RESUMEN
Thrombosis represents the leading cause of death and disability upon major adverse cardiovascular events (MACEs). Numerous pathological conditions such as COVID-19 and metabolic disorders can lead to a heightened thrombotic risk; however, the underlying mechanisms remain poorly understood. Our study illustrates that 2-methylbutyrylcarnitine (2MBC), a branched-chain acylcarnitine, is accumulated in patients with COVID-19 and in patients with MACEs. 2MBC enhances platelet hyperreactivity and thrombus formation in mice. Mechanistically, 2MBC binds to integrin α2ß1 in platelets, potentiating cytosolic phospholipase A2 (cPLA2) activation and platelet hyperresponsiveness. Genetic depletion or pharmacological inhibition of integrin α2ß1 largely reverses the pro-thrombotic effects of 2MBC. Notably, 2MBC can be generated in a gut-microbiota-dependent manner, whereas the accumulation of plasma 2MBC and its thrombosis-aggravating effect are largely ameliorated following antibiotic-induced microbial depletion. Our study implicates 2MBC as a metabolite that links gut microbiota dysbiosis to elevated thrombotic risk, providing mechanistic insight and a potential therapeutic strategy for thrombosis.
Asunto(s)
COVID-19 , Microbioma Gastrointestinal , Trombosis , Humanos , Ratones , Animales , Integrina alfa2beta1/genética , Integrina alfa2beta1/metabolismo , Colágeno/metabolismo , Plaquetas/metabolismo , COVID-19/metabolismoRESUMEN
Chikungunya virus (CHIKV) and Dengue virus (DENV) are vector-borne diseases transmitted by Aedes aegypti and Aedes albopictus that pose a significant threat to global public health. Cases of acute Chikungunya fever often present similar clinical symptoms to other vector-borne diseases, such as Dengue fever. In regions where multiple vector-borne diseases coexist, CHIKV is often overlooked or misdiagnosed as Dengue virus, West Nile virus, Zika virus or other viral infections, which delays its prevention and control. However, IgM antibodies directed against the E2 protein of CHIKV have not yet been generalized to clinical settings due to the low sensitivity and high cost in commercial kits. Indirect ELISA with peptides provides an effective supplementary tool for detecting CHIKV IgM antibodies. Our study aims at examining the potential of linear epitopes on the E2 glycoprotein that specifically bind to IgM antibodies as serodiagnostic tool for CHIKV. The sensitivity of the established peptide indirect ELISA method for detecting clinical samples is significantly better than that of commercial kits, realizing a beneficial supplement to the existing IgM antibody assay. It also established the groundwork for comprehending the biological mechanisms of the CHIKV E2 protein and the advancement of innovative epitope peptide vaccines.
Asunto(s)
Fiebre Chikungunya , Virus Chikungunya , Dengue , Infección por el Virus Zika , Virus Zika , Humanos , Fiebre Chikungunya/diagnóstico , Epítopos , Pruebas Serológicas , Proteínas Virales , Infección por el Virus Zika/diagnóstico , Anticuerpos Antivirales , Inmunoglobulina MRESUMEN
Bismuth-based metal-organic frameworks (Bi-MOFs) have received attention in electrochemical CO2-to-formate conversion. However, the low conductivity and saturated coordination of Bi-MOFs usually lead to poor performance, which severely limits their widespread application. Herein, a conductive catecholate-based framework with Bi-enriched sites (HHTP, 2,3,6,7,10,11-hexahydroxytriphenylene) is constructed and the zigzagging corrugated topology of Bi-HHTP is first unraveled via single-crystal X-ray diffraction. Bi-HHTP possesses excellent electrical conductivity (1.65 S m-1) and unsaturated coordination Bi sites are confirmed by electron paramagnetic resonance spectroscopy. Bi-HHTP exhibited an outstanding performance for selective formate production of 95% with a maximum turnover frequency of 576 h-1 in a flow cell, which surpassed most of the previously reported Bi-MOFs. Significantly, the structure of Bi-HHTP could be well maintained after catalysis. In situ attenuated total reflectance Fourier transform infrared spectroscopy (ATR-FTIR) confirms that the key intermediate is *COOH species. Density functional theory (DFT) calculations reveal that the rate-determining step is *COOH species generation, which is consistent with the in situ ATR-FTIR results. DFT calculations confirmed that the unsaturated coordination Bi sites acted as active sites for electrochemical CO2-to-formate conversion. This work provides new insights into the rational design of conductive, stable, and active Bi-MOFs to improve their performance towards electrochemical CO2 reduction.
RESUMEN
The SARS-CoV-2 Omicron variant evades most currently approved neutralizing antibodies (nAbs) and caused drastic decrease of plasma neutralizing activity elicited by vaccination or prior infection, urging the need for the development of pan-variant antivirals. Breakthrough infection induces a hybrid immunological response with potentially broad, potent and durable protection against variants, therefore, convalescent plasma from breakthrough infection may provide a broadened repertoire for identifying elite nAbs. We performed single-cell RNA sequencing (scRNA-seq) and BCR sequencing (scBCR-seq) of B cells from BA.1 breakthrough-infected patients who received 2 or 3 previous doses of inactivated vaccine. Elite nAbs, mainly derived from the IGHV2-5 and IGHV3-66/53 germlines, showed potent neutralizing activity across Wuhan-Hu-1, Delta, Omicron sublineages BA.1 and BA.2 at picomolar NT50 values. Cryo-EM analysis revealed diverse modes of spike recognition and guides the design of cocktail therapy. A single injection of paired antibodies cocktail provided potent protection in the K18-hACE2 transgenic female mouse model of SARS-CoV-2 infection.
Asunto(s)
COVID-19 , SARS-CoV-2 , Femenino , Animales , Ratones , SARS-CoV-2/genética , Infección Irruptiva , Sueroterapia para COVID-19 , Anticuerpos Neutralizantes , Ratones Transgénicos , Anticuerpos AntiviralesRESUMEN
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has caused a global coronavirus disease 2019 (COVID-19) pandemic that has affected the lives of billions of individuals. However, the host-virus interactions still need further investigation to reveal the underling mechanism of SARS-CoV-2 pathogenesis. Here, transcriptomics analysis of SARS-CoV-2 infection highlighted possible correlation between host-associated signaling pathway and virus. In detail, cAMP-protein kinase (PKA) pathway has an essential role in SARS-CoV-2 infection, followed by the interaction between cyclic AMP response element binding protein (CREB) and CREB-binding protein (CBP) could be induced and leading to the enhancement of CREB/CBP transcriptional activity. The replication of Delta and Omicron BA.5 were inhibited by about 49.4% and 44.7% after knockdown of CREB and CBP with small interfering RNAs, respectively. Furthermore, a small organic molecule naphthol AS-E (nAS-E), which targets on the interaction between CREB and CBP, potently inhibited SARS-CoV-2 wild-type (WT) infection with comparable the half-maximal effective concentration (EC50 ) 1.04 µM to Remdesivir 0.57 µM. Compared with WT virus, EC50 in Calu-3 cells against Delta, Omicron BA.2, and Omicron BA.5 were, on average, 1.5-fold, 1.1-fold, and 1.5-fold higher, respectively, nAS-E had a satisfied antiviral effect against Omicron variants. Taken together, our study demonstrated the importance of CREB/CBP induced by cAMP-PKA pathway during SARS-CoV-2 infection, and further provided a novel CREB/CBP interaction therapeutic drug targets for COVID-19.
Asunto(s)
COVID-19 , Proteína de Unión a Elemento de Respuesta al AMP Cíclico , Interacciones Huésped-Patógeno , Humanos , COVID-19/metabolismo , AMP Cíclico/metabolismo , Proteína de Unión a Elemento de Respuesta al AMP Cíclico/metabolismo , Proteínas Quinasas Dependientes de AMP Cíclico/metabolismo , SARS-CoV-2/metabolismo , SARS-CoV-2/patogenicidad , Proteína de Unión a CREB/metabolismo , Interacciones Huésped-Patógeno/genética , Interacciones Huésped-Patógeno/fisiologíaRESUMEN
This article addresses the event-triggered coordinated control problem for multiple Euler-Lagrange systems subject to parameter uncertainties and external disturbances. Based on the event-triggered technique, a distributed coordinated control scheme is first proposed, where the neural network-based estimation method is incorporated to compensate for parameter uncertainties. Then, an input-based continuous event-triggered (CET) mechanism is developed to schedule the triggering instants, which ensures that the control command is activated only when some specific events occur. After that, by analyzing the possible finite-time escape behavior of the triggering function, the real-time data sampling and event monitoring requirement in the CET strategy is tactfully ruled out, and the CET policy is further transformed into a periodic event-triggered (PET) one. In doing so, each agent only needs to monitor the triggering function at the preset periodic sampling instants, and accordingly, frequent control updating is further relieved. Besides, a parameter selection criterion is provided to specify the relationship between the control performance and the sampling period. Finally, a numerical example of attitude synchronization for multiple satellites is performed to show the effectiveness and superiority of the proposed coordinated control scheme.
RESUMEN
There is an urgent need for a broad-spectrum and protective vaccine due to the emergence and rapid spreading of more contagious SARS-CoV-2 strains. We report the development of RBMRNA-176, a pseudouridine (Ψ) nucleoside-modified mRNA-LNP vaccine encoding pre-fusion stabilized trimeric SARS-CoV-2 spike protein ectodomain, and evaluate its immunogenicity and protection against virus challenge in mice and nonhuman primates. A prime-boost immunization with RBMRNA-176 at intervals of 21 days resulted in high IgG titers (over 1:819,000 endpoint dilution) and a CD4+ Th1-biased immune response in mice. RBMRNA-176 vaccination induced pseudovirus-neutralizing antibodies with IC50 ranging from 1:1020 to 1:2894 against SARS-CoV-2 spike pseudotyped wild-type and variant viruses, including Alpha, Beta, Gamma, and Kappa. Moreover, significant control of viral replication and histopathology in lungs was observed in vaccinated mice. In nonhuman primates, a boost given by RBMRNA-176 on day 21 after the prime induced a persistent and sustained IgG response. RBMRNA-176 vaccination also protected macaques against upper and lower respiratory tract infection, as well as lung injury. Altogether, these findings support RBMRNA-176 as a vaccine candidate for prevention of COVID-19.
RESUMEN
Infection by severe acute respiratory syndrome-related coronavirus 2 (SARS-CoV-2) has posed a severe threat to global public health. The current study revealed that several inhibitors of protein kinases C (PKCs) possess protective activity against SARS-CoV-2 infection. Four pan-PKC inhibitors, Go 6983, bisindolylmaleimide I, enzastaurin, and sotrastaurin, reduced the replication of a SARS-CoV-2 replicon in both BHK-21 and Huh7 cells. A PKCδ-specific inhibitor, rottlerin, was also effective in reducing viral infection. The PKC inhibitors acted at an early step of SARS-CoV-2 infection. Finally, PKC inhibitors blocked the replication of wild-type SARS-CoV-2 in ACE2-expressing A549 cells. Our work highlights the importance of the PKC signaling pathway in infection by SARS-CoV-2 and provides evidence that PKC-specific inhibitors are potential therapeutic agents against SARS-CoV-2. IMPORTANCE There is an urgent need for effective therapeutic drugs to control the pandemic caused by severe acute respiratory syndrome-related coronavirus 2 (SARS-CoV-2). We found that several inhibitors of protein kinases C (PKCs) dramatically decrease the replication of SARS-CoV-2 in cultured cells. These PKC inhibitors interfere with an early step of viral infection. Therefore, the rapid and prominent antiviral effect of PKC inhibitors underscores that they are promising antiviral agents and suggests that PKCs are important host factors involved in infection by SARS-CoV-2.
Asunto(s)
Antivirales , Proteína Quinasa C , SARS-CoV-2 , Humanos , Enzima Convertidora de Angiotensina 2 , Antivirales/farmacología , Células Cultivadas , Proteína Quinasa C/farmacología , SARS-CoV-2/efectos de los fármacos , Tratamiento Farmacológico de COVID-19RESUMEN
Intestinal ischemia/reperfusion (II/R) is a common acute and critical condition in clinical practice with a high mortality rate. However, there is still a lack of effective prevention and treatment measures for II/R injury. The role of the gut microbiota in II/R has attracted widespread attention. Recent evidence has demonstrated that the gut microbiota plays a pivotal role in the occurrence, development, and prognosis of II/R. Therefore, maintaining the homeostasis of gut microbiota and its metabolites may be a potential strategy for the treatment of II/R. This review focuses on the importance of crosstalk between the gastrointestinal ecosystem and II/R to highlight II/R-induced gut microbiota signatures and potential applications of microbial-based therapies in II/R. This will also provide potentially effective biomarkers for the prediction, diagnosis and treatment of II/R.
Asunto(s)
Microbioma Gastrointestinal , Daño por Reperfusión , Disbiosis/complicaciones , Ecosistema , Humanos , Isquemia/complicaciones , Daño por Reperfusión/complicaciones , Daño por Reperfusión/tratamiento farmacológicoRESUMEN
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), the virus driving the ongoing coronavirus disease 2019 (COVID-19) pandemic, continues to rapidly evolve. Because of the limited efficacy of vaccination in prevention of SARS-CoV-2 transmission and continuous emergence of variants of concern (VOCs), orally bioavailable and broadly efficacious antiviral drugs are urgently needed. Previously, we showed that the parent nucleoside of remdesivir, GS-441524, has potent anti-SARS-CoV-2 activity. Here, we report that esterification of the 5'-hydroxyl moieties of GS-441524 markedly improved antiviral potency. This 5'-hydroxyl-isobutyryl prodrug, ATV006, demonstrated excellent oral bioavailability in rats and cynomolgus monkeys and exhibited potent antiviral efficacy against different SARS-CoV-2 VOCs in vitro and in three mouse models. Oral administration of ATV006 reduced viral loads and alleviated lung damage when administered prophylactically and therapeutically to K18-hACE2 mice challenged with the Delta variant of SARS-CoV-2. These data indicate that ATV006 represents a promising oral antiviral drug candidate for SARS-CoV-2.
Asunto(s)
Tratamiento Farmacológico de COVID-19 , Profármacos , Adenosina/uso terapéutico , Adenosina Monofosfato/análogos & derivados , Animales , Antivirales/farmacología , Antivirales/uso terapéutico , Ratones , Profármacos/farmacología , Profármacos/uso terapéutico , Ratas , SARS-CoV-2RESUMEN
HCoV-OC43 is one of the mildly pathogenic coronaviruses with high infection rates in common population. Here, 43 HCoV-OC43 related cases with pneumonia were reported, corresponding genomes of HCoV-OC43 were obtained. Phylogenetic analyses based on complete genome, orf1ab and spike genes revealed that two novel genotypes of HCoV-OC43 have emerged in China. Obvious recombinant events also can be detected in the analysis of the evolutionary dynamics of novel HCoV-OC43 genotypes. Estimated divergence time analysis indicated that the two novel genotypes had apparently independent evolutionary routes. Efforts should be conducted for further investigation of genomic diversity and evolution analysis of mildly pathogenic coronaviruses.
Asunto(s)
Resfriado Común/epidemiología , Infecciones por Coronavirus/epidemiología , Coronavirus Humano OC43/genética , Genoma Viral , Genotipo , Neumonía Viral/epidemiología , Secuencia de Bases , Teorema de Bayes , Niño , Niño Hospitalizado , Preescolar , China/epidemiología , Resfriado Común/patología , Resfriado Común/transmisión , Resfriado Común/virología , Infecciones por Coronavirus/patología , Infecciones por Coronavirus/transmisión , Infecciones por Coronavirus/virología , Coronavirus Humano OC43/clasificación , Coronavirus Humano OC43/patogenicidad , Monitoreo Epidemiológico , Femenino , Humanos , Lactante , Masculino , Método de Montecarlo , Mutación , Filogenia , Neumonía Viral/patología , Neumonía Viral/transmisión , Neumonía Viral/virología , Recombinación GenéticaRESUMEN
This article considers the connectivity preservation coordinated control problem for multiple spacecraft systems subject to limited communication resources and sensing capability. By constructing a novel bump function, a distance-dependent dynamic communication network model is first presented, which characterizes the interaction strength as a nonlinear smooth function varying with the relative distance of spacecraft continuously. Subsequently, based on an edge-tension potential function, a distributed event-driven coordinated control scheme is proposed to achieve formation consensus, while ensuring that adjacent spacecraft is always within the allowable connectivity range. Meanwhile, to avoid redundant data transmissions, a hybrid dynamic event-triggered mechanism with maximum triggering interval is developed to schedule the communication frequency among spacecraft. It is proven that the onboard communication resources occupation can be reduced significantly and the Zeno phenomenon is strictly excluded. Finally, the efficiency of the proposed method for, as an example, four-spacecraft formation system is substantiated.
RESUMEN
The current COVID-19 pandemic, caused by SARS-CoV-2, poses a serious public health threat. Effective therapeutic and prophylactic treatments are urgently needed. Angiotensin-converting enzyme 2 (ACE2) is a functional receptor for SARS-CoV-2, which binds to the receptor binding domain (RBD) of SARS-CoV-2 spike protein. Here, we developed recombinant human ACE2-Fc fusion protein (hACE2-Fc) and a hACE2-Fc mutant with reduced catalytic activity. hACE2-Fc and the hACE2-Fc mutant both efficiently blocked entry of SARS-CoV-2, SARS-CoV, and HCoV-NL63 into hACE2-expressing cells and inhibited SARS-CoV-2 S protein-mediated cell-cell fusion. hACE2-Fc also neutralized various SARS-CoV-2 strains with enhanced infectivity including D614G and V367F mutations, as well as the emerging SARS-CoV-2 variants, B.1.1.7 (Alpha), B.1.351 (Beta), B.1.617.1 (Kappa), and B.1.617.2 (Delta), demonstrating its potent and broad-spectrum antiviral effects. In addition, hACE2-Fc proteins protected HBE from SARS-CoV-2 infection. Unlike RBD-targeting neutralizing antibodies, hACE2-Fc treatment did not induce the development of escape mutants. Furthermore, both prophylactic and therapeutic hACE2-Fc treatments effectively protected mice from SARS-CoV-2 infection, as determined by reduced viral replication, weight loss, histological changes, and inflammation in the lungs. The protection provided by hACE2 showed obvious dose-dependent efficacy in vivo. Pharmacokinetic data indicated that hACE2-Fc has a relative long half-life in vivo compared to soluble ACE2, which makes it an excellent candidate for prophylaxis and therapy for COVID-19 as well as for SARS-CoV and HCoV-NL63 infections.
RESUMEN
Virus-specific T cells play essential roles in protection against multiple virus infections, including SARS-CoV and MERS-CoV. While SARS-CoV-2-specific T cells have been identified in COVID-19 patients, their role in the protection of SARS-CoV-2-infected mice is not established. Here, using mice sensitized for infection with SARS-CoV-2 by transduction with an adenovirus expressing the human receptor (Ad5-hACE2), we identified SARS-CoV-2-specific T cell epitopes recognized by CD4+ and CD8+ T cells in BALB/c and C57BL/6 mice. Virus-specific T cells were polyfunctional and were able to lyse target cells in vivo. Further, type I interferon pathway was proved to be critical for generating optimal antiviral T cell responses after SARS-CoV-2 infection. T cell vaccination alone partially protected SARS-CoV-2-infected mice from severe disease. In addition, the results demonstrated cross-reactive T cell responses between SARS-CoV and SARS-CoV-2, but not MERS-CoV, in mice. Understanding the role of the T cell response will guide immunopathogenesis studies of COVID-19 and vaccine design and validation.
Asunto(s)
COVID-19/inmunología , Epítopos de Linfocito T/inmunología , Interacciones Huésped-Patógeno/fisiología , Linfocitos T/inmunología , Linfocitos T/virología , Enzima Convertidora de Angiotensina 2/genética , Enzima Convertidora de Angiotensina 2/metabolismo , Animales , Anticuerpos Neutralizantes/sangre , Linfocitos T CD4-Positivos/virología , Linfocitos T CD8-positivos/virología , Chlorocebus aethiops , Reacciones Cruzadas , Mapeo Epitopo , Interferón Tipo I/inmunología , Interferón Tipo I/metabolismo , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Coronavirus del Síndrome Respiratorio de Oriente Medio/inmunología , Coronavirus Relacionado al Síndrome Respiratorio Agudo Severo/inmunología , SARS-CoV-2/inmunología , SARS-CoV-2/patogenicidad , Células VeroRESUMEN
BACKGROUND: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) has extensively and rapidly spread in the world, causing an outbreak of acute infectious pneumonia. However, no specific antiviral drugs or vaccines can be used. Phillyrin (KD-1), a representative ingredient of Forsythia suspensa, possesses anti-inflammatory, anti-oxidant, and antiviral activities. However, little is known about the antiviral abilities and mechanism of KD-1 against SARS-CoV-2 and human coronavirus 229E (HCoV-229E). PURPOSE: The study was designed to investigate the antiviral and anti-inflammatory activities of KD-1 against the novel SARS-CoV-2 and HCoV-229E and its potential effect in regulating host immune response in vitro. METHODS: The antiviral activities of KD-1 against SARS-CoV-2 and HCoV-229E were assessed in Vero E6 cells using cytopathic effect and plaque-reduction assay. Proinflammatory cytokine expression levels upon infection with SARS-CoV-2 and HCoV-229E infection in Huh-7 cells were measured by real-time quantitative PCR assays. Western blot assay was used to determine the protein expression of nuclear factor kappa B (NF-κB) p65, p-NF-κB p65, IκBα, and p-IκBα in Huh-7 cells, which are the key targets of the NF-κB pathway. RESULTS: KD-1 could significantly inhibit SARS-CoV-2 and HCoV-229E replication in vitro. KD-1 could also markedly reduce the production of proinflammatory cytokines (TNF-α, IL-6, IL-1ß, MCP-1, and IP-10) at the mRNA levels. Moreover, KD-1 could significantly reduce the protein expression of p-NF-κB p65, NF-κB p65, and p-IκBα, while increasing the expression of IκBα in Huh-7 cells. CONCLUSIONS: KD-1 could significantly inhibit virus proliferation in vitro, the up-regulated expression of proinflammatory cytokines induced by SARS-CoV-2 and HCoV-229E by regulating the activity of the NF-кB signaling pathway. Our findings indicated that KD-1 protected against virus attack and can thus be used as a novel strategy for controlling the coronavirus disease 2019.
Asunto(s)
Antiinflamatorios/farmacología , Antivirales/farmacología , Betacoronavirus/efectos de los fármacos , Coronavirus Humano 229E/efectos de los fármacos , Infecciones por Coronavirus , Glucósidos/farmacología , FN-kappa B/metabolismo , Pandemias , Neumonía Viral , Animales , COVID-19 , Chlorocebus aethiops , Coronavirus/efectos de los fármacos , Infecciones por Coronavirus/metabolismo , Infecciones por Coronavirus/virología , Citocinas/metabolismo , Forsythia/química , Humanos , Fitoterapia , Extractos Vegetales/farmacología , Neumonía Viral/metabolismo , Neumonía Viral/virología , SARS-CoV-2 , Síndrome Respiratorio Agudo Grave/virología , Transducción de Señal/efectos de los fármacos , Células Vero , Replicación Viral/efectos de los fármacosRESUMEN
COVID-19, caused by SARS-CoV-2, is a virulent pneumonia, with >4,000,000 confirmed cases worldwide and >290,000 deaths as of May 15, 2020. It is critical that vaccines and therapeutics be developed very rapidly. Mice, the ideal animal for assessing such interventions, are resistant to SARS-CoV-2. Here, we overcome this difficulty by exogenous delivery of human ACE2 with a replication-deficient adenovirus (Ad5-hACE2). Ad5-hACE2-sensitized mice developed pneumonia characterized by weight loss, severe pulmonary pathology, and high-titer virus replication in lungs. Type I interferon, T cells, and, most importantly, signal transducer and activator of transcription 1 (STAT1) are critical for virus clearance and disease resolution in these mice. Ad5-hACE2-transduced mice enabled rapid assessments of a vaccine candidate, of human convalescent plasma, and of two antiviral therapies (poly I:C and remdesivir). In summary, we describe a murine model of broad and immediate utility to investigate COVID-19 pathogenesis and to evaluate new therapies and vaccines.
Asunto(s)
Betacoronavirus/inmunología , Infecciones por Coronavirus/patología , Infecciones por Coronavirus/prevención & control , Modelos Animales de Enfermedad , Pandemias/prevención & control , Neumonía Viral/patología , Neumonía Viral/prevención & control , Vacunación , Enzima Convertidora de Angiotensina 2 , Animales , COVID-19 , Chlorocebus aethiops , Infecciones por Coronavirus/virología , Evaluación Preclínica de Medicamentos/métodos , Femenino , Humanos , Interferón gamma/genética , Interferón gamma/metabolismo , Pulmón/patología , Pulmón/virología , Masculino , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Noqueados , Peptidil-Dipeptidasa A/genética , Peptidil-Dipeptidasa A/metabolismo , Neumonía Viral/virología , Receptor de Interferón alfa y beta/genética , Receptor de Interferón alfa y beta/metabolismo , SARS-CoV-2 , Factor de Transcripción STAT1/genética , Factor de Transcripción STAT1/metabolismo , Organismos Libres de Patógenos Específicos , Transducción Genética , Células Vero , Carga Viral , Replicación ViralRESUMEN
Severe Acute Respiratory Syndrome Coronavirus 2 (SARS-CoV-2) has spread worldwide through person-to-person contact, causing a public health emergency of international concern. At present, there is no specific antiviral treatment recommended for SARS-CoV-2 infection. Liu Shen capsule (LS), a traditional Chinese medicine, has been proven to have a wide spectrum of pharmacological properties, such as anti-inflammatory, antiviral and immunomodulatory activities. However, little is known about the antiviral effect of LS against SARS-CoV-2. Herein, the study was designed to investigate the antiviral activity of SARS-CoV-2 and its potential effect in regulating the host's immune response. The inhibitory effect of LS against SARS-CoV-2 replication in Vero E6 cells was evaluated by using the cytopathic effect (CPE) and plaque reduction assay. The number of virions of SARS-CoV-2 was observed under transmission electron microscope after treatment with LS. Proinflammatory cytokine expression levels upon SARS-CoV-2 infection in Huh-7 cells were measured by real-time quantitative PCR assays. The results showed that LS could significantly inhibit SARS-CoV-2 replication in Vero E6 cells, and reduce the number of virus particles and it could markedly reduce pro-inflammatory cytokines (TNF-α, IL-6, IL-1ß, IL-8, CCL-2/MCP-1 and CXCL-10/IP-10) production at the mRNA levels. Moreover, the expression of the key proteins in the NF-κB/MAPK signaling pathway was detected by western blot and it was found that LS could inhibit the expression of p-NF-κB p65, p-IκBα and p-p38 MAPK, while increasing the expression of IκBα. These findings indicate that LS could inhibit SARS-CoV-2 virus infection via downregulating the expression of inflammatory cytokines induced virus and regulating the activity of NF-κB/MAPK signaling pathway in vitro, making its promising candidate treatment for controlling COVID-19 disease.
Asunto(s)
Betacoronavirus/efectos de los fármacos , Mezclas Complejas/farmacología , FN-kappa B/antagonistas & inhibidores , FN-kappa B/metabolismo , Transducción de Señal/efectos de los fármacos , Adenosina Monofosfato/análogos & derivados , Adenosina Monofosfato/farmacología , Alanina/análogos & derivados , Alanina/farmacología , Animales , Antiinflamatorios/farmacología , Antivirales/farmacología , COVID-19 , Proliferación Celular/efectos de los fármacos , Células Cultivadas , Chlorocebus aethiops , Infecciones por Coronavirus/virología , Humanos , Mediadores de Inflamación/metabolismo , Pandemias , Neumonía Viral/virología , SARS-CoV-2 , Virión/efectos de los fármacosRESUMEN
Middle East respiratory syndrome coronavirus (MERS-CoV) causes severe acute respiratory disease in humans. MERS-CoV strains from early epidemic clade A and contemporary epidemic clade B have not been phenotypically characterized to compare their abilities to infect cells and mice. We isolated the clade B MERS-CoV ChinaGD01 strain from a patient infected during the South Korean MERS outbreak in 2015 and compared the phylogenetics and pathogenicity of MERS-CoV EMC/2012 (clade A) and ChinaGD01 (clade B) in vitro and in vivo Genome alignment analysis showed that most clade-specific mutations occurred in the orf1ab gene, including mutations that were predicted to be potential glycosylation sites. Minor differences in viral growth but no significant differences in plaque size or sensitivity to beta interferon (IFN-ß) were detected between these two viruses in vitro ChinaGD01 virus infection induced more weight loss and inflammatory cytokine production in human DPP4-transduced mice. Viral titers were higher in the lungs of ChinaGD01-infected mice than with EMC/2012 infection. Decreased virus-specific CD4+ and CD8+ T cell numbers were detected in the lungs of ChinaGD01-infected mice. In conclusion, MERS-CoV evolution induced changes to reshape its pathogenicity and virulence in vitro and in vivo and to evade adaptive immune response to hinder viral clearance.IMPORTANCE MERS-CoV is an important emerging pathogen and causes severe respiratory infection in humans. MERS-CoV strains from early epidemic clade A and contemporary epidemic clade B have not been phenotypically characterized to compare their abilities to infect cells and mice. In this study, we showed that a clade B virus ChinaGD01 strain caused more severe disease in mice, with delayed viral clearance, increased inflammatory cytokines, and decreased antiviral T cell responses, than the early clade A virus EMC/2012. Given the differences in pathogenicity of different clades of MERS-CoV, periodic assessment of currently circulating MERS-CoV is needed to monitor potential severity of zoonotic disease.
Asunto(s)
Infecciones por Coronavirus/virología , Genotipo , Interacciones Huésped-Patógeno , Coronavirus del Síndrome Respiratorio de Oriente Medio/fisiología , Adulto , Animales , Modelos Animales de Enfermedad , Genoma Viral , Interacciones Huésped-Patógeno/inmunología , Humanos , Interferón Tipo I/farmacología , Masculino , Ratones , Coronavirus del Síndrome Respiratorio de Oriente Medio/clasificación , Coronavirus del Síndrome Respiratorio de Oriente Medio/aislamiento & purificación , Coronavirus del Síndrome Respiratorio de Oriente Medio/patogenicidad , Filogenia , ARN Viral , Linfocitos T/inmunología , Linfocitos T/metabolismo , Virulencia , Replicación Viral/efectos de los fármacos , Replicación Viral/genética , Secuenciación Completa del GenomaRESUMEN
SARS-CoV-2 caused a major outbreak of severe pneumonia (COVID-19) in humans. Viral RNA was detected in multiple organs in COVID-19 patients. However, infectious SARS-CoV-2 was only isolated from respiratory specimens. Here, infectious SARS-CoV-2 was successfully isolated from urine of a COVID-19 patient. The virus isolated could infect new susceptible cells and was recognized by its' own patient sera. Appropriate precautions should be taken to avoid transmission from urine.