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1.
J Mol Graph Model ; 118: 108345, 2023 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-36308946

RESUMO

Human norovirus (HuNoV) causes acute viral gastroenteritis in all age groups, and dehydration and severe diarrhea in the elderly. The World Health Organization reports ∼1.45 million deaths from acute gastroenteritis annually in the world. Rupintrivir, an inhibitory medicine against the human rhinovirus C3 protease, has been reported to inhibit HuNoV 3C protease. However, several HuNoV 3C protease mutations have been revealed to reduce the susceptibility of HuNoV to rupintrivir. The structural details behind rupintrivir-resistance of these single-point mutations (A105V and I109V) are not still clear. Hence, in this study, a combination of computational techniques were used to determine the rupintrivir-resistance mechanism and to propose an inhibitor against wild-type and mutant HuNoV 3C protease through structure-based virtual screening. Dynamic structural results indicated the unstable binding of rupintrivir at the cleft binding site of the wild-type and mutant 3C proteases, leading to its detachment. Our findings presented that the domain II of the HuNoV 3C protease had a critical role in binding of inhibitory molecules. Binding energy computations, steered molecular dynamics and umbrella sampling simulations confirmed that amentoflavone, the novel suggested inhibitor, strongly binds to the cleft site of all protease models and has a good structural stability in the complex system along the molecular dynamic simulations. Our in silico study proposed the selected compound as a potential inhibitor against the HuNoV 3C protease. However, additional experimental and clinical studies are required to corroborate the therapeutic efficacy of the compound.


Assuntos
Antivirais , Norovirus , Inibidores de Proteases , Humanos , Antivirais/química , Antivirais/farmacologia , Gastroenterite/tratamento farmacológico , Gastroenterite/virologia , Norovirus/efeitos dos fármacos , Norovirus/metabolismo , Peptídeo Hidrolases , Inibidores de Proteases/química , Inibidores de Proteases/farmacologia , Proteínas Virais/antagonistas & inibidores , Proteínas Virais/química
2.
Biomolecules ; 12(11)2022 11 17.
Artigo em Inglês | MEDLINE | ID: mdl-36421714

RESUMO

Lipid metabolic reprogramming is one of the hallmarks of hepatocarcinogenesis and development. Therefore, lipid-metabolism-related genes may be used as potential biomarkers for hepatocellular carcinoma (HCC). This study aimed to screen for genes with dysregulated expression related to lipid metabolism in HCC and explored the clinical value of these genes. We screened differentially expressed proteins between tumorous and adjacent nontumorous tissues of hepatitis B virus (HBV)-related HCC patients using a Nanoscale Liquid Chromatography-Tandem Mass Spectrometry platform and combined it with transcriptomic data of lipid-metabolism-related genes from the GEO and HPA databases to identify dysregulated genes that may be involved in lipid metabolic processes. The potential clinical values of these genes were explored by bioinformatics online analysis tools (GEPIA, cBioPortal, SurvivalMeth, and TIMER). The expression levels of the secreted protein (angiopoietin-like protein 6, ANGPTL6) in serum were analyzed by ELISA. The ability of serum ANGPTL6 to diagnose early HCC was assessed by ROC curves. The results showed that serum ANGPTL6 could effectively differentiate between HBV-related early HCC patients with normal serum alpha-fetoprotein (AFP) levels and the noncancer group (healthy participants and chronic hepatitis B patients) (AUC = 0.717, 95% CI: from 0.614 to 0.805). Serum ANGPTL6 can be used as a potential second-line biomarker to supplement serum AFP in the early diagnosis of HBV-related HCC.


Assuntos
Proteínas Semelhantes a Angiopoietina , Carcinoma Hepatocelular , Hepatite B , Neoplasias Hepáticas , Humanos , alfa-Fetoproteínas/análise , Proteínas Semelhantes a Angiopoietina/genética , Carcinoma Hepatocelular/diagnóstico , Carcinoma Hepatocelular/genética , Carcinoma Hepatocelular/virologia , Metabolismo dos Lipídeos/genética , Neoplasias Hepáticas/diagnóstico , Neoplasias Hepáticas/genética , Neoplasias Hepáticas/virologia , Hepatite B/complicações , Hepatite B/diagnóstico , Biomarcadores Tumorais/genética
3.
J Nat Prod ; 85(11): 2667-2674, 2022 Nov 25.
Artigo em Inglês | MEDLINE | ID: mdl-36346918

RESUMO

Chromatographic separation on the liquid-state fermented products produced by the fungal strain Alternaria alstroemeriae Km2286 isolated from the littoral medicinal herb Atriplex maximowicziana Makino resulted in the isolation of compounds 1-9. Structures were determined by spectroscopic analysis as four undescribed perylenequinones, altertromins A-D (1-4), along with altertoxin IV (5), altertoxin VIII (6), stemphyperylenol (7), tenuazonic acid (8), and allo-tenuazonic acid (9). Compounds 1-6 exhibited antiviral activities against Epstein-Barr virus (EBV) with EC50 values ranging from 0.17 ± 0.07 to 3.13 ± 0.31 µM and selectivity indices higher than 10. In an anti-neuroinflammatory assay, compounds 1-4, 6, and 7 showed inhibitory activity of nitric oxide production in lipopolysaccharide-induced microglial BV-2 cells, with IC50 values ranging from 0.33 ± 0.04 to 4.08 ± 0.53 µM without significant cytotoxicity. This is the first report to describe perylenequinone-type compounds with potent anti-EBV and anti-neuroinflammatory activities.


Assuntos
Alternaria , Anti-Inflamatórios , Antivirais , Atriplex , Infecções por Vírus Epstein-Barr , Herpesvirus Humano 4 , Perileno , Plantas Medicinais , Quinonas , Humanos , Alternaria/química , Alternaria/isolamento & purificação , Atriplex/microbiologia , Infecções por Vírus Epstein-Barr/virologia , Herpesvirus Humano 4/efeitos dos fármacos , Estrutura Molecular , Perileno/química , Perileno/isolamento & purificação , Perileno/farmacologia , Plantas Medicinais/microbiologia , Quinonas/química , Quinonas/isolamento & purificação , Quinonas/farmacologia , Ácido Tenuazônico/química , Anti-Inflamatórios/química , Anti-Inflamatórios/isolamento & purificação , Anti-Inflamatórios/farmacologia , Antivirais/química , Antivirais/isolamento & purificação , Antivirais/farmacologia
4.
BMJ Case Rep ; 15(11)2022 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-36379626

RESUMO

SARS-CoV-2 is now a major global health issue and manifests mainly as a respiratory disorder. Several other complications involving hypercoagulability, cardiovascular system and central nervous system have been described in the literature. Among these atypical presentations, encephalopathy associated with SARS-CoV-2 is a rare entity with heterogenous clinical and radiological findings. The direct presence of SARS-CoV-2 in cerebrospinal fluid (CSF) was rarely found in encephalopathy patients with acute SARS-Cov-2 infection.Here, we report a case of myeloencephalitis with positive real-time PCR for SARS-CoV-2 in CSF in a young woman presenting exclusively with neurological symptoms. Other differential diagnosis were extensively pursued by a comprehensive aetiological workup. To our knowledge, this is the first case report in the Omicron era. In the context of recent global explosion of SARS-Cov-2 infections, clinicians should consider this pathogen among other possible neurotropic agents and be familiar with its radiological and clinical presentations.


Assuntos
COVID-19 , Encefalomielite , Feminino , Humanos , Encefalopatias/virologia , COVID-19/complicações , Encefalomielite/diagnóstico , Encefalomielite/virologia , SARS-CoV-2/genética , SARS-CoV-2/isolamento & purificação , Reação em Cadeia da Polimerase em Tempo Real
7.
Oxid Med Cell Longev ; 2022: 2520348, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-36425057

RESUMO

Influenza virus infection is one of the strongest pathogenic factors for the development of acute lung injury (ALI)/ acute respiratory distress syndrome (ARDS). However, the underlying cellular and molecular mechanisms have not been clarified. In this study, we aim to investigate whether melatonin modulates macrophage polarization, oxidative stress, and pyroptosis via activating Apolipoprotein E/low-density lipoprotein receptor (ApoE/LDLR) pathway in influenza A-induced ALI. Here, wild-type (WT) and ApoE-/- mice were instilled intratracheally with influenza A (H3N2) and injected intraperitoneally with melatonin for 7 consecutive days. In vitro, WT and ApoE-/- murine bone marrow-derived macrophages (BMDMs) were pretreated with melatonin before H3N2 stimulation. The results showed that melatonin administration significantly attenuated H3N2-induced pulmonary damage, leukocyte infiltration, and edema; decreased the expression of proinflammatory M1 markers; enhanced anti-inflammatory M2 markers; and switched the polarization of alveolar macrophages (AMs) from M1 to M2 phenotype. Additionally, melatonin inhibited reactive oxygen species- (ROS-) mediated pyroptosis shown by downregulation of malonaldehyde (MDA) and ROS levels as well as inhibition of the NLRP3/GSDMD pathway and lactate dehydrogenase (LDH) release. Strikingly, the ApoE/LDLR pathway was activated when melatonin was applied in H3N2-infected macrophages and mice. ApoE knockout mostly abrogated the protective impacts of melatonin on H3N2-induced ALI and its regulatory ability on macrophage polarization, oxidative stress, and pyroptosis. Furthermore, recombinant ApoE3 (re-ApoE3) inhibited H3N2-induced M1 polarization of BMDMs with upregulation of MT1 and MT2 expression, but re-ApoE2 and re-ApoE4 failed to do this. Melatonin combined with re-ApoE3 played more beneficial protective effects on modulating macrophage polarization, oxidative stress, and pyroptosis in H3N2-infected ApoE-/- BMDMs. Our study indicated that melatonin attenuated influenza A- (H3N2-) induced ALI by inhibiting the M1 polarization of pulmonary macrophages and ROS-mediated pyroptosis via activating the ApoE/LDLR pathway. This study suggested that melatonin-ApoE/LDLR axis may serve as a novel therapeutic strategy for influenza virus-induced ALI.


Assuntos
Lesão Pulmonar Aguda , Melatonina , Infecções por Orthomyxoviridae , Animais , Camundongos , Lesão Pulmonar Aguda/tratamento farmacológico , Lesão Pulmonar Aguda/metabolismo , Lesão Pulmonar Aguda/virologia , Apolipoproteína E3/farmacologia , Apolipoproteínas E/metabolismo , Vírus da Influenza A Subtipo H3N2 , Macrófagos/metabolismo , Melatonina/uso terapêutico , Camundongos Knockout para ApoE , Piroptose , Espécies Reativas de Oxigênio/metabolismo , Infecções por Orthomyxoviridae/tratamento farmacológico
8.
Genes Genomics ; 44(12): 1499-1507, 2022 12.
Artigo em Inglês | MEDLINE | ID: mdl-36331778

RESUMO

BACKGROUND: Ebola virus (EBOV) causes a serious hemorrhagic disease in humans, with a mortality rate of up to 80%. Despite significant achievements in the past decades elucidating the pathogenesis of EBOV, there is still much to be elucidated about the cell type-specific host response and their functional roles during infection. OBJECTIVE: This study aimed to gain insight into cell type-specific host responses to EBOV infection. METHODS: Real-time RT-qPCR analysis was used to identify host transcriptional changes in epithelial Caco-2 cells and endothelial HUVECs by EBOV infection. RESULTS: EBOV efficiently infected to both Caco-2 cells and HUVECs, depending on the time of infection. However, changes in the transcriptional levels of several host cellular genes following viral infection showed significant differences between Caco-2 cells and HUVECs. EBOV infection increases the transcription of TGF-ß1, a key factor in epithelium-to-mesenchyme transition (EMT), only in HUVECs, but not in Caco-2 cells. This upregulation in turn induces the transcription of other EMT signaling molecules such as snail, slug and MMP9, ultimately leading to endothelial-to-mesenchymal transition (EndMT). Furthermore, this EndMT process appears to be associated with increased transcription of stem-cell markers such as Klf4, Sox2 and Oct4. However, most of these transcriptional changes due to EBOV infection did not occur in Caco-2 cells, suggesting that EMT or EndMT by EBOV infection is cell type-specific. CONCLUSION: We propose that EBOV infection induces the expression of TGF-ß1-mediated signals in endothelial HUVECs, resulting in EndMT. This could provide broader information to elucidate the pathogenesis of Ebola virus disease.


Assuntos
Transição Epitelial-Mesenquimal , Doença pelo Vírus Ebola , Células Endoteliais da Veia Umbilical Humana , Fator de Crescimento Transformador beta1 , Humanos , Células CACO-2 , Doença pelo Vírus Ebola/metabolismo , Células Endoteliais da Veia Umbilical Humana/metabolismo , Células Endoteliais da Veia Umbilical Humana/virologia , Fator de Crescimento Transformador beta1/genética , Fator de Crescimento Transformador beta1/metabolismo
9.
Vet Microbiol ; 275: 109583, 2022 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-36332299

RESUMO

A DNA fragment containing CpG motifs (CpG ODN) is one of the potent immunopotentiators used to improve vaccine efficacy. It can enhance a protective immunity by stimulating both innate and adaptive immune responses. In this study, we designed and constructed a recombinant plasmid carrying the combined CpG ODN to generate an immunopotentiator for boosting the immunogenicity of porcine circovirus type 2 (PCV2) virus-like particles (VLPs). The capsid protein of PCV2b was expressed in insect cells and purified by affinity chromatography. The purified capsid protein was incubated with the CpG ODN in the reaction that allowed VLPs formation and encapsidation of the CpG ODN to occur simultaneously. Morphology of the reassembled VLPs was similar to the PCV2 virions as observed using an electron microscope. When the CpG ODN-encapcidated VLPs was treated with DNase I, the VLPs could protect the packaged CpG ODN from the enzyme digestion. Moreover, we immunized mice subcutaneously with VLPs, CpG ODN-loaded VLPs, or phosphate buffer saline for three times at two-week intervals. The results showed that the CpG ODN-loaded VLPs could elicit significantly higher levels of PCV2-specific neutralizing antibodies and interferon gamma (IFN-γ) expression in the immunized mice compared to those conferred by the VLPs alone. Conclusively, we have proved that the CpG ODN incorporated in VLPs can serve as a potent immunopotentiator for PCV2 vaccine development.


Assuntos
Infecções por Circoviridae , Vacinas Virais , Animais , Camundongos , Adjuvantes Imunológicos , Anticorpos Antivirais , Proteínas do Capsídeo , Infecções por Circoviridae/prevenção & controle , Infecções por Circoviridae/veterinária , Circovirus , Suínos , Doenças dos Suínos/prevenção & controle , Doenças dos Suínos/virologia , Ilhas de CpG
11.
Viruses ; 14(11)2022 Nov 03.
Artigo em Inglês | MEDLINE | ID: mdl-36366540

RESUMO

Monitoring virus infections can be an important selection tool in honey bee breeding. A recent study pointed towards an association between the virus-free status of eggs and an increased virus resistance to deformed wing virus (DWV) at the colony level. In this study, eggs from both naturally surviving and traditionally managed colonies from across Europe were screened for the prevalence of different viruses. Screenings were performed using the phenotyping protocol of the 'suppressed in ovo virus infection' trait but with qPCR instead of end-point PCR and a primer set that covers all DWV genotypes. Of the 213 screened samples, 109 were infected with DWV, 54 were infected with black queen cell virus (BQCV), 3 were infected with the sacbrood virus, and 2 were infected with the acute bee paralyses virus. It was demonstrated that incidences of the vertical transmission of DWV were more frequent in naturally surviving than in traditionally managed colonies, although the virus loads in the eggs remained the same. When comparing virus infections with queen age, older queens showed significantly lower infection loads of DWV in both traditionally managed and naturally surviving colonies, as well as reduced DWV infection frequencies in traditionally managed colonies. We determined that the detection frequencies of DWV and BQCV in honey bee eggs were lower in samples obtained in the spring than in those collected in the summer, indicating that vertical transmission may be lower in spring. Together, these patterns in vertical transmission show that honey bee queens have the potential to reduce the degree of vertical transmission over time.


Assuntos
Vírus de RNA , Viroses , Vírus , Animais , Abelhas/virologia , Prevalência , Vírus de RNA/genética
13.
Viruses ; 14(11)2022 Nov 11.
Artigo em Inglês | MEDLINE | ID: mdl-36423106

RESUMO

While some companion animals have been shown to be susceptible to SARS-CoV-2, their role in the COVID-19 pandemic has remained poorly investigated. Equids are susceptible to SARS-CoV-2 based on the similarity of the human ACE-2 receptor and reports of infection. Clinical disease and prevalence factors associated with SARS-CoV-2 infection in equids have not yet been investigated. The aim of this study was to determine the seroprevalence of SARS-CoV-2 and selected prevalence factors in 1186 equids presented for various conditions to a Veterinary Medical Teaching Hospital over a two-year period. Blood samples were tested for SARS-CoV-2 antibodies using an ELISA targeting the receptor binding domain (RBD) of the SARS-CoV-2 spike protein. Further, selected prevalence factors (season, age, breed, sex, presenting complaint) were retrieved from the medical records. No information was available on whether the horses had come into contact with COVID-19-positive individuals. Among the study animals, 42/1186 (3.5%) horses had detectable SARS-CoV-2 antibodies. Amongst the prevalence factors investigated, only seasonality (spring) was associated with a greater frequency of seropositivity to SARS-CoV-2. Horses with medical and surgical complaints were more likely to test seropositive to SARS-CoV-2 compared to horses presented for routine health care procedures, suggesting more frequent and/or longer interactions with individuals with COVID-19. While horses can become infected with SARS-CoV-2 via the occasional spillover from COVID-19 individuals, clinical disease expression remains subclinical, making horses an unlikely contributor to the spread of SARS-CoV-2.


Assuntos
COVID-19 , SARS-CoV-2 , Animais , Anticorpos Antivirais , California , COVID-19/epidemiologia , COVID-19/veterinária , Cavalos/virologia , Hospitais de Ensino , Pandemias , Estudos Soroepidemiológicos , Hospitais Veterinários
14.
Viruses ; 14(11)2022 Nov 14.
Artigo em Inglês | MEDLINE | ID: mdl-36423126

RESUMO

Varicella-zoster virus (VZV) infection of neuronal cells and the activation of cell-intrinsic antiviral responses upon infection are still poorly understood mainly due to the scarcity of suitable human in vitro models that are available to study VZV. We developed a compartmentalized human-induced pluripotent stem cell (hiPSC)-derived neuronal culture model that allows axonal VZV infection of the neurons, thereby mimicking the natural route of infection. Using this model, we showed that hiPSC-neurons do not mount an effective interferon-mediated antiviral response following VZV infection. Indeed, in contrast to infection with Sendai virus, VZV infection of the hiPSC-neurons does not result in the upregulation of interferon-stimulated genes (ISGs) that have direct antiviral functions. Furthermore, the hiPSC-neurons do not produce interferon-α (IFNα), a major cytokine that is involved in the innate antiviral response, even upon its stimulation with strong synthetic inducers. In contrast, we showed that exogenous IFNα effectively limits VZV spread in the neuronal cell body compartment and demonstrated that ISGs are efficiently upregulated in these VZV-infected neuronal cultures that are treated with IFNα. Thus, whereas the cultured hiPSC neurons seem to be poor IFNα producers, they are good IFNα responders. This could suggest an important role for other cells such as satellite glial cells or macrophages to produce IFNα for VZV infection control.


Assuntos
Varicela , Herpes Zoster , Células-Tronco Pluripotentes Induzidas , Interferon-alfa , Neurônios , Humanos , Herpesvirus Humano 3/fisiologia , Células-Tronco Pluripotentes Induzidas/virologia , Interferon-alfa/imunologia , Neurônios/virologia , Células Cultivadas
15.
Viruses ; 14(11)2022 Nov 14.
Artigo em Inglês | MEDLINE | ID: mdl-36423127

RESUMO

Influenza virus infection may cause endothelial activation and dysfunction. However, it is still not known to what extent the influenza virus can dysregulate the expression of various endothelial proteins. The aim of the study is to identify the level of expression of endothelial nitric oxide synthase (eNOS), plasminogen activator inhibitor-1 (PAI-1), and tissue plasminogen activator (tPA) in the pulmonary vascular endothelium, as well as the concentration of PAI-1 and tPA in the blood plasma in Wistar rats. Animals were intranasally infected with rat-adapted influenza A(H1N1)pdm09 virus. The expression of eNOS, PAI-1 and tPA in the pulmonary vascular endothelium was determined by immunohistochemistry; the concentration of PAI-1 and tPA was analyzed by ELISA at 24 and 96 h post infection (hpi). Thus, the expression of eNOS in the pulmonary vascular endothelium decreased by 1.9-fold at 24 hpi and increased by 2-fold at 96 hpi. The expression of PAI-1 in the pulmonary vascular endothelium increased by 5.23-fold and 6.54-fold at 24 and 96 hpi, respectively. The concentration of PAI-1 in the blood plasma of the rats decreased by 3.84-fold at 96 hpi, but not at 24 hpi. The expression of tPA in the pulmonary vascular endothelium was increased 2.2-fold at 96 hpi. The obtained data indicate the development of endothelial dysfunction that is characterized by the dysregulation of endothelial protein expression in non-lethal and clinically non-severe experimental influenza virus infection.


Assuntos
Endotélio Vascular , Vírus da Influenza A Subtipo H1N1 , Infecções por Orthomyxoviridae , Animais , Ratos , Endotélio Vascular/metabolismo , Endotélio Vascular/virologia , Inibidor 1 de Ativador de Plasminogênio/metabolismo , Ratos Wistar , Ativador de Plasminogênio Tecidual/análise , Ativador de Plasminogênio Tecidual/metabolismo , Infecções por Orthomyxoviridae/metabolismo
16.
J Virol ; 96(22): e0148022, 2022 Nov 23.
Artigo em Inglês | MEDLINE | ID: mdl-36317880

RESUMO

Influenza A viruses (FLUAV) cause respiratory diseases in many host species, including humans and pigs. The spillover of FLUAV between swine and humans has been a concern for both public health and the swine industry. With the emergence of the triple reassortant internal gene (TRIG) constellation, establishment of human-origin FLUAVs in pigs has become more common, leading to increased viral diversity. However, little is known about the adaptation processes that are needed for a human-origin FLUAV to transmit and become established in pigs. We generated a reassortant FLUAV (VIC11pTRIG) containing surface gene segments from a human FLUAV strain and internal gene segments from the 2009 pandemic and TRIG FLUAV lineages and demonstrated that it can replicate and transmit in pigs. Sequencing and variant analysis identified three mutants that emerged during replication in pigs, which were mapped near the receptor binding site of the hemagglutinin (HA). The variants replicated more efficiently in differentiated swine tracheal cells compared to the virus containing the wildtype human-origin HA, and one of them was present in all contact pigs. These results show that variants are selected quickly after replication of human-origin HA in pigs, leading to improved fitness in the swine host, likely contributing to transmission. IMPORTANCE Influenza A viruses cause respiratory disease in several species, including humans and pigs. The bidirectional transmission of FLUAV between humans and pigs plays a significant role in the generation of novel viral strains, greatly impacting viral epidemiology. However, little is known about the evolutionary processes that allow human FLUAV to become established in pigs. In this study, we generated reassortant viruses containing human seasonal HA and neuraminidase (NA) on different constellations of internal genes and tested their ability to replicate and transmit in pigs. We demonstrated that a virus containing a common internal gene constellation currently found in U.S. swine was able to transmit efficiently via the respiratory route. We identified a specific amino acid substitution that was fixed in the respiratory contact pigs that was associated with improved replication in primary swine tracheal epithelial cells, suggesting it was crucial for the transmissibility of the human virus in pigs.


Assuntos
Glicoproteínas de Hemaglutininação de Vírus da Influenza , Vírus da Influenza A , Influenza Humana , Infecções por Orthomyxoviridae , Doenças dos Suínos , Animais , Humanos , Glicoproteínas de Hemaglutininação de Vírus da Influenza/genética , Vírus da Influenza A/genética , Influenza Humana/transmissão , Mutação , Infecções por Orthomyxoviridae/transmissão , Vírus Reordenados/genética , Suínos , Doenças dos Suínos/virologia
17.
J Virol ; 96(22): e0155522, 2022 Nov 23.
Artigo em Inglês | MEDLINE | ID: mdl-36317879

RESUMO

Porcine epidemic diarrhea virus (PEDV) is a re-emerging enteric coronavirus currently spreading in several nations and inflicting substantial financial damages on the swine industry. The currently available coronavirus vaccines do not provide adequate protection against the newly emerging viral strains. It is essential to study the relationship between host antiviral factors and the virus and to investigate the mechanisms underlying host immune response against PEDV infection. This study shows that heterogeneous nuclear ribonucleoprotein K (hnRNP K), the host protein determined by the transcription factor KLF15, inhibits the replication of PEDV by degrading the nucleocapsid (N) protein of PEDV in accordance with selective autophagy. hnRNP K was found to be capable of recruiting the E3 ubiquitin ligase, MARCH8, aiming to ubiquitinate N protein. Then, it was found that the ubiquitinated N protein could be delivered into autolysosomes for degradation by the cargo receptor NDP52, thereby inhibiting PEDV proliferation. Moreover, based on the enhanced MyD88 expression, we found that hnRNP K activated the interferon 1 (IFN-1) signaling pathway. Overall, the data obtained revealed a new mechanism of hnRNP K-mediated virus restriction wherein hnRNP K suppressed PEDV replication by degradation of viral N protein using the autophagic degradation pathway and by induction of IFN-1 production based on upregulation of MyD88 expression. IMPORTANCE The spread of the highly virulent PEDV in many countries is still leading to several epidemic and endemic outbreaks. To elucidate effective antiviral mechanisms, it is important to study the relationship between host antiviral factors and the virus and to investigate the mechanisms underlying host immune response against PEDV infection. In the work, we detected hnRNP K as a new host restriction factor which can hinder PEDV replication through degrading the nucleocapsid protein based on E3 ubiquitin ligase MARCH8 and the cargo receptor NDP52. In addition, via the upregulation of MyD88 expression, hnRNP K could also activate the interferon (IFN) signaling pathway. This study describes a previously unknown antiviral function of hnRNP K and offers a new vision toward host antiviral factors that regulate innate immune response as well as a protein degradation pathway against PEDV infection.


Assuntos
Infecções por Coronavirus , Ribonucleoproteínas Nucleares Heterogêneas Grupo K , Interferon Tipo I , Vírus da Diarreia Epidêmica Suína , Replicação Viral , Animais , Antivirais , Chlorocebus aethiops , Infecções por Coronavirus/veterinária , Ribonucleoproteínas Nucleares Heterogêneas Grupo K/genética , Interferons , Fator 88 de Diferenciação Mieloide , Proteínas do Nucleocapsídeo/fisiologia , Vírus da Diarreia Epidêmica Suína/fisiologia , Suínos , Doenças dos Suínos/virologia , Ubiquitina-Proteína Ligases , Células Vero , Interferon Tipo I/imunologia
18.
N Engl J Med ; 387(21): 1957-1968, 2022 11 24.
Artigo em Inglês | MEDLINE | ID: mdl-36346079

RESUMO

BACKGROUND: Bepirovirsen is an antisense oligonucleotide that targets all hepatitis B virus (HBV) messenger RNAs and acts to decrease levels of viral proteins. METHODS: We conducted a phase 2b, randomized, investigator-unblinded trial involving participants with chronic HBV infection who were receiving or not receiving nucleoside or nucleotide analogue (NA) therapy. Participants were randomly assigned (in a 3:3:3:1 ratio) to receive weekly subcutaneous injections of bepirovirsen at a dose of 300 mg for 24 weeks (group 1), bepirovirsen at a dose of 300 mg for 12 weeks then 150 mg for 12 weeks (group 2), bepirovirsen at a dose of 300 mg for 12 weeks then placebo for 12 weeks (group 3), or placebo for 12 weeks then bepirovirsen at a dose of 300 mg for 12 weeks (group 4). Groups 1, 2, and 3 received loading doses of bepirovirsen. The composite primary outcome was a hepatitis B surface antigen (HBsAg) level below the limit of detection and an HBV DNA level below the limit of quantification maintained for 24 weeks after the planned end of bepirovirsen treatment, without newly initiated antiviral medication. RESULTS: The intention-to-treat population comprised 457 participants (227 receiving NA therapy and 230 not receiving NA therapy). Among those receiving NA therapy, a primary-outcome event occurred in 6 participants (9%; 95% credible interval, 0 to 31) in group 1, in 6 (9%; 95% credible interval, 0 to 43) in group 2, in 2 (3%; 95% credible interval, 0 to 16) in group 3, and 0 (0%; post hoc credible interval, 0 to 8) in group 4. Among participants not receiving NA therapy, a primary-outcome event occurred in 7 participants (10%; 95% credible interval, 0 to 38), 4 (6%; 95% credible interval, 0 to 25), 1 (1%; post hoc credible interval, 0 to 6), and 0 (0%; post hoc credible interval, 0 to 8), respectively. During weeks 1 through 12, adverse events, including injection-site reactions, pyrexia, fatigue, and increased alanine aminotransferase levels, were more common with bepirovirsen (groups 1, 2, and 3) than with placebo (group 4). CONCLUSIONS: In this phase 2b trial, bepirovirsen at a dose of 300 mg per week for 24 weeks resulted in sustained HBsAg and HBV DNA loss in 9 to 10% of participants with chronic HBV infection. Larger and longer trials are required to assess the efficacy and safety of bepirovirsen. (Funded by GSK; B-Clear ClinicalTrials.gov number, NCT04449029.).


Assuntos
Antivirais , Hepatite B Crônica , Oligonucleotídeos Antissenso , RNA Viral , Humanos , Antivirais/efeitos adversos , Antivirais/uso terapêutico , DNA Viral/sangue , Antígenos E da Hepatite B/sangue , Antígenos de Superfície da Hepatite B/sangue , Vírus da Hepatite B/genética , Hepatite B Crônica/tratamento farmacológico , Hepatite B Crônica/virologia , Oligonucleotídeos Antissenso/administração & dosagem , Oligonucleotídeos Antissenso/efeitos adversos , Oligonucleotídeos Antissenso/uso terapêutico , Resultado do Tratamento , RNA Viral/efeitos dos fármacos , RNA Mensageiro/efeitos dos fármacos , Injeções Subcutâneas
19.
Nature ; 611(7936): 570-577, 2022 11.
Artigo em Inglês | MEDLINE | ID: mdl-36352231

RESUMO

Expanding our global testing capacity is critical to preventing and containing pandemics1-9. Accordingly, accessible and adaptable automated platforms that in decentralized settings perform nucleic acid amplification tests resource-efficiently are required10-14. Pooled testing can be extremely efficient if the pooling strategy is based on local viral prevalence15-20; however, it requires automation, small sample volume handling and feedback not available in current bulky, capital-intensive liquid handling technologies21-29. Here we use a swarm of millimetre-sized magnets as mobile robotic agents ('ferrobots') for precise and robust handling of magnetized sample droplets and high-fidelity delivery of flexible workflows based on nucleic acid amplification tests to overcome these limitations. Within a palm-sized printed circuit board-based programmable platform, we demonstrated the myriad of laboratory-equivalent operations involved in pooled testing. These operations were guided by an introduced square matrix pooled testing algorithm to identify the samples from infected patients, while maximizing the testing efficiency. We applied this automated technology for the loop-mediated isothermal amplification and detection of the SARS-CoV-2 virus in clinical samples, in which the test results completely matched those obtained off-chip. This technology is easily manufacturable and distributable, and its adoption for viral testing could lead to a 10-300-fold reduction in reagent costs (depending on the viral prevalence) and three orders of magnitude reduction in instrumentation cost. Therefore, it is a promising solution to expand our testing capacity for pandemic preparedness and to reimagine the automated clinical laboratory of the future.


Assuntos
Automação , Teste para COVID-19 , Imãs , Técnicas de Diagnóstico Molecular , Técnicas de Amplificação de Ácido Nucleico , Robótica , SARS-CoV-2 , Humanos , COVID-19/diagnóstico , COVID-19/virologia , Teste para COVID-19/métodos , Técnicas de Diagnóstico Molecular/economia , Técnicas de Diagnóstico Molecular/métodos , Técnicas de Amplificação de Ácido Nucleico/economia , Técnicas de Amplificação de Ácido Nucleico/métodos , Pandemias/prevenção & controle , RNA Viral/análise , RNA Viral/genética , SARS-CoV-2/genética , SARS-CoV-2/isolamento & purificação , Sensibilidade e Especificidade , Algoritmos , Automação/economia , Automação/métodos , Robótica/métodos , Indicadores e Reagentes/economia
20.
Viruses ; 14(11)2022 Oct 31.
Artigo em Inglês | MEDLINE | ID: mdl-36366518

RESUMO

The Omicron variant of SARS-CoV-2 spreads more easily than earlier variants, possibly as a result of a higher viral load in the upper respiratory tract and oral cavity. Hence, we investigated whether the Omicron variant generates a higher viral load than that of the Delta variant in saliva and nasopharynx. Both specimens were collected from 52 Omicron and 17 Delta cases at two time points one week apart and analyzed by qRT-PCR. Viral load was measured as 10 log RNA genome copies per 1000 human cells according to the WHO reference standard. We found that Omicron cases carried a higher viral load and had more sustained viral shedding compared to the Delta cases, especially in the nasopharynx.


Assuntos
COVID-19 , Saliva , Humanos , Nasofaringe/virologia , RNA Viral/genética , RNA Viral/análise , Saliva/virologia , SARS-CoV-2/genética , Carga Viral
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