RESUMO
Adeno-associated virus (AAV)-mediated gene delivery holds great promise for gene therapy. However, the non-invasive delivery of AAV for lung tissues has not been adequately established. Here, we revealed that the intratracheal administration of an appropriate amount of AAV2/8 predominantly targets lung tissue. AAV-mediated gene delivery that we used in this study induced the expression of the desired protein in lung parenchymal cells, including alveolar type II cells. We harnessed the technique to develop severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-susceptible mice. Three kinds of immune function-relevant gene knockout (KO) mice were transduced with AAV encoding human angiotensin-converting enzyme 2 (hACE2) and then injected with SARS-CoV-2. Among these mice, type I interferon receptor (IFNAR) KO mice showed increased viral titer in the lungs compared to that in the other KO mice. Moreover, nucleocapsid protein of SARS-CoV-2 and multiple lesions in the trachea and lung were observed in AAV-hACE2-transduced, SARS-CoV-2-infected IFNAR KO mice, indicating the involvement of type I interferon signaling in the protection of SARS-CoV-2. In this study, we demonstrate the ease and rapidness of the intratracheal administration of AAV for targeting lung tissue in mice, and this can be used to study diverse pulmonary diseases.
Assuntos
COVID-19 , SARS-CoV-2 , Animais , COVID-19/terapia , Dependovirus/genética , Modelos Animais de Doenças , Suscetibilidade a Doenças , Pulmão/patologia , Camundongos , Camundongos Transgênicos , SARS-CoV-2/genéticaRESUMO
Severe diarrhea was reported in goat kids in Chungcheongbuk-do, Korea, from 2021 to 2023, and Cryptosporidium infection was suspected. To confirm the cause of this outbreak, fecal samples were collected from goat farms where diarrhea had been reported and analyzed for Cryptosporidium infection using a molecular assay. A total of 65 fecal samples, including 37 from goats with diarrhea and 28 from goats without diarrhea, were collected from six goat farms. Forty-eight of the goats were kids (<2 months) and 17 were adults (>1 year). Cryptosporidium was identified in 53.8% (35/65) of total samples. Overall, 86.5% (32/37) of the diarrheic fecal samples tested positive; however, Cryptosporidium was not detected in any fecal sample from non-diarrheic adult goats. Therefore, cryptosporidiosis was significantly associated with diarrhea in goat kids, and adult goats were not responsible for transmission of Cryptosporidium to them. Phylogenetic analysis and molecular characterization revealed two Cryptosporidium species, namely, C. parvum (n = 28) and C. xiaoi (n = 7). In the C. parvum-positive samples, gp60 gene analysis revealed three zoonotic subtypes-IIaA18G3R1, IIdA15G1, and IIdA16G1. To the best of our knowledge, this study is the first to identify C. parvum IIaA18G3R1 and IIdA16G1 in goats, as well as the first to identify C. xiaoi in goats in Korea. These results suggest that goat kids play an important role as reservoir hosts for different Cryptosporidium species and that continuous monitoring with biosecurity measures is necessary to control cryptosporidiosis outbreaks.
Assuntos
Criptosporidiose , Cryptosporidium parvum , Cryptosporidium , Doenças das Cabras , Doenças dos Ovinos , Animais , Ovinos , Criptosporidiose/epidemiologia , Cryptosporidium parvum/genética , Cabras , Filogenia , Doenças das Cabras/epidemiologia , Doenças dos Ovinos/epidemiologia , Cryptosporidium/genética , Diarreia/epidemiologia , Diarreia/veterinária , Fezes , Surtos de Doenças/veterinária , República da Coreia/epidemiologia , GenótipoRESUMO
BACKGROUND: Binding IgE to a cognate allergen causes aggregation of Fcε receptor I (FcεRI) in mast cells, resulting in activation of receptor-associated Src family tyrosine kinases, including Lyn and Syk. Protein tyrosine phosphatase, receptor type C (PTPRC), also known as CD45, has emerged as a positive regulator of FcεRI signaling by dephosphorylation of the inhibitory tyrosine of Lyn. OBJECTIVE: Sirtuin 6 (Sirt6), a NAD+-dependent deacetylase, exhibits an anti-inflammatory property. It remains to be determined, however, whether Sirt6 attenuates mast cell-associated diseases, including anaphylaxis. METHODS: FcεRI signaling and mast cell degranulation were measured after IgE cross-linking in murine bone marrow-derived mast cells (BMMCs) and human cord blood-derived mast cells. To investigate the function of Sirt6 in mast cell activation in vivo, we used mast cell-dependent animal models of passive systemic anaphylaxis (PSA) and passive cutaneous anaphylaxis (PCA). RESULTS: Sirt6-deficient BMMCs augmented IgE-FcεRI-mediated signaling and degranulation compared to wild-type BMMCs. Reconstitution of mast cell-deficient KitW-sh/W-sh mice with BMMCs received from Sirt6 knockout mice developed more severe PSA and PCA compared to mice engrafted with wild-type BMMCs. Similarly, genetic overexpression or pharmacologic activation of Sirt6 suppressed mast cell degranulation and blunted responses to PCA. Mechanistically, Sirt6 deficiency increased PTPRC transcription via acetylating histone H3, leading to enhanced aggregation of FcεRI in BMMCs. Finally, we recapitulated the Sirt6 regulation of PTPRC and FcεRI signaling in human mast cells. CONCLUSIONS: Sirt6 acts as a negative regulator of FcεRI signaling cascade in mast cells by suppressing PTPRC transcription. Activation of Sirt6 may therefore represent a promising and novel therapeutic strategy for anaphylaxis.
Assuntos
Anafilaxia/imunologia , Mastócitos/imunologia , Receptores de IgE/imunologia , Sirtuínas/imunologia , Animais , Células da Medula Óssea/citologia , Sangue Fetal/citologia , Humanos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Transdução de Sinais , Sirtuínas/genéticaRESUMO
Diplogasteroides sp., a cryptic population of D. haslacheri, and Parasitorhabditis terebranus were reported from the frass of Monochamus alternatus galleries in dead Pinus thunbergii for the first time in Korea. Females and males are morphologically characterized and their linked DNA barcodes (18S-rRNA, 28S-rRNA, ITS-rRNA and COI) supplied. Females and males of the two species from Korea conform to the original species descriptions from Europe and the USA, with variations in a few details in morphometrics. Specifically, Diplogasteroides sp. is morphologically very similar to D. haslacheri. However, it cannot be designated as D. haslacheri due to the existence of cryptic species complex within the haslacheri group (D. haslacheri, D. asiaticus, D. nix, D. andrassyi, and D. carinthiacus), a condition requiring hybridization studies to test species identity within the group. Based on analysis of COI sequences, differences among these cryptic species are evident. Thus, in addition to hybridization tests, the COI might be a powerful DNA barcoding marker for the precise identification of these cryptic species within the genus. Additionally, this is the first molecular characterization of P. terebranus, and the species is herein recorded for the first time outside its type locality.
RESUMO
This study aimed to examine the distribution of gastrointestinal parasitic infections in domestic pigs in the Republic of Korea. From May 2020 to October 2021, 364 pig fecal samples were collected from 75 farms in 7 Provinces and microscopically examined. A total of 170 (46.7%) pigs were infected with at least one of the following parasites: Balantioides coli, strongyles, Ascaris suum, Trichuris suis, and coccidia. By parasite species, B. coli, strongyles, A. suum, T. suis, and coccidia oocysts or eggs were detected in 144 (39.6%), 24 (6.6%), 14 (3.8%), 4 (1.1%), and 1 (0.3%) samples, respectively. One hundred fifty-four, 15, and 1 cases showed single, double, and triple infections, respectively. Of the swine fecal samples from 75 farms, 69 specimens (92.0%) were infected with 1 or more parasites. All surveyed farms across the country exhibited a positive rate of over 30%, among which the highest positive rate was 65.0% in Chungcheongnam-do, and Jeollabuk-do was followed by 61.9%. Winter showed a statistically lower prevalence than other seasons. This study showed that gastrointestinal parasites are prevalent in pigs in Korea, although the diversity of parasites is low.
Assuntos
Enteropatias Parasitárias/veterinária , Parasitos/classificação , Doenças dos Suínos/epidemiologia , Doenças dos Suínos/parasitologia , Animais , Fezes/parasitologia , Enteropatias Parasitárias/epidemiologia , Enteropatias Parasitárias/parasitologia , Parasitos/isolamento & purificação , Prevalência , República da Coreia/epidemiologia , Estações do Ano , Sus scrofa , SuínosRESUMO
Chios mastic gum (CMG), a resin of the mastic tree (Pistacia lentiscus var. chia), has been used to treat multiple disorders caused by gastrointestinal malfunctions and bacterial infections for more than 2500 years. However, little is known about CMG's antiviral activity. CMG is known to influence multiple cellular processes such as cell proliferation, differentiation and apoptosis. As virus replication is largely dependent on the host cellular metabolism, it is conceivable that CMG regulates virus infectivity. Therefore, in this study, we evaluated CMG's potential as an antiviral drug to treat influenza A virus (IAV) infection. CMG treatment dramatically reduced the cytopathogenic effect and production of RNAs, proteins and infectious particles of IAV. Interestingly, CMG interfered with the early stage of the virus life cycle after viral attachment. Importantly, the administration of CMG greatly ameliorated morbidity and mortality in IAV-infected mice. The results suggest that CMG displays a potent anti-IAV activity by blocking the early stage of viral replication. Thus, mastic gum could be exploited as a novel therapeutic agent against IAV infection.
Assuntos
Vírus da Influenza A Subtipo H1N1/efeitos dos fármacos , Vírus da Influenza A Subtipo H1N1/patogenicidade , Resina Mástique/farmacologia , Infecções por Orthomyxoviridae/tratamento farmacológico , Animais , Antivirais/farmacologia , Antivirais/uso terapêutico , Sobrevivência Celular/efeitos dos fármacos , Efeito Citopatogênico Viral/efeitos dos fármacos , Cães , Células HEK293 , Humanos , Vírus da Influenza A Subtipo H1N1/fisiologia , Células Madin Darby de Rim Canino , Resina Mástique/uso terapêutico , Infecções por Orthomyxoviridae/virologia , Virulência/efeitos dos fármacos , Ligação Viral , Replicação Viral/efeitos dos fármacosRESUMO
The severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) pandemic is causing a global crisis. It is still unresolved. Although many therapies and vaccines are being studied, they are still in their infancy. As this pandemic continues, rapid and accurate research for the development of therapies and vaccines is needed. Therefore, it is necessary to understand characteristics of diseases caused by SARS-CoV-2 through animal models. Syrian hamsters are known to be susceptible to SARS-CoV-2. They were intranasally inoculated with SARS-CoV-2. At 2, 4, 8, 12, and 16 days post-infection (dpi), these hamsters were euthanized, and tissues were collected for ultrastructural and microstructural examinations. Microscopic lesions were prominent in the upper and lower respiratory tracts from 2 and 4 dpi groups, respectively. The respiratory epithelium in the trachea, bronchiole, and alveolar showed pathological changes. Inflammatory cells including neutrophils, lymphocytes, macrophages, and eosinophils were infiltrated in/around tracheal lamina propria, pulmonary vessels, alveoli, and bronchiole. In pulmonary lesions, alveolar wall was thickened with infiltrated inflammatory cells, mainly neutrophils and macrophages. In the trachea, epithelial damages started from 2 dpi and recovered from 8 dpi, consistent with microscopic results, High levels of SARS-CoV-2 nucleoprotein were detected at 2 dpi and 4 dpi. In the lung, lesions were most severe at 8 dpi. Meanwhile, high levels of SARS-CoV-2 were detected at 4 dpi. Electron microscopic examinations revealed cellular changes in the trachea epithelium and alveolar epithelium such as vacuolation, sparse micro-organelle, and poor cellular margin. In the trachea epithelium, the number of cytoplasmic organelles was diminished, and small vesicles were prominent from 2 dpi. Some of these electron-lucent vesicles were filled with virion particles. From 8 dpi, the trachea epithelium started to recover. Because of shrunken nucleus and swollen cytoplasm, the N/C ratio of type 2 pneumocyte decreased at 8 and 12 dpi. From 8 dpi, lamellar bodies on type 2 pneumocyte cytoplasm were increasingly observed. Their number then decreased from 16 dpi. However, there was no significant change in type 1 pneumocyte. Viral vesicles were only observed in the cytoplasm of type 2 pneumocyte. In conclusion, ultra- and micro-structural changes presented in this study may provide useful information for SARS-CoV-2 studies in various fields.
Assuntos
COVID-19/patologia , Sistema Respiratório/patologia , SARS-CoV-2/patogenicidade , Animais , Cricetinae , Imuno-Histoquímica/veterinária , Masculino , Mesocricetus , Projetos Piloto , RNA Viral/química , RNA Viral/isolamento & purificação , Reação em Cadeia da Polimerase em Tempo Real/veterinária , Sistema Respiratório/química , Sistema Respiratório/ultraestrutura , Sistema Respiratório/virologia , Fatores de Tempo , Traqueia/patologia , Traqueia/ultraestrutura , Traqueia/virologia , Redução de PesoRESUMO
Agrimonia pilosa (AP), Galla rhois (RG), and their mixture (APRG64) strongly inhibited SARS-CoV-2 by interfering with multiple steps of the viral life cycle including viral entry and replication. Furthermore, among 12 components identified in APRG64, three displayed strong antiviral activity, ursolic acid (1), quercetin (7), and 1,2,3,4,6-penta-O-galloyl-ß-d-glucose (12). Molecular docking analysis showed these components to bind potently to the spike receptor-binding-domain (RBD) of the SARS-CoV-2 and its variant B.1.1.7. Taken together, these findings indicate APRG64 as a potent drug candidate to treat SARS-CoV-2 and its variants.
Assuntos
Agrimonia/química , Antivirais/química , Produtos Biológicos/química , Tratamento Farmacológico da COVID-19 , Extratos Vegetais/química , SARS-CoV-2/efeitos dos fármacos , Sequência de Aminoácidos , Antivirais/farmacologia , Produtos Biológicos/farmacologia , Descoberta de Drogas , Humanos , Taninos Hidrolisáveis/química , Simulação de Acoplamento Molecular , Extratos Vegetais/farmacologia , Ligação Proteica , Quercetina/química , Glicoproteína da Espícula de Coronavírus/química , Triterpenos/química , Internalização do Vírus/efeitos dos fármacos , Ácido UrsólicoRESUMO
The influenza virus is one of the major public health threats. However, the development of efficient vaccines and therapeutic drugs to combat this virus is greatly limited by its frequent genetic mutations. Because of this, targeting the host factors required for influenza virus replication may be a more effective strategy for inhibiting a broader spectrum of variants. Here, we demonstrated that inhibition of a motor protein kinesin family member 18A (KIF18A) suppresses the replication of the influenza A virus (IAV). The expression of KIF18A in host cells was increased following IAV infection. Intriguingly, treatment with the selective and ATP-competitive mitotic kinesin KIF18A inhibitor BTB-1 substantially decreased the expression of viral RNAs and proteins, and the production of infectious viral particles, while overexpression of KIF18A enhanced the replication of IAV. Importantly, BTB-1 treatment attenuated the activation of AKT, p38 MAPK, SAPK and Ran-binding protein 3 (RanBP3), which led to the prevention of the nuclear export of viral ribonucleoprotein complexes. Notably, administration of BTB-1 greatly improved the viability of IAV-infected mice. Collectively, our results unveiled a beneficial role of KIF18A in IAV replication, and thus, KIF18A could be a potential therapeutic target for the control of IAV infection.
Assuntos
Resistência à Doença , Interações Hospedeiro-Patógeno , Vírus da Influenza A/fisiologia , Influenza Humana/metabolismo , Influenza Humana/virologia , Cinesinas/metabolismo , Replicação Viral , Trifosfato de Adenosina/metabolismo , Animais , Linhagem Celular , Células Cultivadas , Efeito Citopatogênico Viral , Modelos Animais de Doenças , Resistência à Doença/genética , Expressão Gênica , Regulação Viral da Expressão Gênica , Interações Hospedeiro-Patógeno/genética , Humanos , Cinesinas/genética , Masculino , Camundongos , Infecções por Orthomyxoviridae/metabolismo , Infecções por Orthomyxoviridae/virologia , Proteínas Proto-Oncogênicas c-akt/metabolismoRESUMO
The host-associated defence system responsible for the clearance of porcine reproductive and respiratory syndrome virus (PRRSV) from infected pigs is currently poorly understood. To better understand the dynamics of host-pathogen interactions, seventy-five of 100 pigs infected with PRRSV-JA142 and 25 control pigs were euthanized at 3, 10, 21, 28 and 35 days post-challenge (dpc). Blood, lung, bronchoalveolar lavage (BAL) and bronchial lymph node (BLN) samples were collected to evaluate the cellular immune responses. The humoral responses were evaluated by measuring the levels of anti-PRRSV IgG and serum virus-neutralizing (SVN) antibodies. Consequently, the highest viral loads in the sera and lungs of the infected pigs were detected between 3 and 10 dpc, and these resulted in moderate to mild interstitial pneumonia, which resolved accompanied by the clearance of most of the virus by 28 dpc. At peak viremia, the frequencies of alveolar macrophages in infected pigs were significantly decreased, whereas the monocyte-derived DC/macrophage and conventional DC frequencies were increased, and these effects coincided with the early induction of local T-cell responses and the presence of proinflammatory cytokines/chemokines in the lungs, BAL, and BLN as early as 10 dpc. Conversely, the systemic T-cell responses measured in the peripheral blood mononuclear cells were delayed and significantly induced only after the peak viremic stage between 3 and 10 dpc. Taken together, our results suggest that activation of immune responses in the lung could be the key elements for restraining PRRSV through the early induction of T-cell responses at the sites of virus replication.
Assuntos
Imunidade Adaptativa , Líquido da Lavagem Broncoalveolar/imunologia , Imunidade Inata , Pulmão/imunologia , Linfonodos/imunologia , Síndrome Respiratória e Reprodutiva Suína/imunologia , Vírus da Síndrome Respiratória e Reprodutiva Suína/fisiologia , Animais , Brônquios/imunologia , Brônquios/virologia , Líquido da Lavagem Broncoalveolar/virologia , Pulmão/virologia , Linfonodos/virologia , Tecido Parenquimatoso/imunologia , Tecido Parenquimatoso/virologia , Sus scrofa , SuínosRESUMO
Porcine reproductive and respiratory syndrome virus (PRRSV) infection is the most important viral disease causing severe economic losses in the swine industry. However, mechanisms underlying gene expression control in immunity-responsible tissues at different time points during PRRSV infection are poorly understood. We constructed an integrated gene co-expression network and identified tissue- and time-dependent biological mechanisms of PRRSV infection through bioinformatics analysis using three tissues (lungs, bronchial lymph nodes [BLNs], and tonsils) via RNA-Seq. Three groups with specific expression patterns (i.e., the 3-dpi, lung, and BLN groups) were discovered. The 3 dpi-specific group showed antiviral and innate-immune signalling similar to the case for influenza A infection. Moreover, we observed adaptive immune responses in the lung-specific group based on various cytokines, while the BLN-specific group showed down-regulated AMPK signalling related to viral replication. Our study may provide comprehensive insights into PRRSV infection, as well as useful information for vaccine development.
Assuntos
Imunidade Adaptativa/genética , Imunidade Inata/genética , Síndrome Respiratória e Reprodutiva Suína/imunologia , Vírus da Síndrome Respiratória e Reprodutiva Suína/fisiologia , Transcriptoma/imunologia , Animais , Síndrome Respiratória e Reprodutiva Suína/genética , Síndrome Respiratória e Reprodutiva Suína/virologia , Sus scrofa , SuínosRESUMO
Guanylate-binding proteins (GBP1 and GBP5) are known to be important for host resistance against porcine reproductive and respiratory syndrome virus (PRRSV) infection. In this study, the effects of polymorphisms in GBP1 (GBP1E2 and WUR) and GBP5 on host immune responses against PRRSV were investigated to elucidate the mechanisms governing increased resistance to this disease. Seventy-one pigs [pre-genotyped based on three SNP markers (GBP1E2, WUR, and GBP5)] were assigned to homozygous (n = 36) and heterozygous (n = 35) groups and challenged with the JA142 PRRSV strain. Another group of nineteen pigs was kept separately as a negative control group. Serum and peripheral blood mononuclear cells (PBMCs) were collected at 0, 3, 7, 14, 21 and 28 days post-challenge (dpc). Viremia and weight gain were measured in all pigs at each time point, and a flow cytometry analysis of PBMCs was performed to evaluate T cell activation. In addition, 15 pigs (5 pigs per homozygous, heterozygous and negative groups) were sacrificed at 3, 14 and 28 dpc, and the local T cell responses were evaluated in the lungs, bronchoalveolar lavage cells (BALc), lymph nodes and tonsils. The heterozygous pigs showed lower viral loads in the serum and lungs and higher weight gains than the homozygous pigs based on the area under the curve calculation. Consistently, compared with the homozygous pigs, the heterozygous pigs exhibited significantly higher levels of IFN-α in the serum, proliferation of various T cells (γδT, Th1, and Th17) in PBMCs and tissues, and cytotoxic T cells in the lungs and BALc. These results indicate that the higher resistance in the pigs heterozygous for the GBP1E2, WUR and GBP5 markers could be mediated by increased antiviral cytokine (IFN-α) production and T cell activation.
Assuntos
Resistência à Doença , Proteínas de Ligação ao GTP/genética , Polimorfismo Genético , Síndrome Respiratória e Reprodutiva Suína/virologia , Vírus da Síndrome Respiratória e Reprodutiva Suína/genética , Animais , Feminino , Proteínas de Ligação ao GTP/metabolismo , Masculino , SuínosRESUMO
The NAD-dependent histone deacetylase Sirt1 antagonizes p53 transcriptional activity to regulate cell-cycle progression and apoptosis. We have identified a ubiquitin-specific peptidase, USP22, one of the 11 death-from-cancer signature genes that are critical in controlling cell growth and death, as a positive regulator of Sirt1. USP22 interacts with and stabilizes Sirt1 by removing polyubiquitin chains conjugated onto Sirt1. The USP22-mediated stabilization of Sirt1 leads to decreasing levels of p53 acetylation and suppression of p53-mediated functions. In contrast, depletion of endogenous USP22 by RNA interference destabilizes Sirt1, inhibits Sirt1-mediated deacetylation of p53 and elevates p53-dependent apoptosis. Genetic deletion of the usp22 gene results in Sirt1 instability, elevated p53 transcriptional activity and early embryonic lethality in mice. Our study elucidates a molecular mechanism in suppression of cell apoptosis by stabilizing Sirt1 in response to DNA damage and reveals a critical physiological function of USP22 in mouse embryonic development.
Assuntos
Desenvolvimento Embrionário/fisiologia , Endopeptidases/metabolismo , Sirtuína 1/metabolismo , Proteína Supressora de Tumor p53/metabolismo , Animais , Apoptose/genética , Apoptose/fisiologia , Dano ao DNA , Desenvolvimento Embrionário/genética , Endopeptidases/deficiência , Endopeptidases/genética , Estabilidade Enzimática , Feminino , Células HEK293 , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Gravidez , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Sirtuína 1/genética , Ativação Transcricional , Proteases Específicas de Ubiquitina , UbiquitinaçãoRESUMO
Patients with non-small-cell lung cancer (NSCLC) containing epidermal growth factor receptor (EGFR) amplification or sensitive mutations initially respond to tyrosine kinase inhibitor gefitinib; however, the treatment is less effective over time. Gefitinib resistance mechanisms include MET gene amplification. A therapeutic strategy targeting MET as well as EGFR can overcome resistance to gefitinib. In the present study we identified Echinatin (Ecn), a characteristic chalcone in licorice, which inhibited both EGFR and MET and strongly altered NSCLC cell growth. The antitumor efficacy of Ecn against gefitinib-sensitive or -resistant NSCLC cells with EGFR mutations and MET amplification was confirmed by suppressing cell proliferation and anchorage-independent colony growth. During the targeting of EGFR and MET, Ecn significantly blocked the kinase activity, which was validated with competitive ATP binding. Inhibition of EGFR and MET by Ecn decreases the phosphorylation of downstream target proteins ERBB3, AKT and ERK compared with total protein expression or control. Ecn induced the G2/M cell cycle arrest, and apoptosis via the intrinsic pathway of caspase-dependent activation. Ecn induced ROS production and GRP78, CHOP, DR5 and DR4 expression as well as depolarized the mitochondria membrane potential. Therefore, our results suggest that Ecn is a promising therapeutic agent in NSCLC therapy.
Assuntos
Carcinoma Pulmonar de Células não Pequenas/patologia , Chalconas/farmacologia , Gefitinibe/farmacologia , Neoplasias Pulmonares/patologia , Proteínas Proto-Oncogênicas c-met/antagonistas & inibidores , Apoptose/efeitos dos fármacos , Carcinoma Pulmonar de Células não Pequenas/tratamento farmacológico , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Resistencia a Medicamentos Antineoplásicos , Chaperona BiP do Retículo Endoplasmático , Receptores ErbB/antagonistas & inibidores , Receptores ErbB/genética , Glycyrrhiza/química , Humanos , Neoplasias Pulmonares/tratamento farmacológico , Simulação de Acoplamento Molecular , Raízes de Plantas/química , Inibidores de Proteínas Quinases , Proteínas Proto-Oncogênicas c-met/genética , Quinazolinas/farmacologiaRESUMO
As agonists of TLR7/8, single-stranded RNAs (ssRNAs) are safe and promising adjuvants that do not cause off-target effects or innate immune overactivation. However, low stability prevents them from mounting sufficient immune responses. This study evaluates the adjuvant effects of ssRNA derived from the cricket paralysis virus intergenic region internal ribosome entry site, formulated as nanoparticles with a coordinative amphiphile, containing a zinc/dipicolylamine complex moiety as a coordinative phosphate binder, as a stabilizer for RNA-based adjuvants. The nanoformulated ssRNA adjuvant was resistant to enzymatic degradation in vitro and in vivo, and that with a coordinative amphiphile bearing an oleyl group (CA-O) was approximately 100â nm, promoted effective recognition, and improved activation of antigen-presenting cells, leading to better induction of neutralizing antibodies following single immunization. Hence, CA-O may increase the efficacy of ssRNA-based adjuvants, proving useful to meet the urgent need for vaccines during pathogen outbreaks.
Assuntos
Adjuvantes Imunológicos/farmacologia , Células Apresentadoras de Antígenos/imunologia , Composição de Medicamentos , Imunidade Humoral/efeitos dos fármacos , Nanotecnologia , RNA/química , Adjuvantes Imunológicos/química , Animais , HumanosRESUMO
Omega-3 (n-3) polyunsaturated fatty acids (PUFAs) have been known to exert anti-inflammatory effects on various disease states. However, its effect on CD8+ T cell-mediated immunopathology upon viral infection has not been well elucidated yet. In this study, we investigated the possible implication of n-3 PUFAs in CD8+ T cell responses against an acute viral infection. Infection of FAT-1 transgenic mice that are capable of synthesizing n-3 PUFAs from n-6 PUFAs with lymphocytic choriomeningitis virus (LCMV) resulted in significant reduction of anti-viral CD8+ T cell responses. Interestingly, expansion of adoptively transferred wild-type (WT) LCMV-specific T cell receptor (TCR) transgenic CD8+ (P14) T cells into FAT-1 mice was significantly decreased. Also, activation of anti-viral CD4+ helper T cells was reduced in FAT-1 mice. Importantly, P14 cells carrying the fat-1 gene that were adoptively transferred into WT mice exhibited a substantially decreased ability to proliferate and produce cytokines against LCMV infection. Together, n-3 PUFAs attenuated anti-viral CD8+ T cell responses against an acute viral infection and thus could be used to alleviate immunopathology mediated by the viral infection.
Assuntos
Linfócitos T CD8-Positivos/imunologia , Ácidos Graxos Ômega-3/imunologia , Inflamação/etiologia , Coriomeningite Linfocítica/complicações , Vírus da Coriomeningite Linfocítica/imunologia , Animais , Linfócitos T CD8-Positivos/virologia , Inflamação/imunologia , Inflamação/virologia , Coriomeningite Linfocítica/imunologia , Coriomeningite Linfocítica/virologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos TransgênicosRESUMO
The inflammasome is a multiprotein signaling complex that mediates inflammatory innate immune responses through caspase 1 activation and subsequent IL-1ß secretion. However, because its aberrant activation often leads to inflammatory diseases, targeting the inflammasome holds promise for the treatment of inflammation-related diseases. In this study, it was found that a hot-water extract of Sanguisorba officinalis (HSO) suppresses inflammasome activation triggered by adenosine 5'-triphosphate, nigericin, microbial pathogens, and double stranded DNA in bone marrow-derived macrophages. HSO was found to significantly suppress IL-1ß production in a dose-dependent manner; this effect correlated well with small amounts of caspase 1 and little ASC pyroptosome formation in HSO-treated cells. The anti-inflammatory activity of HSO was further confirmed in a mouse model of endotoxin-induced septic shock. Oral administration of HSO reduced IL-1ß titers in the serum and peritoneal cavity, increasing the survival rate. Taken together, our results suggest that HSO is an inhibits inflammasome activation through nucleotide-binding domain and leucine-rich repeat pyrin domain 3, nucleotide-binding domain and leucine-rich repeat caspase recruitment domain 4 and absent in melanoma 2 pathways, and may be useful for treatment of inflammasome-mediated diseases.
Assuntos
Inflamassomos/efeitos dos fármacos , Extratos Vegetais/antagonistas & inibidores , Sanguisorba/química , Choque Séptico/tratamento farmacológico , Animais , Anti-Inflamatórios/farmacologia , Caspase 1/metabolismo , Citocinas/metabolismo , Modelos Animais de Doenças , Endotoxinas/efeitos adversos , Feminino , Medicina Herbária , Inflamação , Interleucina-1beta/sangue , Interleucina-1beta/metabolismo , Lipopolissacarídeos/efeitos adversos , Listeria monocytogenes/patogenicidade , Macrófagos/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Proteínas NLR/metabolismo , Nigericina/farmacologia , Extratos Vegetais/administração & dosagem , Raízes de Plantas/química , República da Coreia , Salmonella typhimurium/patogenicidade , Choque Séptico/microbiologia , Taxa de SobrevidaRESUMO
Middle East respiratory syndrome coronavirus (MERS-CoV) is a single-stranded RNA virus that causes severe respiratory disease in humans with a high fatality rate. Binding of the receptor binding domain (RBD) of the spike (S) glycoprotein to dipeptidyl peptidase 4 is the critical step in MERS-CoV infection of a host cell. No vaccines or clinically applicable treatments are currently available for MERS-CoV. Therefore, rapid diagnosis is important for improving patient outcomes through prompt treatment and protection against viral outbreaks. In this study, the aim was to establish two ELISA systems for detecting antigens and antibodies against MERS-CoV. Using a recombinant full-length S protein, an indirect ELISA was developed and found to detect MERS-CoV-specific antibodies in animal sera and sera of patient with MERS. Moreover, MAbs were induced with the recombinant S protein and RBD and used for sandwich ELISA to detect the MERS-CoV S protein. Neither ELISA system exhibited significant intra-assay or inter-assay variation, indicating good reproducibility. Moreover, the inter-day precision and sensitivity were adequate for use as a diagnostic kit. Thus, these ELISAs can be used clinically to diagnose MERS-CoV.
Assuntos
Anticorpos Antivirais/isolamento & purificação , Antígenos Virais/isolamento & purificação , Infecções por Coronavirus/diagnóstico , Infecções por Coronavirus/imunologia , Testes Imunológicos/métodos , Coronavírus da Síndrome Respiratória do Oriente Médio/imunologia , Glicoproteína da Espícula de Coronavírus/imunologia , Animais , Anticorpos Monoclonais , Anticorpos Antivirais/sangue , Anticorpos Antivirais/imunologia , Antígenos Virais/imunologia , Baculoviridae/genética , Infecções por Coronavirus/virologia , Ensaio de Imunoadsorção Enzimática/métodos , Escherichia coli/genética , Expressão Gênica , Humanos , Coronavírus da Síndrome Respiratória do Oriente Médio/genética , Ligação Proteica/genética , Ligação Proteica/imunologia , Domínios e Motivos de Interação entre Proteínas , Ratos , Ratos Wistar , Receptores Virais/imunologia , Proteínas Recombinantes/genética , Proteínas Recombinantes/imunologia , Reprodutibilidade dos Testes , Sensibilidade e Especificidade , Células Sf9 , Glicoproteína da Espícula de Coronavírus/genética , Proteínas da Matriz Viral/genética , Proteínas da Matriz Viral/imunologiaRESUMO
Host factors such as nutritional status and immune cell state are important for vaccine efficacy. Inflammasome activation may be important for triggering vaccine-induced humoral and cell-mediated immune responses. Formulations with alum as a typical adjuvant to overcome the effects of host factors have recently been shown to induce inflammasome activation, which augments vaccine efficacy. Apoptosis-associated speck-like protein containing a caspase recruitment domain (ASC) is one of the main components of inflammasomes, but it is not clear whether ASC affects the vaccine-induced immune response. Herein, we used two types of vaccines: inactivated influenza vaccine not formulated with alum, and HPV vaccine formulated with alum. We gave the vaccines to ASC knockout (ASC-/- ) mice to investigate the role of ASC in vaccine efficacy. Influenza vaccine-immunized ASC-/- mice did not show antibody titers in week 2 after the first vaccination. After boosting, the antibody titer in ASC-/- mice was about half that in wild type (WT) mice. Furthermore, a cytotoxic T-lymphocyte response against influenza vaccine was not induced in ASC-/- mice. Therefore, vaccinated ASC-/- mice did not show effective protection against viral challenge. ASC-/- mice immunized with alum-formulated HPV vaccine showed similar antibody titers and T-cell proliferation compared with immunized WT mice. However, the HPV vaccine without alum induced up to threefold lower titers of HPV-specific antibody titers in ASC-/- mice compared with those in WT mice. These findings suggest that alum in vaccine can overcome the ASC-deficient condition.
Assuntos
Adjuvantes Imunológicos/uso terapêutico , Hidróxido de Alumínio/imunologia , Apoptose/imunologia , Domínio de Ativação e Recrutamento de Caspases/imunologia , Domínio de Ativação e Recrutamento de Caspases/fisiologia , Vacinas contra Influenza/imunologia , Vacinas contra Papillomavirus/imunologia , Compostos de Alúmen , Animais , Anticorpos Antivirais , Domínio de Ativação e Recrutamento de Caspases/genética , Proliferação de Células/efeitos dos fármacos , Feminino , Humanos , Imunidade Humoral , Inflamassomos , Vacinas contra Influenza/uso terapêutico , Influenza Humana/prevenção & controle , Pulmão/patologia , Pulmão/virologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Testes de Neutralização , Orthomyxoviridae , Vacinas contra Papillomavirus/farmacologia , Vacinas contra Papillomavirus/uso terapêutico , Linfócitos T/efeitos dos fármacos , Vacinação , Vacinas de Produtos Inativados/imunologia , Vacinas de Produtos Inativados/uso terapêuticoRESUMO
Inhibitor K562 (IK) protein was first isolated from the culture medium of K562, a leukemia cell line. It is known to be an inhibitory regulator of interferon-γ-induced major histocompatibility complex class (MHC) II expression. Previously, we found that transgenic (Tg) mice constitutively expressing truncated IK (tIK) showed reduced numbers of pathogenic Th1 and Th17 cells, which are known to be involved in the development of rheumatoid arthritis (RA). Here, we investigated whether exogenous tIK protein has a therapeutic effect in arthritis in disease models and analyzed its mechanism. Exogenous tIK protein was produced in an insect expression system and applied to the collagen antibody-induced arthritis (CAIA) mouse disease model. Injection of tIK protein alleviated the symptoms of arthritis in the CAIA model and reduced Th1 and Th17 cell populations. In addition, treatment of cultured T cells with tIK protein induced expression of A20, a negative regulator of nuclear factor-κB (NFκB)-induced inflammation, and reduced expression of several transcription factors related to T cell activation. We conclude that exogenous tIK protein has the potential to act as a new therapeutic agent for RA patients, because it has a different mode of action to biopharmaceutical agents, such as tumor necrosis factor antagonists, that are currently used to treat RA.