RESUMEN
PURPOSE: The role of the nitric oxide synthases (NOSs) system in cerebral infarction has been examined in pharmacological studies with non-selective NOSs inhibitors. However, due to the non-specificity of the non-selective NOSs inhibitors, its role remains to be fully elucidated. We addressed this issue in mice in which neuronal, inducible, and endothelial NOS isoforms were completely disrupted. METHODS AND RESULTS: We newly generated mice lacking all three NOSs by crossbreeding each single NOS-/- mouse. In the male, cerebral infarct size at 24 h after middle cerebral artery occlusion (MCAO) was significantly smaller in the triple n/i/eNOSs-/- genotype as compared with wild-type genotype. Neurological deficit score and mortality rate were also significantly lower in the triple n/i/eNOSs-/- than in the WT genotype. In contrast, in the female, there was no significant difference in the cerebral infarct size in the two genotypes. In the male triple n/i/eNOSs-/- genotype, orchiectomy significantly increased the cerebral infarct size, and in the orchiectomized male triple n/i/eNOSs-/- genotype, treatment with testosterone significantly reduced it. Cyclopaedic and quantitative comparisons of mRNA expression levels in cerebral infarct lesions between the male wild-type and triple n/i/eNOSs-/- genotypes at 1 h after MCAO revealed significant involvements of decreased oxidative stress and mitigated mitochondrial dysfunction in the alleviated cerebral infarction in the male triple n/i/eNOSs-/- genotype. CONCLUSIONS: These results provide the first evidence that the NOSs system exerts a deleterious effect against acute ischemic brain injury in the male.
Asunto(s)
Infarto de la Arteria Cerebral Media , Óxido Nítrico Sintasa , Ratones , Masculino , Femenino , Animales , Óxido Nítrico Sintasa/genética , Óxido Nítrico Sintasa/metabolismo , Isoformas de Proteínas/metabolismo , Estrés Oxidativo , Óxido Nítrico , Ratones NoqueadosRESUMEN
Lung infection during severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) via the angiotensin-I-converting enzyme 2 (ACE2) receptor induces a cytokine storm. However, the precise mechanisms involved in severe COVID-19 pneumonia are unknown. Here, we showed that interleukin-10 (IL-10) induced the expression of ACE2 in normal alveolar macrophages, causing them to become vectors for SARS-CoV-2. The inhibition of this system in hamster models attenuated SARS-CoV-2 pathogenicity. Genome-wide association and quantitative trait locus analyses identified a IFNAR2-IL10RB readthrough transcript, COVID-19 infectivity-enhancing dual receptor (CiDRE), which was highly expressed in patients harboring COVID-19 risk variants at the IFNAR2 locus. We showed that CiDRE exerted synergistic effects via the IL-10-ACE2 axis in alveolar macrophages and functioned as a decoy receptor for type I interferons. Collectively, our data show that high IL-10 and CiDRE expression are potential risk factors for severe COVID-19. Thus, IL-10R and CiDRE inhibitors might be useful COVID-19 therapies.
Asunto(s)
COVID-19 , Humanos , COVID-19/genética , SARS-CoV-2 , Enzima Convertidora de Angiotensina 2/genética , Interleucina-10/genética , Macrófagos Alveolares/metabolismo , Estudio de Asociación del Genoma Completo , Peptidil-Dipeptidasa A/metabolismoRESUMEN
Chronic obstructive pulmonary disease (COPD/emphysema) is a life-threatening disorder and there are few effective therapies. Cigarette smoke-induced oxidative stress, airway inflammation, and apoptosis of lung cells have been reported to be involved in the pathogenesis of COPD/emphysema and lead to alveolar septal destruction. Here we show that the expression level of FCH and double SH3 domains 1 (FCHSD1) was drastically increased in mice in response to elastase instillation, an experimental model of COPD. FCHSD1 is a member of the F-BAR family with two SH3 domains. We found that Fchsd1 knockout (Fchsd1-/-) mice were protected against airspace enlargement induced by elastase. Elastase-instilled lungs of Fchsd1-/- mice showed reduced inflammation and apoptosis compared with WT mice. We also found that elastase-induced reduction of Sirtuin 1 (SIRT1) levels, a histone deacetylase reported to protect against emphysema, was attenuated in the lungs of Fchsd1-/- mice. Furthermore, FCHSD1 deficiency enhanced nuclear translocation of nuclear factor-like 2 (NRF2), a redox-sensitive transcription factor, following H2O2 stimulation. Conversely, Fchsd1 overexpression inhibited NRF2 nuclear translocation and increased the reduction of SIRT1 levels. Notably, FCHSD1 interacted with NRF2 and SNX9. Our results show that FCHSD1 forms a multicomplex with NRF2 and SNX9 in the cytosol that prevents NRF2 from translocating to the nucleus. We propose that FCHSD1 promotes initiation of emphysema development by inhibiting nuclear translocation of NRF2, which leads to down-regulation of SIRT1.
Asunto(s)
Proteínas de la Membrana/genética , Enfermedad Pulmonar Obstructiva Crónica/patología , Transporte Activo de Núcleo Celular/efectos de los fármacos , Animales , Apoptosis , Muerte Celular , Núcleo Celular/efectos de los fármacos , Núcleo Celular/metabolismo , Peróxido de Hidrógeno/toxicidad , Carioferinas , Pulmón/patología , Masculino , Ratones , Ratones Noqueados , Factor 2 Relacionado con NF-E2/metabolismo , Estrés Oxidativo/efectos de los fármacos , Elastasa Pancreática , Neumonía/complicaciones , Neumonía/patología , Unión Proteica/efectos de los fármacos , Enfermedad Pulmonar Obstructiva Crónica/complicaciones , Enfisema Pulmonar/prevención & control , Sirtuina 1/metabolismo , Nexinas de Clasificación/metabolismoRESUMEN
Schlafen-8 (Slfn8) is a member of the Schlafen family of proteins, which harbor helicase domains and are induced by LPS and interferons. It has been reported that the Schlafen family are involved in various cellular functions, including proliferation, differentiation and regulation of virus replication. Slfn8 has been implicated in T-cell differentiation in the thymus. However, the roles of Slfn8 in the immune system remains unclear. In this study, we generated Slfn8 knockout mice (Slfn8-/-) and investigated the immunological role of Slfn8 using the T-cell-mediated autoimmune model experimental autoimmune encephalomyelitis (EAE). We found that the clinical score was reduced in Slfn8-/- mice. IL-6 and IL-17A cytokine production, which are associated with EAE onset and progression, were decreased in the lymph nodes of Slfn8-/- mice. Immune cell populations in Slfn8-/- mice, including macrophages, neutrophils, T cells and B cells, did not reveal significant differences compared with wild-type mice. In vitro activation of Slfn8-/- T cells in response to TCR stimulation also did not reveal significant differences. To confirm the involvement of non-hematopoietic cells, we isolated CD45- CD31+ endothelial cells and CD45-CD31- gp38+ fibroblastic reticular cells by FACS sorting. We showed that the levels of IL-6 and Slfn8 mRNA in CD45- CD31+ endothelial cells were increased after EAE induction. In contrast, the level of IL-6 mRNA after EAE induction was markedly decreased in CD31+ endothelial cells from Slfn8-/- mice. These results indicate that Slfn8 may play a role in EAE by regulating inflammation in endothelial cells.
Asunto(s)
Encefalomielitis Autoinmune Experimental/genética , Encefalomielitis Autoinmune Experimental/inmunología , Células Endoteliales/metabolismo , Animales , Biomarcadores , Citocinas/metabolismo , Encefalomielitis Autoinmune Experimental/metabolismo , Encefalomielitis Autoinmune Experimental/patología , Inmunidad Innata , Mediadores de Inflamación/metabolismo , Macrófagos/inmunología , Macrófagos/metabolismo , Ratones , Ratones Noqueados , Glicoproteína Mielina-Oligodendrócito/efectos adversos , Fragmentos de Péptidos/efectos adversos , Índice de Severidad de la Enfermedad , Linfocitos T/inmunología , Linfocitos T/metabolismoRESUMEN
The development of effective treatments against cancers is urgently needed, and the accumulation of CD8+ T cells within tumors is especially important for cancer prognosis. Although their mechanisms are still largely unknown, growing evidence has indicated that innate immune cells have important effects on cancer progression through the production of various cytokines. Here, we found that basic leucine zipper transcription factor ATF-like 2 (Batf2) has an antitumor effect. An s.c. inoculated tumor model produced fewer IL-12 p40+ macrophages and activated CD8+ T cells within the tumors of Batf2-/- mice compared with WT mice. In vitro studies also revealed that the IL-12 p40 expression was significantly lower in Batf2-/- macrophages following their stimulation by toll-like receptor ligands, such as R848. Additionally, we found that BATF2 interacts with p50/p65 and promotes IL-12 p40 expression. In conclusion, Batf2 has an antitumor effect through the up-regulation of IL-12 p40 in tumor-associated macrophages, which eventually induces CD8+ T-cell activation and accumulation within the tumor.
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Factores de Transcripción con Cremalleras de Leucina de Carácter Básico/fisiología , Proteínas Supresoras de Tumor/fisiología , Animales , Antineoplásicos/metabolismo , Factores de Transcripción con Cremalleras de Leucina de Carácter Básico/genética , Factores de Transcripción con Cremalleras de Leucina de Carácter Básico/metabolismo , Factores de Transcripción con Cremalleras de Leucina de Carácter Básico/farmacología , Linfocitos T CD8-positivos/metabolismo , Regulación de la Expresión Génica/genética , Humanos , Interleucina-12/metabolismo , Interleucina-12/fisiología , Macrófagos/metabolismo , Glicoproteínas de Membrana/metabolismo , Ratones , Ratones Transgénicos , FN-kappa B/metabolismo , Subunidades de Proteína/metabolismo , Células RAW 264.7 , Receptor Toll-Like 7/metabolismo , Factores de Transcripción/metabolismo , Proteínas Supresoras de Tumor/genética , Proteínas Supresoras de Tumor/farmacología , Regulación hacia ArribaRESUMEN
The RIG-I-like receptors, retinoic acid inducible gene-1 (RIG-I), melanoma differentiation-associated protein 5, and laboratory of genetics and physiology-2, are cytoplasmic sensors for RNA viruses that mediate the antiviral innate immune responses. We demonstrate that really interesting new gene-finger domain- and K homology domain-containing MEX3C regulates RIG-I function. MEX3C colocalizes with RIG-I in the stress granules of virally infected cells, and its overexpression causes the lysine-63-linked ubiquitination of RIG-I and activates IFN-ß promoter. Embryonic fibroblast cells, macrophages, and conventional dendritic cells derived from Mex3c-deficient mice showed defective production of type I IFN after infection with RNA viruses that are recognized by RIG-I. These results demonstrate that MEX3C is an E3 ubiquitin ligase that modifies RIG-I in stress granules and plays a critical role in eliciting antiviral immune responses.
Asunto(s)
Inmunidad Innata/inmunología , Proteínas de Unión al ARN/inmunología , Receptores de Ácido Retinoico/inmunología , Virosis/inmunología , Animales , Citocinas/biosíntesis , Cartilla de ADN/genética , Ensayo de Inmunoadsorción Enzimática , Células HEK293 , Humanos , Immunoblotting , Inmunoprecipitación , Luciferasas , Ratones , Ratones Noqueados , Mutagénesis Sitio-Dirigida , Proteínas de Unión al ARN/genética , Proteínas de Unión al ARN/metabolismo , Reacción en Cadena en Tiempo Real de la Polimerasa , Receptores de Ácido Retinoico/metabolismo , UbiquitinaciónRESUMEN
Regnase-1 (also known as Zc3h12a and MCPIP1) is an RNase that destabilizes a set of mRNAs, including Il6 and Il12b, through cleavage of their 3' UTRs. Although Regnase-1 inactivation leads to development of an autoimmune disease characterized by T cell activation and hyperimmunoglobulinemia in mice, the mechanism of Regnase-1-mediated immune regulation has remained unclear. We show that Regnase-1 is essential for preventing aberrant effector CD4(+) T cell generation cell autonomously. Moreover, in T cells, Regnase-1 regulates the mRNAs of a set of genes, including c-Rel, Ox40, and Il2, through cleavage of their 3' UTRs. Interestingly, T cell receptor (TCR) stimulation leads to cleavage of Regnase-1 at R111 by Malt1/paracaspase, freeing T cells from Regnase-1-mediated suppression. Furthermore, Malt1 protease activity is critical for controlling the mRNA stability of T cell effector genes. Collectively, these results indicate that dynamic control of Regnase-1 expression in T cells is critical for controlling T cell activation.
Asunto(s)
Caspasas/metabolismo , Activación de Linfocitos , Proteínas de Neoplasias/metabolismo , Ribonucleasas/metabolismo , Linfocitos T Colaboradores-Inductores/inmunología , Animales , Enfermedades Autoinmunes/inmunología , Humanos , Interleucina-2/genética , Células Jurkat , Glicoproteínas de Membrana/genética , Ratones , Proteína 1 de la Translocación del Linfoma del Tejido Linfático Asociado a Mucosas , Ligando OX40 , Proteínas Proto-Oncogénicas c-rel/genética , Receptores de Antígenos de Linfocitos T/metabolismo , Linfocitos T Colaboradores-Inductores/metabolismo , Factores de Necrosis Tumoral/genéticaRESUMEN
Toll-like receptor (TLR) signaling activates the inhibitor of transcription factor NF-κB (IκB) kinase (IKK) complex, which governs NF-κB-mediated transcription during inflammation. The RNase regnase-1 serves a critical role in preventing autoimmunity by controlling the stability of mRNAs that encode cytokines. Here we show that the IKK complex controlled the stability of mRNA for interleukin 6 (IL-6) by phosphorylating regnase-1 in response to stimulation via the IL-1 receptor (IL-1R) or TLR. Phosphorylated regnase-1 underwent ubiquitination and degradation. Regnase-1 was reexpressed in IL-1R- or TLR-activated cells after a period of lower expression. Regnase-1 mRNA was negatively regulated by regnase-1 itself via a stem-loop region present in the regnase-1 3' untranslated region. Our data demonstrate that the IKK complex phosphorylates not only IκBα, thereby activating transcription, but also regnase-1, thereby releasing a 'brake' on IL-6 mRNA expression.