Dataset Information

MicroRNA 876-5p modulates EV-A71 replication through downregulation of host antiviral factors.

ABSTRACT: BACKGROUND:Human enterovirus 71 (EV-A71) is a non-enveloped virus that has a single stranded positive sense RNA genome. In a previous study, we showed that miR-876-5p upregulation was observed in the serum of patients with severe EV-A71 infection. Micro-876-5p (miR-876-5p) is a circulating miRNA that can be identified to modulate EV-A71 infections through both in vitro and in vivo studies. However, the regulatory mechanisms that involve miR-876-5p in the EV-A71 infection cycle remain unclear. METHODS:We demonstrated that miR-876-5p facilitated EV-A71 replication and expression by overexpression and knocking-down of miR-876-5p through the transfection of miR-876-5p plasmid and miR-876-5p inhibitor. Although miR-876-5p suppressed CREB5 expression, luciferase reporter assay confirmed this. We also evaluated the role of miR-876-5p in the EV-A71 infection cycle by CREB5 mediated by transfection with an anti-miR-876-5P inhibitor or in combination with an si-CREB5 plasmid. RESULTS:MicroR-876-5p was upregulated in EV-A71-infected neuroblastoma cells. Overexpression of miR-876-5p or knockdown of cyclic-AMP responsive element binding protein 5 (CREB5) promoted EV-A71 replication. The downregulation of miR-876-5p inhibited the accumulation of viral RNA and the production of viral proteins. Interestingly, CREB5 overexpression also suppressed EV-A71 replication. Our in vitro studies reveal that miR-876-5p directly targets CREB5. Finally, downregulation of CREB5 protein abated the inhibitory effect of anti-miR-876-5p and induced inhibitory effect of EV-A71 replication. CONCLUSIONS:Our results suggest that intracellular miR-876-5p promotes EV-A71 replication indirectly by targeting the host CREB5 protein.

SUBMITTER: Xu P

PROVIDER: S-EPMC7003331 | biostudies-literature | 2020 Feb

REPOSITORIES: biostudies-literature

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MicroRNA 876-5p modulates EV-A71 replication through downregulation of host antiviral factors.

Xu Peng P Xu Hwa H Cheng Hsu Sheng HS Chan Han-Hsiang HH Wang Robert Y L RYL

Virology journal 20200205 1

<h4>Background</h4>Human enterovirus 71 (EV-A71) is a non-enveloped virus that has a single stranded positive sense RNA genome. In a previous study, we showed that miR-876-5p upregulation was observed in the serum of patients with severe EV-A71 infection. Micro-876-5p (miR-876-5p) is a circulating miRNA that can be identified to modulate EV-A71 infections through both in vitro and in vivo studies. However, the regulatory mechanisms that involve miR-876-5p in the EV-A71 infection cycle remain unc ...[more]

PMID: 32024541

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Project description:BACKGROUND:The hand, foot and mouth disease (HFMD) is a febrile and exanthematous childhood disease mainly caused by Enterovirus 71 (EV-A71). In severe HFMD, virulent EV-A71 strains can cause acute flaccid paralysis and cardiopulmonary edema leading to death. Currently, no FDA approved antiviral treatment or vaccine is available for EV-A71. Flavonoids such as silymarin and baicalein are known to possess in vitro antiviral properties against viruses. In this study, the cytotoxicity and antiviral activity of silymarin, baicalein and baicalin were investigated. METHODS:The cytotoxic effects of three flavonoids towards rhabdomyosarcoma (RD) cells were first examined using cell proliferation MTS [3-(4,5-dimethylthiazol-2-yl)-5-(3-carboxymethoxyphenyl)-2-(4-sulfophenyl)-2H-tetrazolium] assay. Compounds found to be non-cytotoxic in RD cells were evaluated for their in vitro antiviral properties against the EV-A71 subgenotype B4 strain 41 (5865/SIN/000009) using antiviral assays. Viral infectivity was determined by reduction of the formation of plaques in RD cells. For the measurement of RNA copy number, the real time quantitative reverse transcription PCR (qRT-PCR) was used. The most potent compound was further evaluated to determine the mode of action of inhibition by time course, virus attachment and entry assays in Vero cells. RESULTS:Silymarin was shown to exert direct extracellular virucidal effects against EV-A71 at 50% inhibitory concentration (IC50) of 15.2 ± 3.53 μg/mL with SI of 10.53. Similarly, baicalein exhibited direct extracellular virucidal effects against EV-A71 at a higher IC50 value of 30.88 ± 5.50 μg/mL with SI of 13.64. Besides virucidal activity, silymarin was shown to block both viral attachment and entry of EV-A71 to inhibit infection in Vero cells. CONCLUSIONS:Silymarin has a stronger inhibition activity against EV-A71 in comparison to baicalein. It could serve as a promising antiviral drug to treat EV-A71 infections.

Project description:Backgroundβ-Amyloid (Aβ) protein is a pivotal pathogenetic factor in Alzheimer's disease (AD). However, increasing evidence suggests that the brain has to continuously produce excessive Aβ to efficaciously prevent pathogenic micro-organism infections, which induces and accelerates the disease process of AD. Meanwhile, Aβ exhibits activity against herpes simplex virus type 1 (HSV-1) and influenza A virus (IAV) replication, but not against other neurotropic viruses. Enterovirus A71 (EV-A71) is the most important neurotropic enterovirus in the post-polio era. Given the limitation of existing research on the relationship between Aβ and other virus infections, this study aimed to investigate the potent activity of Aβ on EV-A71 infection and extended the potential function of Aβ in other unenveloped viruses may be linked to Alzheimer's disease or infectious neurological diseases.MethodsAβ peptides 1-42 are a major pathological factor of senile plaques in Alzheimer's disease (AD). Thus, we utilized Aβ1-42 as a test subject to perform our study. The production of monomer Aβ1-42 and their high-molecular oligomer accumulations in neural cells were detected by immunofluorescence assay, ELISA, or Western blot assay. The inhibitory activity of Aβ1-42 peptides against EV-A71 in vitro was detected by Western blot analysis or qRT-PCR. The mechanism of Aβ1-42 against EV-A71 replication was analyzed by time-of-addition assay, attachment inhibition assay, pre-attachment inhibition analysis, viral-penetration inhibition assay, TEM analysis of virus agglutination, and pull-down assay.ResultsWe found that EV-A71 infection induced Aβ production and accumulation in SH-SY5Y cells. We also revealed for the first time that Aβ1-42 efficiently inhibited the RNA level of EV-A71 VP1, and the protein levels of VP1, VP2, and nonstructural protein 3AB in SH-SY5Y, Vero, and human rhabdomyosarcoma (RD) cells. Mechanistically, we demonstrated that Aβ1-42 primarily targeted the early stage of EV-A71 entry to inhibit virus replication by binding virus capsid protein VP1 or scavenger receptor class B member 2. Moreover, Aβ1-42 formed non-enveloped EV-A71 particle aggregates within a certain period and bound to the capsid protein VP1, which partially caused Aβ1-42 to prevent viruses from infecting cells.ConclusionsOur findings unveiled that Aβ1-42 effectively inhibited nonenveloped EV-A71 by targeting the early phase of an EV-A71 life cycle, thereby extending the potential function of Aβ in other non-envelope viruses linked to infectious neurological diseases.

Project description:BackgroundHuman enteroviruses A71 (EV-A71) and D68 (EV-D68) are the suspected causative agents of hand-foot-and-mouth disease, aseptic meningitis, encephalitis, acute flaccid myelitis, and acute flaccid paralysis in children. Until now, no cure nor mucosal vaccine existed for EV-A71 and EV-D68. Novel mucosal bivalent vaccines are highly important for preventing EV-A71 and EV-D68 infections.MethodsIn this study, formalin-inactivated EV-A71 and EV-D68 were used as antigens, while PS-G, a polysaccharide from Ganoderma lucidum, was used as an adjuvant. Natural polysaccharides have the characteristics of intrinsic immunomodulation, biocompatibility, low toxicity, and safety. Mice were immunized intranasally with PBS, EV-A71, EV-D68, or EV-A71 + EV-D68, with or without PS-G as an adjuvant.ResultsThe EV-A71 + EV-D68 bivalent vaccine generated considerable EV-A71- and EV-D68-specific IgG and IgA titres in the sera, nasal washes, saliva, bronchoalveolar lavage fluid, and feces. These antibodies neutralized EV-D68 and EV-A71 infectivity. They also cross-neutralized infections by different EV-D68 and EV-A71 sub-genotypes. Furthermore, compared with the PBS group, EV-A71 + EV-D68 + PS-G-vaccinated mice exhibited an increased number of EV-D68- and EV-A71-specific IgA- and IgG-producing cells. In addition, T-cell proliferative responses, and IFN-γ and IL-17 secretion in the spleen were substantially induced when PS-G was used as an adjuvant with EV-A71 + EV-D68. Finally, in vivo challenge experiments demonstrated that the immune sera induced by EV-A71 + EV-D68 + PS-G conferred protection in neonate mice against lethal EV-A71 and EV-D68 challenges as indicated by the increased survival rate and decreased clinical score and viral RNA tissue expression. Taken together, all EV-A71/EV-D68 + PS-G-immunized mice developed potent specific humoral, mucosal, and cellular immune responses to EV-D68 and EV-A71 and were protected against them.ConclusionsThese findings demonstrated that PS-G can be used as a potential adjuvant for EV-A71 and EV-D68 bivalent mucosal vaccines. Our results provide useful information for the further preclinical and clinical development of a mucosal bivalent enterovirus vaccine against both EV-A71 and EV-D68 infections.

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MicroRNA 876-5p modulates EV-A71 replication through downregulation of host antiviral factors.

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MicroRNA 876-5p modulates EV-A71 replication through downregulation of host antiviral factors.

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