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Urgent research into innovative severe acute respiratory coronavirus-2 (SARS-CoV-2) vaccines that may successfully prevent various emerging emerged variants, particularly the Omicron variant and its subvariants, is necessary. Here, we designed a chimeric adenovirus-vectored vaccine named Ad5-Beta/Delta. This vaccine was created by incorporating the receptor-binding domain from the Delta variant, which has the L452R and T478K mutations, into the complete spike protein of the Beta variant. Both intramuscular (IM) and intranasal (IN) vaccination with Ad5-Beta/Deta vaccine induced robust broad-spectrum neutralization against Omicron BA.5-included variants. IN immunization with Ad5-Beta/Delta vaccine exhibited superior mucosal immunity, manifested by higher secretory IgA antibodies and more tissue-resident memory T cells (TRM) in respiratory tract. The combination of IM and IN delivery of the Ad5-Beta/Delta vaccine was capable of synergically eliciting stronger systemic and mucosal immune responses. Furthermore, the Ad5-Beta/Delta vaccination demonstrated more effective boosting implications after two dosages of mRNA or subunit recombinant protein vaccine, indicating its capacity for utilization as a booster shot in the heterologous vaccination. These outcomes quantified Ad5-Beta/Delta vaccine as a favorable vaccine can provide protective immunity versus SARS-CoV-2 pre-Omicron variants of concern and BA.5-included Omicron subvariants.
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Neutrophil is a pathophysiological character in Alzheimer's disease. The pathogen for neutrophil activation in cerebral tissue is the accumulated amyloid protein. In our present study, neutrophils infiltrate into the cerebra in two models (transgenic model APP/PS1 and stereotactic injection model) and promote neuron apoptosis, releasing their cellular constituents, including mitochondria and mitochondrial DNA (mtDNA). We found that both Aß1-42 and mtDNA could provoke neutrophil infiltration into the cerebra, and they had synergistic effects when they presented together. This neutrophillic neuroinflammation upregulates expressions of STING, NLRP3 and IL-1ß. These inflammatory cytokines with mtDNA constitute the mtDNA-STING-NLRP3/IL-1ß axis, which is the prerequisite for neutrophil infiltration. When any factor in this pathway is depleted, the migration of neutrophils into cerebral tissue is ceased, with neurons and cognitive function being protected. Thus, we provide a novel perspective to alleviate the progression of Alzheimer's disease.
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Enfermedad de Alzheimer , Animales , Ratones , Enfermedad de Alzheimer/metabolismo , Proteína con Dominio Pirina 3 de la Familia NLR/metabolismo , Péptidos beta-Amiloides/metabolismo , Inflamasomas/metabolismo , ADN Mitocondrial/metabolismo , Infiltración Neutrófila , Mitocondrias/metabolismo , Neuronas/metabolismo , Ratones TransgénicosRESUMEN
Molecular target inhibitors have been regularly approved by Food and Drug Administration (FDA) for tumor treatment, and most of them intervene in tumor cell proliferation and metabolism. The RAS-RAF-MEK-ERK pathway is a conserved signaling pathway that plays vital roles in cell proliferation, survival, and differentiation. The aberrant activation of the RAS-RAF-MEK-ERK signaling pathway induces tumors. About 33% of tumors harbor RAS mutations, while 8% of tumors are driven by RAF mutations. Great efforts have been dedicated to targeting the signaling pathway for cancer treatment in the past decades. In this review, we summarized the development of inhibitors targeting the RAS-RAF-MEK-ERK pathway with an emphasis on those used in clinical treatment. Moreover, we discussed the potential combinations of inhibitors that target the RAS-RAF-MEK-ERK signaling pathway and other signaling pathways. The inhibitors targeting the RAS-RAF-MEK-ERK pathway have essentially modified the therapeutic strategy against various cancers and deserve more attention in the current cancer research and treatment.
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BACKGROUND: Mitochondrial transcription factor A (TFAM) is a transcription factor that maintains mitochondrial DNA (mtDNA) stabilization and initiates mtDNA replication. However, little is known about the immune regulation function and TFAM expression in immune cells in the tumors. METHODS: Mouse tumor models were applied to analyze the effect of TFAM deficiency in myeloid cell lineage on tumor progression and tumor microenvironment (TME) modification. In vitro, primary mouse bone marrow-derived dendritic cells (BMDCs) were used in the investigation of the altered function and the activated pathway. OVA was used as the model antigen to validate the activation of immune responses in vivo. STING inhibitors were used to confirm the STING activation provoked by Tfam deficient in DCs. RESULTS: The deletion of TFAM in DCs led to mitochondrial dysfunction and mtDNA cytosolic leakage resulting in the cGAS-STING pathway activation in DCs, which contributed to the enhanced antigen presentation. The deletion of TFAM in DCs has interestingly reversed the immune suppressive TME and inhibited tumor growth and metastasis in tumor models. CONCLUSIONS: We have revealed that TFAM knockout in DCs ameliorated immune-suppressive microenvironment in tumors through STING pathway. Our work suggests that specific TFAM knockout in DCs might be a compelling strategy for designing novel immunotherapy methods in the future.
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Proteínas de Unión al ADN , Células Dendríticas , Proteínas del Grupo de Alta Movilidad , Mitocondrias , Neoplasias , Animales , Ratones , Presentación de Antígeno , Modelos Animales de Enfermedad , ADN Mitocondrial , Proteínas de Unión al ADN/genética , Proteínas del Grupo de Alta Movilidad/genética , Mitocondrias/patología , Neoplasias/patologíaRESUMEN
PRMT6 is a member of the protein arginine methyltransferase family, which is involved in a variety of physiological processes and plays an important role in the occurrence and development of tumors. Due to the high homology of type â PRMTs and the two close binding sites of the SAM pocket and the substrate pocket, selective PRMT6 inhibitors have rarely been reported. In this study, a series of (5-phenylpyridin-3-yl)methanamine derivatives were designed and synthesized, which could form hydrogen bonding interactions with the unique Glu49 of PRMT6, thereby improving the selectivity of the compounds for PRMT6. Among them, a25 had the best activity and selectivity, with more than 25-fold selectivity for PRMT1/8 and more than 50-fold selectivity for PRMT3/4/5/7, which was superior to these reported SAM competitive and substrate competitive PRMT6 inhibitors. Importantly, a25 could significantly inhibit the proliferation of various tumor cells and effectively induce apoptosis of cancer cells. Our data clarified that a25 is a promising selective PRMT6 inhibitor for cancer therapy which is worthy of further evaluation.
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Neoplasias , Proteínas Nucleares , Humanos , Proteínas Nucleares/metabolismo , Metilación , Proteína-Arginina N-Metiltransferasas , Proteínas Represoras/metabolismoRESUMEN
For patients with esophageal squamous cell carcinoma (ESCC), standard therapeutic methods (cisplatin and radiotherapy) have been found to be ineffective and severely toxic. Targeted therapy emerges as a promising solution for this dilemma. It has been reported that targeted therapies are applied alone or in combination with standard conventional therapies for the treatment of a variety of cancers. To the best of our knowledge, in patients with ESCC, the combinational methods containing standard therapy and ERK-targeted therapy have yet to be explored. To analyze the prognostic role of p-ERK in ESCC patients, the Kaplan-Meier analysis and Cox regression model were used. To assess the effects of ERK-targeted therapy (GDC0994) on ESCC cells, in vitro studies including CCK-8 assay, colony formation assay, and scratch wound healing assay were conducted. In addition, the changes in cell cycle distribution and apoptosis were analyzed by flow cytometry. Besides, to assess the efficacy of different therapies in vivo, the xenograft tumor models were established by subcutaneously inoculating tumor cells into the flank/leg of mice. In patients with ESCC, a strong correlation between the high expression level of p-ERK and the poor prognosis (p < 0.01, Log-Rank test) has been identified. By analyzing the results from CCK-8 and scratch wound healing assays, we demonstrated that the ERK inhibitor repressed the viability and migration of ESCC cells. In addition, following the treatment of GDC0994, the volumes of xenograft tumors significantly decreased (p < 0.001, one-way ANOVA). Furthermore, blocking the mitogen-activated protein kinase (MAPK/ERK) pathway enhanced the therapeutic efficacy of both cisplatin and radiotherapy (p < 0.05). These findings imply the role of p-ERK in the prognosis of ESCC patients and the therapeutic value of ERK inhibitors in ESCC.
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Carcinoma de Células Escamosas , Neoplasias Esofágicas , Carcinoma de Células Escamosas de Esófago , Animales , Humanos , Ratones , Apoptosis , Carcinoma de Células Escamosas/tratamiento farmacológico , Carcinoma de Células Escamosas/genética , Carcinoma de Células Escamosas/radioterapia , Línea Celular Tumoral , Proliferación Celular , Quimioradioterapia/métodos , Cisplatino , Neoplasias Esofágicas/tratamiento farmacológico , Neoplasias Esofágicas/genética , Neoplasias Esofágicas/radioterapia , Carcinoma de Células Escamosas de Esófago/tratamiento farmacológico , Carcinoma de Células Escamosas de Esófago/radioterapia , Quinasas MAP Reguladas por Señal Extracelular/antagonistas & inhibidores , Inhibidores de Proteínas Quinasas/farmacologíaRESUMEN
For coronavirus disease 2019 (COVID-19), caused by severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), 15-30% of patients are likely to develop COVID-19-related acute respiratory distress syndrome (ARDS). There are still few effective and well-understood therapies available. Novel variants and short-lasting immunity are posing challenges to vaccine efficacy, so finding antiviral and antiinflammatory treatments remains crucial. Here, tripterin (TP), a traditional Chinese medicine, was encapsulated into liposome (TP lipo) to investigate its antiviral and antiinflammatory effects in severe COVID-19. By using two severe COVID-19 models in human ACE2-transgenic (hACE2) mice, an analysis of TP lipo's effects on pulmonary immune responses was conducted. Pulmonary pathological alterations and viral burden were reduced by TP lipo treatment. TP lipo inhibits SARS-CoV-2 replication and hyperinflammation in infected cells and mice, two crucial events in severe COVID-19 pathophysiology, it is a promising drug candidate to treat SARS-CoV-2-induced ARDS.
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COVID-19 , Síndrome de Dificultad Respiratoria , Humanos , Animales , Ratones , SARS-CoV-2 , Liposomas , Tratamiento Farmacológico de COVID-19 , Antivirales/farmacología , Síndrome de Dificultad Respiratoria/tratamiento farmacológicoRESUMEN
BACKGROUND: Emerging evidence provides mechanistic insights into the pathogenesis of pulmonary fibrosis (PF), and rare anti-PF therapeutic method has promising effect in its treatment. Rho-associated coiled-coil kinases (ROCK) inhibition significantly ameliorates bleomycin-induced PF and decreases macrophage infiltration, but the mechanism remains unclear. We established bleomycin and radiation-induced PF to identify the activity of WXWH0265, a newly designed unselective ROCK inhibitor in regulating macrophages. METHODS: Bleomycin-induced PF was induced by intratracheal instillation and radiation-induced PF was induced by bilateral thoracic irradiation. Histopathological techniques (haematoxylin and eosin, Masson's trichrome and immunohistochemistry) and hydroxyproline were used to evaluate PF severity. Western blot, quantitative real-time reverse transcription-polymerase chain reaction and flow cytometry were performed to explore the underlying mechanisms. Bone marrow-derived macrophages (BMDMs) were used to verify their therapeutic effect. Clodronate liposomes were applied to deplete macrophages and to identify the therapeutic effect of WXWH0265. RESULTS: Therapeutic administration of ROCK inhibitor ameliorates bleomycin-induced PF by inhibiting M2 macrophages polarisation. ROCK inhibitor showed no significant anti-fibrotic effect in macrophages-depleted mice. Treatment with WXWH0265 demonstrated superior protection effect in bleomycin-induced PF compared with positive drugs. In radiation-induced PF, ROCK inhibitor effectively ameliorated PF. Fibroblasts co-cultured with supernatant from various M2 macrophages phenotypes revealed that M2 macrophages stimulated by interleukin-4 promoted extracellular matrix production. Polarisation of M2 macrophages was inhibited by ROCK inhibitor treatment in vitro. The p-signal transducer and activator of transcription 3 (STAT3) in lung tissue and BMDMs was significantly decreased in PF in vivo and vitro after treated with ROCK inhibitors. CONCLUSION: Inhibiting ROCK could significantly attenuate bleomycin- and radiation-induced PF by regulating the macrophages polarisation via phosphorylation of STAT3. WXWH0265 is a kind of efficient unselective ROCK inhibitor in ameliorating PF. Furthermore, the results provide empirical evidence that ROCK inhibitor, WXWH0265 is a potential drug to prevent the development of PF.
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Fibrosis Pulmonar , Factor de Transcripción STAT3 , Quinasas Asociadas a rho , Animales , Bleomicina/efectos adversos , Ácido Clodrónico/metabolismo , Interleucina-4/metabolismo , Liposomas/metabolismo , Macrófagos/metabolismo , Ratones , Fosforilación , Fibrosis Pulmonar/inducido químicamente , Fibrosis Pulmonar/tratamiento farmacológico , Fibrosis Pulmonar/patología , Factor de Transcripción STAT3/metabolismo , Quinasas Asociadas a rho/antagonistas & inhibidoresRESUMEN
The recently emerged Omicron (B.1.1.529) variant has rapidly surpassed Delta to become the predominant circulating SARS-CoV-2 variant, given the higher transmissibility rate and immune escape ability, resulting in breakthrough infections in vaccinated individuals. A new generation of SARS-CoV-2 vaccines targeting the Omicron variant are urgently needed. Here, we developed a subunit vaccine named RBD-HR/trimer by directly linking the sequence of RBD derived from the Delta variant (containing L452R and T478K) and HR1 and HR2 in SARS-CoV-2 S2 subunit in a tandem manner, which can self-assemble into a trimer. In multiple animal models, vaccination of RBD-HR/trimer formulated with MF59-like oil-in-water adjuvant elicited sustained humoral immune response with high levels of broad-spectrum neutralizing antibodies against Omicron variants, also inducing a strong T cell immune response in vivo. In addition, our RBD-HR/trimer vaccine showed a strong boosting effect against Omicron variants after two doses of mRNA vaccines, featuring its capacity to be used in a prime-boost regimen. In mice and non-human primates, RBD-HR/trimer vaccination could confer a complete protection against live virus challenge of Omicron and Delta variants. The results qualified RBD-HR/trimer vaccine as a promising next-generation vaccine candidate for prevention of SARS-CoV-2, which deserved further evaluation in clinical trials.
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COVID-19 , Vacunas Virales , Animales , Anticuerpos Neutralizantes , COVID-19/prevención & control , Vacunas contra la COVID-19 , Humanos , Ratones , Ratones Endogámicos BALB C , Subunidades de Proteína , SARS-CoV-2 , Vacunas de Subunidad , AguaRESUMEN
Neutrophil extracellular traps (NETs) can capture and kill viruses, such as influenza viruses, human immunodeficiency virus (HIV), and respiratory syncytial virus (RSV), thus contributing to host defense. Contrary to our expectation, we show here that the histones released by NETosis enhance the infectivity of SARS-CoV-2, as found by using live SARS-CoV-2 and two pseudovirus systems as well as a mouse model. The histone H3 or H4 selectively binds to subunit 2 of the spike (S) protein, as shown by a biochemical binding assay, surface plasmon resonance and binding energy calculation as well as the construction of a mutant S protein by replacing four acidic amino acids. Sialic acid on the host cell surface is the key molecule to which histones bridge subunit 2 of the S protein. Moreover, histones enhance cell-cell fusion. Finally, treatment with an inhibitor of NETosis, histone H3 or H4, or sialic acid notably affected the levels of sgRNA copies and the number of apoptotic cells in a mouse model. These findings suggest that SARS-CoV-2 could hijack histones from neutrophil NETosis to promote its host cell attachment and entry process and may be important in exploring pathogenesis and possible strategies to develop new effective therapies for COVID-19.
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COVID-19 , SARS-CoV-2 , Animales , Histonas , Ratones , Ácido N-Acetilneuramínico , Subunidades de Proteína/metabolismo , Glicoproteína de la Espiga del Coronavirus/química , Internalización del VirusRESUMEN
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) is the etiology of coronavirus disease 2019 (COVID-19) pandemic. Current variants including Alpha, Beta, Gamma, Delta, and Lambda increase the capacity of infection and transmission of SARS-CoV-2, which might disable the in-used therapies and vaccines. The COVID-19 has now put an enormous strain on health care system all over the world. Therefore, the development of animal models that can capture characteristics and immune responses observed in COVID-19 patients is urgently needed. Appropriate models could accelerate the testing of therapeutic drugs and vaccines against SARS-CoV-2. In this review, we aim to summarize the current animal models for SARS-CoV-2 infection, including mice, hamsters, nonhuman primates, and ferrets, and discuss the details of transmission, pathology, and immunology induced by SARS-CoV-2 in these animal models. We hope this could throw light to the increased usefulness in fundamental studies of COVID-19 and the preclinical analysis of vaccines and therapeutic agents.
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The development of animal models for COVID-19 is essential for basic research and drug/vaccine screening. Previously reported COVID-19 animal models need to be established under a high biosafety level condition for the utilization of live SARS-CoV-2, which greatly limits its application in routine research. Here, we generate a mouse model of COVID-19 under a general laboratory condition that captures multiple characteristics of SARS-CoV-2-induced acute respiratory distress syndrome (ARDS) observed in humans. Briefly, human ACE2-transgenic (hACE2) mice were intratracheally instilled with the formaldehyde-inactivated SARS-CoV-2, resulting in a rapid weight loss and detrimental changes in lung structure and function. The pulmonary pathologic changes were characterized by diffuse alveolar damage with pulmonary consolidation, hemorrhage, necrotic debris, and hyaline membrane formation. The production of fatal cytokines (IL-1ß, TNF-α, and IL-6) and the infiltration of activated neutrophils, inflammatory monocyte-macrophages, and T cells in the lung were also determined, suggesting the activation of an adaptive immune response. Therapeutic strategies, such as dexamethasone or passive antibody therapy, could effectively ameliorate the disease progression in this model. Therefore, the established mouse model for SARS-CoV-2-induced ARDS in the current study may provide a robust tool for researchers in the standard open laboratory to investigate the pathological mechanisms or develop new therapeutic strategies for COVID-19 and ARDS.
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Enzima Convertidora de Angiotensina 2/inmunología , COVID-19/inmunología , Pulmón/inmunología , Síndrome de Dificultad Respiratoria/inmunología , SARS-CoV-2/inmunología , Enzima Convertidora de Angiotensina 2/genética , Animales , COVID-19/genética , Modelos Animales de Enfermedad , Humanos , Ratones , Ratones Transgénicos , Síndrome de Dificultad Respiratoria/genéticaRESUMEN
Mesenchymal stem cell (MSC)-mediated immunomodulation has been harnessed for the treatment of human diseases, but its underlying mechanism has not been fully understood. Dead cells, including apoptotic cells have immunomodulatory properties. It has been repeatedly reported that the proportion of nonviable MSCs in a MSC therapeutic preparation varied from 5~50% in the ongoing clinical trials. It is conceivable that the nonviable cells in a MSC therapeutic preparation may play a role in the therapeutic effects of MSCs. We found that the MSC therapeutic preparation in the present study had about 5% dead MSCs (DMSCs), characterized by apoptotic cells. Namely, 1 × 106 MSCs in the preparation contained about 5 × 104 DMSCs. We found that the treatment with even 5 × 104 DMSCs alone had the equal therapeutic effects as with 1 × 106 MSCs. This protective effect of the dead MSCs alone was confirmed in four mouse models, including concanavalin A (ConA)- and carbon tetrachloride (CCl4)-induced acute liver injury, LPS-induced lung injury and spinal cord injury. We also found that the infused MSCs died by apoptosis in vivo. Furthermore, the therapeutic effect was attributed to the elevated level of phosphatidylserine (PS) upon the injection of MSCs or DMSCs. The direct administration of PS liposomes (PSLs) mimic apoptotic cell fragments also exerted the protective effects as MSCs and DMSCs. The Mer tyrosine kinase (MerTK) deficiency or the knockout of chemokine receptor C-C motif chemokine receptor 2 (CCR2) reversed these protective effects of MSCs or DMSCs. These results revealed that DMSCs alone in the therapeutic stem cell preparation or the apoptotic cells induced in vivo may exert the same immunomodulatory property as the "living MSCs preparation" through releasing PS, which was further recognized by MerTK and participated in modulating immune cells.
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Apoptosis , Intoxicación por Tetracloruro de Carbono , Trasplante de Células Madre Mesenquimatosas , Células Madre Mesenquimatosas/metabolismo , Fosfatidilserinas/metabolismo , Animales , Intoxicación por Tetracloruro de Carbono/metabolismo , Intoxicación por Tetracloruro de Carbono/terapia , Femenino , Ratones , Ratones NoqueadosAsunto(s)
COVID-19 , Modelos Animales de Enfermedad , Síndrome de Dificultad Respiratoria , SARS-CoV-2 , Animales , COVID-19/genética , COVID-19/metabolismo , COVID-19/patología , Humanos , Ratones , Ratones Transgénicos , Síndrome de Dificultad Respiratoria/genética , Síndrome de Dificultad Respiratoria/metabolismo , Síndrome de Dificultad Respiratoria/patología , Síndrome de Dificultad Respiratoria/virología , SARS-CoV-2/genética , SARS-CoV-2/metabolismoRESUMEN
Owing to the excellent antibacterial and antiviral activity, silver nanoparticles have a widespread use in the food and pharmaceutical industries. With the increase in the production and use of the related products, the potential hazard of silver nanoparticles has aroused public attention. The main purpose of this study is to explore the toxicity of silver nanoparticles and induction of lung inflammation in vitro and in vivo. Here, we validated that small amounts of silver ions dissolved from silver nanoparticles caused the depolarization of plasma membrane, resulting in an overload of intracellular sodium and calcium, and eventually led to the cell necrosis. The blockers of calcium or sodium channels inversed the toxicity of silver ions. Then, we instilled silver nanoparticles or silver nitrate (50 µg per mouse) into the lungs of mice, and this induced pulmonary injury and mitochondrial content release, led to the recruitment of neutrophils to the lung tissue via p38 MAPK pathway. Altogether, these data show that released silver ions from nanoparticles induced cell necrosis through Na+ and Ca2+ influx and triggered pulmonary inflammation through elevating mitochondrial-related contents released from these necrotic cells.
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Nanopartículas del Metal/efectos adversos , Mitocondrias/metabolismo , Neumonía/patología , Plata/efectos adversos , Células A549 , Animales , Calcio/metabolismo , ADN Mitocondrial/metabolismo , Proteínas Activadoras de GTPasa/metabolismo , Humanos , Iones , Nanopartículas del Metal/ultraestructura , Ratones Endogámicos C57BL , Mitocondrias/efectos de los fármacos , N-Formilmetionina Leucil-Fenilalanina/farmacología , Necroptosis/efectos de los fármacos , Necrosis , Infiltración Neutrófila/efectos de los fármacos , Proteínas Quinasas/metabolismo , Sodio/metabolismoRESUMEN
Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) causes a respiratory disease called coronavirus disease 2019 (COVID-19), the spread of which has led to a pandemic. An effective preventive vaccine against this virus is urgently needed. As an essential step during infection, SARS-CoV-2 uses the receptor-binding domain (RBD) of the spike protein to engage with the receptor angiotensin-converting enzyme 2 (ACE2) on host cells1,2. Here we show that a recombinant vaccine that comprises residues 319-545 of the RBD of the spike protein induces a potent functional antibody response in immunized mice, rabbits and non-human primates (Macaca mulatta) as early as 7 or 14 days after the injection of a single vaccine dose. The sera from the immunized animals blocked the binding of the RBD to ACE2, which is expressed on the cell surface, and neutralized infection with a SARS-CoV-2 pseudovirus and live SARS-CoV-2 in vitro. Notably, vaccination also provided protection in non-human primates to an in vivo challenge with SARS-CoV-2. We found increased levels of RBD-specific antibodies in the sera of patients with COVID-19. We show that several immune pathways and CD4 T lymphocytes are involved in the induction of the vaccine antibody response. Our findings highlight the importance of the RBD domain in the design of SARS-CoV-2 vaccines and provide a rationale for the development of a protective vaccine through the induction of antibodies against the RBD domain.
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Anticuerpos Antivirales/inmunología , Betacoronavirus/inmunología , Infecciones por Coronavirus/inmunología , Infecciones por Coronavirus/prevención & control , Pandemias/prevención & control , Neumonía Viral/inmunología , Neumonía Viral/prevención & control , Glicoproteína de la Espiga del Coronavirus/química , Glicoproteína de la Espiga del Coronavirus/inmunología , Vacunas Virales/inmunología , Animales , Anticuerpos Neutralizantes/inmunología , COVID-19 , Vacunas contra la COVID-19 , Humanos , Macaca mulatta/inmunología , Macaca mulatta/virología , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Modelos Animales , Modelos Moleculares , Dominios Proteicos , SARS-CoV-2 , Suero/inmunología , Bazo/citología , Bazo/inmunología , Linfocitos T/inmunología , VacunaciónRESUMEN
Rationale: The adverse health effects of nano-particulate pollutants have attracted much attention in recent years. Carbon nanomaterials are recognized as risk factors for prolonged inflammatory responses and diffuse alveolar injury. Previous research indicated a central role of alveolar macrophages in the pathogenesis of particle-related lung disease, but the underlying mechanism remains largely unknown. Methods: C57BL/6 mice were intratracheally instilled with carbon black nanoparticles (CBNPs). Cell necrosis and the infiltrated neutrophils in the lungs were detected by flow cytometry. Release of mitochondria was observed with Mito Tracker and mitochondrial DNA (mtDNA) was quantified by qPCR via Taqman probes. TLR9-p38 MAPK signaling pathway was detected by Western blotting. The production of lipid chemoattractant leukotriene B4 (LTB4) in the supernatant and bronchoalveolar lavage fluid (BALF) was quantitated using an enzyme immunoassay (EIA). Results: In the present study, we found that a single instillation of CBNPs induced neutrophil influx in C57BL/6 mice as early as 4 h post-exposure following the rapid appearance of cell damage indicators in BALF at 30 min. Macrophages exposed to CBNPs showed necrotic features and were characterized by lysosome rupture, cathepsin B release, reactive oxygen species generation, and reduced intracellular ATP level. Necrosis was partly inhibited by a specific lysosomal cathepsin B inhibitor CA074 Me. Further analyses suggested that the resulting leakage of mtDNA from the necrotic cells activated neutrophils and triggered severe inflammation in vivo. Pulmonary neutrophilic inflammation induced by mtDNA was reduced in TLR9-/- mice. Additionally, mtDNA induced LTB4 production from macrophages, which may contribute to neutrophil recruitment. Conclusion: We demonstrated here that CBNPs induce acute cell necrosis through lysosomal rupture and that mtDNA released from necrotic cells functions as a key event mediating pulmonary neutrophilic inflammation. This study described a novel aspect of the pathogenesis of particle-induced inflammatory response and provided a possible therapeutic target for the regulation of inflammation.
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Pulmón , Macrófagos/efectos de los fármacos , Nanopartículas/efectos adversos , Neutrófilos/efectos de los fármacos , Neumonía/inducido químicamente , Hollín/efectos adversos , Animales , Células Cultivadas , Femenino , Pulmón/efectos de los fármacos , Pulmón/patología , Lisosomas/efectos de los fármacos , Macrófagos/patología , Ratones , Ratones Endogámicos C57BL , Necrosis/inducido químicamente , Neutrófilos/patologíaRESUMEN
[This corrects the article DOI: 10.1371/journal.pone.0176823.].