A study of antigen selection by extracellular vesicles as vaccine candidates against Mycobacterium tuberculosis infection.

Introduction. Tuberculosis (TB), an infectious disease caused by Mycobacterium tuberculosis (M. tb), remains a significant global public health concern. It is crucial to develop more effective vaccines for TB in order to achieve global control of the disease. Extracellular vesicles (EVs) are spherical membrane-bound structures released by pathogens and host cells. During the course of an infection, both pathogen- and host-derived EVs are produced and play important roles in determining the course of the infection. EVs offer intriguing tools as potential vaccines due to their ability to deliver multiple pathogen or host antigens.Hypothesis /Gap Statement. We hypothesized that EVs derived from M. tb and EVs from M. tb-infected macrophages may serve as potential vaccine candidates against M. tb infection.Aim. This study aims to compare the immunogenicity and immune protection between M. tb EVs and M. tb-infected macrophage-derived EVs.Methodology. In this study, EVs were extracted from culture supernatants of M. tb and M. tb-infected macrophages, respectively. Mass spectrometry was employed to explore the antigen composition of H37Rv-Mφ-EVs and H37Rv-EVs. Cytokine profiling and antibody response assays were used to analyse the immunogenicity offered by EVs. Additionally, we used histological examination to evaluate and protective efficacy of the EVs.Results. Our results demonstrated that mice immunized by EVs released from M. tb-infected macrophages induced stronger inflammatory cytokine response than M. tb. Moreover, EVs from M. tb-infected macrophages reinforced T-cell activation and antibody response compared to M. tb EVs. Proteomic analysis revealed that EVs from M. tb-infected macrophages containing immunodominant cargos have stronger binding ability with major histocompatibility complex molecules, which may contribute to the protection from M. tb infection. Indeed, immunization of EVs released from M. tb-infected macrophages significantly reduced the bacterial load and better protection against M. tb infection than EVs from M. tb. Importantly, the selected antigens (Ag85B, ESAT-6 and the Rv0580c) from EVs of M. tb-infected macrophages exhibited effective immunogenicity.Conclusion. Our results suggested that EVs derived from M. tb-infected macrophages might serve as a proper antigenic library for vaccine candidates against M. tb challenge.

Effect of desflurane anesthesia on flash visual evoked potential monitoring in patients undergoing spine surgery: study protocol for a randomized controlled trial.

BACKGROUND: Flash visual evoked potentials (FVEPs) are a reliable method for protecting visual function during spine surgery in prone position. However, the popularization and application of FVEPs remain limited due to the unclear influence of various anesthetics on FVEPs. Exploring the effects of anesthetic drugs on FVEP and establishing appropriate anesthesia maintenance methods are particularly important for promoting and applying FVEP. According to the conventional concept, inhaled narcotic drugs significantly affect the success of FVEP monitoring, FVEP extraction, and interpretation. Nonetheless, our previous study demonstrated that sevoflurane-propofol balanced anesthesia was a practicable regimen for FVEPs. Desflurane is widely used in general anesthesia for its rapid recovery properties. As the effect of desflurane on FVEP remains unclear, this trial will investigate the effect of different inhaled concentrations of desflurane anesthesia on amplitude of FVEPs during spine surgery, aiming to identify more feasible anesthesia schemes for the clinical application of FVEP. METHODS/ DESIGN: A total of 70 patients undergoing elective spinal surgery will be enrolled in this prospective, randomized controlled, open-label, patient-assessor-blinded, superiority trial and randomly assigned to the low inhaled concentration of desflurane group (LD group) maintained with desflurane-propofolremifentanil-balanced anesthesia or high inhaled concentration of desflurane group (HD group) maintained with desflurane-remifentanil anesthesia maintenance group at a ratio of 1:1. All patients will be monitored for intraoperative FVEPs, and the baseline will be measured half an hour after induction under total intravenous anesthesia (TIVA). After that, patients will receive 0.5 minimum alveolar concentration (MAC) of desflurane combined with propofol and remifentanil for anesthesia maintenance in the LD group, while 0.7-1.0 MAC of desflurane and remifentanil will be maintained in the HD group. The primary outcome is the N75-P100 amplitude 1 h after the induction of anesthesia. We intend to use the dual measure evaluation, dual data entry, and statistical analysis by double trained assessors to ensure the reliability and accuracy of the results. DISCUSSION: This randomized controlled trial aims to explore the superiority effect of low inhaled concentration of desflurane combined with propofolremifentanil-balanced anesthesia versus high inhaled concentration of desflurane combined with remifentanil anesthesia on amplitude of FVEPs. The study is meant to be published in a peer-reviewed journal and might guide the anesthetic regimen for FVEPs. The conclusion is expected to provide high-quality evidence for the effect of desflurane on FVEPs and aim to explore more feasible anesthesia schemes for the clinical application of FVEPs and visual function protection. TRIAL REGISTRATION: This study was registered on clinicaltrials.gov on July 15, 2022. CLINICALTRIALS: gov Identifier: NCT05465330.

Effect of esketamine combined with pregabalin on acute postsurgical pain in patients who underwent resection of spinal neoplasms: a randomized controlled trial.

ABSTRACT: Moderate-to-severe acute postsurgical pain (APSP) can prolong the recovery and worsen the prognosis of patients who undergo spinal surgery. Esketamine and pregabalin may resolve APSP without causing hyperpathia or respiratory depression after surgery. However, there are other risks, such as dissociative symptoms. We designed a randomized controlled trial to investigate the effect of the combination of these 2 drugs on the incidence of APSP in patients who underwent resection of spinal neoplasms. Patients aged 18 to 65 years were randomized to receive esketamine (a bolus dose of 0.5 mg·kg -1 and an infusion dose of 0.12 mg·kg -1 ·h -1 for 48 hours after surgery) combined with oral pregabalin (75-150 mg/day, starting 2 hours before surgery and ending at 2 weeks after surgery) or an identical volume of normal saline and placebo capsules. The primary outcome was the proportion of patients with moderate-to-severe APSP (visual analog scale score ≥ 40) during the first 48 hours after surgery. Secondary outcomes included the incidence of drug-related adverse events. A total of 90 patients were randomized. The incidence of moderate-to-severe APSP in the combined group (27.3%) was lower than that in the control group (60.5%) during the first 48 hours after surgery (odds ratio = 0.25, 95% CI = 0.10-0.61; P = 0.002). The occurrence of mild dissociative symptoms was higher in the combined group than in the control group (18.2% vs 0%). In conclusion, esketamine combined with pregabalin could effectively alleviate APSP after spinal surgery, but an analgesic strategy might increase the risk of mild dissociative symptoms.

Differential traits between microvesicles and exosomes in enterovirus infection.

Extracellular vesicles (EVs), including exosomes and microvesicles (MVs), are released by most cell types into the extracellular space and represent the pathophysiological condition of their source cells. Recent studies demonstrate that EVs derived from infected cells and tumors contribute to disease pathogenesis. However, very few studies have rigorously characterized exosomes and microvesicles in infectious diseases. In this study, we focused on subpopulations of EVs during the human enterovirus infection and explored the distinct traits and functions of EVs. We construct an effective immunomagnetic method to isolate exosomes and MVs from enterovirus-infected cells excluding virion. The morphology and sizes of exosomes and MVs have no significant alteration after enterovirus infection. Meanwhile, our study observed that the enterovirus infection could induce exosome secretion but not MVs. In vivo study showed that there was differential biodistribution between exosomes and MVs. Using deep RNA sequencing, we found that the cargo information in MVs rather than in exosomes could accurately reflect pathological condition of original cells. Our study demonstrated that it should be considered to use MVs as clinical diagnostics during in enterovirus infection because their composition is reflective of pathological changes.

Keratinocyte derived HMGB1 aggravates psoriasis dermatitis via facilitating inflammatory polarization of macrophages and hyperproliferation of keratinocyte.

Psoriasis is one of the most common immune-mediated chronic inflammatory skin diseases, involving excessive proliferation of keratinocyte and infiltration of immune cells. There are many factors that cause the onset of psoriasis, so the exact pathogenesis of psoriasis still needs to be determined. High mobility group box-1 (HMGB1), a pro-inflammatory cytokine, is closely related to the pathogenesis of various inflammatory diseases. However, there are few studies investigating the effects of HMGB1 on inflammatory dermatoses. Here, we found that keratinocyte in the the IMQ-treated skin lesions of psoriasis model mice expressed more HMGB1. Notably, HMGB1 produced by keratinocyte could promote the activation of inflammatory type macrophages without affecting the polarization of anti-inflammatory type macrophages. Meanwhile, the proportion of M1 type macrophages in the skin lesions is significantly increased. Moreover, local clearance of macrophages in the skin could alleviate psoriasis like inflammation. Finally, keratinocyte-derived HMGB1 could also act on itself in turn, promoting the excessive proliferation and the mRNA expression of inflammatory cytokines of keratinocyte. Therefore, this study not only found the effect of HMGB1 on the hyperproliferation of keratinocyte, but also revealed that keratinocyte could communicate with macrophages through HMGB1, thereby facilitating macrophage inflammatory polarization. Collectively, these findings have clinical significance for the research and treatment of psoriasis, HMGB1 may become a potential target for the treatment of psoriasis.

Caffeic acid phenethyl ester suppresses metastasis of breast cancer cells by inactivating FGFR1 via MD2.

BACKGROUND: Tumor metastasis is the main cause of death for breast cancer patients. Caffeic acid phenethyl ester (CAPE) has strong anti-tumor effects with very low toxicity and may be a potential candidate drug. However, the anti-metastatic effect and molecular mechanism of CAPE on breast cancer need more research. METHODS: MCF-7 and MDA-MB-231 breast cancer cells were used here. Wound healing and Transwell assay were used for migration and invasion detection. Western blot and RT-qPCR were carried out for the epithelial-to-myofibroblast transformation (EMT) process investigation. Western blot and immunofluorescence were performed for fibroblast growth factor receptor1 (FGFR1) phosphorylation and nuclear transfer detection. Co-immunoprecipitation was used for the FGFR1/myeloid differentiation protein2 (MD2) complex investigation. RESULTS: Our results suggested that CAPE blocks the migration, invasion, and EMT process of breast cancer cells. Mechanistically, CAPE inhibits FGFR1 phosphorylation and nuclear transfer while overexpression of FGFR1 reduces the anti-metastasis effect of CAPE. Further, we found that FGFR1 is bound to MD2, and silencing MD2 inhibits FGFR1 phosphorylation and nuclear transfer as well as cell migration and invasion. CONCLUSION: This study illustrated that CAPE restrained FGFR1 activation and nuclear transfer through MD2/FGFR1 complex inhibition and showed good inhibitory effects on the metastasis of breast cancer cells.

Identification and validation of the microRNAs and hub genes for pancreatic ductal adenocarcinoma by an integrated bioinformatic analysis.

Background: In the progression of pancreatic ductal adenocarcinoma (PDAC), aberrant micro RNAs (miRNAs) expression plays a crucial role. This study sought to identify and validate the key miRNAs and potential target genes involved in PDAC. A bioinformatic analysis was conducted to determine their potential use as biomarkers and therapeutic targets. Methods: Gene profiling data sets (GSE41372 and GSE32688) were retrieved from the Gene Expression Omnibus database. Differentially expressed miRNAs (DEMs) with a P value <0.05, and |fold change| >2 was identified. The prognostic value of the DEMs was accessed using the online server Kaplan-Meier plotter. Further, gene ontology terms and Kyoto Encyclopedia of Genes and Genomes pathway analyses were performed using DAVID 6.7. The protein-protein interaction analyses were conducted with STRING, and miRNA-hub gene networks were constructed using Cytoscape software. The PDAC cells were transfected with miRNA inhibitors or mimics. Cell Counting Kit-8 assays and terminal deoxynucleotidyl transferase dUTP nick end labeling (TUNEL) staining were used to examine cell proliferation and apoptosis, respectively. Wound-healing assays were performed to evaluate cell migration. Results: Three DEMs (hsa-miR-21-5p, hsa-miR-135b-5p, and hsa-miR-222-3p) were identified. High expression levels of hsa-miR-21-5p, hsa-miR-135b-5p, or hsa-miR-222-3p predicted poor overall survival in PDAC patients. The pathway analysis revealed that the predicted target genes of the DEMs were closely related to several signaling pathways (including 'pathways in cancer', 'miRNAs in cancer', 'platinum drug resistance', 'lipid and atherosclerosis', and 'MAPK signaling pathway'). The MYC proto-oncogene (MYC), phosphate and tensin homolog gene (PTEN), poly(ADP-ribose) polymerase 1 (PARP1), von Hippel-Lindau (VHL), and fork head box p3 (FOXP3) were identified as potential target genes. The inhibition of hsa-miR-21-5p, hsa-miR-135b-5p, or hsa-miR-222-3p expression decreased cell proliferation. The overexpression of hsa-miR-21-5p, hsa-miR-135b-5p, or hsa-miR-222-3p facilitated PDAC cell migration. Conclusions: This study constructed the miRNA-hub gene network, which provides novel insights into the PDAC progression. Although further research is required, our results offer clues for new potential prognostic markers and therapeutic targets of PDAC.

Effects of Ketamine on Chronic Postsurgical Pain in Patients Undergoing Surgery: A Systematic Review and Meta-analysis.

BACKGROUND: Chronic postsurgical pain (CPSP) has become a common complication during the perioperative period. The efficacy of one of the most potent strategies, ketamine, remains unclear. OBJECTIVES: The aim of this meta-analysis was to evaluate the effect of ketamine on CPSP in patients undergoing common surgeries.. STUDY DESIGN: Systematic review and meta-analysis. METHODS: English-language randomized controlled trials (RCTs) published in MEDLINE, Cochrane Library, and EMBASE from 1990 through 2022 were screened. RCTs with a placebo control group that evaluated the effect of intravenous ketamine on CPSP in patients undergoing common surgeries were included. The primary outcome was the proportion of patients who experienced CPSP 3 - 6 months postsurgery. The secondary outcomes included adverse events, emotional evaluation, and 48 hour postoperative opioid consumption. We followed the Preferred Reporting Items for Systematic Reviews and Meta-analyses (PRISMA) guidelines. Pooled effect sizes were measured using the common-effects model or random-effects model, and several subgroup analyses were conducted. RESULTS: Twenty RCTs were included with 1,561 patients. Our pooled meta-analysis showed a significant difference between ketamine and placebo in the treatment of CPSP (Relative Risk [RR] = 0.86; 95% CI, 0.77 - 0.95; P = 0.02; I2 = 44%). In the subgroup analyses, our results indicated that compared with placebo, intravenous ketamine might decrease the prevalence of CPSP 3 - 6 months postsurgery (RR = 0.82; 95% CI, 0.72 - 0.94; P = 0.03; I2 = 45%). For adverse events, we observed that intravenous ketamine might lead to hallucinations (RR = 1.61; 95% CI, 1.09 - 2.39; P = 0.27; I2 = 20%) but did not increase the incidence of postoperative nausea and vomiting (RR = 0.98; 95% CI, 0.86 - 1.12; P = 0.66; I2 = 0%). LIMITATIONS: Inconsistent assessment tools and follow-up for chronic pain may contribute to the high heterogeneity and limitation of this analysis. CONCLUSIONS: We discovered that intravenous ketamine may reduce the incidence of CPSP in patients undergoing surgery, especially 3 - 6 months postsurgery. Because of the small sample size and high heterogeneity of the included studies, the effect of ketamine in the treatment of CPSP still needs to be explored in future large-sample, standardized-assessment studies.

miR-9a-5p Protects Ischemic Stroke by Regulating Oxidative Stress and Mitochondrial Autophagy.

Purpose: Present research is aimed at exploring the effect of miR-9a-5p on mitochondrial autophagy and alleviating cellular oxidative stress injury in ischemic stroke. Methods: SH-SY5Y cells were cultured with oxygen-glucose deprivation/reoxygenation (OGD/R) to simulate ischemia/reperfusion. The cells were treated in an anaerobic incubator (95% N2, 5% CO2) for 2 h and then reoxygenated in the normoxic condition for 24 h with 2 ml of normal medium. Cells were transfected with miR-9a-5p mimic/inhibitor or negative control. The RT-qPCR assay was utilized to measure the mRNA expression. Western blot was utilized to evaluate the protein expression. The CCK-8 assay was conducted to detect cell viability. Flow cytometry was applied to examine apoptosis and the cell cycle. The ELISA assay was applied to measure the contents of SOD and MDA in mitochondria. Autophagosomes were observed via electron microscopy. Results: By comparison with the control group, the miR-9a-5p expression in the OGD/R group obviously declined. Mitochondrial crista breaks, vacuole-like changes, and increased autophagosome formation were observed in the OGD/R group. OGD/R injury enhanced oxidative stress damage and mitophagy. When transfected with the miR-9a-5p mimic, mitophagosome production of SH-SY5Y cells decreased and oxidative stress injury was inhibited. However, the miR-9a-5p inhibitor obviously increased mitophagosome production and enhanced oxidative stress injury. Conclusion: miR-9a-5p protects against ischemic stroke by inhibiting OGD/R-induced mitochondrial autophagy and alleviating cellular oxidative stress injury.

Exosomes mediate Coxsackievirus B3 transmission and expand the viral tropism.

Specific virus-receptor interactions are important determinants in viral host range, tropism and pathogenesis, influencing the location and initiation of primary infection as well as viral spread to other target organs/tissues in the postviremic phase. Coxsackieviruses of Group B (CVB) and its six serotypes (CVB1-6) specifically interact with two receptor proteins, coxsackievirus-adenovirus receptor (CAR) and decay-accelerating factor (DAF), and cause various lesions in most permissive tissues. However, our previous data and other studies revealed that virus receptor-negative cells or tissues can be infected with CVB type 3 (CVB3), which can also effectively replicate. To study this interesting finding, we explored the possibility that exosomes are involved in CVB3 tropism and that exosomes functionally enhance CVB3 transmission. We found that exosomes carried and delivered CVB3 virions, resulting in efficient infection in receptor-negative host cells. We also found that delivery of CVB3 virions attached to exosomes depended on the virus receptor CAR. Importantly, exosomes carrying CVB3 virions exhibited greater infection efficiency than free virions because they accessed various entry routes, overcoming restrictions to viral tropism. In vivo experiments demonstrated that inhibition of exosome coupling with virions attenuated CVB3-induced immunological system dysfunction and reduced mortality. Our study describes a new mechanism in which exosomes contribute to viral tropism, spread, and pathogenesis.

Factors associated with artificial airway retention after skull base chordoma resection: A retrospective cohort study.

Background: Chordoma is a malignant bone and soft tissue tumor derived from embryonic notochord remnants, and skull base chordoma accounts for ~1/3 of all chordoma cases. Skull base chordoma is closely related to the brainstem and cranial nerves and has a high recurrence rate. The purpose of this study was to investigate the influence of the timing of tracheal extubation on perioperative pulmonary complications. We also aimed to explore predictors of postoperative artificial airway (AA) retention in patients with skull base chordoma. Methods: This was a single-center, retrospective cohort study. The study population included all skull base chordoma patients undergoing surgical treatment between January 2019 and December 2021 at Beijing Tiantan Hospital. The primary outcome was the incidence of postoperative pulmonary complications. Several patient characteristics were evaluated for potential associations with AA retention. Results: A total of 310 patients with skull base chordoma were enrolled. The frequency of AA retention after surgery for skull base chordoma was 30.97%. The incidence of postoperative pulmonary complications was much lower in those without AA retention (3.74 vs. 39.58%, P < 0.001). Factors with the highest point estimates for the odds of AA retention included body mass index, cranial nerve involvement, maximum tumor diameter, operative method, hemorrhage volume, operative duration and intraoperative mechanical ventilation duration. Conclusions: In this retrospective cohort study, most of the factors associated with postoperative airway retention were closely related to the patient's tumor characteristics. These data demonstrate that respiratory management in patients with skull base chordoma remains an ongoing concern.

Effect of esketamine on perioperative depressive symptoms in major surgery patients (PASSION II): study protocol for a randomised controlled trial.

INTRODUCTION: Depressive symptoms are common for patients undergoing major surgery and may worsen their mental health and lead to poor clinical outcomes. It is essential to seek a safe rapid-acting treatment for relieving moderate-to-severe depressive symptoms in patients undergoing major surgery. METHODS AND ANALYSIS: This study is a randomised, placebo-controlled and double-blinded trial aiming to determine the effect of esketamine on moderate-to-severe depressive symptoms in patients undergoing major surgery. Five hundred and sixty-four participants, aged 18-65 years old, undergoing major surgery will be randomly allocated into the esketamine and placebo groups at a 1:1 ratio. Esketamine or placebo will be given intravenously at the same speed on suturing the incision by anaesthesiologists in charge who are blinded to the randomisation. In the esketamine group, the total dosage of esketamine will be 0.2 mg/kg body weight. To estimate the efficacy and safety endpoints, blinded evaluation by trained researchers will be completed at 3 days, 5 days, 1 month, 3 months and 6 months after surgery. The primary outcome is the remission rate at the third postoperative day. The secondary outcomes include depression-related scores, severe pain events and safety-related endpoints such as psychotic symptoms, manic symptoms and dissociative symptoms. ETHICS AND DISSEMINATION: This study was approved by the Institutional Review Board of Beijing Tiantan Hospital, Capital Medical University, Beijing, China on 30 October 2020 (KY-2020-058-02). This trial is designed to explore whether the administration of esketamine could improve the mental health of patients with depressive symptoms undergoing major surgery. The conclusions of this study will be published in peer-reviewed journals. TRIAL REGISTRATION NUMBER: NCT04425473.

Nonstructural Protein NSs Hampers Cellular Antiviral Response through LSm14A during Severe Fever with Thrombocytopenia Syndrome Virus Infection.

The nonstructural protein (NSs) of severe fever with thrombocytopenia syndrome virus (SFTSV) plays multiple functions in the virus life cycle. Proteomic screening for host proteins interacting with NSs identified the cellular protein LSm14A. LSm14A, a member of the LSm family involved in RNA processing in the processing bodies, binds to viral RNA or synthetic homolog and mediates IFN regulatory factor 3 activation and IFN-ß induction. NSs interacted with and colocalized with LSm14A, and this interaction effectively inhibited downstream phosphorylation and dimerization of IFN regulatory factor 3, resulting in the suppression of antiviral signaling and IFN induction in several cell types of human origin. Knockdown of NSs resulted in the suppression of SFTSV replication in host cells. Viral RNA bound to LSm14A-NSs protein complex during the interaction. A newly discovered LRRD motif of NSs functioned to interact with LSm14A. Altogether, our data demonstrated a mechanism used by SFTSV to inhibit host innate immune response.

Cathelicidin antimicrobial peptides suppress EV71 infection via regulating antiviral response and inhibiting viral binding.

Cathelicidin antimicrobial peptides (human LL-37 and mouse CRAMP) are mainly virucidal to enveloped virus. However, the effects and relative mechanisms of LL-37 and CRAMP on non-enveloped virus are elusive. We herein found that CRAMP expression was significantly up-regulated post non-enveloped Enterovirus 71 (EV71) infection in different tissues of newborn ICR mice, while EV71 replication gradually declined post CRAMP up-regulation, indicating the antiviral potential of cathelicidin against EV71. In vitro antiviral assay showed that LL-37 and CRAMP markedly reduced cytopathic effects (CPE), intracellular viral RNA copy numbers, viral VP1 protein levels, and extracellular virons in U251 cells post EV71 infection, indicating that LL-37 and CRAMP significantly inhibited EV71 replication. Mechanism of action assay showed that LL-37 and CRAMP were not virucidal to EV71, but markedly regulated antiviral immune response in U251 cells. Co-incubation of LL-37 or CRAMP with U251 cells markedly increased the basal interferon-ß (IFN-ß) expression and interferon regulatory transcription factor 3 (IRF3) phosphorylation, modestly enhanced IFN-ß production and IRF3 phosphorylation upon EV71 infection, and significantly reduced interleukin-6 (IL-6) production and p38 mitogen-activated protein kinase (MAPK) activation post EV71 infection. Additionally, LL-37 and CRAMP directly inhibited viral binding to U251 cells. Collectively, LL-37 and CRAMP markedly inhibited EV71 replication via regulating antiviral response and inhibiting viral binding, providing potent candidates for peptide drug development against EV71 infection.

USP39 Serves as a Deubiquitinase to Stabilize STAT1 and Sustains Type I IFN-Induced Antiviral Immunity.

Deubiquitinating enzymes (DUBs) are cysteine proteases that reverse the ubiquitination by removing ubiquitins from the target protein. The human genome encodes ∼100 potential DUBs, which can be classified into six families, influencing multiple cellular processes, such as antiviral responses, inflammatory responses, apoptosis, etc. To systematically explore the role of DUBs involved in antiviral immunity, we performed an RNA interference-based screening that contains 97 human DUBs. We identified that ubiquitin-specific protease (USP) 39 expression modulates the antiviral activity, which is, to our knowledge, a previously unknown function of this enzyme. Small interfering RNA knockdown of USP39 significantly enhanced viral replication, whereas overexpression of USP39 had an opposite effect. Mechanistically, USP39 does not affect the production of type I IFN but significantly promotes JAK/STAT downstream of type I signaling by enhancing IFN-stimulated response elements promoter activity and expression of IFN-stimulated genes. Interestingly, USP39, previously considered not to have the deubiquitinase activity, in this study is proved to interact with STAT1 and sustain its protein level by deubiqutination. Furthermore, we found that through novel mechanism USP39 can significantly decrease K6-linked but not K48-linked ubiquitination of STAT1 for degradation. Taken together, these findings uncover that USP39 is, to our knowledge, a new deubiquitinase that positively regulates IFN-induced antiviral efficacy.

Severe Fever With Thrombocytopenia Syndrome Virus-Induced Macrophage Differentiation Is Regulated by miR-146.

Severe fever with thrombocytopenia syndrome (SFTS) is an emerging hemorrhagic fever with a high mortality rate in humans, which is caused by SFTS virus (SFTSV), a novel phlebovirus in the Bunyaviridae family, is tick borne and endemic in Eastern Asia. Previous study found that SFTSV can infect and replicate in macrophages in vivo and in vitro. However, the role of macrophages in virus replication and the potential pathogenic mechanisms of SFTSV in macrophage remain unclear. In this study, we provided evidence that the SFTSV infection drove macrophage differentiation skewed to M2 phenotype, facilitated virus shedding, and resulted in viral spread. We showed evidence that miR-146a and b were significantly upregulated in macrophages during the SFTSV infection, driving the differentiation of macrophages into M2 cells by targeting STAT1. Further analysis revealed that the elevated miR-146b but not miR-146a was responsible for IL-10 stimulation. We also found that SFTSV increased endogenous miR-146b-induced differentiation of macrophages into M2 cells mediated by viral non-structural protein (NSs). The M2 skewed differentiation of macrophages may have important implication to the pathogenesis of SFTS.

MOV10 interacts with Enterovirus 71 genomic 5'UTR and modulates viral replication.

As a cytoplasmic parasite, RNA virus develops sophisticated mechanisms to counter host defense and utilize host proteins to facilitate its replication. Here we found Moloney leukemia virus 10 (MOV10), a highly conserved cellular protein belonging to SF1 helicase family, played critical roles in EV71 infection. Silencing cellular MOV10 could restrict EV71 replication, while over-expressing MOV10 resulted in increased viral replication at low dosage and repressed viral replication at high dosage. Further investigation showed that MOV10 exhibited dual functions in EV71 regulation, its C-terminus positively regulated viral replication by binding to EV71 cloverleaf-like structure and the internal ribosome entry site while the N-terminus showed a potential antiviral activity when individually overexpressed. In addition, RNA-dependent interaction between MOV10 and HuR as well as the co-localization of MOV10 and processing bodies were also observed post infection. Taken together, our data indicate a crucial role of MOV10 in EV71 infection for the first time, providing new insights for its roles in EV71 infection.

Enterovirus 71 induces autophagy by regulating has-miR-30a expression to promote viral replication.

Enterovirus 71 (EV71), the etiological agent of hand-foot-and-mouth disease, has increasingly become a public health challenge around the world. Previous studies reported that EV71 infection can induce autophagic machinery to enhance viral replication in vitro and in vivo, but did not address the underlying mechanisms. Increasing evidence suggests that autophagy, in a virus-specific manner, may function to degrade viruses or facilitate viral replication. In this study, we reported that EV71 infection of human epidermoid carcinoma (Hep2) and African green monkey kidney cells (Vero) induced autophagy, which is beneficial for viral replication. Our investigation of the mechanisms revealed that EV71 infection resulted in the reduction of cellular miR-30a, which led to the inhibition of Beclin-1, a key autophagy-promoting gene that plays important roles at the early phase of autophagosome formation. We provided further evidence that by modulating cellular miR-30a level through either overexpression or inhibition, one can inhibit or promote EV71 replication, respectively, through regulating autophagic activity.

Harmine blocks herpes simplex virus infection through downregulating cellular NF-κB and MAPK pathways induced by oxidative stress.

Herpes simplex virus types 1 and 2 (HSV-1 and -2) are highly prevalent in many populations and therapeutic options are limited. Both viruses can establish latency by maintaining viral genomes in neurons of sensory ganglia. Primary or recurrent HSV infections may lead to deleterious outcomes: HSV-1 infection may result in corneal blindness and encephalitis and HSV-2 infection leads to herpes genitalis. While no effective vaccine is available, acyclovir is widely used for therapy, which targets and inhibits viral DNA polymerase. Although acyclovir is of low toxicity, resistant strains arise due to persistent use, mainly in immune compromised patients. In our effort to identify new HSV inhibitory molecules, harmine was found to potently inhibit HSV infection. Harmine, a beta-carbon alkaloid with an indole core structure and a pyridine ring, is widely distributed in plants. Earlier studies showed that harmine exhibited pharmacological activities such as antifungal, antimicrobial, antitumor, antiplasmodial and antioxidants. In the current study, we showed that harmine was a potent inhibitor of HSV-2 infection in vitro assays with EC50 value at around 1.47µM and CC50 value at around 337.10µM. The HSV RNA transcription, protein synthesis, and virus titers were reduced by the presence of harmine in a dose dependent manner. Further study on the mechanism of the anti-HSV activity showed that harmine blocked HSV-induced ROS production and the upregulated cytokine/chemokine expression, but our evidence showed that the inhibition of viral replication was unlikely mediated by the blocking of ROS production. We demonstrated that harmine significantly reduced HSV-2-induced NF-κB activation, as well as IκB-α degradation and p65 nuclear translocation. We found that harmine also inhibited HSV-2-mediated p38 kinase and c-Jun N-terminal kinases (JNK) phosphorylation.