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BACKGROUND: The function of kallistatin in airway inflammation, particularly chronic rhinosinusitis with nasal polyps (CRSwNP), has not been elucidated. OBJECTIVE: We sought to investigate the role of kallistatin in airway inflammation. METHODS: Kallistatin and proinflammatory cytokine expression levels were detected in nasal polyps. For the in vivo studies, we constructed the kallistatin-overexpressing transgenic mice to elucidate the role of kallistatin in airway inflammation. Furthermore, the levels of plasma IgE and proinflammatory cytokines in the airways were evaluated in the kallistatin-/- rat in vivo model under a type 2 inflammatory background. Finally, the Notch signaling pathway was explored to understand the role of kallistatin in CRSwNP. RESULTS: We showed that the expression of kallistatin was significantly higher in nasal polyps than in the normal nasal mucosa and correlated with IL-4 expression. We also discovered that the nasal mucosa of kallistatin-overexpressing transgenic mice expressed higher levels of IL-4 expression, associating to TH2-type inflammation. Interestingly, we observed lower IL-4 levels in the nasal mucosa and lower total plasma IgE of the kallistatin-/- group treated with house dust mite allergen compared with the wild-type house dust mite group. Finally, we observed a significant increase in the expression of Jagged2 in the nasal epithelium cells transduced with adenovirus-kallistatin. This heightened expression correlated with increased secretion of IL-4, attributed to the augmented population of CD4+CD45+Notch1+ T cells. These findings collectively may contribute to the induction of TH2-type inflammation. CONCLUSIONS: Kallistatin was demonstrated to be involved in the CRSwNP pathogenesis by enhancing the TH2 inflammation, which was found to be associated with more expression of IL-4, potentially facilitated through Jagged2-Notch1 signaling in CD4+ T cells.
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Linfócitos T CD4-Positivos , Mucosa Nasal , Rinossinusite , Serpinas , Células Th2 , Animais , Feminino , Humanos , Masculino , Camundongos , Ratos , Linfócitos T CD4-Positivos/imunologia , Quimiotaxia de Leucócito/imunologia , Doença Crônica , Citocinas/metabolismo , Imunoglobulina E/imunologia , Imunoglobulina E/sangue , Inflamação/imunologia , Interleucina-4/imunologia , Interleucina-4/metabolismo , Camundongos Transgênicos , Mucosa Nasal/imunologia , Mucosa Nasal/metabolismo , Pólipos Nasais/imunologia , Rinossinusite/imunologia , Serpinas/imunologia , Serpinas/genética , Serpinas/metabolismo , Transdução de Sinais , Células Th2/imunologiaRESUMO
Due to the extensive genetic and antigenic variation in Porcine Reproductive and Respiratory Syndrome Virus (PRRSV), as well as its rapid mutability and evolution, PRRS prevention and control can be challenging. An expeditious and sensitive neutralization assay for PRRSV is presented to monitor neutralizing antibodies (NAbs) in serum during vaccine research. Here, a PRRSV expressing eGFP was successfully rescued with reverse genetics based on the infectious clone HuN4-F112-eGFP which we constructed. The fluorescent protein expressions of the reporter viruses remained stable for at least five passages. Based on this reporter virus, the neutralization assay can be easily used to evaluate the level of NAbs by counting cells with green fluorescence. Compared with the classical CPE assay, the newly developed assay increases sensitivity by one- to four-fold at the early antibody response stage, thus saving 2 days of assay waiting time. By using this assay to unveil the dynamics of neutralizing antibodies against PRRSV, priming immunity through either a single virulent challenge or only vaccination could produce limited NAbs, but re-infection with PRRSV would induce a faster and stronger NAb response. Overall, the novel HuN4-F112-eGFP-based neutralization assay holds the potential to provide a highly efficient platform for evaluating the next generation of PRRS vaccines.
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BACKGROUND: Transbronchial cryoablation shows potential as a local therapy for inoperable peripheral lung cancer. However, its clinical application for peripheral pulmonary lesions has not been reported yet. METHODS: An improved cryoprobe with an 8-mm-long, 1.9-mm-wide cryotip was used. Initially, the safety and effectiveness of this cryoprobe were assessed in an in vivo porcine model. Transbronchial cryoablation with 2 or 3 freeze-thaw cycles (10 min or 15 min in each freezing time) was performed in 18 pigs under CT monitoring. Radiological and pathological examinations were performed to evaluate the extent of cryoablation. Subsequently, nine patients with stage IA peripheral lung cancer or metastases underwent transbronchial cryoablation with this cryoprobe under the guidance of navigation bronchoscopy and cone-beam CT. Technical success, safety and outcomes were assessed. RESULTS: 36 cryoablation procedures were performed successfully without any major complications in the porcine model. The extent of cryoablation increased with freezing time and the number of freeze-thaw cycles, which peaked at 24 hours and then gradually decreased. Pathological results showed a change from massive haemorrhage at 24 hours to fibrous hyperplasia with chronic inflammation after 4 weeks. In the clinical trial, 10 cryoablations were performed on 9 tumours with a technical success rate of 100%. One mild treatment-related complication occurred. Of the nine tumours, seven achieved complete ablation, while two exhibited incomplete ablation and subsequent local progression at 6 months. CONCLUSION: Our initial experience indicated that transbronchial cryoablation was a safe and feasible procedure for non-surgical peripheral stage IA lung cancer or pulmonary metastases. TRIAL REGISTRATION NUMBER: ChiCTR2200061544.
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Broncoscopia , Criocirurgia , Neoplasias Pulmonares , Criocirurgia/métodos , Criocirurgia/instrumentação , Animais , Neoplasias Pulmonares/cirurgia , Neoplasias Pulmonares/patologia , Suínos , Humanos , Broncoscopia/métodos , Masculino , Feminino , Idoso , Pessoa de Meia-Idade , Resultado do Tratamento , Tomografia Computadorizada de Feixe CônicoRESUMO
The nonstructural proteins (Nsps) of porcine reproductive and respiratory syndrome virus (PRRSV) play essential roles in virus replication-a multistep process that requires the participation of host factors. It is of great significance for the development of antiviral drugs to characterize the host proteins that interact with PRRSV Nsps and their functions in PRRSV replication. Here, we determined that proteasome subunit ß type 1 (PSMB1) interacted with viral Nsp12 to inhibit PRRSV replication in target and permissive cells. PSMB1 could be downregulated by PRRSV infection through interaction with the transcription factor EBF1. Proteasome and autophagy inhibitor assays showed that PSMB1 was regulated by the autophagic pathway to degrade Nsp12. Cotransfection of PSMB1 and Nsp12 increased the level of intracellular autophagy; both molecules were colocated in lysosomes. We also found that the selective autophagy cargo receptor protein NBR1 and E3 ubiquitin ligase STUB1 interacted with PSMB1 and Nsp12, respectively, in the autophagic degradation of Nsp12. Furthermore, the degradation of Nsp12 by PSMB1 was mainly dependent on the ubiquitination of Nsp12 at lysine site 130. Our results indicate for the first time that PSMB1 is an anti-PRRSV host protein that inhibits the replication of PRRSV by degradation of Nsp12 through the selective autophagy pathway. IMPORTANCE PRRS is a major threat to the global pig industry and urgently requires an effective and sustainable control strategy. PRRSV Nsps have important roles in viral RNA synthesis, proteinase activity, induction of replication-associated membrane rearrangements, replicative endoribonuclease activity, determination of virulence, and regulation of host immune response. Research associated with PRRSV Nsps can provide vital guidance to modify the PRRSV genome through reverse genetics in the development of vaccines and diagnostics. The function of Nsp12, which generally plays essential roles in virus replication, remains unclear. We demonstrated that PSMB1 interacted with and degraded Nsp12 through an autophagic pathway to inhibit PRRSV replication. Our data confirmed a novel antiviral function of PSMB1 and allowed us to elaborate on the roles of Nsp12 in PRRSV pathogenesis. These findings suggest a valid and highly conserved candidate target for the development of novel therapies and more effective vaccines and demonstrate the complex cross talk between selective autophagy and PRRSV infection.
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Autofagia , Vírus da Síndrome Respiratória e Reprodutiva Suína , Proteínas não Estruturais Virais , Replicação Viral , Animais , Antivirais , Síndrome Respiratória e Reprodutiva Suína/imunologia , Síndrome Respiratória e Reprodutiva Suína/prevenção & controle , Síndrome Respiratória e Reprodutiva Suína/virologia , Vírus da Síndrome Respiratória e Reprodutiva Suína/fisiologia , Complexo de Endopeptidases do Proteassoma/metabolismo , Suínos , Ubiquitinação , Proteínas não Estruturais Virais/metabolismo , Interações entre Hospedeiro e Microrganismos/imunologiaRESUMO
BACKGROUND: Occupational Bloodborne Exposures (OBEs) are incidents where healthcare workers come into contact with blood or other potentially infectious materials, leading to risks of transmitting bloodborne pathogens. Nursing students, often in direct contact with patients, face heightened risks due to their duties. METHODS: First, we conducted a cross-sectional survey using a OBEs questionnaire to explore the knowledge, attitudes, practices, and needs regarding OBEs among nursing students. Subsequently, we used a randomized controlled trial (RCT) to compare the impact of the Presentation-Assimilation-Discussion (PAD) method with the traditional lecture-based learning (LBL) method on OBEs education for nursing students. Pre-test, post-test, and retention test were used to observe the teaching effectiveness, and the students' feedback on the teaching method was also observed. RESULTS: In the cross-sectional survey, we found that nursing students lacked sufficient knowledge and management skills regarding OBEs but recognized the importance of standard precautions and expressed a desire for systematic OBEs training during their education and internships. In the RCT, the total, theoretical, and practical scores of the PAD and LBL groups were comparable in the pre-test (56.70 ± 3.47 vs. 56.40 ± 3.95, 33.09 ± 3.39 vs. 33.33 ± 2.44, 23.61 ± 4.66 vs. 23.07 ± 4.84, p > 0.05). After training, the PAD model demonstrated an advantage over the LBL model in immediate total (84.25 ± 4.06 vs. 78.95 ± 4.23, p < 0.001), theoretical (54.32 ± 2.43 vs. 51.44 ± 2.58, p < 0.001), and practical scores (29.93 ± 3.90 vs. 27.51 ± 4.33, p < 0.01). It also showed superior retention of total (69.05 ± 3.87 vs. 65.77 ± 2.94, p < 0.001) and theoretical scores (39.05 ± 3.05 vs. 36.23 ± 3.18, p < 0.001). However, there was no significant difference in the retention of practical scores between the two groups (30.00 ± 4.76 vs. 29.53 ± 3.73, p > 0.05). The PAD group benefited more across various learning dimensions but reported a higher study load. CONCLUSIONS: Our study reveals that the PAD model could be a valuable approach for teaching OBEs to nursing students.
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Cryo-electron tomography (cryoET) is a powerful technology that allows in-situ observation of the molecular structure of tissues and cells. Cryo-focused ion beam (cryoFIB) milling plays an important role in the preparation of high-quality thin lamellar samples for cryoET studies, thus, promoting the rapid development of cryoET in recent years. However, locating the regions of interest in a large cell or tissue during cryoFIB milling remains a major challenge limiting cryoET applications on arbitrary biological samples. Here, we report an on-the-fly localization method based on cellular secondary electron imaging (CSEI), which is derived from a basic imaging function of the cryoFIB instruments and enables high-contrast imaging of the cellular contents of frozen-hydrated biological samples. Moreover, CSEI does not require fluorescent labels and additional devices. The present study discusses the imaging principles and settings for optimizing CSEI. Tests on several commercially available cryoFIB instruments demonstrated that CSEI was feasible on mainstream instruments to observe all types of cellular contents and reliable under different milling conditions. We established a simple milling-localization workflow and tested it using the basal body of Chlamydomonas reinhardtii.
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Tomografia com Microscopia Eletrônica , Elétrons , Microscopia Crioeletrônica/métodos , Tomografia com Microscopia Eletrônica/métodos , ÍonsRESUMO
Femtosecond time-resolved X-ray absorption (XANES) at the Co K-edge, X-ray emission (XES) in the Co Kß and valence-to-core regions, and broadband UV-vis transient absorption are combined to probe the femtosecond to picosecond sequential atomic and electronic dynamics following photoexcitation of two vitamin B12 compounds, hydroxocobalamin and aquocobalamin. Polarized XANES difference spectra allow identification of sequential structural evolution involving first the equatorial and then the axial ligands, with the latter showing rapid coherent bond elongation to the outer turning point of the excited state potential followed by recoil to a relaxed excited state structure. Time-resolved XES, especially in the valence-to-core region, along with polarized optical transient absorption suggests that the recoil results in the formation of a metal-centered excited state with a lifetime of 2-5 ps. This combination of methods provides a uniquely powerful tool to probe the electronic and structural dynamics of photoactive transition-metal complexes and will be applicable to a wide variety of systems.
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New synthetic hybrid materials and their increasing complexity have placed growing demands on crystal growth for single-crystal X-ray diffraction analysis. Unfortunately, not all chemical systems are conducive to the isolation of single crystals for traditional characterization. Here, small-molecule serial femtosecond crystallography (smSFX) at atomic resolution (0.833 Å) is employed to characterize microcrystalline silver n-alkanethiolates with various alkyl chain lengths at X-ray free electron laser facilities, resolving long-standing controversies regarding the atomic connectivity and odd-even effects of layer stacking. smSFX provides high-quality crystal structures directly from the powder of the true unknowns, a capability that is particularly useful for systems having notoriously small or defective crystals. We present crystal structures of silver n-butanethiolate (C4), silver n-hexanethiolate (C6), and silver n-nonanethiolate (C9). We show that an odd-even effect originates from the orientation of the terminal methyl group and its role in packing efficiency. We also propose a secondary odd-even effect involving multiple mosaic blocks in the crystals containing even-numbered chains, identified by selected-area electron diffraction measurements. We conclude with a discussion of the merits of the synthetic preparation for the preparation of microdiffraction specimens and compare the long-range order in these crystals to that of self-assembled monolayers.
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African swine fever virus (ASFV) is a highly contagious and deadly virus that leads to high mortality rates in domestic swine populations. Although the envelope protein CD2v of ASFV has been implicated in immunomodulation, the molecular mechanisms underlying CD2v-mediated immunoregulation remain unclear. In this study, we generated a stable CD2v-expressing porcine macrophage (PAM-CD2v) line and investigated the CD2v-dependent transcriptomic landscape using RNA-seq. GO terms enrichment analysis and gene set enrichment analysis revealed that CD2v predominantly affected the organization and assembly process of the extracellular matrix. Wound healing and Transwell assays showed that CD2v inhibited swine macrophage migration. Further investigation revealed a significant decrease in the expression of transcription factor early growth response 1 (EGR1) through inhibiting the activity of extracellular signal-regulated kinase 1 and 2 (ERK1/2). Notably, EGR1 knockout in swine macrophages restricted cell migration, whereas EGR1 overexpression in PAM-CD2v restored the ability of macrophage migration, suggesting that CD2v inhibits swine macrophage motility by downregulating EGR1 expression. Furthermore, we performed chromatin immunoprecipitation and sequencing for EGR1 and the histone mark H3K27 acetylation (H3K27ac), and we found that EGR1 co-localized with the activated histone modification H3K27ac neighboring the transcriptional start sites. Further analysis indicated that EGR1 and H3K27ac co-occupy the promoter regions of cell locomotion-related genes. Finally, by treating various derivatives of swine macrophages with lipopolysaccharides, we showed that depletion of EGR1 decreased the expression of inflammatory cytokines including TNFα, IL1α, IL1ß, IL6, and IL8, which play essential roles in inflammation and host immune response. Collectively, our results provide new insights into the immunomodulatory mechanism of ASFV CD2v.
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Vírus da Febre Suína Africana , Febre Suína Africana , Doenças dos Suínos , Suínos , Animais , Vírus da Febre Suína Africana/genética , Citocinas/genética , Citocinas/metabolismo , Sistema de Sinalização das MAP Quinases , Proteínas Virais/metabolismo , Macrófagos , Movimento CelularRESUMO
A chiral Brønsted acid catalysed phosphine-mediated deoxygenation protocol is reported. This metal-free method provides a precise kinetic resolution platform for azaarylethynyl tertiary alcohols, which are a broad category of biologically and synthetically important azaarene derivatives. In addition to providing an efficient method for the first asymmetric preparation of these tertiary alcohols, the strategy facilitates the construction of azaaryl-functionalized allenes with good to excellent enantioselectivities. The high selectivity factors (s up to 235), broad substrate scope, and ability to convert azaaryl compounds into both chiral tertiary alcohols and allenes robustly underscore the efficiency and promising utility of this method. The practicability is further validated by the successful synthesis of deuterated allenes with high ee values and substantial incorporation of deuterium using inexpensive D2 O as the deuterium source.
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OBJECTIVES: To assess the impact of risk factors on the disease control among chronic rhinosinusitis (CRS) patients, following 1 year of functional endoscopic sinus surgery (FESS), and combining the risk factors to formulate a convenient, visualised prediction model. DESIGN: A retrospective and nonconcurrent cohort study. SETTING AND PARTICIPANTS: A total of 325 patients with CRS from June 2018 to July 2020 at the First Affiliated Hospital of Sun Yat-sen University, the Third Affliated Hospital of Sun Yat-sen University, the Seventh Affiliated Hospital of Sun Yat-sen University. MAIN OUTCOMES MEASURES: Outcomes were time to event measures: the disease control of CRS after surgery 1 year. The presence of nasal polyps, smoking habits, allergic rhinitis (AR), the ratio of tissue eosinophil (TER) and peripheral blood eosinophil count (PBEC) and asthma was assessed. The logistic regression models were used to conduct multivariate and univariate analyses. Asthma, TER, AR, PBEC were also included in the nomogram. The calibration curve and area under curve (AUC) were used to evaluate the forecast performance of the model. RESULTS: In univariate analyses, most of the covariates had significant associations with the endpoints, except for age, gender and smoking. The nomogram showed the highest accuracy with an AUC of 0.760 (95% CI, 0.688-0.830) in the training cohort. CONCLUSIONS: In this cohort study that included the asthma, AR, TER, PBEC, which had significantly affected the disease control of CRS after surgery. The model provided relatively accurate prediction in the disease control of CRS after FESS and served as a visualised reference for daily diagnosis and treatment.
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Asma , Pólipos Nasais , Rinite Alérgica , Rinite , Sinusite , Asma/complicações , Doença Crônica , Estudos de Coortes , Humanos , Pólipos Nasais/complicações , Pólipos Nasais/diagnóstico , Pólipos Nasais/cirurgia , Estudos Retrospectivos , Rinite/complicações , Rinite/diagnóstico , Rinite/cirurgia , Rinite Alérgica/complicações , Fatores de Risco , Sinusite/diagnóstico , Sinusite/etiologia , Sinusite/cirurgiaRESUMO
The smut fungus Ustilago esculenta infects Zizania latifolia and induces stem expansion to form a unique vegetable named Jiaobai. Although previous studies have demonstrated that hormonal control is essential for triggering stem swelling, the role of hormones synthesized by Z. latifolia and U. esculenta and the underlying molecular mechanism are not yet clear. To study the mechanism that triggers swollen stem formation, we analyzed the gene expression pattern of both interacting organisms during the initial trigger of culm gall formation, at which time the infective hyphae also propagated extensively and penetrated host stem cells. Transcriptional analysis indicated that abundant genes involving fungal pathogenicity and plant resistance were reprogrammed to maintain the subtle balance between the parasite and host. In addition, the expression of genes involved in auxin biosynthesis of U. esculenta obviously decreased during stem swelling, while a large number of genes related to the synthesis, metabolism and signal transduction of hormones of the host plant were stimulated and showed specific expression patterns, particularly, the expression of ZlYUCCA9 (a flavin monooxygenase, the key enzyme in indole-3-acetic acid (IAA) biosynthesis pathway) increased significantly. Simultaneously, the content of IAA increased significantly, while the contents of cytokinin and gibberellin showed the opposite trend. We speculated that auxin produced by the host plant, rather than the fungus, triggers stem swelling. Furthermore, from the differently expressed genes, two candidate Cys2-His2 (C2H2) zinc finger proteins, GME3058_g and GME5963_g, were identified from U. esculenta, which may conduct fungus growth and infection at the initial stage of stem-gall formation.
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Basidiomycota/genética , Resistência à Doença/genética , Perfilação da Expressão Gênica/métodos , Doenças das Plantas/genética , Tumores de Planta/genética , Poaceae/genética , Sequência de Aminoácidos , Basidiomycota/metabolismo , Basidiomycota/patogenicidade , Proteínas Fúngicas/classificação , Proteínas Fúngicas/genética , Proteínas Fúngicas/metabolismo , Regulação Fúngica da Expressão Gênica , Regulação da Expressão Gênica de Plantas , Interações Hospedeiro-Patógeno/genética , Hifas/genética , Hifas/metabolismo , Hifas/patogenicidade , Ácidos Indolacéticos/metabolismo , Oxigenases/genética , Oxigenases/metabolismo , Filogenia , Doenças das Plantas/microbiologia , Reguladores de Crescimento de Plantas/biossíntese , Proteínas de Plantas/classificação , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Caules de Planta/genética , Caules de Planta/metabolismo , Caules de Planta/microbiologia , Tumores de Planta/microbiologia , Poaceae/metabolismo , Poaceae/microbiologia , Homologia de Sequência de Aminoácidos , Virulência/genéticaRESUMO
Oral squamous cell carcinoma (OSCC) is a prevalent cancer that develops in the head and neck area and has high annual mortality despite optimal treatment. microRNA-218 (miR-218) is a tumour inhibiting non-coding RNA that has been reported to suppress the cell proliferation and invasion in various cancers. Thus, our study aims to determine the mechanism underlying the inhibitory role of miR-218 in OSCC. We conducted a bioinformatics analysis to screen differentially expressed genes in OSCC and their potential upstream miRNAs. After collection of surgical OSCC tissues, we detected GREM1 expression by immunohistochemistry, RT-qPCR and Western blot analysis, and miR-218 expression by RT-qPCR. The target relationship between miR-218 and GREM1 was assessed by dual-luciferase reporter gene assay. After loss- and gain-of-function experiments, OSCC cell proliferation, migration and invasion were determined by MTT assay, scratch test and Transwell assay, respectively. Expression of TGF-ß1, Smad4, p21, E-cadherin, Vimentin and Snail was measured by RT-qPCR and Western blot analysis. Finally, effects of miR-218 and GREM1 on tumour formation and liver metastasis were evaluated in xenograft tumour-bearing nude mice. GREM1 was up-regulated, and miR-218 was down-regulated in OSCC tissues, and GREM1 was confirmed to be the target gene of miR-218. Furthermore, after up-regulating miR-218 or silencing GREM1, OSCC cell proliferation, migration and invasion were reduced. In addition, expression of TGF-ß signalling pathway-related genes was diminished by overexpressing miR-218 or down-regulating GREM1. Finally, up-regulated miR-218 or down-regulated GREM1 reduced tumour growth and liver metastasis in vivo. Taken together, our findings suggest that the overexpression of miR-218 may inhibit OSCC progression by inactivating the GREM1-dependent TGF-ß signalling pathway.
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Carcinoma de Células Escamosas/genética , Transição Epitelial-Mesenquimal/genética , Regulação Neoplásica da Expressão Gênica , Peptídeos e Proteínas de Sinalização Intercelular/genética , MicroRNAs/genética , Neoplasias Bucais/genética , Interferência de RNA , Adulto , Idoso , Animais , Carcinoma de Células Escamosas/diagnóstico , Linhagem Celular Tumoral , Movimento Celular/genética , Proliferação de Células/genética , Biologia Computacional/métodos , Bases de Dados Genéticas , Modelos Animais de Doenças , Feminino , Perfilação da Expressão Gênica , Genes Reporter , Xenoenxertos , Humanos , Peptídeos e Proteínas de Sinalização Intercelular/metabolismo , Masculino , Camundongos , Pessoa de Meia-Idade , Modelos Biológicos , Neoplasias Bucais/diagnóstico , Gradação de Tumores , Metástase Neoplásica , Estadiamento de Neoplasias , Transdução de Sinais , Fator de Crescimento Transformador beta/metabolismoRESUMO
Pseudorabies virus (PRV), the agent of pseudorabies, has raised considerable attention since 2011 due to the outbreak of emerging PRV variants in China. In the present study, we obtained two monoclonal antibodies (mAbs) known as 2E5 and 5C3 against the glycoprotein E (gE) of a PRV variant (JS-2012 strain). The two mAbs reacted with wild PRV but not the vaccine strain (gE-deleted virus). The 2E5 was located in 161RLRRE165, which was conserved in almost of all PRV strains, while 5C3 in 148EMGIGDY154 was different from almost of all genotype I PRV, in which the 149th amino acid is methionine (M) instead of arginine (R). The two epitopes peptides located in the hydrophilic region and reacted with positive sera against genotype II PRV (JS-2012), which suggests they were likely dominant B-cell epitopes. Furthermore, the mutant peptide 148ERGIGDY154 (genotype I) did not react with the mAb 5C3 or positive sera against genotype II PRV (JS-2012). In conclusion, both mAb 2E5 and 5C3 could be used to identify wild PRV strains from vaccine strains, and mAb 5C3 and the epitope peptide of 5C3 might be used for epidemiological investigation to distinguish genotype II from genotype I PRV.
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Anticorpos Monoclonais/imunologia , Epitopos de Linfócito B/imunologia , Herpesvirus Suídeo 1/química , Proteínas do Envelope Viral/imunologia , Animais , Linhagem Celular , Chlorocebus aethiops , Herpesvirus Suídeo 1/efeitos dos fármacos , Herpesvirus Suídeo 1/imunologia , Camundongos , Peptídeos/farmacologia , Suínos , Células Vero , Proteínas do Envelope Viral/antagonistas & inibidoresRESUMO
Porcine reproductive and respiratory syndrome virus (PRRSV) has been a major threat to global industrial pig farming ever since its emergence in the late 1980s. Identification of sustainable and effective control measures against PRRSV transmission is a pressing problem. The nucleocapsid (N) protein of PRRSV is specifically localized in the cytoplasm and nucleus of virus-infected cells which is important for PRRSV replication. In the current study, a new host restricted factor, Moloney leukemia virus 10-like protein (MOV10), was identified as an inhibitor of PRRSV replication. N protein levels and viral replication were significantly reduced in Marc-145â¯cells stably overexpressing MOV10 compared with those in wild-type Marc-145â¯cells. Adsorption experiments revealed that MOV10 did not affect the attachment and internalization of PRRSV. Co-immunoprecipitation and immunofluorescence co-localization analyses showed that MOV10 interacted and co-localized with the PRRSV N protein in the cytoplasm. Notably, MOV10 affected the distribution of N protein in the cytoplasm and nucleus, leading to the retention of N protein in the former. Taken together, these findings demonstrate for the first time that MOV10 inhibits PRRSV replication by restricting the nuclear import of N protein. These observations have great implications for the development of anti-PRRSV drugs and provide new insight into the role of N protein in PRRSV biology.
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Citoplasma/metabolismo , Proteínas do Nucleocapsídeo/química , Vírus da Síndrome Respiratória e Reprodutiva Suína/fisiologia , RNA Helicases/metabolismo , Replicação Viral , Criação de Animais Domésticos , Animais , Linhagem Celular , Chlorocebus aethiops , Replicação do DNA , Células HEK293 , Humanos , Vírus da Leucemia Murina de Moloney/metabolismo , Síndrome Respiratória e Reprodutiva Suína/metabolismo , Ligação Proteica , Suínos , Proteínas não Estruturais Virais/metabolismoRESUMO
Significant differences exist between the highly pathogenic (HP) porcine reproductive and respiratory syndrome virus (PRRSV) and its attenuated pathogenic (AP) strain in the ability to infect host cells. The mechanisms by which different virulent strains invade host cells remain relatively unknown. In this study, pulmonary alveolar macrophages (PAMs) are infected with HP-PRRSV (HuN4) and AP-PRRSV (HuN4-F112) for 24 h, then harvested and subjected to label-free quantitative MS. A total of 2849 proteins are identified, including 95 that are differentially expressed. Among them, 26 proteins are located on the membrane. The most differentially expressed proteins are involved in response to stimulus, metabolic process, and immune system process, which mainly have the function of binding and catalytic activity. Cluster of differentiation CD163, vimentin (VIM), and nmII as well as detected proteins are assessed together by string analysis, which elucidated a potentially different infection mechanism. According to the function annotations, PRRSV with different virulence may mainly differ in immunology, inflammation, immune evasion as well as cell apoptosis. This is the first attempt to explore the differential characteristics between HP-PRRSV and its attenuated PRRSV infected PAMs focusing on membrane proteins which will be of great help to further understand the different infective mechanisms of HP-PRRSV and AP-PRRSV.
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Macrófagos Alveolares/metabolismo , Proteínas de Membrana/análise , Síndrome Respiratória e Reprodutiva Suína/metabolismo , Vírus da Síndrome Respiratória e Reprodutiva Suína/patogenicidade , Proteoma/análise , Proteômica/métodos , Alvéolos Pulmonares/metabolismo , Animais , Células Cultivadas , Interações Hospedeiro-Patógeno , Macrófagos Alveolares/virologia , Síndrome Respiratória e Reprodutiva Suína/virologia , Vírus da Síndrome Respiratória e Reprodutiva Suína/classificação , Vírus da Síndrome Respiratória e Reprodutiva Suína/fisiologia , Alvéolos Pulmonares/virologia , Suínos , VirulênciaRESUMO
The affected infraorbital nerve (IFBN) and inferior alveolar nerve (IFAN) status in patients with jaw fibrous dysplasia has not been definitely depicted. In this study, the authors try to explore the status of affected IFBN and IFAN in patients with jaw fibrous dysplasia. Ten patients with jaw fibrous dysplasia were included in this study. The complaints of numbness in the IFBA and IFAN innervated area were asked and recorded, and careful clinical examination was performed to evaluate the touch sense, pain sense, pressure sense, and temperature sense in the IFBA and IFAN innervated areas. Computed tomography scans also were performed to evaluate the imaging characteristics of affected IFBA and IFAN. The results showed that 1 patient with maxillary lesion showed complaints of slight numbness, and clinical examination showed that the patient exhibited slight insensitive in pain sense. In addition, 1 patient with mandibular lesion showed relative obvious complaints of numbness, and clinical examination showed that the patient exhibited slight insensitive in pain sense and temperature sense, but not serious. All other patients exhibited no numbness in the IFBA and IFAN innervated area. Although the position and morphology changed in some patients, all neural canal of affected IFBA or IFAN existed and showed no invasion of lesion. Taking these findings together, it further confirmed that evaluation of the function of IFBAN and IFAN is necessary for patients with jaw fibrous dysplasia, and the affected IFBAN and IFAN may should be reserved in most patients with jaw fibrous dysplasia when resecting or recontouring the lesion.
Assuntos
Doenças dos Nervos Cranianos/etiologia , Displasia Fibrosa Óssea/complicações , Doenças Mandibulares/complicações , Doenças Maxilares/complicações , Adolescente , Adulto , Doenças dos Nervos Cranianos/fisiopatologia , Feminino , Displasia Fibrosa Óssea/fisiopatologia , Humanos , Hipestesia/etiologia , Hipestesia/fisiopatologia , Masculino , Mandíbula/inervação , Doenças Mandibulares/fisiopatologia , Nervo Mandibular/fisiologia , Maxila/inervação , Doenças Maxilares/fisiopatologia , Nervo Maxilar/fisiologia , Pressão , Distúrbios Somatossensoriais/etiologia , Distúrbios Somatossensoriais/fisiopatologia , Sensação Térmica/fisiologia , Tomografia Computadorizada por Raios X/métodos , Tato/fisiologia , Adulto JovemRESUMO
Photoexcitation of spin crossover (SCO) complexes can trigger extensive electronic spin transitions and transformation of molecular structure. However, the precise nature of the associated ultrafast structural dynamics remains elusive, especially in the solid state. Here, we studied a single-crystal SCO material with femtosecond electron diffraction (FED). The unique capability of FED allows us to directly probe atomic motions and to track ultrafast structural changes within a crystal lattice. By monitoring the time-dependent changes of the Bragg reflections, we observed the formation of a photoinduced structure similar to the thermally induced high-spin state. The data and refinement calculations indicate the global structural reorganization within 2.3â ps, as the metal-ligand bond distribution narrows during intramolecular vibrational energy redistribution (IVR) driving the intermolecular rearrangement. Three independent dynamical group are identified to model the structural dynamics upon photoinduced SCO.
RESUMO
Since the highly pathogenic porcine reproductive and respiratory syndrome virus (HP-PRRSV) variant emerged in 2006, it has caused death in more than 20 million pigs in China and other Southeast Asian countries, making it the most destructive swine pathogen currently in existence. To characterize the cellular responses to HP-PRRSV infection, the gene expression profile of porcine alveolar macrophage (PAM) cells, the primary target cells of PRRSV, was analyzed in HP-PRRSV-infected and uninfected PAMs by suppression subtractive hybridization. After confirmation by Southern blot, genes that were differentially expressed in the HP-PRRSV-infected and uninfected PAMs were sequenced and annotated. Genes that were upregulated mainly in HP-PRRSV-infected PAM cells were related to immunity and cell signaling. Among the differentially expressed genes, Mx1 and HSP70 protein expression was confirmed by western blotting, and IL-8 expression was confirmed by ELISA. In PAM cells isolated from HP-PRRSV-infected piglets, the differential expression of 21 genes, including IL-16, TGF-beta type 1 receptor, epidermal growth factor, MHC-I SLA, Toll-like receptor, hepatoma-derived growth factor, FTH1, and MHC-II SLA-DRB1, was confirmed by real-time PCR. To our knowledge, this is the first study to demonstrate differential gene expression between HP-PRRSV-infected and uninfected PAMs in vivo. The results indicate that HP-PRRSV infection excessively stimulates genes involved in the innate immune response, including proinflammatory cytokines and chemokines.