Inhibit Enterovirus 71 Replication Research Articles

High mobility group box 1 knockdown inhibits EV71 replication and attenuates cell pyroptosis through TLR4/NF-κB/NLRP3 axis.

Enterovirus 71 (EV71) is a major causative agent of hand, foot, and mouth disease (HFMD) in children. Nowadays, there are still no effective antiviral drugs for EV71 infection. High mobility group box 1 (HMGB1) is reported to be highly expressed in HFMD patients. However, therole and underlying mechanism of HMGB1 in EV71-associated HFMD are still unclear. HMGB1 expression was detected using RT-qPCR and western blot assays. Loss- and gain-function experiments were performed to evaluate the effects of HMGB1 on EV71-infected cells. The virus titer was examined by TCID50. CCK-8 and flow cytometry assays were applied to detect the cell viability and cell cycle. Oxidative stress was determined by relative commercial kits. HMGB1 level was elevated in the serum of EV71-infected patients with HFMD and EV71-induced RD cells. EV71 infection induced the transfer of HMGB1 from the nucleus into the cytoplasm. HMGB1 knockdown inhibited virus replication,viral protein (VP1) expression and promoted antiviral factor expression. In addition, the inhibition of HMGB1 improved cell viability, protected against S phase arrest, and inhibited EV71-induced cell injury and oxidative stress, whereasHMGB1 overexpression showed the opposite effects. In terms of mechanism, HMGB1 overexpression activated the TLR4/NF-κB/NLRP3 signaling pathway and promoted cell pyroptosis. The inhibition of TLR4 and NF-κB reversed the effects of HMGB1 overexpression on virus replication, oxidative stress, and pyroptosis. In conclusion, HMGB1 knockdown inhibits EV71 replication and attenuates pyroptosis through TLR4/NF-κB/NLRP3 axis.

TLR4 signalling: the key to controlling EV71 replication and inflammatory response.

Hand, foot, and mouth disease (HFMD) is a common infectious disease caused by enterovirus 71 (EV71) that frequently affects children, leading to severe infections in some cases. In general, when infection occurs, the body upregulates inflammatory responses to eliminate pathogenic microorganisms to protect the host from infection. However, EV71 may inhibit host's innate immunity to promote virus infection. At present, it is not fully understood how EV71 hijack the host cells for its own replication. Toll-like receptor 4 (TLR4), a natural immune receptor, historically associated with bacterial endotoxin-induced inflammatory responses. However, it is still unclear whether and how TLR4 is altered during EV71 infection. In this study, we observed a reduction in both TLR4 protein and gene transcript levels in RD, GES-1, and Vero cells following EV71 infection, as detected by RT-qPCR, immunofluorescence staining and western blot. Furthermore, we observed that the TLR4 downstream molecules of MYD88, p-NF-κB p65, p-TBK1 and related inflammatory cytokines were also reduced, suggesting that antiviral innate immune and inflammatory response were suppressed. To determine the impact of TLR4 changes on EV71 infection, we interfered EV71-infected RD cells with TLR4 agonist or inhibitor and the results showed that activation of TLR4 inhibited EV71 replication, while inhibition of TLR4 promote EV71 replication. Besides, EV71 replication was also promoted in TLR4 siRNA-transfected and EV71-infected RD cells. This suggests that down-regulation the expression of TLR4 by EV71 can inhibit host immune defense to promote EV71 self-replication. This novel mechanism may be a strategy for EV71 to evade host immunity.

Targeting HNRNPA2B1 inhibits enterovirus 71 replication in SK-N-SH cells

ObjectiveTo investigate the effect of heterogeneous nuclear ribonucleoprotein A2B1 (HNRNPA2B1) on the replication of enterovirus 71 (EV-71) in SK-N-SH cells. MethodsThe mRNA and protein expression of HNRNPA2B1 in SK-N-SH cells were detected by real-time quantitative PCR (qRT-PCR) and western blotting (WB), respectively. WB was used to detect HNRNPA2B1 protein expression in the nucleus and cytosol. The localization of HNRNPA2B1 protein in the nucleus and cytosol was detected by immunofluorescence (IF). The expression of HNRNPA2B1 was inhibited by small interfering RNA (si-HNRNPA2B1). Viral RNA, viral structural protein VP1, and viral titer were detected by qRT-PCR, WB, and viral dilution counting, respectively. ResultsEV-71 infection significantly upregulates the expression of HNRNPA2B1 in SK-N-SH cells. EV-71 infection promotes HNRNPA2B1 nucleus-cytoplasm redistribution. Down-regulation of HNRNPA2B1 expression significantly inhibited EV-71 replication. ConclusionHNRNPA2B1 protein redistributed from nucleus to cytoplasm and is highly expressed in the cytoplasm during EV-71 infection. Inhibition of HNRNPA2B1 levels effectively inhibits EV-71 replication in SK-N-SH cells.

Cell membrane-bound toll-like receptor-1/2/4/6 monomers and -2 heterodimer inhibit enterovirus 71 replication by activating the antiviral innate response.

Host immune activation is critical for enterovirus 71 (EV71) clearance and immunopathogenesis. However, the mechanism of innate immune activation, especially of cell membrane-bound toll-like receptors (TLRs), against EV71 remains unknown. We previously demonstrated that TLR2 and its heterodimer inhibit EV71 replication. In this study, we systematically investigated the effects of TLR1/2/4/6 monomers and TLR2 heterodimer (TLR2/TLR1, TLR2/TLR6, and TLR2/TLR4) on EV71 replication and innate immune activation. We found that the overexpression of human- or mouse-derived TLR1/2/4/6 monomers and TLR2 heterodimer significantly inhibited EV71 replication and induced the production of interleukin (IL)-8 via activation of the phosphoinositide 3-kinase/protein kinase B (PI3K/AKT) and mitogen-activated protein kinase (MAPK) pathways. Furthermore,human-mouse chimeric TLR2 heterodimer inhibited EV71 replication and activated innate immunity. Dominant-negative TIR-less (DN)-TLR1/2/4/6 did not exert any inhibitory effects, whereas DN-TLR2 heterodimer inhibited EV71 replication. Prokaryotic expression of purified recombinant EV71 capsid proteins (VP1, VP2, VP3, and VP4) or overexpression of EV71 capsid proteins induced the production of IL-6 and IL-8 via activation of the PI3K/AKT and MAPK pathways. Notably, two types of EV71 capsid proteins served as pathogen-associated molecular patterns for TLR monomers (TLR2 and TLR4) and TLR2 heterodimer (TLR2/TLR1, TLR2/TLR6, and TLR2/TLR4) and activated innate immunity. Collectively, our results revealed that membrane TLRs inhibited EV71 replication via activation of the antiviral innate response, providing insights into the EV71 innate immune activation mechanism.

ML390 inhibits enterovirus 71 replication by targeting de novo pyrimidine biosynthesis pathway

Enterovirus 71 (EV71), a small, single-stranded, positive-sense RNA virus belonging to the enterovirus genus in the family Picornaviridae, causes hand, foot, and mouth disease. Although EV71 seriously threatens to public health, no effective antiviral drugs are available for treating this disease. In this study, we found that ML390, a dihydroorotate dehydrogenase inhibitor, has potential anti-EV71 activity. ML390 dose-dependently inhibited EV71 replication with IC50 and selectivity index values of 0.06601 μM and 156.5, respectively. Supplementation with the downstream product orotate significantly suppressed the ability of ML390 to inhibit EV71 replication. Moreover, an adequate supply of exogenous uridine and cytosine suppressed the anti-EV71 activity of ML390. Thus, the antiviral activity of ML390 is mediated by the inhibition of the pyrimidine synthesis pathway. In an EV71-infected mouse model, ML390 reduced the load of EV71 in the brain, liver, heart, spleen, front legs, and hind legs, and significantly increased the survival rate of the mice infected by EV71. ML390 shows potential for the treatment of hand, foot, and mouth disease caused by EV71 infection.

IMB-0523 Inhibits Enterovirus 71 Replication by Activating Signal Transducer and Activator of Transcription 3 Signaling to Upregulate Interferon-Stimulated Genes Expression

Abstract Background Hand, foot, and mouth disease caused by enterovirus 71 (EV71) infection is prevalent in the Asia-Pacific region in recent years. Currently, no drug is available for the prevention and treatment of EV71 infection. IMB-0523, a N-phenylbenzamide derivative, inhibits hepatitis B virus replication by upregulating the expression of APOBEC3G. In the present study, the effect of IMB-0523 on EV71 replication and related mechanism were investigated. Methods The cytotoxicity of IMB-0523 was determined by cell counting kit. Quantitative real-time polymerase chain reaction and Western blot assay were used to detect the effect of IMB-0523 on EV71 replication and related mechanism. Cytopathic effect assay was used to investigate the effect of IMB-0523 on different EV71 strains, coxsackievirus A16, and coxsackieviruses of group B. Results The results showed that IMB-0523 could dose-dependently inhibit EV71 replication. Preliminary mechanism studies showed that IMB-0523 could activate STAT3 signaling to upregulate the expression of interferon-stimulated genes to play an antiviral role. In addition, IMB-0523 inhibited the replication of different EV71 strains, coxsackievirus A16, and coxsackieviruses of group B. Conclusions IMB-0523 inhibits EV71 replication by activating the STAT3 signaling pathway to upregulate interferon-stimulated gene expression. IMB-0523 has broad-spectrum antiviral potential and may be used as a lead compound for the development of broad spectrum antiviral drugs.

TRAF3IP3 Is Cleaved by EV71 3C Protease and Exhibits Antiviral Activity.

Enterovirus 71 (EV71) is one of the major pathogens of hand, foot, and mouth disease, which poses a major risk to public health and infant safety. 3C protease (3Cpro), a non-structural protein of EV71, promotes viral protein maturation by cleaving polyprotein precursors and facilitates viral immune escape by cleaving host proteins. In this study, we screened for human proteins that could interact with EV71 3Cpro using a yeast two-hybrid assay. Immune-associated protein TRAF3 Interacting Protein 3 (TRAF3IP3) was selected for further study. The results of co-immunoprecipitation and immunofluorescence demonstrated the interaction between TRAF3IP3 and EV71 3Cpro. A cleavage band was detected, indicating that both transfected 3Cpro and EV71 infection could cleave TRAF3IP3. 87Q-88G was identified as the only 3Cpro cleavage site in TRAF3IP3. In Jurkat and rhabdomyosarcoma (RD) cells, TRAF3IP3 inhibited EV71 replication, and 3Cpro cleavage partially resisted TRAF3IP3-induced inhibition. Additionally, the nuclear localization signal (NLS) and nuclear export signal (NES) of TRAF3IP3 were identified. The NES contributed to TRAF3IP3 alteration of 3Cpro localization and inhibition of EV71 replication. Together, these results indicate that TRAF3IP3 inhibits EV71 replication and 3Cpro resists such inhibition via proteolytic cleavage, providing a new example of virus-host interaction.

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

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

Small Molecule Inhibits Enterovirus 71 Replication by Targeting the IRES Domain

Human Enterovirus 71 (EV71), one of the major etiological agents of the hand, foot, and mouth disease (HFMD), represents a serious global health threat against which there is no FDA‐approved treatment. Enteroviruses utilize a highly structured RNA domain, known as a type I Internal Ribosome Entry Site (IRES), to facilitate cap‐independent translation and genome replication by recruiting multiple host RNA‐binding proteins (ITAFs) that fine‐tune translation. Of particular interest, the stem‐loop II (SLII) IRES domain has been shown to specifically interact with several ITAFs such as hnRNP A1 and AUF1, having the SLII‐bulge as the binding surface to effectively modulate viral translation. Further studies showed that deletions and mutations to the bulge motif inhibit EV71 replication by offsetting IRES‐driven translation. Altogether, the results provide strong basis to classify the EV71 SLII domain as a promising target to develop novel antiviral agents. Here, we screened an RNA focused small molecule (rfsm) library against the EV71 SLII domain which yielded in the identification of six rfms that specifically bind to the bulge motif when detected by NMR spectroscopy and titration calorimetry. One of the aforementioned rfms impairs IRES‐dependent translation and displays dose‐dependent antiviral properties in cell culture. Furthermore, the NMR‐derived 3D structure of the SLII‐rfsm complex provides evidence that the small molecule binds to the bulge to induce a conformational change that exposes key nucleotides on the binding site for AUF1. This was further validated both in an in vitro biochemical assay and in vivo. Collectively, these results suggest an allosteric mechanism of inhibition in which the rfsm stabilizes the SLII‐AUF1 complex to repress translation. Therefore, this study provides structural and functional insight to develop novel RNA‐focused small molecule inhibitors of the Enterovirus genus.Support or Funding InformationNIH R01GM126833

Lycorine Derivative LY-55 Inhibits EV71 and CVA16 Replication Through Downregulating Autophagy.

Hand, foot, and mouth disease (HFMD) is a global health concern, especially in the Asia-Pacific region. HFMD caused by Enterovirus 71 (EV71) and Coxsackievirus A16 (CVA16) infection is usually self-limited but occasionally leads to severe pulmonary edema, neurological complications, and even death. Unfortunately, no effective drugs are currently available in clinical practice for the prevention and treatment of HFMD. Thus, anti-HFMD drugs must be urgently developed. A previous study had reported that lycorine could inhibit EV71 replication. In the present study, we found that LY-55, a lycorine derivative, inhibited the replication of EV71 and CVA16 in vitro and provided partial protection to mice from EV71 infection, as indicated by the decreased viral load and protein expression levels in muscles, clinical scores, and increased survival rates of infected mice. Mechanistically, LY-55 was not directly viricidal. Instead, the LY-55-mediated inhibition of EV71 and CVA16 was found to be mechanistically possible, at least in part, through downregulating autophagy, which plays an important role for EV71 and CVA16 replication. These findings suggest that LY-55 could be a potential lead or supplement for the development of anti-HFMD agents in the future.

APOBEC3G is a restriction factor of EV71 and mediator of IMB-Z antiviral activity

Enterovirus 71 (EV71), a single-stranded positive-sense RNA virus, is the causative agent of hand, foot, and mouth disease (HFMD), for which no effective antiviral therapy is currently available. Apolipoprotein B messenger RNA-editing enzyme catalytic polypeptide-like 3G (APOBEC3G or A3G) is a cytidine deaminase that inhibits the replication of several viruses, such as human immunodeficiency virus-1, hepatitis B virus and hepatitis C virus. In our efforts toward understanding the antiviral spectrum and mechanism of A3G, we found that ectopic expression of A3G inhibited EV71 replication, whereas knockdown of endogenous A3G expression promoted EV71 replication. Moreover, inhibition of EV71 replication by IMB-Z, a N-phenylbenzamide derivative, is associated with increased levels of intracellular A3G, but reducing the level of A3G by RNA interference diminished the antiviral activity of IMB-Z. Mechanistically, we obtained evidence suggesting that the cytidine deaminase activity is not required for A3G inhibition of EV71 replication. Instead, we demonstrated that A3G can interact with viral 3D RNA-dependent RNA polymerase (RdRp) and viral RNA and be packaged into progeny virions to reduce its infectivity. Taken together, our results indicate that A3G is a cellular restriction factor of EV71 and mediator of the antiviral activity of IMB-Z. Pharmacological induction and/or stabilization of A3G is a potential therapeutic approach to treat diseases caused by EV71 infection, such as HFMD.

The inactivation of the non-enveloped enterovirus 71 (EV71) by a novel disinfectant gel formulation for topical use

Background: Hand-foot-mouth disease may cause severe central nervous system complications and even death, that is induced mainly by enterovirus 71 (EV71), which is a non-enveloped virus. Inactivation of the EV71 on hands could effectively inhibit the transmission. However, the inactivations of the EV71 by conventional disinfectants including the alcohols are poor, due to the high stability of the EV71. A novel pyridyl imidazolidinone compound (TJAB1099) was designed to specifically inhibit EV71 replication in vitro. It may potentially be developed as formulations applied on hands for EV71 transmission control.Methods: The stress stability of TJAB1099 was first evaluated after storing in high temperature (60 °C, RH 10%), high humidity (25 °C, RH90%), and the high-intensity photolysis (4500 Lx ± 500 Lx) for 15 days, respectively. A wash-free antimicrobial gel containing the TJAB1099 was developed using the copolymer carrier. The antiviral activity, the acute oral toxicity, and the local irritation of the antimicrobial gel were evaluated accordingly.Results: The results indicated that the TJAB1099 was stable during the storage in high temperature and humidity. However, a significant change (p < .0001) was detected when TJAB1099 stored in the high-intensity photolysis. The antimicrobial gel containing 1 μM TJAB1099 could inhibit EV71 significantly higher than the ethanol (75%) (p < .0001) and commercialized disinfectant products (p < .0001). The results of acute oral toxicity and the local irritation suggest that the TJAB1099 containing antimicrobial gel was not causing skin irritations and acute oral toxicity symptoms.Conclusions: The results suggest that the antimicrobial gel containing TJAB1099 was safe and could effectively inhibit EV71 transmission in vitro.

Harmine, a small molecule derived from natural sources, inhibits enterovirus 71 replication by targeting NF-κB pathway

Enterovirus 71 (EV71) infection of young children can cause neurological manifestations, which is mainly responsible for the fatality. Although a vaccine is recently available for preventing enterovirus 71 infection, its efficacy remains to be seen. Therefore, there is a pressing need for anti-viral agents for the treatment of EV71 infection. By screening a natural compound library for inhibitory activity of EV71 replication, we identified a small molecule, harmine, that inhibited EV71 replication by targeting NF-κB signaling pathway. Harmine is a β-carboline alkaloid found in the medicinal plant Peganum harmala, which is used as a folk antitumor medicine in China and other parts of the Asia. The estimated EC50 value for harmine to block EV71 infection was 20 μM, while the CC50 was estimated at 500 μM in vitro. Harmine inhibited replication of EV71, as evidenced by its ability to diminish plague formation induced by EV71 and to reduce the level of viral RNA and protein. Mechanistic studies indicated that harmine suppressed EV71 replication through inhibition of NF-κB signaling pathway. Harmine treatment also reduced EV71-induced reactive oxygen species (ROS) formation, which was associated with a decline in EV71-associated NF-κB activation. In addition, the harmine treatment could protect AG129 mice against EV71 replication in vivo. These findings suggest that harmine may present as a candidate antiviral drug for the treatment of EV71 infection.

Fibronectin Facilitates Enterovirus 71 Infection by Mediating Viral Entry.

Fibronectin (FN) is a high-molecular-weight extracellular matrix protein that contains the RGDS motif, which is required to bind to integrins. Synthetic RGDS peptides have been reported to compete with FN to bind to the cell surface and inhibit the function of FN. Here, we identified that synthetic RGDS peptides significantly inhibit human enterovirus 71 (EV71) infection in cell cultures. In addition, mice treated with RGDS peptides and infected with EV71 had a significantly higher survival rate and a lower viral load than the control group. Because RGDS peptides affect the function of FN, we questioned whether FN may play a role in virus infection. Our study indicates that overexpression of FN enhanced EV71 infection. In contrast, knockout of FN significantly reduced viral yield and decreased the viral binding to host cells. Furthermore, EV71 entry, rather than intracellular viral replication, was blocked by FN inhibitor pretreatment. Next, we found that FN could interact with the EV71 capsid protein VP1, and further truncated-mutation assays indicated that the D2 domain of FN could interact with the N-terminal fragment of VP1. Taken together, our results demonstrate that the host factor FN binds to EV71 particles and facilitates EV71 entry, providing a potential therapy target for EV71 infection.IMPORTANCE Hand, foot, and mouth disease outbreaks have occurred frequently in recent years, sometimes causing severe neurological complications and even death in infants and young children worldwide. Unfortunately, no effective antiviral drugs are available for human enterovirus 71 (EV71), one of the viruses that cause hand, foot, and mouth disease. The infection process and the host factors involved remain unknown, although several receptors have been identified. In this study, we found that the host factor fibronectin (FN) facilitated EV71 replication by interacting with EV71 particles and further mediated their entry. The RGDS peptide, an FN inhibitor, significantly inhibited EV71 replication in both RD cells and mice. In conclusion, our research identified a new host factor involved in EV71 infection, providing a new potential antiviral target for EV71 treatment.

Honeysuckle-encoded microRNA2911 inhibits Enterovirus 71 replication via targeting VP1 gene

Enterovirus 71 (EV71) is the primary pathogen of hand-foot-and-mouth disease (HFMD) in children and virus infections are associated with severe neurological dysfunctions and even death. MIR2911 is a honeysuckle-encoded atypical microRNA with extreme stability. Here, we report that MIR2911 directly inhibits EV71 replication by targeting the VP1 gene. Bioinformatics prediction and luciferase reporter assay showed that MIR2911 could target the VP1 gene of EV71. Transfection experiments using synthetic MIR2911 and extracted RNA from HS decoction shown that each of these preparations was capable of inhibiting EV71 VP1 protein expression; however, these preparations did not impact EV71 mutants in which the MIR2911-binding sites were mutated. Furthermore, EV71 replication was increased by antagomirs against MIR2911 in the HS decoction, implying that MIR2911 was physiologically functional in controlling EV71 replication in vitro. These results indicated that, by targeting VP1 gene, MIR2911 may effectively inhibit EV71 replication. Our results also provide a potential novel strategy on the therapy and/or prevention of HFMD originating from EV71 virus infection.

GS-9620 inhibits enterovirus 71 replication mainly through the NF-κB and PI3K-AKT signaling pathways

Human enterovirus 71 (EV71) is the second most common cause of hand, foot, and mouth disease (HFMD), which can occur as a severe epidemic especially among children under 5-years old. New and improved treatment strategies to control EV71 infection are therefore urgently required. The heterocyclic compound GS-9620, a potent and selective agonist of Toll-like receptor 7 (TLR7), has been reported to activate plasmacytoid dendritic cells (pDCs), and suppress HBV as well as HIV replication. In this study, we indicated that GS-9620 also could inhibit EV71 replication in the mouse model of EV71 infection. With three-days treatment after EV71 infection, the levels of proinflammatory cytokines/chemokines, like IFN-α, IFN-γ and MCP-1, were sharply reduced in serum compared to those without treatment. Furthermore, GS-9620 activated TLR7 in the limb muscle cells, which stimulated the NF-κB and PI3K/AKT signaling pathways. When NF-κB or PI3K/AKT inhibitors were used, the antiviral effect of the GS-9620 was impacted. Overall, our data implied GS-9620 probably activates NF-κB and PI3K/AKT signaling pathways to clear the virus.

MiR-16-5p mediates a positive feedback loop in EV71-induced apoptosis and suppresses virus replication

Enterovirus 71 (EV71) is the predominant causative pathogen of hand-foot-and-mouth disease (HFMD). Contrary to other HFMD-causing enterovirus, EV71 can lead to severe neurological complications, even death. MicroRNAs (miRNAs) are small non-coding RNAs that constitute the largest family of gene regulators participating in numerous biological or pathological processes. We previously reported that miR-16-5p increases with severity of HFMD by investigating the expression patterns of host miRNAs in patients with HFMD. However, the mechanisms by which EV71 induces miR-16-5p expression are not clear, and the interaction between EV71 and miR-16-5p is not yet fully understood. Here, we confirmed EV71-induced expression of miR-16-5p both in vitro and in vivo and show that upregulation of miR-16-5p by EV71 infection may occur at the posttranscriptional level. Moreover, EV71-induced caspase activation facilitates the processing of pri-miR-16-1. We also revealed that miR-16-5p can promote EV71-induced nerve cells apoptosis through activating caspase-3. In addition, we found that miR-16-5p can inhibit EV71 replication. CCNE1 and CCND1, two important cell cycle regulators, play an important role in the suppression of EV71 replication by miR-16-5p. Therefore, miR-16-5p is a positive feedback regulator in EV71-induced apoptosis and a suppressor of virus replication. These results help in understanding the interaction network between miRNA and EV71 infection and provide a potential target for the development of antiviral therapy.

Corydaline inhibits enterovirus 71 replication by regulating COX-2 expression

Enterovirus 71 (EV71) is a huge threat to the worldwide public health and there is no approved antiviral drug for EV71-induced disease therapy. Corydaline exists antiallergic and antinociceptive activities, but the anti-EV71 activity of corydaline is still not reported. In this study, corydaline could suppress the expression of viral structural and non-structural proteins. Furthermore, corydaline inhibits EV71 replication by suppressing the COX-2 expression and the phosphorylation of JNK MAPK and P38 MAPK but not ERK MAPK in vitro. Based on these findings, corydaline could be a potential lead or supplement for the development of new anti-EV71 agents in the future.

MiR-127-5p negatively regulates enterovirus 71 replication by directly targeting SCARB2.

Enterovirus 71 (EV71) is the major causative agent of hand‐foot‐and‐mouth disease in young children and can cause severe cerebral and pulmonary complications and even fatality. This study aimed at elucidating whether and how EV71 infection is regulated by a cellular microRNA, miR‐127‐5p. We found that miR‐127‐5p can downregulate the expression of SCARB2, a main receptor of EV71, by targeting two potential sites in its 3′ UTR region and inhibit EV71 infection. Meanwhile, miR‐127‐5p expression was upregulated during EV71 infection. Notably, transfecting cells with miR‐127‐5p mimics led to a significant decrease in viral replication, while inhibition of endogenous miR‐127‐5p facilitated viral replication. Furthermore, our evidence showed that miR‐127‐5p did not affect postentry viral replication. Taken together, these results indicated that miR‐127‐5p inhibited EV71 replication by targeting the SCARB2 mRNA.

Berberine inhibits enterovirus 71 replication by downregulating the MEK/ERK signaling pathway and autophagy

BackgroundThe MEK-ERK signaling pathway and autophagy play an important role for enterovirus71(EV71) replication. Inhibition of MEK-ERK signaling pathway and autophagy is shown to impair EV71 replication. Berberine (BBR), an isoquinoline alkaloid isolated from Berberis vulgaris L., has been reported to have ability to regulate this signaling pathway and autophagy. Herein, we want to determine whether berberine can inhibit EV71 infection by downregulating the MEK/ERK signaling pathway and autophagy.MethodsThe antiviral effect of berberine was determined by cytopathic effect (CPE) assay, western blotting assay and qRT-PCR assay. The mechanism of BBR anti-virus was determined by western blotting assay and immunofluorescence assay.ResultsWe showed that berberine does-dependently reduced EV71 RNA and protein synthesis, which was, at least in part, the result of inhibition of activation of MEK/ERK signaling pathway. Furthermore, we found that berberine suppressed the EV71-induced autophagy by activating AKT protein and inhibiting the phosphorylation of JNK and PI3KIII.ConclusionsBBR inhibited EV71 replication by downregulating autophagy and MEK/ERK signaling pathway. These findings suggest that BBR may be a potential agent or supplement against EV71 infection.