Liver-specific Mettl14 deletion induces nuclear heterotypia and dysregulates RNA export machinery.

Modification of RNA with N6-methyladenosine (m6A) has gained attention in recent years as a general mechanism of gene regulation. In the liver, m6A, along with its associated machinery, has been studied as a potential biomarker of disease and cancer, with impacts on metabolism, cell cycle regulation, and pro-cancer state signaling. However these observational data have yet to be causally examined in vivo. For example, neither perturbation of the key m6A writers Mettl3 and Mettl14, nor the m6A readers Ythdf1 and Ythdf2 have been thoroughly mechanistically characterized in vivo as they have been in vitro. To understand the functions of these machineries, we developed mouse models and found that deleting Mettl14 led to progressive liver injury characterized by nuclear heterotypia, with changes in mRNA splicing, processing and export leading to increases in mRNA surveillance and recycling.

From ground to gut: Evaluating the human health risk of potentially toxic elements in soil, groundwater, and their uptake by Cocos nucifera in arsenic-contaminated environments.

This study aimed to gauge the toxicity of potentially toxic elements (PTEs) in coconut crops cultivated in arsenic-contaminated areas while offering a global perspective encompassing more than 100 impacted countries. The current investigation provides crucial insights into the assessment of PTEs pollution using the Bioaccumulation factor, Geo-accumulation index, Potential ecological risk index, Hazardous index, and Lifetime cancer risk (LCR) and highlights the potential human health risks posed by contaminated food, water, and soil. From 22 severely polluted sites in West Bengal, India, soil, groundwater (GW), and coconut water (CW) samples were collected, acidified, and digested using microwave digestion, for PTEs quantification using inductively coupled plasma-optical emission spectroscopy (ICP-OES). Results revealed that despite high concentrations of arsenic in soils (4.6 ± 3.4 mg kg-1), and GW (22.2 ± 150.9 µg L-1), CW (0.7 ± 3.1 µg L-1) levels were within permissible limits. Groups of PTEs with comparable sources and distributions were discovered through Principal Component Analysis (PCA). A speciation diagram was used to predict the prevalence of arsenic species in all three matrices. The Hazardous Index (HI < 1) indicated no probability of non-carcinogenic diseases for children and adults in all the compartments. However, exposure to GW and soil contaminated with Cr, As, and Cd by children (9.02 × 10-13 to 2.77 × 10-4) and adults (6.51 × 10-14 to 1.18 × 10-4) would increase their susceptibility to cancer (LCR >10-6). The study concluded that moderate lifetime consumption of CW is safe and has no significant impact on healthy individuals. Additionally, CW is a rich source of essential micronutrients such as Zn, Fe, Mn, and B. Overall, the findings of this study could help in developing appropriate strategies for reducing PTEs contamination and protecting human health.

Region-specific reversal of epidermal planar polarity in the rosette fancy mouse.

The planar cell polarity (PCP) pathway collectively orients cells with respect to a body axis. Hair follicles of the murine epidermis provide a striking readout of PCP activity in their uniform alignment across the skin. Here, we characterize, from the molecular to tissue-scale, PCP establishment in the rosette fancy mouse, a natural variant with posterior-specific whorls in its fur, to understand how epidermal polarity is coordinated across the tissue. We find that rosette hair follicles emerge with reversed orientations specifically in the posterior region, creating a mirror image of epidermal polarity. The rosette trait is associated with a missense mutation in the core PCP gene Fzd6, which alters a consensus site for N-linked glycosylation, inhibiting its membrane localization. Unexpectedly, the Fzd6 trafficking defect does not block asymmetric localization of the other PCP proteins. Rather, the normally uniform axis of PCP asymmetry rotates where the PCP-directed cell movements that orient follicles are reversed, suggesting the PCP axis rotates 180°. Collectively, our multiscale analysis of epidermal polarity reveals PCP patterning can be regionally decoupled to produce posterior whorls in the rosette fancy mouse.

Region-specific reversal of epidermal planar polarity in the fancy rosette mouse.

The planar cell polarity (PCP) pathway collectively orients thousands of cells with respect to a body axis to direct cellular behaviors that are essential for embryonic morphogenesis. Hair follicles of the murine epidermis provide a striking readout of PCP activity in their uniform alignment along the entire skin surface. Here, we characterize, from the molecular to tissue-scale, PCP establishment in the rosette fancy mouse, a natural variant with posterior-specific whorls in its fur, to understand how epidermal polarity is coordinated across the tissue. We find that embryonic hair follicles of rosette mutants emerge with reversed orientations specifically in the posterior region, creating a mirror image of epidermal polarity. The rosette trait is associated with a missense mutation in the core PCP gene Fzd6 , which alters a consensus site for N-linked glycosylation and inhibits its membrane localization. Unexpectedly, this defect in Fzd6 trafficking, observed across the entire dorsal epidermis, does not interfere with the ability of other core PCP proteins to localize asymmetrically. Rather, the normally uniform axis of PCP asymmetry is disrupted and rotated in the posterior region such that polarity is reflected on either side of a transition zone. The result is a reversal of polarized cell movements that orient nascent follicles, specifically in the posterior of the embryo. Collectively, our multiscale analysis of epidermal polarity reveals PCP patterning can be regionally decoupled to produce the unique posterior whorls of the fancy rosette mouse. Summary: Region-specific rotation of the Planar Cell Polarity axis reverses posterior hair follicles in the fancy rosette mouse.

Targeted viral adaptation generates a simian-tropic hepatitis B virus that infects marmoset cells.

Hepatitis B virus (HBV) only infects humans and chimpanzees, posing major challenges for modeling HBV infection and chronic viral hepatitis. The major barrier in establishing HBV infection in non-human primates lies at incompatibilities between HBV and simian orthologues of the HBV receptor, sodium taurocholate co-transporting polypeptide (NTCP). Through mutagenesis analysis and screening among NTCP orthologues from Old World monkeys, New World monkeys and prosimians, we determined key residues responsible for viral binding and internalization, respectively and identified marmosets as a suitable candidate for HBV infection. Primary marmoset hepatocytes and induced pluripotent stem cell-derived hepatocyte-like cells support HBV and more efficient woolly monkey HBV (WMHBV) infection. Adapted chimeric HBV genome harboring residues 1-48 of WMHBV preS1 generated here led to a more efficient infection than wild-type HBV in primary and stem cell derived marmoset hepatocytes. Collectively, our data demonstrate that minimal targeted simianization of HBV can break the species barrier in small NHPs, paving the path for an HBV primate model.

Mechanical Behavior of Austenitic Steel under Multi-Axial Cyclic Loading.

Low-nickel austenitic steel is subjected to high-pressure torsion fatigue (HPTF) loading, where a constant axial compression is overlaid with a cyclic torsion. The focus of this work lies on investigating whether isotropic J2 plasticity or crystal plasticity can describe the mechanical behavior during HPTF loading, particularly focusing on the axial creep deformation seen in the experiment. The results indicate that a J2 plasticity model with an associated flow rule fails to describe the axial creep behavior. In contrast, a micromechanical model based on an empirical crystal plasticity law with kinematic hardening described by the Ohno-Wang rule can match the HPTF experiments quite accurately. Hence, our results confirm the versatility of crystal plasticity in combination with microstructural models to describe the mechanical behavior of materials under reversing multiaxial loading situations.

Structural features stabilized by divalent cation coordination within hepatitis E virus ORF1 are critical for viral replication.

Hepatitis E virus (HEV) is an RNA virus responsible for over 20 million infections annually. HEV's open reading frame (ORF)1 polyprotein is essential for genome replication, though it is unknown how the different subdomains function within a structural context. Our data show that ORF1 operates as a multifunctional protein, which is not subject to proteolytic processing. Supporting this model, scanning mutagenesis performed on the putative papain-like cysteine protease (pPCP) domain revealed six cysteines essential for viral replication. Our data are consistent with their role in divalent metal ion coordination, which governs local and interdomain interactions that are critical for the overall structure of ORF1; furthermore, the 'pPCP' domain can only rescue viral genome replication in trans when expressed in the context of the full-length ORF1 protein but not as an individual subdomain. Taken together, our work provides a comprehensive model of the structure and function of HEV ORF1.

Micromechanical Modeling of AlSi10Mg Processed by Laser-Based Additive Manufacturing: From as-Built to Heat-Treated Microstructures.

The unique microstructure of the alloy AlSi10Mg produced by the laser-based powder bed fusion of metals (PBF-LB/M) provides high-strength and high-strain-hardening capabilities of the material. The microstructure and mechanical properties of 3D-printed, i.e., additively manufactured, AlSi10Mg are significantly altered by post-building heat-treatment processes applied in order to tailor the final properties of the parts. Using an accurate computational model to predict and improve the mechanical performance of 3D-printed samples considering their microstructural features can accelerate their employment in envisaged applications. The present study aims to investigate the correlation between microstructural features and the mechanical behavior of as-built, direct-aged, and T6 heat-treated samples of PBF-LB/M AlSi10Mg under tensile loading using experiment and microstructure-sensitive modeling approaches. Nanoindentation tests are used to calibrate the parameters of the constitutive models for the Al and Si-rich phases. The experimental investigations revealed that heat treatment significantly changes the sub-grain morphology of the Si-rich phase, and this can have a considerable effect on the mechanical behavior of the components. The effect of the modeling of the Si-rich phase in the representative volume elements on the prediction of mechanical behavior is investigated using the J2 plasticity model. The combination of the crystal plasticity model for Al and the J2 plasticity model for the Si-rich phase is used to predict the tensile properties of the as-built and heat-treated states. The predicted results are in good agreement with the experimental results. This approach can be used to understand the microstructure-property relationship of PBF-LB/M AlSi10Mg and eventually tailor heat treatment for PBF-LB/M AlSi10Mg based on the requirement of the application.

Food gels: principles, interaction mechanisms and its microstructure.

Food hydrogels are important materials having great scientific interest due to biocompatibility, safety and environment-friendly characteristics. In the food industry, hydrogels are widely used due to their three-dimensional crosslinked networks. Furthermore, they have attracted great attention due to their wide range of applications in the food industry, such as fat replacers, encapsulating agents, target delivery vehicles, and many more. In addition to basic and recent knowledge on food hydrogels, this review exclusively focuses on sensorial perceptions, nutritional significance, body interactions, network structures, mechanical properties, and potential hydrogel applications in food and food-based matrices. Additionally, this review highlights the structural design of hydrogels, which provide the forward-looking idea for future applications of food hydrogels (e.g., 3D or 4D printing).

Generation and characterization of genetically and antigenically diverse infectious clones of dengue virus serotypes 1-4.

Dengue is caused by four genetically distinct viral serotypes, dengue virus (DENV) 1-4. Following transmission by Aedes mosquitoes, DENV can cause a broad spectrum of clinically apparent disease ranging from febrile illness to dengue hemorrhagic fever and dengue shock syndrome. Progress in the understanding of different dengue serotypes and their impacts on specific host-virus interactions has been hampered by the scarcity of tools that adequately reflect their antigenic and genetic diversity. To bridge this gap, we created and characterized infectious clones of DENV1-4 originating from South America, Africa, and Southeast Asia. Analysis of whole viral genome sequences of five DENV isolates from each of the four serotypes confirmed their broad genetic and antigenic diversity. Using a modified circular polymerase extension reaction (CPER), we generated de novo viruses from these isolates. The resultant clones replicated robustly in human and insect cells at levels similar to those of the parental strains. To investigate in vivo properties of these genetically diverse isolates, representative viruses from each DENV serotype were administered to NOD Rag1-/-, IL2rgnull Flk2-/- (NRGF) mice, engrafted with components of a human immune system. All DENV strains tested resulted in viremia in humanized mice and induced cellular and IgM immune responses. Collectively, we describe here a workflow for rapidly generating de novo infectious clones of DENV - and conceivably other RNA viruses. The infectious clones described here are a valuable resource for reverse genetic studies and for characterizing host responses to DENV in vitro and in vivo.

The human microbiome encodes resistance to the antidiabetic drug acarbose.

The human microbiome encodes a large repertoire of biochemical enzymes and pathways, most of which remain uncharacterized. Here, using a metagenomics-based search strategy, we discovered that bacterial members of the human gut and oral microbiome encode enzymes that selectively phosphorylate a clinically used antidiabetic drug, acarbose1,2, resulting in its inactivation. Acarbose is an inhibitor of both human and bacterial α-glucosidases3, limiting the ability of the target organism to metabolize complex carbohydrates. Using biochemical assays, X-ray crystallography and metagenomic analyses, we show that microbiome-derived acarbose kinases are specific for acarbose, provide their harbouring organism with a protective advantage against the activity of acarbose, and are widespread in the microbiomes of western and non-western human populations. These results provide an example of widespread microbiome resistance to a non-antibiotic drug, and suggest that acarbose resistance has disseminated in the human microbiome as a defensive strategy against a potential endogenous producer of a closely related molecule.

New mouse models for high resolution and live imaging of planar cell polarity proteins in vivo.

The collective polarization of cellular structures and behaviors across a tissue plane is a near universal feature of epithelia known as planar cell polarity (PCP). This property is controlled by the core PCP pathway, which consists of highly conserved membrane-associated protein complexes that localize asymmetrically at cell junctions. Here, we introduce three new mouse models for investigating the localization and dynamics of transmembrane PCP proteins: Celsr1, Fz6 and Vangl2. Using the skin epidermis as a model, we characterize and verify the expression, localization and function of endogenously tagged Celsr1-3xGFP, Fz6-3xGFP and tdTomato-Vangl2 fusion proteins. Live imaging of Fz6-3xGFP in basal epidermal progenitors reveals that the polarity of the tissue is not fixed through time. Rather, asymmetry dynamically shifts during cell rearrangements and divisions, while global, average polarity of the tissue is preserved. We show using super-resolution STED imaging that Fz6-3xGFP and tdTomato-Vangl2 can be resolved, enabling us to observe their complex localization along junctions. We further explore PCP fusion protein localization in the trachea and neural tube, and discover new patterns of PCP expression and localization throughout the mouse embryo.

Light promoted brown staining of protoplasm by Ag+ is ideal to test wheat pollen viability rapidly.

Pollen viability is crucial for wheat breeding programs. The unique potential of the protoplasm of live cells to turn brown due to the synthesis of silver nanoparticles (AgNPs) through rapid photoreduction of Ag+, was exploited for testing wheat pollen viability. Ag+-viability test medium (consisting of 0.5 mM AgNO3 and 300 mM KNO3) incubated with wheat pollen turned brown within 2 min under intense light (~600 µmol photon flux density m-2s-1), but not in dark. The brown medium displayed AgNPs-specific surface plasmon resonance band in its absorption spectrum. Light microscopic studies showed the presence of uniformly stained brown protoplasm in viable pollen incubated with Ag+-viability medium in the presence of light. Investigations with transmission electron microscope coupled with energy dispersive X-ray established the presence of distinct 5-35 nm NPs composed of Ag. Powder X-ray diffraction analysis revealed that AgNPs were crystalline and biphasic composed of Ag0 and Ag2O. Conversely, non-viable pollen and heat-killed pollen did not turn brown on incubation with Ag+-medium in light. We believe that the viable wheat pollen turn brown rapidly by bio-transforming Ag+ to AgNPs through photoreduction. Our findings furnish a novel simplest and rapid method for testing wheat pollen viability.

Repositioning of migrated self-expanding metallic tracheobronchial stent: predictors of a successful maneuver and its impact on survival.

BACKGROUND: Endobronchial stents that are used to treat airway obstruction may migrate over time. These stents can be repositioned. However, not much has been reported about this technique. We retrospectively reviewed our experience with self-expanding metallic stents (SEMS) and attempted to determine-(I) factors related to successful stent repositioning; (II) determine its impact on survival. METHODS: Demographic, medical history, and stent-related procedure factors were extracted from the electronic health record. Primary outcomes were bronchial stent repositioning success and survival (days until death). As validation of successful repositioning, the durations of successful and failed repositioning procedures were compared using an independent t-test. RESULTS: Seventy-six patients underwent stent repositioning, of which, 55.3% (n=42) were successfully repositioned. The probability of success in repositioning procedures was accounted for by patient sex, stent location, and stent diameter. Females were more likely to have a successful repositioning compared to males. Stent repositioning in the LMS was more likely to be successful and stents larger in diameter tended to increase the likelihood of successful repositioning. Long-term survival was higher for those who had a successful procedure. Stent location and disease subgroups predicted average length of survival. CONCLUSIONS: Repositioning of migrated stents can be successfully performed regardless of the reasons for initial placement, duration of stenting and degree of original obstruction. Larger stents are easier to reposition and so were stents in the left main stem (LMS) airway. A successful stent repositioning maneuver improved long-term survival although did not have any impact survival in the immediate post-procedural period.

Optimized reconstruction of the crystallographic orientation density function based on a reduced set of orientations.

Crystallographic textures, as they develop for example during cold forming, can have a significant influence on the mechanical properties of metals, such as plastic anisotropy. Textures are typically characterized by a non-uniform distribution of crystallographic orientations that can be measured by diffraction experiments like electron backscatter diffraction (EBSD). Such experimental data usually contain a large number of data points, which must be significantly reduced to be used for numerical modeling. However, the challenge in such data reduction is to preserve the important characteristics of the experimental data, while reducing the volume and preserving the computational efficiency of the numerical model. For example, in micromechanical modeling, representative volume elements (RVEs) of the real microstructure are generated and the mechanical properties of these RVEs are studied by the crystal plasticity finite element method. In this work, a new method is developed for extracting a reduced set of orientations from EBSD data containing a large number of orientations. This approach is based on the established integer approximation method and it minimizes its shortcomings. Furthermore, the L 1 norm is applied as an error function; this is commonly used in texture analysis for quantitative assessment of the degree of approximation and can be used to control the convergence behavior. The method is tested on four experimental data sets to demonstrate its capabilities. This new method for the purposeful reduction of a set of orientations into equally weighted orientations is not only suitable for numerical simulation but also shows improvement in results in comparison with other available methods.

Change in urodynamic pattern and incidence of urinary tract infection in patients with traumatic spinal cord injury practicing clean self-intermittent catheterization.

Objective: To observe changes in cystometric parameters in individuals with spinal cord injury (SCI) with neurogenic bladder practicing clean intermittent self-catheterization (CIC) and incidence of urinary tract infection (UTI) in such patients.Design: Prospective, observational study.Setting: Tertiary Urban Rehabilitation Hospital.Participants: Persons with neurogenic bladder caused by traumatic SCI and practicing CIC.Interventions: Clinical evaluation, complete urine analysis, urine culture and sensitivity, ultrasonography of the abdomen and urodynamic study were evaluated at baseline and at follow-up (6 months to 1 year).Outcome Measures: Detrusor pattern, cystometric capacity, detrusor compliance, detrusor leak point pressure, residual urine, incidence of UTI.Results: Thirty-one participants were included in the study. The baseline cystometric study showed that 15 had overactive detrusor and 16 had detrusor areflexia. The mean cystometric capacity decreased significantly between baseline and follow-up in both the groups but remained within the normal threshold limit, decline being more marked in the overactive detrusor group, who also had more marked decrease in compliance. Mean detrusor leak point pressure was below 40 cm H2O in all participants in both groups at baseline and follow-up. Mean residual urine improved at follow-up in both groups. Incidence of UTI was 2.29 episodes per patient per year, and more frequent in the overactive detrusor group. Escherichia coli was the causative agent in 45%.Conclusion: The cystometric capacity and compliance decreased significantly though patients were doing regular CIC and managed on antimuscarinics for detrusor overactivity (DO). UTI is more common in individuals with SCI with DO and E. coli is the most common cause of UTI.

A metagenomic strategy for harnessing the chemical repertoire of the human microbiome.

Extensive progress has been made in determining the effects of the microbiome on human physiology and disease, but the underlying molecules and mechanisms governing these effects remain largely unexplored. Here, we combine a new computational algorithm with synthetic biology to access biologically active small molecules encoded directly in human microbiome-derived metagenomic sequencing data. We discover that members of a clinically used class of molecules are widely encoded in the human microbiome and that they exert potent antibacterial activities against neighboring microbes, implying a possible role in niche competition and host defense. Our approach paves the way toward a systematic unveiling of the chemical repertoire encoded by the human microbiome and provides a generalizable platform for discovering molecular mediators of microbiome-host and microbiome-microbiome interactions.

Comparison of Endobronchial Ultrasound-guided Transbronchial Needle Aspiration With Stylet Retracted Partially Versus Completely for Molecular Testing.

BACKGROUND: Endobronchial ultrasound-guided transbronchial needle aspiration (EBUS-TBNA) is considered to be the initial diagnostic modality for most patients with lung cancer. However, the optimal technique for maximizing yield continues to vary in the real-world setting. OBJECTIVES: To evaluate the diagnostic yield of EBUS-TBNA with capillary sampling compared with complete stylet removal for molecular testing. METHODS: Retrospective study, data from patients between January to May 2017 with indication of EBUS-TBNA whom ancillary testing, that is, next-generation sequencing, anaplastic lymphoma kinase (ALK), and programed death ligand-1 (PD-L1) expression was reviewed. The yield of 2 techniques, stylet retracted halfway (group 1) versus complete retraction (group 2), was compared. RESULTS: A total of 24/27 (88.88%) samples were adequate for next-generation sequencing analysis in group 1 and 21/23 (91.30%) in group 2. For other molecular analyses, 24/27 (88.88%) samples in group 1 and 20/23 (86.95%) samples in group 2 were adequate for ALK analysis. 23/27 (85.18%) samples for group 1 and 20/23 (86.95%) samples for group 2 were adequate for PD-L1 analysis. Positive expression of PD-L1>50% was achieved in 9/23 (39.13%) of group 1 and 5/20 (25%) of group 2. There was no statistical difference in the yield between the 2 groups. CONCLUSION: EBUS-TBNA using either capillary sampling or complete stylet removal are effective and has a high proportion of satisfactory results for ancillary testing.

Diagnostic Yield of the Virtual Bronchoscopic Navigation System Guided Sampling of Peripheral Lung Lesions using Ultrathin Bronchoscope and Protected Bronchial Brush.

OBJECTIVES: The use of an ultrathin bronchoscope (UB) to diagnose peripheral pulmonary lesions is described. A virtual bronchoscopic navigation system was used to direct the ultrathin scope to the nodule. One of the constraints of this technique was the inability to confirm the target lesion position during biopsy by using a conventional linear endobronchial ultrasound probe, since the probe does not fit into a 1.2 mm working channel of this bronchoscope. The aim of the study was to review our institutional experience with the use of a UB for sampling peripheral pulmonary lesions using the transbronchial brush guided by virtual bronchoscopy. We describe a technique wherein we attempt to brush all the visible bronchial sub-segments once the bronchoscope has reached close to the nodule. MATERIALS AND METHODS: In total, 52 patients underwent the procedure between 2010 and 2017. A multiplanar computed tomography (CT) scan of the chest was obtained and subsequently uploaded to the Lung Point Virtual bronchoscopy navigation software. The UB was parked close to the lesion. All visible airway branches were then brushed using a protected bronchial brush. The data were retrospectively abstracted from the electronic medical records using standardized forms. RESULTS: A total of 52 lesions (40 solid, 8 part-solid, 3 cavitary, and 1 ground-glass) were sampled using a transbronchial brush (median, 2; range, 1-8). Twenty-four lesions were under 2 cm in size. The overall success rates were 67.3%. The average diameter of nodules was 2.7±1.01 cm; 65% lesions were in the outer-third of the lungs. The cancer-specific sensitivity was 72.5%. The presence of bronchus sign; location of the lesion; and the characteristics, size, and stage of cancer did not have any impact on the diagnostic yield. CONCLUSION: Virtual bronchoscopy-guided ultrathin bronchoscopy with bronchial brushing is safe and has a diagnostic yield comparable to other described techniques for evaluating peripheral pulmonary nodules.