analysis. The proposed scheduling strategy is thus able to achieve much better schedulability, particularly for systems with more than two criticality levels, without any increase in the runtimes.
Tumor Priming by SMO Inhibition Enhances Antibody Delivery and Efficacy in a Pancreatic Ductal Adenocarcinoma Model

Despite frequent overexpression of numerous growth factor receptors by pancreatic ductal adenocarcinomas (PDAC), such as EGFR, therapeutic antibodies have not proven effective. Desmoplasia, hypovascularity, and hypoperfusion create a functional drug delivery barrier that contributes to treatment resistance. Drug combinations that target tumor/stroma interactions could enhance tumor deposition of therapeutic antibodies, although clinical trials have yet to support this strategy. We hypothesize that macromolecular or nanoparticulate therapeutic agents may best exploit stroma-targeting “tumor priming” strategies, based on the fundamental principles of the Enhanced Permeability and Retention phenomenon. Therefore, we investigated the molecular and pharmacologic tumor responses to NVP-LDE225, an SMO inhibitor of sonic hedgehog signaling (sHHI), of patient-derived xenograft models that recapitulate the desmoplasia and drug delivery barrier properties of PDAC. Short-term sHHI exposure mediated dose- and time-dependent changes in tumor microvessel patency, extracellular matrix architecture, and interstitial pressure, which waned with prolonged sHHI exposure, and increased nanoparticulate permeability probe deposition in multiple PDAC patient-derived xenograft isolates. During sHHI-mediated priming, deposition and intratumor distribution of both a nontargeted mAb and a mAb targeting EGFR, cetuximab, were enhanced. Sequencing the sHH inhibitor with cetuximab administration resulted in marked tumor growth inhibition compared with cetuximab alone. These studies suggest that PDAC drug delivery barriers confound efforts to employ mAb against targets in PDAC, and that short-term, intermittent exposure to stromal modulators can increase tumor cell exposure to therapeutic antibodies, improving their efficacy, and potentially minimize adverse effects that may accompany longer-term, continuous sHHI treatment.
Blast Induced Thresholds for Neuronal Networks (BITNeT)

16. SECURITY CLASSIFICATION OF: Connecting the human brain response to blast loading across multiple length scales – from synapses through to circuit pathways and neurobehavior – remains a key challenge for developing countermeasures to limit the effects of blast in the military. Our MURI intends to help fill this gap by conducting a series of experiments connecting cell, tissue, and animal responses, developing scaling rules to predict the human experience. We examine circuits constructed in vitro and in vivo, and assess the effect of blast-induced circuit changes on the subsequent neurological impairment. We recruited and collaborate with experts in each of the necessary knowledge 1. REPORT DATE (DD-MM-YYYY)
A review of dengue diagnostics and implications for surveillance and control

Abstract Dengue is the world’s most common arboviral infection, with almost 4 billion people estimated to be living at risk of dengue infection. A recently introduced vaccine is currently recommended only for seropositive individuals in a restricted age range determined by transmission intensity. With no effective dengue vaccine for the general population or any antiviral therapy, dengue control continues to rely heavily on vector control measures. Early and accurate diagnosis is important for guiding appropriate management and for disease surveillance to guide prompt dengue control interventions. However, major uncertainties exist in dengue diagnosis and this has important implications for all three. Dengue can be diagnosed clinically against predefined lists of signs and symptoms and by detection of dengue-specific antibodies, non-structural 1 antigen or viral RNA by reverse transcriptase–polymerase chain reaction. All of these methods have their limitations. This review aims to describe and quantify the advantages, uncertainties and variability of the various diagnostic methods used for dengue and discuss their implications and applications for dengue surveillance and control.
In vitro and in silico studies on membrane interactions of diverse Capsicum annuum flower γ‐thionin peptides

Thionins are small, cysteine‐rich peptides that play an important role in plant defense, primarily through their interactions with membranes. Eight novel γ‐thionin peptides (CanThio1‐8) were isolated from the flower of Capsicum annuum. Sequence analysis revealed that the peptides cluster into three groups. A representative peptide from each group (CanThio1, 2, and 3) was used for experimental characterization. Interestingly, peptides were found to possess some cytotoxic activity against normal human embryonic kidney cell line but higher cytotoxicity against cancer cell line MCF‐7. CanThio3 peptide was chosen as a representative peptide to study the molecular mechanism of action on membranes. Microsecond timescale atomistic simulations of CanThio3 were performed in the presence of a POPC (1‐palmitoyl‐2‐oleoyl‐sn‐glycero‐3‐phosphocholine) lipid bilayer. Simulations revealed that CanThio3 interacts with the bilayer and causes lipid thinning in the vicinity. Nonpolar amino acids specific to the α‐core region of CanThio3 along with nonpolar residues in the γ‐core region are seen to interact with the lipid tails. The differences in the amino acid sequence of CanThio peptides in these regions explain the variability in cytotoxic activities. In summary, our results demonstrate the membrane‐mediated activity of a novel series of γ‐thionin peptides from C. annuum.