Inhibition of S6K lowers age-related inflammation and increases lifespan through the endolysosomal system.

Suppression of target of rapamycin complex 1 (TORC1) by rapamycin ameliorates aging in diverse species. S6 kinase (S6K) is an essential mediator, but the mechanisms involved are unclear. Here we show that activation of S6K specifically in Drosophila fat-body blocked extension of lifespan by rapamycin, induced accumulation of multilamellar lysosomes and blocked age-associated hyperactivation of the NF-κB-like immune deficiency (IMD) pathway, indicative of reduced inflammaging. Syntaxin 13 mediated the effects of TORC1-S6K signaling on lysosome morphology and inflammaging, suggesting they may be linked. Inflammaging depended on the IMD receptor regulatory isoform PGRP-LC, and repression of the IMD pathway from midlife extended lifespan. Age-related inflammaging was higher in females than in males and was not lowered in males by rapamycin treatment or lowered S6K. Rapamycin treatment also elevated Syntaxin 12/13 levels in mouse liver and prevented age-related increase in noncanonical NF-κB signaling, suggesting that the effect of TORC1 on inflammaging is conserved from flies to mammals.

Repeat length of C9orf72-associated glycine-alanine polypeptides affects their toxicity.

G4C2 hexanucleotide repeat expansions in a non-coding region of the C9orf72 gene are the most common cause of familial amyotrophic lateral sclerosis (ALS) and frontotemporal dementia (FTD). G4C2 insertion length is variable, and patients can carry up to several thousand repeats. Dipeptide repeat proteins (DPRs) translated from G4C2 transcripts are thought to be a main driver of toxicity. Experiments in model organisms with relatively short DPRs have shown that arginine-rich DPRs are most toxic, while polyGlycine-Alanine (GA) DPRs cause only mild toxicity. However, GA is the most abundant DPR in patient brains, and experimental work in animals has generally relied on the use of low numbers of repeats, with DPRs often tagged for in vivo tracking. Whether repeat length or tagging affect the toxicity of GA has not been systematically assessed. Therefore, we generated Drosophila fly lines expressing GA100, GA200 or GA400 specifically in adult neurons. Consistent with previous studies, expression of GA100 and GA200 caused only mild toxicity. In contrast, neuronal expression of GA400 drastically reduced climbing ability and survival of flies, indicating that long GA DPRs can be highly toxic in vivo. This toxicity could be abolished by tagging GA400. Proteomics analysis of fly brains showed a repeat-length-dependent modulation of the brain proteome, with GA400 causing earlier and stronger changes than shorter GA proteins. PolyGA expression up-regulated proteins involved in ER to Golgi trafficking, and down-regulated proteins involved in insulin signalling. Experimental down-regulation of Tango1, a highly conserved regulator of ER-to Golgi transport, partially rescued GA400 toxicity, suggesting that misregulation of this process contributes to polyGA toxicity. Experimentally increasing insulin signaling also rescued GA toxicity. In summary, our data show that long polyGA proteins can be highly toxic in vivo, and that they may therefore contribute to ALS/FTD pathogenesis in patients.

Toxicity of C9orf72-associated dipeptide repeat peptides is modified by commonly used protein tags.

Hexanucleotide repeat expansions in the C9orf72 gene are the most prevalent genetic cause of amyotrophic lateral sclerosis and frontotemporal dementia. Transcripts of the expansions are translated into toxic dipeptide repeat (DPR) proteins. Most preclinical studies in cell and animal models have used protein-tagged polyDPR constructs to investigate DPR toxicity but the effects of tags on DPR toxicity have not been systematically explored. Here, we used Drosophila to assess the influence of protein tags on DPR toxicity. Tagging of 36 but not 100 arginine-rich DPRs with mCherry increased toxicity, whereas adding mCherry or GFP to GA100 completely abolished toxicity. FLAG tagging also reduced GA100 toxicity but less than the longer fluorescent tags. Expression of untagged but not GFP- or mCherry-tagged GA100 caused DNA damage and increased p62 levels. Fluorescent tags also affected GA100 stability and degradation. In summary, protein tags affect DPR toxicity in a tag- and DPR-dependent manner, and GA toxicity might be underestimated in studies using tagged GA proteins. Thus, including untagged DPRs as controls is important when assessing DPR toxicity in preclinical models.

Research on Automatic Identification and Rating of Ferrite-Pearlite Grain Boundaries Based on Deep Learning.

Grain size has a significant effect on the mechanical properties of metals. It is very important to accurately rate the grain size number of steels. This paper presents a model for automatic detection and quantitative analysis of the grain size of ferrite-pearlite two-phase microstructure to segment ferrite grain boundaries. In view of the challenging problem of hidden grain boundaries in pearlite microstructure, the number of hidden grain boundaries is inferred by detecting them with the confidence of average grain size. The grain size number is then rated using the three-circle intercept procedure. The results show that grain boundaries can be accurately segmented by using this procedure. According to the rating results of grain size number of four types of ferrite-pearlite two-phase microstructure samples, the accuracy of this procedure is greater than 90%. The grain size rating results deviate from those calculated by experts using the manual intercept procedure by less than Grade 0.5-the allowable detection error specified in the standard. In addition, the detection time is shortened from 30 min of the manual intercept procedure to 2 s. The procedure presented in this paper allows automatic rating of grain size number of ferrite-pearlite microstructure, thereby effectively improving the detection efficiency and reducing the labor intensity.

Research on Intelligent Identification and Grading of Nonmetallic Inclusions in Steels Based on Deep Learning.

Non-metallic inclusions are unavoidable defects in steel, and their type, quantity, size, and distribution have a great impact on the quality of steel. At present, non-metallic inclusions are mainly detected manually, which features high work intensity, low efficiency, proneness to misjudgment, and low consistency of results. In this paper, based on deep neural network algorithm, a small number of manually labeled, low-resolution metallographic images collected by optical microscopes are used as the dataset for intelligent boundary extraction, classification, and rating of non-metallic inclusions. The training datasets are cropped into those containing only a single non-metallic inclusion to reduce the interference of background information and improve the accuracy. To deal with the unbalanced distribution of each category of inclusions, the reweighting cross entropy loss and focal loss are respectively used as the category prediction loss and boundary prediction loss of the DeepLabv3+ semantic segmentation model. Finally, the length and width of the minimum enclosing rectangle of the segmented inclusions are measured to calculate the grade of inclusions. The resulting accuracy is 90.34% in segmentation and 90.35% in classification. As is verified, the model-based rating results are consistent with those of manual labeling. For a single sample, the detection time is reduced from 30 min to 15 s, significantly improving the detection efficiency.

Effects of Cold Rolling on the Microstructure and Corrosion Resistance of the Double-Glow Plasma Ni-Cr Alloying Layer on Q235 Steel.

A Ni-Cr alloyed layer was prepared on the surface of Q235 steel using double-glow plasma surface alloying (DGPSA) technology and the alloyed layer was cold-rolled with different deformation rates. The microstructure, composition distribution and phase composition of the alloyed layer were characterized using a scanning electron microscope (SEM), an energy dispersive spectrometer (EDS), X-ray diffraction (XRD) and an electrochemical workstation. On this basis, the corrosion resistance of the alloyed layer was analyzed. The results showed that a Ni-Cr alloyed layer formed on the surface of Q235 steel following double-glow plasma nickel-chromium alloying. The alloy elements of Ni and Cr were distributed in a gradient from the outside to the inside and the main phases were FeCr0.29Ni0.16C0.06, Cr23C6 and γ solid solution. The alloyed layer, once cold-rolled with different deformation rates, underwent synchronous plastic deformation with the substrate, with no fracturing and spalling. The self-corrosion potential of the cold-rolled specimens in 5% H2SO4 and 3.5% NaCl solution is close to that of 304L stainless steel, and the corrosion currents are much lower. The corrosion resistance of the cold-rolled specimens is comparable to the original specimens, with no significant changes.

Hot Deformation Behavior of Cu-Sn-La Polycrystalline Alloy Prepared by Upcasting.

In this study, the hot deformation of a Cu-0.55Sn-0.08La (wt.%) alloy was studied using a Gleeble-3180 testing machine at deformation temperatures of 400-700 °C and various strain rates. The stress-strain curve showed that the hot deformation behavior of the Cu-0.55Sn-0.08La (wt.%) alloy was significantly affected by work hardening, dynamic recovery, and dynamic recrystallization. The activation energy Q was 261.649 kJ·mol-1 and hot compression constitutive equation was determined as ε=sinh0.00651σ10.2378∙exp33.6656-261.649RT. The microstructural evolution of the alloy during deformation at 400 °C revealed the presence of both slip and shear bands in the grains. At 700 °C, dynamic recrystallization grains were observed, but recrystallization was incomplete. In summary, these results provide the theoretical basis for the continuous extrusion process of alloys with promising application prospects in the future.

Enhanced Replication of Virulent Newcastle Disease Virus in Chicken Macrophages Is due to Polarized Activation of Cells by Inhibition of TLR7.

Newcastle disease (ND), caused by infections with virulent strains of Newcastle disease virus (NDV), is one of the most important infectious disease affecting wild, peridomestic, and domestic birds worldwide. Vaccines constructed from live, low-virulence (lentogenic) viruses are the most accepted prevention and control strategies for combating ND in poultry across the globe. Avian macrophages are one of the first cell lines of defense against microbial infection, responding to signals in the microenvironment. Although macrophages are considered to be one of the main target cells for NDV infection in vivo, very little is known about the ability of NDV to infect chicken macrophages, and virulence mechanisms of NDV as well as the polarized activation patterns of macrophages and correlation with viral infection and replication. In the present study, a cell culture model (chicken bone marrow macrophage cell line HD11) and three different virulence and genotypes of NDV (including class II virulent NA-1, class II lentogenic LaSota, and class I lentogenic F55) were used to solve the above underlying questions. Our data indicated that all three NDV strains had similar replication rates during the early stages of infection. Virulent NDV titers were shown to increase compared to the other lentogenic strains, and this growth was associated with a strong upregulation of both pro-inflammatory M1-like markers/cytokines and anti-inflammatory M2-like markers/cytokines in chicken macrophages. Virulent NDV was found to block toll-like receptor (TLR) 7 expression, inducing higher expression of type I interferons in chicken macrophages at the late stage of viral infection. Only virulent NDV replication can be inhibited by pretreatment with TLR7 ligand. Overall, this study demonstrated that virulent NDV activates a M1-/M2-like mixed polarized activation of chicken macrophages by inhibition of TLR7, resulting in enhanced replication compared to lentogenic viruses.

Dispersal and Transmission of Avian Paramyxovirus Serotype 4 among Wild Birds and Domestic Poultry.

Avian paramyxovirus serotype 4 (APMV-4) is found sporadically in wild birds worldwide, and it is an economically important poultry pathogen. Despite the existence of several published strains, very little is known about the distribution, host species, and transmission of APMV-4 strains. To better understand the relationships among these factors, we conducted an APMV-4 surveillance of wild birds and domestic poultry in six provinces of China suspected of being intercontinental flyways and sites of interspecies transmission. APMV-4 surveillance was conducted in 9,160 wild birds representing seven species, and 1,461 domestic poultry in live bird markets (LMBs) from December 2013 to June 2016. The rate of APMV-4 isolation was 0.10% (11/10,621), and viruses were isolated from swan geese, bean geese, cormorants, mallards, and chickens. Sequencing and phylogenetic analyses of the 11 isolated viruses indicated that all the isolates belonging to genotype I were epidemiologically connected with wild bird-origin viruses from the Ukraine and Italy. Moreover, chicken-origin APMV-4 strains isolated from the LBMs were highly similar to wild bird-origin viruses from nearby lakes with free-living wild birds. In additional, a hemagglutination-negative APMV-4 virus was identified. These findings, together with recent APMV-4 studies, suggest potential virus interspecies transmission between wild birds and domestic poultry, and reveal possible epidemiological intercontinental connections between APMV-4 transmission by wild birds.

A Raf kinase inhibitor demonstrates antiviral activities both in vitro and in vivo against different genotypes of virulent Newcastle disease virus.

Newcastle disease (ND) is still one of the major plagues of birds worldwide. Combat actions are limited to vaccines, highlighting the urgent need for new and amply available antiviral drugs. Previous results have shown that Newcastle disease virus (NDV) downregulates the intracellular Raf kinase inhibitor protein (RKIP) expression for efficient replication, suggesting that this molecular may be a suitable target for antiviral intervention. In the present work, we investigated whether or not the Raf kinase inhibitor V (RKIV), which functions in the same way as RKIP by targeting the intracellular Raf kinase, is able to suppress the propagation of enzootic virulent NDV in vitro and in vivo. In vitro antiviral activity of RKIV was assessed by cell-based assay, and in vivo activity was determined in the chicken model. Our results clearly showed that RKIV treatment protected the cells from NDV-induced CPE with the effective concentrations on nM level, and inhibited virus replication in the lungs of infected chickens in a dose-dependent manner and protected chickens from the lethal infection by NDV. Thus, we conclude that the Raf kinase inhibitor compound RKIV, by inhibiting the host cellular target Raf kinase, might be very promising as a new class of antivirals against the enzootic virulent NDV infection.

High Genetic Diversity of Newcastle Disease Virus in Wild and Domestic Birds in Northeastern China from 2013 to 2015 Reveals Potential Epidemic Trends.

Newcastle disease (ND), caused by the virulent Newcastle disease virus (NDV), is one of the most important viral diseases of birds globally, but little is currently known regarding enzootic trends of NDV in northeastern China, especially for class I viruses. Thus, we performed a surveillance study for NDV in northeastern China from 2013 to 2015. A total 755 samples from wild and domestic birds in wetlands and live bird markets (LBMs) were collected, and 10 isolates of NDV were identified. Genetic and phylogenetic analyses showed that five isolates from LBMs belong to class I subgenotype 1b, two (one from wild birds and one from LBMs) belong to the vaccine-like class II genotype II, and three (all from wild birds) belong to class II subgenotype Ib. Interestingly, the five class I isolates had epidemiological connections with viruses from southern, eastern, and southeastern China. Our findings, together with recent prevalence trends of class I and virulent class II NDV in China, suggest possible virus transmission between wild and domestic birds and the potential for an NDV epidemic in the future.

Expression of Raf kinase inhibitor protein is downregulated in response to Newcastle disease virus infection to promote viral replication.

Newcastle disease virus (NDV) causes a severe and economically significant disease affecting almost the entire poultry industry worldwide. However, factors that affect NDV replication in host cells are poorly understood. Raf kinase inhibitory protein (RKIP) is a physiological inhibitor of c-RAF kinase and NF-κB signalling, known for their functions in the control of immune response as well as tumour invasion and metastasis. In the present study, we investigated the consequences of overexpression of host RKIP during viral infection. We demonstrate that NDV infection represses RKIP expression thereby promoting virus replication. Experimental upregulation of RKIP in turn acts as a potential antiviral defence mechanism in host cells that restricts NDV replication by repressing the activation of Raf/MEK/ERK and IκBα/NF-κB signalling pathways. Our results not only extend the concept of linking NDV-host interactions, but also reveal RKIP as a new class of protein-kinase-inhibitor protein that affects NDV replication with therapeutic potential.