Expression Of Viral Nucleoprotein Research Articles

Palmatine as a potent immunomodulator: Enhancing resistance to Micropterus salmoides rhabdovirus in largemouth bass through innate immune activation and viral suppression

Micropterus salmoides rhabdovirus (MSRV) poses a significant threat to aquaculture, causing substantial economic losses. In this study, we evaluated the antiviral efficacy and immunomodulatory potential of palmatine, a plant-derived monomer, against MSRV infection in largemouth bass. Our results demonstrated that palmatine significantly inhibited MSRV replication, with a reduction in viral nucleoprotein expression by 85 % at a safe concentration. Additionally, palmatine pre-treatment of EPC cells enhanced their antiviral capacity, with a maximum inhibition rate of 82 % following 24 h pre-incubation. Palmatine also effectively reduced MSRV-induced cytopathic effects, protecting cellular integrity and maintaining mitochondrial membrane potential. In vivo studies revealed that palmatine immersion at 80 mg/L was non-toxic and significantly suppressed MSRV replication in largemouth bass, increasing survival rates by 53 % over 15 d. Furthermore, palmatine pre-treatment enhanced the fish's resistance to MSRV, with a 78 % inhibition rate of viral replication and a 46 % increase in survival rate. Mechanistically, palmatine activated key immune genes, including IRF3, IRF7, and IFN, indicating its role in boosting innate immune responses. The compound also reduced horizontal transmission of MSRV in a cohabitation model, decreasing viral spread by up to 78 % over nine days. These findings highlight palmatine's potential as a small-molecule immunomodulator in aquaculture, offering a sustainable approach to disease management and enhancing fish health and welfare. Integrating palmatine into fish diets as an immunostimulant could provide a continuous, proactive defense against viral outbreaks, promoting more resilient and sustainable aquaculture practices.

Investigation of the Antiviral Mechanism of Curcumin Analog EF-24 against Siniperca cachuatsi Rhabdovirus

Siniperca chuatsi rhabdovirus (SCRV) is a major strain of viral fish virus resulting in multiple transmissions and devastating damage in aquaculture. Currently, there are no available approved therapeutics. In this study, we screened and identified a novel curcumin analog (EF-24) for evaluating its in vitro anti-SCRV properties and potential molecular mechanisms. Present results demonstrated that EF-24 could strongly delay the occurrence of cytopathic effects (CPEs) in epithelioma papulosum cyprinid cells (EPCs) and inhibit SCRV replication and viral nucleoprotein expression in the early stages of infection by the time-of-addition assay. Furthermore, flow cytometry analysis after Annexin V-FITC/PI double staining and immunofluorescence microscopy observation after JC-1 incubation showed that EF-24 downregulated cell mitochondrial apoptosis induced by SCRV. The enzymatic activities of caspase-3 and caspase-9 were also reduced after EF-24 treatment, indicating that EF-24 may protect cells from SCRV infection by decreasing mitochondrial intrinsic apoptosis in infected cells. Collectively, we demonstrated for the first time that the curcumin analog EF-24 possesses antiviral ability against SCRV, suggesting its potential for effective control of fish rhabdovirus spreading.

Host Cell-dependent Modulatory Role of Ras Homolog Enriched in Brain-Like-1 (RhebL1) Protein in Influenza A/NWS/33 Virus-infected Mammalian Cells.

The Mammalian Target of Rapamycin (mTOR) signaling pathway regulates protein phosphorylation and exerts control over major cellular processes. mTOR is activated by the small G-protein Ras Homolog Enriched in Brain (Rheb), which is encoded by the Rheb1 and Rheb-like-1 (RhebL1) genes. There is currently a paucity of information on the role of RhebL1, and specifically its involvement in viral infection. In the present study we investigated the role of RhebL1 during human influenza A/NWS/33 (NWS/33) (H1N1) virus infection of rhesus monkey-kidney (LLC-MK2) cells and human type II alveolar epithelial (A549) cells. To assess the efficiency of NWS/33 virus replication, the expression of viral nucleoprotein was examined by indirect immunofluorescence (IIF) and the viral yield by fifty percent tissue culture infectious dose assay. An RNA-mediated RNA interference approach was used to investigate the role of RhebL1 during NWS/33 infection. RhebL1 expression was evaluated by IIF, Western blotting, and enzyme-linked immunosorbent assays. A two-tailed Student's t-test was applied to evaluate differences between groups. RhebL1 was differentially expressed in the cell models used in this study. Silencing of the RhebL1 gene led to increased NWS/33 virus infection in A549 cells, but not in LLC-MK2 cells. Moreover, the expression of hyperphosphorylated cytokeratin 8, a marker of NWS/33 virus infection efficiency, increased in A549 cells depleted of RhebL1 but remained almost unchanged in LLC-MK2 cells. These are the first results showing involvement of the endogenous RhebL1 protein during viral infection. Our data suggests that RhebL1 exerts a host cell-dependent modulatory role during influenza virus infection. RhebL1 appears to be a restrictive factor against NWS/33 virus replication in A549 cells, but not in LLC-MK2.

Anti-SARS-CoV-2 Activity of Extracellular Vesicle Inhibitors: Screening, Validation, and Combination with Remdesivir.

The coronavirus disease 2019 (COVID-19) pandemic severely impacts health, economy, and society worldwide. Antiviral drugs against SARS-CoV-2 are urgently needed to cope with this global crisis. It has been found that the biogenesis and release mechanisms of viruses share a common pathway with extracellular vesicles (EVs). We hypothesized that small molecule inhibitors of EV biogenesis/release could exert an anti-SARS-CoV-2 effect. Here, we screened 17 existing EV inhibitors and found that calpeptin, a cysteine proteinase inhibitor, exhibited the most potent anti-SARS-CoV-2 activity with no apparent cytotoxicity. Calpeptin demonstrated the dose-dependent inhibition against SARS-CoV-2 viral nucleoprotein expression in the infected cells with a half-maximal inhibitory concentration (IC50) of 1.44 µM in Vero-E6 and 26.92 µM in Calu-3 cells, respectively. Moreover, calpeptin inhibited the production of infectious virions with the lower IC50 of 0.6 µM in Vero E6 cells and 10.12 µM in Calu-3 cells. Interestingly, a combination of calpeptin and remdesivir, the FDA-approved antiviral drug against SARS-CoV-2 viral replication, significantly enhanced the anti-SARS-CoV-2 effects compared to monotherapy. This study discovered calpeptin as a promising candidate for anti-SARS-CoV-2 drug development. Further preclinical and clinical studies are warranted to elucidate the therapeutic efficacy of calpeptin and remdesivir combination in COVID-19.

Differential Cell Line Susceptibility to Crimean-Congo Hemorrhagic Fever Virus.

Crimean-Congo hemorrhagic fever (CCHF) is a severe tick-borne viral disease of global concerns due to the increasing incidence and lack of effective treatments. The causative agent, CCHF virus (CCHFV), has been characterized for years; however, its tropism in cell lines of different host and tissue origins remains unclear. This study characterized the susceptibility of 16 human and 6 animal cell lines to CCHFV. Increased viral load and viral nucleoprotein expression, and productive CCHFV replication were detected in human vascular (HUVEC), renal (SW-13 and HEK-293), hepatic (Huh7), and cerebral (U-87 MG) cell lines, which were considered CCHFV-highly permissive cell lines. Renal cell lines derived from monkey and dog could also support CCHFV replication. This study evaluated the susceptibility of different cell lines to CCHFV and identified CCHFV-permissive cell lines. Our findings raise concerns regarding the use of cell lines in ex vivo studies of CCHFV and may have important implications for further fundamental research, which would promote understanding of CCHFV pathogenesis and transmission, as well as benefit designing strategies for disease prevention and control.

Viral-induced impairment of innate immune response and functional properties of cerebrovascular endothelial cells correlates with brain viral invasion.

Event Abstract Back to Event Viral-induced impairment of innate immune response and functional properties of cerebrovascular endothelial cells correlates with brain viral invasion. Christian Bleau1, Mélanie Burnette1 and Lucie Lamontagne1* 1 Université du Québec à Montréal, Sciences Biologiques, Canada The blood-brain barrier (BBB) provides significant protection against brain invasion by hematogenous spread viruses but some have evolved strategies to bypass this primary endothelial cell (EC) barrier and induce neuropathogenesis. The role of BBB ECs in a rapid early anti-viral immunity limiting viral spread to the brain is largely unknown. Using mouse hepatitis viruses (MHVs) of different neurovirulence, we investigated the viral-induced innate response and structural dysfunctions of ECs according to viral permittivity. C57BL/6 mice were infected i.p. with the highly-neurotropic (L2-MHV3), mildly-neurotropic (MHV-A59) or the attenuated variant 51.6-MHV3 showing low tropism for ECs. Higher intracerebral levels of IFN-b, IL-6, TNF-a chemokines and TLR-2 and lowered ECs junctionnal protein mRNA expression, as assessed by qRT-PCR, correlating with viral nucleoprotein expression, were observed only in brain from L2-MHV3 infected mice. Using EC line Bend.3 as an in vitro BBB model, infection with L2-MHV3 provoked higher cytopathic effects, decreased IFN-b, IL-6, TNF-a and chemokine and transendothelial resistance, correlating with lower junctional protein mRNA expression when compared with lower virulent MHV-A59 and 51.6-MHV3. Such disorders in innate immunity and viral spreading induced by L2-MHV3 in EC depend on both TLR-2- and specific viral receptor-dependent endocytic ways used by L2-MHV3 than lower virulent viral strains. Our findings highlight the importance of innate immunity parameters and functions of BBB ECs that may prevent access of pathogenic neurotropic viruses to the brain. Acknowledgements This work was funded by NSERC-Canada. Keywords: Brain, Endothelial Cells, Coronavirus, blood brain barrier, Cytokines, TLR2 Conference: 15th International Congress of Immunology (ICI), Milan, Italy, 22 Aug - 27 Aug, 2013. Presentation Type: Abstract Topic: Host-pathogen interactions Citation: Bleau C, Burnette M and Lamontagne L (2013). Viral-induced impairment of innate immune response and functional properties of cerebrovascular endothelial cells correlates with brain viral invasion. . Front. Immunol. Conference Abstract: 15th International Congress of Immunology (ICI). doi: 10.3389/conf.fimmu.2013.02.00658 Copyright: The abstracts in this collection have not been subject to any Frontiers peer review or checks, and are not endorsed by Frontiers. They are made available through the Frontiers publishing platform as a service to conference organizers and presenters. The copyright in the individual abstracts is owned by the author of each abstract or his/her employer unless otherwise stated. Each abstract, as well as the collection of abstracts, are published under a Creative Commons CC-BY 4.0 (attribution) licence (https://creativecommons.org/licenses/by/4.0/) and may thus be reproduced, translated, adapted and be the subject of derivative works provided the authors and Frontiers are attributed. For Frontiers’ terms and conditions please see https://www.frontiersin.org/legal/terms-and-conditions. Received: 12 Jun 2013; Published Online: 22 Aug 2013. * Correspondence: Dr. Lucie Lamontagne, Université du Québec à Montréal, Sciences Biologiques, Montréal, Québec, H3C 3P8, Canada, lamontagne.lucie@uqam.ca Login Required This action requires you to be registered with Frontiers and logged in. To register or login click here. Abstract Info Abstract The Authors in Frontiers Christian Bleau Mélanie Burnette Lucie Lamontagne Google Christian Bleau Mélanie Burnette Lucie Lamontagne Google Scholar Christian Bleau Mélanie Burnette Lucie Lamontagne PubMed Christian Bleau Mélanie Burnette Lucie Lamontagne Related Article in Frontiers Google Scholar PubMed Abstract Close Back to top Javascript is disabled. Please enable Javascript in your browser settings in order to see all the content on this page.

Polarized entry and release of Junín virus, a New World arenavirus

Junin virus (JUNV), the causative agent of Argentine haemorrhagic fever, is a human pathogen that naturally enters the body through the epithelial cells of the respiratory and digestive tracts. The interaction of JUNV with two types of polarized epithelial cultures, Vero C1008 and A549, was investigated. Radioactive virus-binding assays showed that JUNV infects polarized lines preferentially through the apical surface. High-level expression of viral nucleoprotein was detected in polarized cell lines infected through the apical domain. Virus production from apical media was about 100-fold higher than that found into the basolateral medium. Confocal-immunofluorescence analysis revealed high-level expression of glycoprotein at the apical-membrane surface. Disruption of the microtubule network by colchicine impaired JUNV vectorial release. This is the first study to analyse the interaction between a member of the virus family Arenaviridae and polarized epithelial cells, showing preferential entry and release from the apical plasma membrane.

Differentiation of strains of equine arteritis virus of differing virulence to horses by growth in equine endothelial cells.

To compare growth characteristics of strains of equine arteritis virus (EAV) of differing virulence to horses in rabbit kidney (RK)-13 cells and equine endothelial cells (EECs) cultured from the pulmonary artery of a foal. 13 strains of EAV, including 11 field isolates of differing virulence to horses; the highly virulent, horse-adapted Bucyrus strain; and the modified-live virus (MLV) vaccine derived from it. The growth characteristics of the 13 strains were compared in EECs and RK-13 cells. Viral nucleoprotein expression, cytopathogenicity, and plaque size were compared to determine whether growth characteristics of the 13 strains were predictive of their virulence to horses. Cytopathogenicity, viral nucleoprotein expression, and plaque size induced by all 13 viruses were similar in RK-13 cells, whereas virulent strains of EAV caused significantly larger plaques in EECs than did the avirulent strains of EAV. Paradoxically, the highly attenuated MLV vaccine and 1 field isolate of EAV caused plaques in EECs that were larger than those caused by any of the other viruses, and sequence analysis confirmed the field isolate of EAV to be indistinguishable from the MLV vaccine. With the notable exception of the MLV vaccine, growth of the various strains of EAV in EECs was predictive of their individual virulence to horses. Thus, EECs provide a relevant and useful model to further characterize determinants of virulence and attenuation amongst strains of EAV.