The internal ribosome entry site (IRES) is a cis-acting element found in certain RNA viruses. In virus-infected cells, the Senecavirus A (SVA) IRES can directly recruit the small ribosomal subunit to an internal initiation codon on the mRNA, enabling translation initiation independently of both the 5' cap structure of the viral genome and the host cell eukaryotic initiation factor 4F (eIF4F). As a small RNA virus, SVA frequently accumulates genetic mutations during transmission. This study aimed to characterize the mutational and evolutionary patterns of the SVA genome during its transmission within host cells exhibiting enhanced innate immunity. SVA was serially passaged for 80 generations in preactivated 3D4/21 cells that had been established in an antiviral innate immune state. During serial passaging, two stable single-nucleotide mutations were consistently identified in the IRES region of the viral genomic RNA from passages 60-80: a uridine (U) insertion at genomic position 109, and a guanine (G)-to-adenine (A) substitution at position 265. Furthermore, the rescued SVA mutants harboring these mutations exhibited significantly higher replication titers than the parental strain. Notably, our results indicated that the mutant virus was unable to evade host antiviral innate immunity, whereas it significantly enhanced the translational activity of the SVA IRES. Collectively, these findings provide a foundation for understanding how single nucleotide polymorphisms (SNPs) in the 5' untranslated region (UTR) region influence viral IRES activity and the replication capacity of recombinant viruses.

Lipid metabolism plays a crucial role in cellular signal transduction, affects the structural integrity of cell membranes, and regulates energy metabolism. However, various viruses, including African swine fever virus (ASFV), usurp lipid metabolism to enhance their replication. The mechanism for the positive role of lipid metabolism in ASFV infection is unclear. Here, we present data that ASFV infection concurrently upregulates both fatty acid synthesis (FAS) and fatty acid β-oxidation (FAO) to enhance viral replication. Pharmacological inhibition of FAS significantly suppresses ASFV replication, an effect that can be markedly reversed by exogenous palmitate (the end product of FAS). Similarly, inhibition of FAO also impairs viral replication. Lipidomic profiling revealed that ASFV infection dramatically alters lipid droplet (LD) lipid composition, particularly triglycerides (TG) and diacylglycerols (DAG). ASFV infection triggers the accumulation of LDs, which in turn promote viral replication. Mechanistically, we discovered that ASFV exploits chaperone-mediated autophagy (CMA) to degrade perilipin 2 (PLIN2), a protein on the LD surface, thereby stimulating lipolysis. Furthermore, ASFV infection induces LD-mitochondrion contacts, facilitating the transfer of LD-derived fatty acids to mitochondria. These data indicate that LDs provide lipids to fuel ASFV-induced FAO upregulation. Collectively, our study reveals that ASFV orchestrates a complex metabolic network involving FAS, LD biogenesis, lipolysis, and FAO to optimize viral replication. These findings elucidate the pivotal role of lipid metabolism in ASFV replication, revealing a mechanism through which the virus manipulates cellular lipid pathways to facilitate its replication. This insight not only advances our understanding of ASFV pathogenesis but also presents potential therapeutic avenues for inhibiting viral production by modulating lipid metabolic processes.IMPORTANCEAfrican swine fever (ASF), caused by African swine fever virus (ASFV), represents a catastrophic threat to the global swine industry, with no safe and effective vaccines or antiviral therapies currently available except in Vietnam. Understanding how ASFV reprograms host lipid metabolism is critical for developing targeted interventions. Our study reveals a novel metabolic hijacking strategy employed by ASFV to reprogram lipid metabolism pathways, including fatty acid synthesis (FAS), lipid droplet (LD) biogenesis, chaperone-mediated autophagy (CMA)-mediated lipolysis, and mitochondrial β-oxidation (FAO), to support viral replication. Notably, we provide evidence that ASFV exploits CMA to degrade perilipin 2 (PLIN2), a key protein stabilizing lipid droplets, thereby promoting lipolysis. This mechanism resolves the paradox of concurrent upregulation of FAS and FAO by facilitating lipid shuttling through LD-mitochondrion contacts. Our findings offer new insights into how ASFV exploits host lipid networks and may pave the way for designing vaccines or targeted drugs to control ASF.

[This corrects the article DOI: 10.3389/fonc.2025.1553626.].

African swine fever virus (ASFV) infection causes a severe hemorrhagic disease in pigs, characterized by excessive inflammatory responses and tissue damage, posing substantial threats to the pig industry worldwide. Given the lack of vaccines and effective antiviral treatments, as well as the largely unknown functions of most ASFV-encoded proteins, it's urgent to study the proteins that are crucial in triggering inflammatory responses and how they do so. This study demonstrated that ASFV exploited the NOD-, LRR- and pyrin domain-containing protein 3 (NLRP3) inflammasome to induce pyroptosis and inflammatory responses, effectively replacing the non-functional porcine AIM2 pseudogene. Screening over 150 proteins encoded by the ASFV genome, EP364R was identified as the viral factor responsible for driving NLRP3-mediated pyroptosis and high-level cytokine production. Ectopic expression of EP364R in mice elicited significant upregulation of serum pro-inflammatory cytokines and splenomegaly, while its expression in bone marrow-derived macrophages (BMDMs) from NLRP3-knockout mice abrogated pyroptosis and related effects. Mechanistic investigation revealed that the helicase DDX3X acted as a molecular bridge, enabling EP364R to interact with NLRP3 to promote the aggregation and activation of inflammasomes. Depletion of DDX3X abolished EP364R's ability to induce NLRP3-dependent pyroptosis and pro-inflammatory cytokine production. We found that the NACHT domain of porcine NLRP3 interacted with DDX3X, and EP364R established a connection with the NACHT and LRR domains of NLRP3 through DDX3X. However, EP364R bound to all the domains of DDX3X. Molecular docking analysis revealed that DDX3X interacted with EP364R through a spatially defined interface, thereby exerting its function. Furthermore, a natural compound library was employed to screen functional compounds targeting EP364R, and HAMNO was identified as an inhibitor that bound to E256, K259, and D260 of EP364R, consequently suppressing ASFV replication. Our findings explain how ASFV triggers pyroptosis and excessive cytokine release, and identify a potent small-molecule inhibitor of ASFV, aiding the development of vaccines and therapies to prevent and control African swine fever (ASF) caused by ASFV infection.

The segmented genome of live-attenuated rotavirus (RV) vaccines provides an ideal platform for developing multivalent vaccines against enteric pathogens. Here, we engineered the commercial human RV vaccine strain LLR as a vector to deliver Clostridium perfringens alpha-toxin (CPA), a critical virulence factor associated with gastroenteritis. A screen of four 2A peptide sequences identified that porcine teschovirus-1 2A (P2A) preceded by an N-terminal GSG spacer enabled optimal cleavage and antigen expression. In mice, the resulting recombinant virus elicited potent systemic antibody responses against both RV and CPA following oral administration. When tested in a maternal immunization model, intramuscular immunization of cows generated high titers of neutralizing antibodies in colostrum, demonstrating its potential utility for passive immunization. By integrating a key bacterial antigen into a licensed RV vaccine backbone, we establish a versatile strategy to combat viral-bacterial co-infections and polymicrobial enteric diseases, offering a promising new approach to reduce the global burden of diarrheal mortality.IMPORTANCEThe lack of vaccines targeting polymicrobial enteric infections represents a critical gap in the fight against diarrheal diseases, a leading cause of infant mortality worldwide. To bridge this gap, we engineered a novel bivalent vaccine designed to provide dual protection against rotavirus and Clostridium perfringens alpha-toxin (CPA)-mediated disease. Leveraging an enhanced reverse genetics system, we successfully utilized the commercialized rotavirus vaccine strain LLR as a viral vector to express the key C. perfringens virulence factor, CPA. This strategy not only offers a path to broader protection against diarrheal disease but also establishes a versatile platform for developing vaccines against other viral-bacterial co-infections.

Viruses, lacking an intrinsic metabolic network, exploit host cell metabolism in order to hijack resources for their own replication. However, the regulatory relationship between foot-and-mouth disease virus (FMDV) and energy metabolism remains incompletely elucidated. Here, metabolomic analysis was performed on PK-15 cells and tonsils infected with FMDV. Metabolites involved in glycolysis were significantly upregulated, whereas metabolites associated with the tricarboxylic acid (TCA) cycle were markedly downregulated in vivo and in vitro, which indicated that FMDV induced reprogramming of host cell energy metabolism, shifting it from oxidative phosphorylation (OXPHOS) to glycolysis. Furthermore, FMDV infection promoted glucose uptake and its subsequent utilization in FMDV-infected PK-15 cells, facilitating viral replication. Mechanistically, on the one hand, FMDV infection activated the AKT-MTOR-dependent macroautophagy/autophagy pathway to suppress the expression of OTUD4 (OTU deubiquitinase 4), upregulating HK2 (hexokinase 2) to facilitate glycolysis. On the other hand, FMDV infection decreased the level of mitochondrial SIRT3 (sirtuin 3) through PINK1-PRKN-dependent mitophagy, leading to the increase of HIF1A (hypoxia inducible factor 1 subunit alpha), HK2, phosphofructokinase 1 (PFK1), and PKM/PKM2 (pyruvate kinases M1/2), promoting glycolysis. Overall, this study elucidates how FMDV modulates glycolysis through two different autophagic pathways, which would contribute to our understanding of how the autophagy-glycolysis axis regulates viral replication, providing new avenues for developing antiviral strategies targeting metabolism.Abbreviations: 3-MA: 3-methyladenine; ACTB: actin beta; ATG: autophagy related; CQ: chloroquine; dpi: days post-infection; EV71: enterovirus 71; FMDV: foot-and-mouth disease virus; HK2: hexokinase 2; HIF1A: hypoxia inducible factor 1 subunit alpha; hpt: hours post-transfection; MOI: multiplicity of infection; MTOR: mechanistic target of rapamycin kinase; OTUD4: OTU deubiquitinase 4; OXPHOS: oxidative phosphorylation; PFK1: phosphofructokinase 1; PFKFB3: 6-phosphofructo-2-kinase/fructose-2,6-bisphosphatase 3; PKM/PKM2: pyruvate kinase M1/2; SIRT3: sirtuin 3; siRNA: small interfering RNA; TCID50: 50% tissue culture infectious doses; TCA: tricarboxylic acid; TUNEL: terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling; WT: wild type.

Foot-and-mouth disease virus (FMDV), a highly contagious picornavirus, employs multifaceted strategies to evade host innate immunity, with viral proteins 3C protease (3Cpro) and 2B serving as key immune antagonists. The stimulator of interferon genes 1 (STING1) is a critical innate immune adaptor; however, its role and regulatory mechanisms during FMDV infection remain incompletely understood. Here, we report that STING1 inhibits FMDV replication through an interferon (IFN)-independent mechanism, while FMDV counteracts this antiviral effect by degrading STING1 via 3Cpro and 2B. Mechanistically, FMDV 3Cpro mediates STING1 degradation in a protease activity-dependent manner; this STING1-degrading activity is conserved among 3Cpro proteins of poliovirus, enterovirus 71, and coxsackievirus, but not senecavirus A. In contrast, FMDV 2B suppresses STING1 expression at the mRNA level, and neither proteasomal, lysosomal, nor caspase pathways are involved in 3Cpro/2B-mediated STING1 downregulation. Furthermore, the STING1 stabilizer SB24011 enhances endogenous STING1 expression, dose-dependently inhibits FMDV replication by targeting viral internal ribosome entry site (IRES)-mediated translation, and exhibits broad-spectrum antiviral activity against multiple picornaviruses. In vivo, SB24011 treatment alleviates virus-induced histopathological lesions. Collectively, our findings reveal a novel IFN-independent antiviral role of STING1 against FMDV, identify 3Cpro and 2B as FMDV-encoded STING1 antagonists, and highlight the potential of SB24011 as a broad-spectrum anti-picornavirus therapeutic agent.

Background and aimsPegylated interferon alpha (Peg-IFN-α) has the potential for eradicating hepatitis B surface antigen (HBsAg). The aim of our study is to investigate whether the expression levels of adenosine deaminase acting on RNA 1 (ADAR1), NEDD4-binding protein 1 (N4BP1), proteasome activator complex subunit 1 (PSME1) mRNAs in peripheral blood mononuclear cells (PBMCs) of HBeAg-negative chronic hepatitis B virus (HBV) patients are associated with the response to Peg-IFN-α treatment and HBsAg clearance.MethodsIn this prospective study, HBeAg-negative chronic HBV patients treated with Peg-IFN-α were followed for 48 weeks. Patients were categorized into the virological response (VR) group and non-virological response (NVR) group based on the observed changes in HBV DNA and HBsAg levels at week 48 of treatment. Additionally, patients were classified into a serological response (SR) group and a non-serological response (NSR) group according to whether serum HBsAg loss or seroconversion occurred. The expression levels of ADAR1, N4BP1, and PSME1 mRNAs in PBMCs were detected by real-time quantitative PCR. The diagnostic performance of ADAR1, N4BP1, and PSME1 was assessed by analyzing the receiver operating characteristic (ROC) curve and calculating the area under the curve (AUC).ResultsAfter the treatment period, the VR and SR rates were 47.25% and 35.16%, respectively. Dynamic changes in ADAR1, N4BP1, and PSME1 mRNA levels differed significantly between the VR and NVR groups, as well as between the SR and NSR groups. Multivariate analysis revealed that ADAR1 was independently associated with VR and SR at weeks 12 and 24; N4BP1 was independently associated with VR at weeks 12 and 24; PSME1 was independently associated with VR and SR at weeks 12 and 24. At week 24, the AUCs for ADAR1 in predicting VR and SR were 0.9230 and 0.8554. N4BP1 had AUCs of 0.7393 for VR at week 12 and 0.7198 for SR at week 24, while PSME1 had AUCs of 0.7418 for VR and 0.7426 for SR at week 12.ConclusionsADAR1, N4BP1, and PSME1 are novel biomarkers for early therapeutic response to Peg-IFN-α and HBsAg clearance.Clinical Trial Registrationhttps://www.medicalresearch.org.cn/login, identifier 2023−311.

Foot-and-mouth disease virus (FMDV) represents a major threat to global livestock production. The capsid protein VP1 is crucial for infection; however, the host factors and mechanisms responsible for VP1 restriction remain poorly understood. We previously identified the host chaperone DNAJA3 as a host restriction factor that inhibits FMDV infection by promoting VP1 degradation through the autophagy-lysosomal pathway. Here, we elucidate the molecular mechanism by which DNAJA3 mediates degradation of VP1. We demonstrate that DNAJA3 recruits the autophagy cargo receptor TOLLIP to facilitate selective autophagic degradation of VP1. Mechanistically, TOLLIP directly interacts with the 1-37 amino acid (aa) region of VP1 through its N-terminal and C-terminal domains, and full-length TOLLIP is required for efficient VP1 degradation. Furthermore, DNAJA3 recruits the E3 ubiquitin ligase TRIM21 to promote VP1 polyubiquitination through K27-, K48-, and K63-linked ubiquitination. TOLLIP additionally restricts FMDV internalization by modulating early endosomal trafficking in an autophagy-dependent manner. In vivo, tollip-deficient suckling mice exhibit increased susceptibility to FMDV infection. Moreover, TOLLIP exerts antiviral activity against multiple picornaviruses, including Senecavirus A and Enterovirus 71. Notably, FMDV downregulates endogenous TOLLIP expression through the protease activity of the viral 3C protein, enabling the virus to evade host autophagic surveillance. Collectively, our study identifies a novel DNAJA3-TRIM21-TOLLIP axis that restricts FMDV infection through selective autophagy, establishes TOLLIP as a host restriction factor against picornaviruses, and reveals potential molecular targets for antiviral intervention.

ABSTRACT Retinoic acid-inducible gene I (RIG-I)-like receptors, including RIG-I and MDA5, are key cytoplasmic pattern-recognition receptors that detect viral RNA in the cytoplasm and trigger the production of interferon (IFN). Among them, MDA5, rather than RIG-I, is considered the primary sensor during picornavirus infection due to the unique features of picornaviral RNA. However, previous studies have indicated that RIG-I also possesses antiviral activity against several picornaviruses, suggesting its potential importance during viral replication. Here, we found that 3A proteins from various picornaviruses, including Senecavirus A (SVA), Enterovirus 71 (EV71), Encephalomyocarditis virus (EMCV), foot-and-mouth disease virus (FMDV), and Coxsackievirus A16 (CA16), directly target RIG-I to suppress RIG-I-like receptor-mediated IFN-β production. Mechanistically, these picornaviral 3A proteins partially share the following similar strategies to dysregulate RIG-I activation: (i) all interact with RIG-I, (ii) EMCV and FMDV 3A reduce RIG-I expression, and (iii) SVA, EV71, EMCV, and FMDV 3A impair the interaction between RIG-I and MAVS by diminishing K63-linked ubiquitination of RIG-I. These findings broaden our understanding of how picornaviruses employ nonstructural proteins to evade innate immune responses during early infection.IMPORTANCEPicornaviruses cause a broad spectrum of human and animal diseases; however, the mechanisms by which they counteract host antiviral defenses remain incompletely understood. Owing to the distinct structural features of picornaviral RNA, MDA5 is widely regarded as the primary sensor mediating antiviral responses during picornavirus infection. However, accumulating evidence suggests that RIG-I also contributes to antiviral defense. Picornaviruses have evolved various means to suppress RIG-I and MDA5 activity, thereby facilitating evasion of the innate immune response and underscoring the importance of RIG-I during picornavirus infection. This study identifies RIG-I as a conserved target of the nonstructural protein 3A from multiple picornaviruses, including Senecavirus A, Enterovirus 71, Encephalomyocarditis virus, foot-and-mouth disease virus, and Coxsackievirus A16, and uncovers both shared and virus-specific strategies that dysregulate RIG-I-mediated interferon production. Collectively, these findings expand our understanding of the antagonistic mechanisms of how picornaviruses manipulate the host innate immune system.

ABSTRACT RIG-I like receptors (RLRs) recognize RNA viruses and induce an innate immune response. Although many host factors strictly regulate the signal transduction of the RLR pathway, the mechanisms remain unclear. In the present study, we demonstrated that virus infection slightly increased the expression of programmed cell death protein 10 (PDCD10). PDCD10 overexpression inhibited interferon beta (IFN-β) promoter activation after Sendai virus (SeV) infection. Moreover, PDCD10 negatively regulated RNA virus-induced IFN-β secretion. These effects were reversed following PDCD10 gene knockout. PDCD10 also interacted with virus-induced signaling adaptor (VISA) and disrupted the formation of the VISA-IRF3 complex to inhibit IFN-β production. Additionally, PDCD10 promoted foot-and-mouth disease virus (FMDV) replication by inhibiting IFN-β production. FMDV is the causative pathogen of foot-and-mouth disease, one of the most destructive and contagious animal diseases in the world. The FMDV 3A protein plays important roles in viral replication, host tropism, and immune regulation. Our experimental results also showed that full-length 3A cooperated with PDCD10 to inhibit IFN-β production by promoting the binding of PDCD10 to VISA. Collectively, the study findings revealed that PDCD10, as a new negative regulator, cooperated with 3A to inhibit viral-induced IFN-β production.IMPORTANCEFoot-and-mouth disease virus (FMDV) is a pathogen that causes a highly contagious and destructive foot-and-mouth disease in animals with cloven hooves. Although the 3A protein of FMDV is involved in viral replication and host tropism, its function remains unclear. PDCD10 plays critical roles in normal cardiovascular development, cell proliferation, and normal structure and assembly of the Golgi complex. The present study showed that PDCD10 expression was slightly increased by virus infection, while PDCD10 promoted FMDV replication. Our results also demonstrated that PDCD10 inhibited Sendai virus-induced interferon beta (IFN-β) production through interaction with virus-induced signaling adaptor (VISA). PDCD10 also disrupted VISA-IRF3 complex formation to impair IFN-β production induced by RNA virus. The FMDV 3A protein bound with PDCD10 to synergistically promote FMDV replication. This study helped to reveal the potential mechanism of FMDV 3A protein and PDCD10 impact on viral replication.

Senecavirus A (SVA) belongs to the picornaviruses and has emerged as a promising candidate for oncolytic virotherapy in humans. Understanding the immune suppression mechanisms employed by SVA can help optimize its therapeutic efficacy as an oncolytic virus while simultaneously minimizing its immune suppressive effects on normal tissues. In this study, we identified a novel function of the SVA structural protein VP2 as a key viral immune suppressive factor during SVA infection. VP2 targets and degrades IKBKE/IKKε, a key component of the innate immune pathway, thereby suppressing host innate immune responses. It preferentially interacts with the selective autophagic receptor CALCOCO2/NDP52 (calcium binding and coiled-coil domain 2), which then recognizes the K33-linked ubiquitinated IKBKE and delivers it to phagophores for degradation. The E3 ligase RNF114 is responsible for catalyzing the K33-linked ubiquitination of IKBKE at Lys490, and VP2 significantly promoted this modification, which further accelerated IKBKE degradation. Importantly, we found that picornavirus VP2 proteins share this conserved mechanism in degradation of IKBKE and suppression of host innate immunity. These data elucidate the negative regulatory mechanism involving the VP2-RNF114-IKBKE/IKKε-CALCOCO2 axis, and reveal an immune evasion strategy employed by picornaviruses. These findings will provide valuable insights for the development of picornaviral vaccines and antiviral/antitumor therapeutics.Abbreviations: 3-MA: 3-methyladenine; ATG5: autophagy related 5; ATG7: autophagy related 7; CALCOCO2/NDP52: calcium binding and coiled-coil domain 2; CQ: chloroquine; co-IP: co-immunoprecipitation; DAPI: 4',6-diamidino-2'-phenylindole; EV71: enterovirus 71; FMDV: foot-and-mouth disease virus; hpi: hours post-infection; IFN: interferon; IKBKE/IKKε: inhibitor of nuclear factor kappa B kinase subunit epsilon); ISGs: IFN-stimulated genes; MAP1LC3/LC3: microtubule associated protein 1 light chain 3; MG132: cbz-leu-leu-leucinal; MOI: multiplicity of infection; NBR1: NBR1 autophagy cargo receptor; OPTN: optineurin; RNF114: ring finger protein 114; RT-PCR: real-time polymerase chain reaction; siRNA: small interfering RNA; SQSTM1/p62: sequestosome 1; SVA: Senecavirus A; TCID50: 50% tissue culture infectious doses. TOLLIP: toll interacting protein; TRIM17: tripartite motif containing 17; TRIM25: tripartite motif containing 25; TRIM28: tripartite motif containing 28; TRIP12/THRI12: thyroid hormone receptor interactor 12; Ub: ubiquitin; Vec: vector; WCL: whole-cell lysate; WT: wild-type.

African swine fever (ASF), caused by the African swine fever virus (ASFV), is a highly contagious and often fatal disease affecting domestic pigs and wild boars, resulting in substantial economic losses globally. The lack of commercially available vaccines or effective antiviral drugs highlights the urgent need for novel therapeutic strategies. In this study, we conducted a cell-based screening of a targeted anti-DNA virus compound library to identify inhibitors of ASFV replication. Among the candidates, Azvudine-a clinically approved nucleoside analog-demonstrated potent dose-dependent suppression of ASFV replication in porcine alveolar macrophages (PAMs), with a half-maximal inhibitory concentration (IC50) of 0.19 μM and minimal cytotoxicity. Further mechanistic studies indicated that Azvudine acts at a post-entry stage of the viral life cycle and does not directly inactivate viral particles. These findings suggest that Azvudine is a promising repurposed candidate for anti-ASFV drug development.

Spinal cord injury (SCI) involves neuronal apoptosis and axonal disruption, leading to severe motor dysfunction. Studies indicate that exosomes transport microRNAs (miRNAs) and play a crucial role in intercellular communication. This study aimed to explore whether the bone marrow mesenchymal stem cell (BMSCs)-exosomal miR-17-92 cluster can protect against SCI and to explain the underlying mechanisms. In vivo and in vitro SCI models were established and treated with control exosomes (con-exo) or exosomes derived from BMSCs transfected with miR-17-92 cluster plasmid (miR-17-92-exo). Rat BMSCs were isolated and positive markers were identified by flow cytometry. BMSC-derived exosomes were extracted and verified using transmission electron microscopy (TEM), nanoparticle tracking analysis (NTA), and western blotting. The expression of the miR-17-92 cluster was validated by quantitative reverse transcription PCR (qRT-PCR). Spinal cord function, histopathological changes, apoptotic cells, and inflammatory cytokines release in spinal cord tissues were assessed using the Basso-Beattie-Bresnahan (BBB) score, hematoxylin and eosin (HE) staining, terminal deoxynucleotide transferase (TdT)-mediated dUTP nick-end labeling (TUNEL) staining, enzyme-linked immunosorbent assay (ELISA), and qRT-PCR. In PC12 cells, cell proliferation, apoptosis, apoptosis-related proteins cleaved-Caspase3 expression, and inflammatory factors secretion were analyzed using a cell counting kit-8 (CCK8) assay, flow cytometry, western blotting, and ELISA. Our data revealed that the exosomes were successfully isolated from rat BMSCs. The BMSC-exosomal miR-17-92 cluster improved neural functional recovery after SCI, as evidenced by an increased BBB score, improved pathological damage, reduced neuronal apoptosis, and decreased inflammatory factors release. Additionally, miR-17-92-exo treatment significantly inhibited lipopolysaccharide (LPS)-induced reduction in cell viability, increase in cell apoptosis, and upregulation of inflammatory factors in PC12 cells. The exosomal miR-17-92 cluster derived from BMSCs improved functional recovery and exhibited neuroprotective effects in SCI by alleviating apoptosis and inflammation.

The infectious bronchitis virus (IBV) causes significant economic losses to the global poultry industry. Recently, there has been a rapid spread of the GVI-1 lineage of IBV in Asia, particularly in China. However, to date there have been few studies that have assessed the immune protection efficacy of commonly used IB vaccines against the GVI-1 lineage strains. In this study, we evaluated the protective efficacy of two commonly used vaccines, H120 and NNA, against the GVI-1 lineage HX strain based on serological neutralization tests and animal challenge protection experiments. The protective efficacy of sera from chickens immunized using different vaccination strategies against the HX strain was evaluated using chicken embryos, with the results indicating that a combined vaccination strategy using H120 and NNA provided better antiviral effects in chicken embryos than those obtained using either of these two vaccines administered alone. In challenge protection experiments on chicks, we assessed clinical symptoms, viral loads in the trachea and kidneys, and histopathological damage levels. The results revealed that when administered alone, the H120 and NNA vaccines were unable to provide complete protection against HX strain infection, whereas the combined vaccination reduced the pathological damage caused by infection. Multiple bioinformatics analyses revealed significant differences in the nucleic acid and amino acid similarities between the GVI-1 lineage strain HX and the attenuated vaccine strains H120 and NNA, particularly in the S1 gene antigenic epitopes. Our findings in this study, in which we examined the differences in immune protection efficacy of two IB vaccines against a GVI-1 lineage strain, can provide a theoretical basis for optimizing vaccine design.

Prediabetic neuropathic pain has been classified as peripheral neuropathic pain associated with polyneuropathy caused by impaired glucose tolerance or impaired fasting glucose, which is a preclinical stage and might develop type 2 diabetes mellitus. Our previous research highlighted that prediabetes is accompanied by dramatic bilateral mechanical hyperalgesia following high energy diet (HED) which results in myelin and axonal degenerations along somatosensory system. However, the pathogenic mechanisms underlying prediabetic neuropathic pain remain unclear. The nuclear sirtuin 6 (SIRT6) is a crucial deacetylase in the regulation of multiple cellular biological processes, such as DNA repair, genome stability, inflammation and metabolic homeostasis. In current study we show that the expressions of SIRT6 were significantly decreased, while its downstream NF-κB and proinflammatory mediator IL-6 and IL-1β were significantly increased in both dorsal root ganglia (DRG) and spinal dorsal horn of rats with prediabetic neuropathic pain induced by HED. Moreover, siRNA-SIRT6 treatment induced a significant reduction in bilateral paw withdrawal mechanical thresholds, indicating that SIRT6 down-regulation contributed to prediabetic neuropathic pain induced by HED. Furthermore, it was also found that SIRT6 reduction induced the activation of HMGB1 via disinhibition of NF-κB in both DRG and spinal dorsal horn of prediabetic rats. In conclusion, prediabetic neuropathic pain is caused by SIRT6 reduction through upregulating HMGB1-RAGE signaling at both peripheral and spinal levels.

Classical swine fever virus (CSFV) spreads in domestic and wild pig populations, causing significant economic losses in the swine industry. Despite the global implementation of live attenuated vaccines, CSFV remains a persistent threat, with sporadic outbreaks reported annually. A major limitation of the current vaccines is safety concerns and the inability to differentiate infected from vaccinated animals (DIVA). The development of DIVA-compliant vaccines is desirable for effectively controlling or eradicating classical swine fever (CSF). Here, we developed two lipid nanoparticle (LNP)-encapsulated mRNA vaccines encoding either the extracellular domain of the CSFV envelope protein E2 (E2-ECD) or its N-terminal 172-amino acid fragment (E2-ECD-N). Immunological assays in mice revealed high antigenicity and long-lasting protective antibody responses from a single dose of either the E2-ECD or E2-ECD-N mRNA vaccine. Notably, both the E2-ECD and E2-ECD-N mRNA vaccines induced robust T cell responses in mice. Furthermore, a single dose (100 μg) of the E2-ECD mRNA vaccine was sufficient to induce long-term (up to 4 months) protective immunity against CSFV infection in rabbits. Our findings highlight the potential of CSFV-E2-based mRNA vaccines as promising strategies for effective CSF prevention and control while enabling DIVA.

The control of foot-and-mouth disease virus (FMDV) primarily relies on vaccine immunization; however, this approach is not always fully effective, underscoring the urgent need for novel antiviral strategies. This study identifies RPL35 as a host antiviral protein that targets FMDV. Further mechanistic investigations demonstrate that RPL35 directly interacts with the FMDV structural protein VP2, mediating its K48-linked polyubiquitination and subsequent degradation. The Lys217 residue of VP2 is critical for RPL35's antiviral activity, as evidenced by the increased viral virulence observed with the rO-VP2K217R mutant virus. Through an unbiased proteomic screen, we revealed that RPL35 recruits the E3 ligase AMFR to ubiquitinate and degrade VP2. Additionally, FMDV induces the degradation of KPNA3, thereby blocking RPL35's nuclear translocation. This study advances our understanding of host-virus interactions and provides new insights into developing antiviral drugs targeting the ubiquitin-proteasome pathway.IMPORTANCEThis investigation elucidated the antiviral role of RPL35 in the context of FMDV infection. Our results indicate that RPL35 facilitates the recruitment of AMFR, which, in turn, promotes K48-linked polyubiquitination and subsequent proteasomal degradation of the viral protein VP2. This process thereby mitigates viral infection. Further analysis identified Lys217 of VP2 as a critical ubiquitination site for RPL35, with the inhibitory effect of RPL35 being abolished in the recombinant mutant virus rO-VP2K217R. Additionally, we found that FMDV induces the degradation of KPNA3, which obstructs the nuclear translocation of RPL35. Collectively, these findings suggest that RPL35 functions as a potent antiviral effector in suppressing FMDV infection.

Foot-and-mouth disease (FMD) is a highly contagious and economically devastating disease. Traditional intramuscular (IM) injection of inactivated FMD vaccines has been crucial for disease control, but intradermal (ID) immunization offers advantages such as reduced vaccine dosage and decreased animal discomfort. The differences in immune efficiency of intradermal and intramuscular delivery of FMD commercial inactivated vaccine in pigs remain unclear. Here, we compared the immune efficacy of ID and IM immunization using the same dose of commercial FMD vaccine. Experimental pigs were administered the vaccine via either ID or IM routes. At 28 days post-immunization (dpi), the ID group produces less uniform levels of FMDV-specific antibodies than IM groups. The detection of IL-4, IL-6, and IFN-γ in porcine serum samples revealed a lower cellular immune response in the ID group as well in the early stages. Meanwhile, the IM group exhibited higher percentages of CD8+T cells producing IFN-γ. These results indicate that, at the same dose of FMDV antigen emulsified with adjuvant ISA 201VG, initial humoral and cellular immune responses were weaker in the ID group compared to the IM group in the early stages, and the uniformity of antibody response levels was poorer in the ID group. While ID immunization holds promise for future vaccine strategies, its application with the current inactivated FMDV vaccine carries a higher risk of immunization failure due to the heterogeneity of immune responses.