VP2 mediates the release of the feline calicivirus RNA genome by puncturing the endosome membrane of infected cells.

Feline calicivirus (FCV) is one of the few members of the Caliciviridae family that grows well in cell lines and, therefore, serves as a surrogate to study the biology of other viruses in the family. Conley et al. (14) demonstrated that upon the receptor engagement to the capsid, FCV VP2 forms a portal-like assembly, which might provide a channel for RNA release. However, the process of calicivirus RNA release is not yet fully understood. Our findings suggest that the separation of the FCV capsid from its genome RNA (gRNA) occurs rapidly in the early endosomes of infected cells. Using a liposome model decorated with the FCV cell receptor fJAM-A, we demonstrate that FCV releases its gRNA into the liposomes by penetrating membranes under low pH conditions. Furthermore, we found that VP2, which is rich in hydrophobic residues at its N-terminus, functions as the pore-forming protein. When we substituted the VP2 N-terminal hydrophobic residues, the gRNA release efficacy of the FCV mutants decreased. In conclusion, our results suggest that in the acidic environment of early endosomes, FCV VP2 functions as the pore-forming protein to mediate gRNA release into the cytoplasm of infected cells. This provides insight into the mechanism of calicivirus genome release.IMPORTANCEResearch on the biology and pathogenicity of certain caliciviruses, such as Norovirus and Sapovirus, is hindered by the lack of easy-to-use cell culture system. Feline calicivirus (FCV), which grows effectively in cell lines, is used as a substitute. At present, there is limited understanding of the genome release mechanism in caliciviruses. Our findings suggest that FCV uses VP2 to pierce the endosome membrane for genome release and provide new insights into the calicivirus gRNA release mechanism.

Simulation Study on the Charge Collection Mechanism of FinFET Devices in Single-Event Upset.

Planar devices and FinFET devices exhibit significant differences in single-event upset (SEU) response and charge collection. However, the charge collection process during SEU in FinFET devices has not been thoroughly investigated. This article addresses this gap by establishing a FinFET SRAM simulation structure and employing simulation software to delve into the charge collection process of FinFET devices during single-event upset. The results reveal substantial differences in charge collection between NMOS and PMOS, and that direct incidence of PMOS leads to the phenomenon of multiple-node charge collection causing SRAM unit upset followed by recovery.

Avian ANP32A incorporated in avian influenza A virions promotes interspecies transmission by priming early viral replication in mammals.

Species-specific differences in acidic nuclear phosphoprotein 32 family member A (ANP32A) determine the restriction of avian-signature polymerase in mammalian cells. Mutations that evade this restriction, such as PB2-E627K, are frequently acquired when avian influenza A viruses jump from avian hosts to mammalian hosts. However, the mechanism underlying this adaptation process is still unclear. Here, we report that host factor ANP32 proteins can be incorporated into influenza viral particles through combination with the viral RNA polymerase (vPol) and then transferred into targeted cells where they support virus replication. The packaging of the ANP32 proteins into influenza viruses is dependent on their affinity with the vPol. Avian ANP32A (avANP32A) delivered by avian influenza A virions primes early viral replication in mammalian cells, thereby favoring the downstream interspecies transmission event by increasing the total amount of virus carrying adaptive mutations. Our study clarifies one role of avANP32A where it is used by avian influenza virus to help counteract the restriction barrier in mammals.

3D-printed coloured tooth model for inlay preparation in pre-clinical dental education.

INTRODUCTION: Accurate inlay preparation is extremely important in pre-clinical training. However, there is a lack of tools to guide students to efficiently practise inlay preparation. Therefore, a 3D-printed coloured tooth model for inlay preparation was designed to guide beginners to practise inlay preparation by themselves according to different colour prompts. This study aimed to evaluate the benefits of using a 3D-printed coloured tooth model in the pre-clinical training on inlay preparation. MATERIALS AND METHODS: Twenty-eight students in their fourth-year undergraduate dental program participated in this study. The participants were randomly assigned to two groups for the inlay preparation. Group 1 prepared a plain tooth model for the first and fourth attempts and a 3D-printed coloured tooth model for the second and third attempts (n = 14). Group 2 prepared four plain tooth models (n = 14). The first and fourth tooth models prepared by both groups were scored using an evaluation system (Fair Grade 2000, NISSIN). Next, questionnaires answered by students were used to evaluate the benefits of using a 3D-printed coloured tooth model and self-evaluate hands-on ability using a grading system (1 = strongly agree, 2 = agree, 3 = neutral, 4 = disagree, and 5 = strongly disagree). The scores were evaluated statistically using the Mann-Whitney U test, and the given grades are displayed as percentages and mean values. RESULTS: There was an overall increase in the clinical confidence of all students after repeated attempts to prepare an inlay; however, students from group 1, who had used the 3D-printed coloured tooth model, had more positive experiences and comments. The 3D-printed coloured tooth model for inlay preparation has been widely praised by participants. Comparing the average score of the first and fourth preparations, the average score of group 1 increased by 12% (Ø 54.46 ± 8.33, Ø 61.11 ± 7.13, p = .090), while that of group 2 increased by 0.72% (Ø 56.39 ± 9.59, Ø 56.80 ± 8.46, p = .925). CONCLUSION: Students favoured the use of the 3D-printed coloured tooth model, and this improved the average score for inlay preparation. The 3D-printed coloured tooth model for inlay preparation is expected to play an important role in dental education in the future.

Blood-brain barrier dysfunction mediated by the EZH2-Claudin-5 axis drives stress-induced TNF-α infiltration and depression-like behaviors.

Growing evidence suggests that neurovascular dysfunction characterized by blood-brain barrier (BBB) breakdown underlies the development of psychiatric disorders, such as major depressive disorder (MDD). Tight junction (TJ) proteins are critical modulators of homeostasis and BBB integrity. TJ protein Claudin-5 is the most dominant BBB component and is downregulated in numerous depression models; however, the underlying mechanisms remain elusive. Here, we demonstrate a molecular basis of BBB breakdown that links stress and depression. We implemented an animal model of depression, chronic unpredictable mild stress (CUMS) in male C57BL/6 mice, and showed that hippocampal BBB breakdown was closely associated with stress vulnerability. Concomitantly, we found that dysregulated Cldn5 level coupled with repression of the histone methylation signature at its promoter contributed to stress-induced BBB dysfunction and depression. Moreover, histone methyltransferase enhancer of zeste homolog 2 (EZH2) knockdown improved Cldn5 expression and alleviated depression-like behaviors by suppressing the tri-methylation of lysine 27 on histone 3 (H3K27me3) in chronically stressed mice. Furthermore, the stress-induced excessive transfer of peripheral cytokine tumor necrosis factor-α (TNF-α) into the hippocampus was prevented by Claudin-5 overexpression and EZH2 knockdown. Interestingly, antidepressant treatment could inhibit H3K27me3 deposition at the Cldn5 promoter, reversing the loss of the encoded protein and BBB damage. Considered together, these findings reveal the importance of the hippocampal EZH2-Claudin-5 axis in regulating neurovascular function and MDD development, providing potential therapeutic targets for this psychiatric illness.

vicR overexpression in Streptococcus mutans causes aggregation and affects interspecies competition.

Streptococcus mutans is considered to be a major causative agent of dental caries. VicRK is a two-component signal transduction system (TCSTS) of S. mutans, which can regulate the virulence of S. mutans, such as biofilm formation, exopolysaccharide production, acid production, and acid resistance. Meanwhile, it can also regulate the production of mutacins (nlmC) through the TCSTS ComDE. In this study, we found that the vicR-overexpressing strain was more likely to aggregate to form cell clusters, leading to the formation of abnormal biofilm; the overexpression of vicR increased the length of the chain of S. mutans. Furthermore, the expression of the mutacins in the vicR overexpression strain was increased under aerobic conditions. Compared with the control strain and the parental strain, the vicR overexpression strain was more competitive against Streptococcus gordonii. But there was no significant difference against Streptococcus sanguinis. In clinical strains, the expression level of vicR was positively correlated with their competitive ability against S. gordonii. Transcriptional profiling revealed 24 significantly upregulated genes in the vicR-overexpressing strain, including nlmA, nlmB, nlmC, and nlmD encoding mutacins. Electrophoretic mobility shift assays and DNase I footprinting assays confirmed that VicR can directly bind to the promoter sequence of nlmD. Taken together, our findings further demonstrate that VicRK, an important TCSTS of S. mutans, is involved in S. mutans cell morphology and biofilm formation. VicRK regulates the production of more mutacins in S. mutans in response to oxygen stimulation. VicR can bind to the promoter sequence of nlmD, thereby directly regulating the production of mutacins NlmD.

Composite Sophora Colon-Soluble Capsule Ameliorates DSS-Induced Ulcerative Colitis in Mice via Gut Microbiota-Derived Butyric Acid and NCR+ ILC3.

OBJECTIVE: To investigate the effects of composite Sophora colon-soluble Capsule (CSCC) on gut microbiota-mediated short-chain fatty acids (SCFAs) production and downstream group 3 innate lymphoid cells (ILC3s) of dextran sulfate sodium (DSS)-induced ulcerative colitis (UC) mice model. METHODS: The main components of CSCC were analyzed by hybrid ultra-high-performance liquid chromatography ion mobility spectromety quadrupole time-of-flight mass spectrometry (UHPLC-IM-QTOF/MS). Twenty-four male BALB/c mice were randomly divided into 4 groups (n=6) by using a computer algorithm-generated random digital, including control, DSS model, mesalazine, and CSCC groups. A DSS-induced colitis mice model was established to determine the effects of CSCC by recording colonic weight, colonic length, index of colonic weight, and histological colonic score. The variations in ILC3s were assessed by immunofluorescence and flow cytometry. The results of gut microbiota and SCFAs were acquired by 16s rDNA and gas chromatography-mass spectrometry (GC-MS) analysis. The expression levels of NCR+ ILC3-, CCR6+ Nkp46- (Lti) ILC3-, and ILCreg-specific markers were detected by enzyme-linked immunosorbent assay, and real-time quantitative polymerase chain reaction and Western blot, respectively. RESULTS: The main components of CSCC were matrine, ammothamnine, Sophora flavescens neoalcohol J, and Sophora oxytol U. After 7 days of treatment, CSCC significantly alleviated colitis by promoting the reproduction of intestinal probiotics manifested as upregulation of the abundance of Bacteroidetes species and specifically the Bacteroidales_S24-7 genus (P<0.05). Among the SCFAs, the content of butyric acid increased the most after CSCC treatment. Meanwhile, compared with the model group, Lti ILC3s and its biomarkers were significantly downregulated and NCR+ ILC3s were significantly elevated in the CSCC group (P<0.01). Further experiments revealed that ILC3s were differentiated from Lti ILC3s to NCR+ ILC3s, resulting in interleukin-22 production which regulates gut epithelial barrier function. CONCLUSION: CSCC may exert a therapeutic effect on UC by improving the gut microbiota, promoting metabolite butyric acid production, and managing the ratio between NCR+ ILC3s and Lti ILC3s.

Spatio-temporal spread and evolution of influenza A (H7N9) viruses.

The influenza A (H7N9) virus has been seriously concerned for its potential to cause an influenza pandemic. To understand the spread and evolution process of the virus, a spatial and temporal Bayesian evolutionary analysis was conducted on 2,052 H7N9 viruses isolated during 2013 and 2018. It revealed that the H7N9 virus was probably emerged in a border area of Anhui Province in August 2012, approximately 6 months earlier than the first human case reported. Two major epicenters had been developed in the Yangtze River Delta and Peral River Delta regions by the end of 2013, and from where the viruses have also spread to other regions at an average speed of 6.57 km/d. At least 24 genotypes showing have been developed and each of them showed a distinct spatio-temporal distribution pattern. Furthermore, A random forest algorithm-based model has been developed to predict the occurrence risk of H7N9 virus. The model has a high overall forecasting precision (> 97%) and the monthly H7N9 occurrence risk for each county of China was predicted. These findings provide new insights for a comprehensive understanding of the origin, evolution, and occurrence risk of H7N9 virus. Moreover, our study also lays a theoretical basis for conducting risk-based surveillance and prevention of the disease.

Effects of Repetitive Transcranial Magnetic Stimulation on Cognitive Function in Patients With Stress-Related Depression: A Randomized Double-Blind fMRI and 1H-MRS Study.

Objectives: To reveal the effects of repetitive transcranial magnetic stimulation (rTMS) on the improvement of cognitive function in patients with stress-related depression, and to enrich the neural mechanism(s) underlying rTMS so as to improve cognitive function in patients with stress-related depression. Methods: We conducted a randomized, double-blind, placebo-controlled study of rTMS in patients with stress-related depression who were 18-40 years of age. Patients were randomly allocated to either a sham or experimental group in a 1:1 ratio. A 10-session rTMS protocol was used with 10-Hz stimulation over the left dorsolateral prefrontal cortex (DLPFC). Clinical assessments (HAMD, HAMA, DASS, MoCA), neuropsychologic (Stroop, WCST), and resting state fMRI and 1H-MRS assessments were executed at two time points-baseline and after the 10th rTMS session. Results: rTMS relieved the mental symptoms of patients in both groups. The MoCA score of patients in the experimental group increased; the number of correct answers increased significantly in Stroop testing, and the number of errors and omissions decreased significantly; the number of persistent errors decreased significantly; and the time used to complete the test decreased to an even greater extent in the WCST experimental group. The ReHo value in the lingual gyrus of the right hemisphere and the cuneus of the left and right hemispheres in the experimental group decreased after treatment. The DC value in the left and right hemispheric cuneus and postcentral gyrus of the left hemisphere in the experimental group diminished after treatment. The functional connections of these brain regions also changed as the Cho and NAA/Cr of the left DLPFC changed, with alterations related to the improvement in cognitive function. The level of choline (Cho) in the left DLPFC of the experimental group was significantly lower than that of the control group, and the level of N-acetylaspartate/creatine (NAA/Cr) in the left DLPFC of the control group was significantly higher than that of the experimental group. These changes were related to the overall improvement in cognitive function. Conclusions: Ten-Hz rTMS over the left DLPFC improved the cognitive function of patients with stress-related depression. The governing mechanism for this phenomenon may be via rTMS effects on multiple visual-related brain regions and their functional connections, and on the somatosensory cortex and its functional connection with visual and auditory cortex, reducing the level of Cho and stabilizing the level of NAA/Cr in the left DLPFC.

The W-Acidic Motif of Histidine Kinase WalK Is Required for Signaling and Transcriptional Regulation in Streptococcus mutans.

In Streptococcus mutans, we find that the histidine kinase WalK possesses the longest C-terminal tail (CTT) among all 14 TCSs, and this tail plays a key role in the interaction of WalK with its response regulator WalR. We demonstrate that the intrinsically disordered CTT is characterized by a conserved tryptophan residue surrounded by acidic amino acids. Mutation in the tryptophan not only disrupts the stable interaction, but also impairs the efficient phosphotransferase and phosphatase activities of WalRK. In addition, the tryptophan is important for WalK to compete with DNA containing a WalR binding motif for the WalR interaction. We further show that the tryptophan is important for in vivo transcriptional regulation and bacterial biofilm formation by S. mutans. Moreover, Staphylococcus aureus WalK also has a characteristic CTT, albeit relatively shorter, with a conserved W-acidic motif, that is required for the WalRK interaction in vitro. Together, these data reveal that the W-acidic motif of WalK is indispensable for its interaction with WalR, thereby playing a key role in the WalRK-dependent signal transduction, transcriptional regulation and biofilm formation.

Homocysteine-Induced Disturbances in DNA Methylation Contribute to Development of Stress-Associated Cognitive Decline in Rats.

Chronic stress is generally accepted as the main risk factor in the development of cognitive decline; however, the underlying mechanisms remain unclear. Previous data have demonstrated that the levels of homocysteine (Hcy) are significantly elevated in the plasma of stressed animals, which suggests that Hcy is associated with stress and cognitive decline. To test this hypothesis, we analyzed the cognitive function, plasma concentrations of Hcy, and brain-derived neurotropic factor (BDNF) levels in rats undergoing chronic unpredicted mild stress (CUMS). The results showed that decreased cognitive behavioral performance and decreased BDNF transcription and protein expression were correlated with hyperhomocysteinemia (HHcy) levels in stressed rats. Diet-induced HHcy mimicked the cognitive decline and BDNF downregulation in the same manner as CUMS, while Hcy reduction (by means of vitamin B complex supplements) alleviated the cognitive deficits and BDNF reduction in CUMS rats. Furthermore, we also found that both stress and HHcy disturbed the DNA methylation process in the brain and induced DNA hypermethylation in the BDNF promoter. In contrast, control of Hcy blocked BDNF promoter methylation and upregulated BDNF levels in the brain. These results imply the possibility of a causal role of Hcy in stress-induced cognitive decline. We also used ten-eleven translocation (TET1), an enzyme that induces DNA demethylation, to verify the involvement of Hcy and DNA methylation in the regulation of BDNF expression and the development of stress-related cognitive decline. The data showed that TET1-expressing viral injection into the hippocampus inhibited BDNF promoter methylation and significantly mitigated the cognitive decline in HHcy rats. Taken together, novel evidence from the present study suggests that Hcy is likely involved in chronic stress-induced BDNF reduction and related cognitive deficits. In addition, the negative side-effects of HHcy may be associated with Hcy-induced DNA hypermethylation in the BDNF promoter. The results also suggest the possibility of Hcy as a target for therapy and the potential value of vitamin B intake in preventing stress-induced cognitive decline.

Artificial intelligence for caries and periapical periodontitis detection.

OBJECTIVES: Periapical periodontitis and caries are common chronic oral diseases affecting most teenagers and adults worldwide. The purpose of this study was to develop an evaluation tool to automatically detect dental caries and periapical periodontitis on periapical radiographs using deep learning. METHODS: A modified deep learning model was developed using a large dataset (4129 images) with high-quality annotations to support the automatic detection of both dental caries and periapical periodontitis. The performance of the model was compared to the classification performance of dentists. RESULTS: The deep learning model automatically distinguished dental caries with an F1-score of 0.829 and periapical periodontitis with an F1-score of 0.828. The comparison of model-only and expert-only detection performance showed that the accuracy of the fully automatic method was significantly higher than that of the young dentists. With deep learning assistance, the experts not only reached a higher diagnostic accuracy with an average F1-score of 0.7844 for dental caries and 0.8208 for periapical periodontitis compared to expert-only scenarios, but also increased inter-observer agreement from 0.585/0.590 to 0.726/0.713 for dental caries and from 0.623/0.563 to 0.752/0.740 for periapical periodontitis. CONCLUSIONS: Based on these experimental results, deep learning can improve the accuracy and consistency of evaluating dental caries and periapical periodontitis on periapical radiographs. CLINICAL SIGNIFICANCE: Deep learning models can improve accuracy and consistency and reduce the workload of dentists, making artificial intelligence a powerful tool for clinical practice.

Norepinephrine promotes glioma cell migration through up-regulating the expression of Twist1.

BACKGROUND: Glioma cells are characterized by high migration ability, resulting in aggressive growth of the tumors and poor prognosis of patients. It has been reported that the stress-induced hormone norepinephrine (NE) contributes to tumor progression through mediating a number of important biological processes in various cancers. However, the role of NE in the regulation of glioma migration is still unclear. Epithelial-to-mesenchymal transition (EMT) is one of the most important steps for tumor migration and metastasis. Twist1, as a key regulator of EMT, has been found to be elevated during glioma migration. But it is still unknown whether Twist1 is involved in the effect of NE on the migration of glioma cells. METHODS: Wound healing assay and transwell assay were conducted to evaluate the migration of glioma cells upon different treatments. The mesenchymal-like phenotype and the expression of Twist1 after NE treatment were assessed by cell diameters, real-time PCR, western blot and immunofluorescence staining. The gain-and loss-of-function experiments were carried out to investigate the biological function of Twist1 in the migration induced by NE. Finally, the clinical significance of Twist1 was explored among three public glioma datasets. RESULTS: In this study, our finding revealed a facilitative effect of NE on glioma cell migration in a ß-adrenergic receptor (ADRB)-dependent way. Mechanistically, NE induced mesenchymal-like phenotype and the expression of Twist1. Twist1 overexpression promoted glioma cells migration, while knockdown of Twist1 abolished the discrepancy in the migration ability between NE treated glioma cells and control cells. In addition, the clinical analysis demonstrated that Twist1 was up-regulated in malignant gliomas and recurrent gliomas, and predicted a poor prognosis of glioma patients. CONCLUSIONS: NE enhanced the migration ability of glioma cells through elevating the expression of Twist1. Our finding may provide potential therapeutic target for protecting patients with glioma from the detrimental effects of stress biology on the tumor progression.

Structural Insights into Alphavirus Assembly Revealed by the Cryo-EM Structure of Getah Virus.

Getah virus (GETV) is a member of the alphavirus genus, and it infects a variety of animal species, including horses, pigs, cattle, and foxes. Human infection with this virus has also been reported. The structure of GETV has not yet been determined. In this study, we report the cryo-EM structure of GETV at a resolution of 3.5 Å. This structure reveals conformational polymorphism of the envelope glycoproteins E1 and E2 at icosahedral 3-fold and quasi-3-fold axes, which is believed to be a necessary organization in forming a curvature surface of virions. In our density map, three extra densities are identified, one of which is believed a "pocket factor"; the other two are located by domain D of E2, and they may maintain the stability of E1/E2 heterodimers. We also identify three N-glycosylations at E1 N141, E2 N200, and E2 N262, which might be associated with receptor binding and membrane fusion. The resolving of the structure of GETV provides new insights into the structure and assembly of alphaviruses and lays a basis for studying the differences of biology and pathogenicity between arthritogenic and encephalitic alphaviruses.

Production and application of mouse monoclonal antibodies targeting linear epitopes in pB602L of African swine fever virus.

African swine fever (ASF) is an acute hemorrhagic disease of domestic pigs. The causative agent of ASF, ASF virus (ASFV), is a double-stranded DNA virus, the sole member in the family Asfarviridae. The non-structural protein pB602L of ASFV is a molecular chaperone of the major capsid protein p72 and plays a key role in icosahedral capsid assembly. This protein is antigenic and is a target for developing diagnostic tools for ASF. To generate monoclonal antibodies (mAbs) against pB602L, a prokaryotically expressed recombinant pB602L protein was produced, purified, and used as an antigen to immunize mice. A total of eight mouse mAbs were obtained, and their binding epitopes were screened by Western blot using an overlapping set of polypeptides from pB602L. Three linear epitopes were identified and designated epitope 1 (366ANRERYNY373), epitope 2 (415GPDAPGLSI423), and epitope 3 (498EMLNVPDD505). Based on the epitope recognized, the eight mAbs were placed into three groups: group 1 (B2A1, B2F1, and B2D10), group 2 (B2H10, B2B2, B2D8, and B2A3), and group 3 (B2E12). The mAbs B2A1, B2H10, and B2E12, each representing one of the groups, were used to detect pB602L in ASFV-infected porcine alveolar macrophages (PAMs) and pig tissues, using an indirect fluorescence assay (IFA) and immunohistochemical staining, respectively. The results showed that pB602L was detectable with all three mAbs in immunohistochemical staining, but only B2H10 was suitable for detecting the proteins in ASFV-infected PAMs by IFA. In summary, we developed eight anti-pB602L mouse mAbs recognizing three linear epitopes in the protein, which can be used as reagents for basic and applied research on ASFV.

Freestanding Graphene Fabric Film for Flexible Infrared Camouflage.

Graphene films, fabricated by chemical vapor deposition (CVD) method, have exhibited superiorities in high crystallinity, thickness controllability, and large-scale uniformity. However, most synthesized graphene films are substrate-dependent, and usually fragile for practical application. Herein, a freestanding graphene film is prepared based on the CVD route. By using the etchable fabric substrate, a large-scale papyraceous freestanding graphene fabric film (FS-GFF) is obtained. The electrical conductivity of FS-GFF can be modulated from 50 to 2800 Ω sq-1 by tailoring the graphene layer thickness. Moreover, the FS-GFF can be further attached to various shaped objects by a simple rewetting manipulation with negligible changes of electric conductivity. Based on the advanced fabric structure, excellent electrical property, and high infrared emissivity, the FS-GFF is thus assembled into a flexible device with tunable infrared emissivity, which can achieve the adaptive camouflage ability in complicated backgrounds. This work provides an infusive insight into the fabrication of large-scale freestanding graphene fabric films, while promoting the exploration on the flexible infrared camouflage textiles.

Leucine mediates cognitive dysfunction in early life stress-induced mental disorders by activating autophagy.

Objectives: To explore the relationship between leucine in cerebrospinal fluid (CSF) and cognitive dysfunction in rats with early life stress (ELS) induced mental illness, and pathophysiological mechanism involved. Methods: The maternal separation (MS), an animal paradigm used widely as a preclinical model of ELS which is one of the important risk factors for mental disorders. Behavioral experiments including open-field test, sucrose preference, object recognition and Morris water maze tests, Nissl staining, transmission electron microscopy and WES were employed in the present study. Results: The behavioral results showed that MS rats were more prone to cognitive impairment and depression-and-anxiety-like behaviors than controls, including spatial self-exploration ability, memory ability, and spatial learning and memory function. Nissl staining analysis indicated that the number of neurons in the CA1 and CA3 regions of the hippocampus significantly decreased and the arrangement of nerve cells was abnormal. The leucine levels were decreased in the CSF of MS rats and highly correlated with the number of hippocampal neurons, and yet leucine supplementation improved the degree of MS-induced cognitive impairment. Furthermore, there were autophagosomes in the hippocampus of the low-leucine diet rats of the control and MS group but not in the high-leucine diet MS group by transmission electron microscopy. The protein expression of Beclin-1 in the hippocampus was significantly increased in the MS normal diet group and MS low-leucine diet group, yet decreased in the MS high-leucine diet group compared with the MS low-leucine diet group. Meanwhile, the Bcl-2/Bax ratio was significantly decreased in the control low-leucine diet group, MS normal diet group and MS low-leucine diet group. Ultimately, in vitro experiments suggested that leucine deficiency could activate neuronal autophagy including enhanced LC3II/LC3I and mRFP-GFP-LC3, which was consistent with the in vivo results, and the cell apoptosis rate and lactate dehydrogenase (LDH) cytotoxicity were also increased with leucine deficiency, while the above effects could be partly reversed by autophagy inhibitor treatment. Conclusions: MS model caused adult male rats to be susceptible to cognitive dysfunction, which may regulate autophagy in hippocampal neurons through leucine metabolism in CSF.

Chronic stress promotes glioma cell proliferation via the PI3K/Akt signaling pathway.

High malignancy and high mortality of glioma render it urgent to elucidate the underlying mechanisms of glioma carcinogenesis and explore novel targets for therapy. Epidemiologic and clinical studies have revealed that chronic stress promotes the progression of various solid tumors and is correlated with poor prognosis; however, findings reporting the involvement of chronic stress in glioma are rare. In the present study, a chronic restraint animal model and a chronic stress cell model were established to explore the effects of chronic stress on glioma and its molecular mechanisms. The results revealed that chronic stress promoted glioma growth in vivo, and the serum levels of the stress hormones glucocorticoid (GC) and noradrenaline (NE) were significantly increased. In addition, GC and NE were verified to accelerate the proliferation of glioma cells in vitro. Mechanistically, the phosphatidylinositol 3­kinase (PI3K)/Akt signaling pathway was revealed to be activated under stress conditions, and inhibition of the expression of p­Akt could restrain the stress hormone­induced glioma cell proliferation. In addition, our data indicated that the GC receptor (GR) and ß­adrenergic receptors (ADRBs) were both required for the biological functions of GC and NE in glioma cells. In conclusion, these results indicated that chronic stress and the stress hormones GC and NE activated PI3K/Akt signaling through binding to GR and ADRBs, thereby promoting glioma cell growth. Our findings may provide potential therapeutic targets and pave the way for the development of new strategies to protect patients with glioma from the detrimental effects of stress on tumor progression.

Integrating Pharmacology and Microbial Network Analysis with Experimental Validation to Reveal the Mechanism of Composite Sophora Colon-Soluble Capsule against Ulcerative Colitis.

Ulcerative colitis (UC) has multifactorial pathogenesis that acts synergistically, such as immune system dysregulation and expansion of infectious gut microbiota. Therefore, a multicomponent treatment derived from Chinese herbal medicine that interacts with multiple targets synergistically is needed. Composite sophora colon-soluble capsule (CSCC) is a Chinese herbal formula that has shown therapeutic efficacy against UC in randomized clinical trials. However, its bioactive components and potential target genes against UC remain unclear. Here, we used a network pharmacology approach to detect component-target-pathway interactions of CSCC against UC. A total of 29 gene targets, 91 bioactive components, and 20 enriched pathways of CSCC were identified. The IL-17 signaling pathway activated by infectious gastrointestinal microbes and predicted by the network analysis to be a major pathway modulated by CSCC against UC was studied in a dextran sulfate sodium-induced colitis model. CSCC showed remarkable efficacy against UC with respect to the attenuation of colon length, body weight loss, and disease activity index through gut microbiota recovery and intestinal immune homeostasis. The rectal administration of CSCC reduced the numbers of Th17 cells isolated from both mesenteric lymph nodes and lamina propria mononuclear cells and the levels of IL-17A, IL-6, IL-1ß, and TNF-α. Additionally, the percentage of Treg cells and the levels of their hallmark cytokines were upregulated. Rectal administration of CSCC led to microbiota regulation with a significant correlation between suppression of Verrucomicrobiaceae and Ruminococcaceae, as well as the elevation of Lactobacillaceae, and CSCC administration via microbiome correlation heatmaps and cooccurrence network analysis at multiple time points. Thus, our study presents an effective herbal formula, CSCC, for UC treatment and explores its components and mechanisms of efficacy through the examination of gut microbiota and hallmark cytokines in the IL-17 pathway.

Implication of the Identification of an Earlier Pseudorabies Virus (PRV) Strain HLJ-2013 to the Evolution of Chinese PRVs.

Pseudorabies viruses (PRVs) pose a great threat to the pig industry of many countries around the world. Human infections with PRV have also been reported occasionally in China. Therefore, understanding the epidemiology and evolution of PRVs is of great importance for disease control in the pig populations and humans as well. In this study, we isolated a PRV designated HLJ-2013 from PRV-positive samples that had been collected in Heilongjiang, China, in 2013. The full genome sequence of the virus was determined to be ∼143 kbp in length using high-throughput sequencing. The genomic sequence identities between this isolate and 21 other previous PRV isolates ranged from 92.4% (with Bartha) to 97.3% (with SC). Phylogenetic analysis based on the full-length genome sequences revealed that PRV HLJ-2013 clustered together with all the Chinese strains in one group belonging to Genotype II, but this virus occurred phylogenetically earlier than all the other Chinese PRV strains. Phylogenetic trees based on both protein-coding genes and non-coding regions revealed that HLJ-2013 probably obtained its genome sequences from three origins: a yet unknown parent virus, the European viruses, and the same ancestor of all Chinese PRVs. Recombination analysis showed that HLJ-2013-like virus possibly donated the main framework of the genome of the Chinese PRVs. HLJ-2013 exhibited cytopathic and growth characteristics similar to that of the Chinese PRV strains SC and HeN1, but its pathogenicity in mice was higher than that of SC and lower than that of HeN1. The identification of HLJ-2013 takes us one step closer to understanding the origin of PRVs in China and provides new knowledge about the evolution of PRVs worldwide.