Your browser doesn't support javascript.
loading
Mostrar: 20 | 50 | 100
Resultados 1 - 20 de 384
Filtrar
1.
Cell Chem Biol ; 29(1): 5-18.e6, 2022 01 20.
Artículo en Inglés | MEDLINE | ID: mdl-34672954

RESUMEN

The global epidemic caused by the coronavirus severe acute respiratory syndrome coronavirus-2 (SARS-CoV-2) has resulted in the infection of over 200 million people. To extend the knowledge of interactions between SARS-CoV-2 and humans, we systematically investigate the interactome of 29 viral proteins in human cells by using an antibody-based TurboID assay. In total, 1,388 high-confidence human proximal proteins with biotinylated sites are identified. Notably, we find that SARS-CoV-2 manipulates the antiviral and immune responses. We validate that the membrane protein ITGB1 associates angiotensin-converting enzyme 2 (ACE2) to mediate SARS-CoV-2 entry. Moreover, we reveal that SARS-CoV-2 proteins inhibit activation of the interferon pathway through the mitochondrial protein mitochondrial antiviral-signaling protein (MAVS) and the methyltransferase SET domain containing 2, histone lysine methyltransferase (SETD2). We propose 111 potential drugs for the clinical treatment of coronavirus disease 2019 (COVID-19) and identify three compounds that significantly inhibit the replication of SARS-CoV-2. The proximity labeling map of SARS-CoV-2 and humans provides a resource for elucidating the mechanisms of viral infection and developing drugs for COVID-19 treatment.


Asunto(s)
Anticuerpos/inmunología , Antivirales/inmunología , SARS-CoV-2/inmunología , Enzima Convertidora de Angiotensina 2/inmunología , Antivirales/farmacología , COVID-19/inmunología , Humanos , Integrina beta1/inmunología , Pruebas de Sensibilidad Microbiana , Tratamiento Farmacológico de COVID-19
2.
Int J Mol Sci ; 22(11)2021 May 30.
Artículo en Inglés | MEDLINE | ID: mdl-34070747

RESUMEN

Colorectal cancer (CRC) is one of the main causes of cancer death in the world. Post-translational modifications (PTMs) have been extensively studied in malignancies due to its relevance in tumor pathogenesis and therapy. This review is focused on the dysregulation of glycosyltransferase expression in CRC and its impact in cell function and in several biological pathways associated with CRC pathogenesis, prognosis and therapeutic approaches. Glycan structures act as interface molecules between cells and their environment and in several cases facilitate molecule function. CRC tissue shows alterations in glycan structures decorating molecules, such as annexin-1, mucins, heat shock protein 90 (Hsp90), ß1 integrin, carcinoembryonic antigen (CEA), epidermal growth factor receptor (EGFR), insulin-like growth factor-binding protein 3 (IGFBP3), transforming growth factor beta (TGF-ß) receptors, Fas (CD95), PD-L1, decorin, sorbin and SH3 domain-containing protein 1 (SORBS1), CD147 and glycosphingolipids. All of these are described as key molecules in oncogenesis and metastasis. Therefore, glycosylation in CRC can affect cell migration, cell-cell adhesion, actin polymerization, mitosis, cell membrane repair, apoptosis, cell differentiation, stemness regulation, intestinal mucosal barrier integrity, immune system regulation, T cell polarization and gut microbiota composition; all such functions are associated with the prognosis and evolution of the disease. According to these findings, multiple strategies have been evaluated to alter oligosaccharide processing and to modify glycoconjugate structures in order to control CRC progression and prevent metastasis. Additionally, immunotherapy approaches have contemplated the use of neo-antigens, generated by altered glycosylation, as targets for tumor-specific T cells or engineered CAR (Chimeric antigen receptors) T cells.


Asunto(s)
Neoplasias Colorrectales/genética , Glicoesfingolípidos/inmunología , Glicosiltransferasas/genética , Mucinas/genética , Proteínas de Neoplasias/genética , Procesamiento Proteico-Postraduccional , Anexina A1/genética , Anexina A1/inmunología , Neoplasias Colorrectales/inmunología , Neoplasias Colorrectales/patología , Neoplasias Colorrectales/terapia , Decorina/genética , Decorina/inmunología , Receptores ErbB/genética , Receptores ErbB/inmunología , Regulación Neoplásica de la Expresión Génica , Glicoesfingolípidos/metabolismo , Glicosilación , Glicosiltransferasas/inmunología , Humanos , Inmunoterapia Adoptiva/métodos , Proteína 3 de Unión a Factor de Crecimiento Similar a la Insulina/genética , Proteína 3 de Unión a Factor de Crecimiento Similar a la Insulina/inmunología , Integrina beta1/genética , Integrina beta1/inmunología , Proteínas de Microfilamentos/genética , Proteínas de Microfilamentos/inmunología , Mucinas/inmunología , Proteínas de Neoplasias/inmunología , Receptor fas/genética , Receptor fas/inmunología
3.
Int J Biochem Cell Biol ; 131: 105905, 2021 02.
Artículo en Inglés | MEDLINE | ID: mdl-33359084

RESUMEN

Beyond its well-known canonical function as a tumor suppressor, p53 is also involved in numerous cellular processes through altered transcription under both normal and pathological conditions. The functional diversity of p53 outputs is complex and dependent on cell context. However, the underlying mechanisms responsible for this diversity remain largely unclear. The emerging evidence of p53 mutations involved in regulating endocytic trafficking and signaling, in tandem to promote malignancy (invasion, exosome biogenesis and immune evasion), sheds light on possible mechanisms behind the p53-driven complexity. The interrelated nature of endocytic trafficking and receptor signaling that form dynamic and adaptable feedback loops - either positive or negative - functions to modulate multiple cellular outputs. Biasing the tunable endocytic trafficking and receptor signaling network by mutant p53 expands the purview of p53, allowing its contribution to diverse and aggressive phenotypes. In this review, we explore recent studies in which the novel role of mutant p53 in altering endocytic trafficking to bias receptor signaling and drive transforming phenotypes is revealed. Understanding the complex crosstalk of mutant p53, endocytic trafficking and receptor signaling will allow the development of therapies to selectively target p53-altered endocytic processes.


Asunto(s)
Endocitosis/genética , Mutación con Ganancia de Función , Integrina beta1/genética , Neoplasias Pulmonares/genética , Proteína p53 Supresora de Tumor/genética , Ciclo Celular/genética , Línea Celular Tumoral , Movimiento Celular , Proliferación Celular , ARN Helicasas DEAD-box/genética , ARN Helicasas DEAD-box/inmunología , Endosomas/genética , Endosomas/metabolismo , Receptores ErbB/genética , Receptores ErbB/inmunología , Regulación Neoplásica de la Expresión Génica , Humanos , Integrina beta1/inmunología , Neoplasias Pulmonares/inmunología , Neoplasias Pulmonares/patología , Ribonucleasa III/genética , Ribonucleasa III/inmunología , Transducción de Señal , Escape del Tumor , Proteína p53 Supresora de Tumor/inmunología
4.
Front Immunol ; 11: 599936, 2020.
Artículo en Inglés | MEDLINE | ID: mdl-33381120

RESUMEN

Multiple sclerosis is a chronic auto-inflammatory disease of the central nervous system affecting patients worldwide. Neuroinflammation in multiple sclerosis is mainly driven by peripheral immune cells which invade the central nervous system and cause neurodegenerative inflammation. To enter the target tissue, immune cells have to overcome the endothelium and transmigrate into the tissue. Numerous molecules mediate this process and, as they determine the tissue invasiveness of immune cells, display great therapeutic potential. Melanoma cell adhesion molecule (MCAM) is a membrane-anchored glycoprotein expressed by a subset of T-cells and MCAM+ T-cells have been shown to contribute to neuroinflammation in multiple sclerosis. The role of the MCAM molecule for brain invasion, however, remained largely unknown. In order to investigate the role of the MCAM molecule on T-cells, we used different in vitro and in vivo assays, including ex vivo flow chambers, biochemistry and microscopy experiments of the mouse brain. We demonstrate that MCAM directly mediates adhesion and that the engagement of MCAM induces intracellular signaling leading to ß1-integrin activation on human T-cells. Furthermore, we show that MCAM engagement triggers the phosphorylation of PLCγ1 which is required for integrin activation and thus amplification of the cellular adhesive potential. To confirm the physiological relevance of our findings in vivo, we demonstrate that MCAM plays an important role in T-cell recruitment into the mouse brain. In conclusion, our data demonstrate that MCAM expressed on T-cells acts as an adhesion molecule and a signaling receptor that may trigger ß1-integrin activation via PLCγ1 upon engagement.


Asunto(s)
Encéfalo/inmunología , Memoria Inmunológica , Integrina beta1/inmunología , Esclerosis Múltiple/inmunología , Fosfolipasa C gamma/inmunología , Linfocitos T/inmunología , Animales , Encéfalo/patología , Antígeno CD146/inmunología , Modelos Animales de Enfermedad , Femenino , Humanos , Masculino , Ratones , Esclerosis Múltiple/patología , Linfocitos T/patología
5.
J Virol ; 94(21)2020 10 14.
Artículo en Inglés | MEDLINE | ID: mdl-32817220

RESUMEN

Lassa fever (LF) is a zoonotic viral hemorrhagic fever caused by Lassa virus (LASV), which is endemic to West African countries. Previous studies have suggested an important role for T-cell-mediated immunopathology in LF pathogenesis, but the mechanisms by which T cells influence disease severity and outcome are not well understood. Here, we present a multiparametric analysis of clinical immunology data collected during the 2017-2018 Lassa fever outbreak in Nigeria. During the acute phase of LF, we observed robust activation of the polyclonal T-cell repertoire, which included LASV-specific and antigenically unrelated T cells. However, severe and fatal LF cases were characterized by poor LASV-specific effector T-cell responses. Severe LF was also characterized by the presence of circulating T cells with homing capacity to inflamed tissues, including the gut mucosa. These findings in LF patients were recapitulated in a mouse model of LASV infection, in which mucosal exposure resulted in remarkably high lethality compared to skin exposure. Taken together, our findings indicate that poor LASV-specific T-cell responses and activation of nonspecific T cells with homing capacity to inflamed tissues are associated with severe LF.IMPORTANCE Lassa fever may cause severe disease in humans, in particular in areas of endemicity like Sierra Leone and Nigeria. Despite its public health importance, the pathophysiology of Lassa fever in humans is poorly understood. Here, we present clinical immunology data obtained in the field during the 2018 Lassa fever outbreak in Nigeria indicating that severe Lassa fever is associated with activation of T cells antigenically unrelated to Lassa virus and poor Lassa virus-specific effector T-cell responses. Mechanistically, we show that these bystander T cells express defined tissue homing signatures that suggest their recruitment to inflamed tissues and a putative role of these T cells in immunopathology. These findings open a window of opportunity to consider T-cell targeting as a potential postexposure therapeutic strategy against severe Lassa fever, a hypothesis that could be tested in relevant animal models, such as nonhuman primates.


Asunto(s)
Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD8-positivos/inmunología , Brotes de Enfermedades , Mucosa Intestinal/inmunología , Fiebre de Lassa/inmunología , Virus Lassa/patogenicidad , Activación de Linfocitos , Adolescente , Adulto , Anciano , Animales , Linfocitos T CD4-Positivos/patología , Linfocitos T CD4-Positivos/virología , Linfocitos T CD8-positivos/patología , Linfocitos T CD8-positivos/virología , Niño , Preescolar , Femenino , Regulación de la Expresión Génica , Antígenos HLA-DR/genética , Antígenos HLA-DR/inmunología , Humanos , Lactante , Recién Nacido , Integrina beta1/genética , Integrina beta1/inmunología , Interferón gamma/genética , Interferón gamma/inmunología , Mucosa Intestinal/patología , Mucosa Intestinal/virología , Fiebre de Lassa/genética , Fiebre de Lassa/mortalidad , Fiebre de Lassa/virología , Virus Lassa/crecimiento & desarrollo , Virus Lassa/inmunología , Proteína 1 de la Membrana Asociada a los Lisosomas/genética , Proteína 1 de la Membrana Asociada a los Lisosomas/inmunología , Masculino , Ratones , Persona de Mediana Edad , Nigeria/epidemiología , Estudios Retrospectivos , Índice de Severidad de la Enfermedad , Piel/inmunología , Piel/patología , Piel/virología , Análisis de Supervivencia , Factor de Necrosis Tumoral alfa/genética , Factor de Necrosis Tumoral alfa/inmunología
6.
Cancer Invest ; 38(4): 228-239, 2020 Apr.
Artículo en Inglés | MEDLINE | ID: mdl-32208057

RESUMEN

The aim of this study was to characterize both by flow cytometry analysis and immunohistochemistry cervix uteri cells of nulliparous women screened for cervical intraepithelial neoplasia (CIN) in comparison to a group without CIN by using mesenchymal stem cell-like and hematopoietic lineage markers. A significant expression for CD29, CD38, HLA-I, and HLA-II was correlated positively to the CIN degree and it was more relevant in patients positive for human papilloma virus (HPV). Thus, identification and detailed characterization of pluripotent resident in uteri cells could be a promising therapeutic target.


Asunto(s)
Cuello del Útero/citología , Células Madre Neoplásicas/patología , Infecciones por Papillomavirus/patología , Displasia del Cuello del Útero/patología , Neoplasias del Cuello Uterino/patología , ADP-Ribosil Ciclasa 1/análisis , ADP-Ribosil Ciclasa 1/inmunología , ADP-Ribosil Ciclasa 1/metabolismo , Adulto , Biopsia , Cuello del Útero/inmunología , Cuello del Útero/patología , Cuello del Útero/virología , Inhibidor p16 de la Quinasa Dependiente de Ciclina/análisis , Inhibidor p16 de la Quinasa Dependiente de Ciclina/inmunología , Inhibidor p16 de la Quinasa Dependiente de Ciclina/metabolismo , Femenino , Citometría de Flujo , Antígenos de Histocompatibilidad Clase I/análisis , Antígenos de Histocompatibilidad Clase I/inmunología , Antígenos de Histocompatibilidad Clase I/metabolismo , Antígenos de Histocompatibilidad Clase II/análisis , Antígenos de Histocompatibilidad Clase II/inmunología , Antígenos de Histocompatibilidad Clase II/metabolismo , Humanos , Inmunohistoquímica , Inmunofenotipificación , Integrina beta1/análisis , Integrina beta1/inmunología , Integrina beta1/metabolismo , Glicoproteínas de Membrana/análisis , Glicoproteínas de Membrana/inmunología , Glicoproteínas de Membrana/metabolismo , Clasificación del Tumor , Células Madre Neoplásicas/inmunología , Células Madre Neoplásicas/virología , Papillomaviridae/inmunología , Papillomaviridae/aislamiento & purificación , Infecciones por Papillomavirus/inmunología , Infecciones por Papillomavirus/virología , Estudios Prospectivos , Neoplasias del Cuello Uterino/inmunología , Neoplasias del Cuello Uterino/prevención & control , Neoplasias del Cuello Uterino/virología , Displasia del Cuello del Útero/inmunología , Displasia del Cuello del Útero/virología
7.
J Clin Invest ; 130(2): 715-732, 2020 02 03.
Artículo en Inglés | MEDLINE | ID: mdl-31661467

RESUMEN

Although the impact of Th17 cells on autoimmunity is undisputable, their pathogenic effector mechanism is still enigmatic. We discovered soluble N-ethylmaleimide-sensitive factor attachment receptor (SNARE) complex proteins in Th17 cells that enable a vesicular glutamate release pathway that induces local intracytoplasmic calcium release and subsequent damage in neurons. This pathway is glutamine dependent and triggered by binding of ß1-integrin to vascular cell adhesion molecule 1 (VCAM-1) on neurons in the inflammatory context. Glutamate secretion could be blocked by inhibiting either glutaminase or KV1.3 channels, which are known to be linked to integrin expression and highly expressed on stimulated T cells. Although KV1.3 is not expressed in CNS tissue, intrathecal administration of a KV1.3 channel blocker or a glutaminase inhibitor ameliorated disability in experimental neuroinflammation. In humans, T cells from patients with multiple sclerosis secreted higher levels of glutamate, and cerebrospinal fluid glutamine levels were increased. Altogether, our findings demonstrate that ß1-integrin- and KV1.3 channel-dependent signaling stimulates glutamate release from Th17 cells upon direct cell-cell contact between Th17 cells and neurons.


Asunto(s)
Integrina beta1/inmunología , Canal de Potasio Kv1.3/inmunología , Esclerosis Múltiple/inmunología , Transducción de Señal/inmunología , Células Th17/inmunología , Animales , Comunicación Celular/genética , Comunicación Celular/inmunología , Ácido Glutámico/genética , Ácido Glutámico/inmunología , Humanos , Integrina beta1/genética , Canal de Potasio Kv1.3/genética , Ratones , Ratones Noqueados , Esclerosis Múltiple/genética , Esclerosis Múltiple/patología , Proteínas SNARE/genética , Proteínas SNARE/inmunología , Transducción de Señal/genética , Células Th17/patología , Molécula 1 de Adhesión Celular Vascular/genética , Molécula 1 de Adhesión Celular Vascular/inmunología
8.
Bull Exp Biol Med ; 167(5): 653-655, 2019 Sep.
Artículo en Inglés | MEDLINE | ID: mdl-31641985

RESUMEN

Expression of CD11, CD29, CD36, and DC-STAMP molecules by macrophages was analyzed in in vitro experiments. These molecules mediate cell fusion, one of the mechanisms underlying the formation of multinuclear macrophages. Macrophages were obtained from intact and BCG-infected male BALB/c mice. In intact cultures, multinuclear macrophages appeared primarily due to amitotic division of cell nuclei, while in macrophage cultures from infected mice, the process of cell fusion predominated. In intact macrophage cultures, bi- and multinuclear cells expressed primarily CD29 and CD36. In cultures from infected mice, macrophages expressing CD29 and DC-STAMP predominated, but bi- and multinuclear macrophages expressing CD11 and CD36 predominated over mononuclear ones. The study of macrophage fusion mechanism can be useful for understanding of this biological phenomenon as the mechanisms of delivery of M. tuberculosis and lysosomotropic anti-tuberculosis drugs into tuberculous granulomas to suppress M. tuberculosis persisting in macrophages and reduce the destructive potential of granulomas.


Asunto(s)
Vacuna BCG/administración & dosificación , Antígenos CD11/genética , Antígenos CD36/genética , Integrina beta1/genética , Macrófagos/microbiología , Proteínas de la Membrana/genética , Proteínas del Tejido Nervioso/genética , Animales , Antígenos CD11/inmunología , Antígenos CD36/inmunología , Fusión Celular , Núcleo Celular/ultraestructura , Expresión Génica , Integrina beta1/inmunología , Macrófagos/inmunología , Macrófagos/patología , Masculino , Proteínas de la Membrana/inmunología , Ratones , Ratones Endogámicos BALB C , Proteínas del Tejido Nervioso/inmunología , Cultivo Primario de Células
9.
J Hepatol ; 71(6): 1193-1205, 2019 12.
Artículo en Inglés | MEDLINE | ID: mdl-31433301

RESUMEN

BACKGROUND & AIMS: Hepatic recruitment of monocyte-derived macrophages (MoMFs) contributes to the inflammatory response in non-alcoholic steatohepatitis (NASH). However, how hepatocyte lipotoxicity promotes MoMF inflammation is unclear. Here we demonstrate that lipotoxic hepatocyte-derived extracellular vesicles (LPC-EVs) are enriched with active integrin ß1 (ITGß1), which promotes monocyte adhesion and liver inflammation in murine NASH. METHODS: Hepatocytes were treated with either vehicle or the toxic lipid mediator lysophosphatidylcholine (LPC); EVs were isolated from the conditioned media and subjected to proteomic analysis. C57BL/6J mice were fed a diet rich in fat, fructose, and cholesterol (FFC) to induce NASH. Mice were treated with anti-ITGß1 neutralizing antibody (ITGß1Ab) or control IgG isotype. RESULTS: Ingenuity® Pathway Analysis of the LPC-EV proteome indicated that ITG signaling is an overrepresented canonical pathway. Immunogold electron microscopy and nanoscale flow cytometry confirmed that LPC-EVs were enriched with activated ITGß1. Furthermore, we showed that LPC treatment in hepatocytes activates ITGß1 and mediates its endocytic trafficking and sorting into EVs. LPC-EVs enhanced monocyte adhesion to liver sinusoidal cells, as observed by shear stress adhesion assay. This adhesion was attenuated in the presence of ITGß1Ab. FFC-fed, ITGß1Ab-treated mice displayed reduced inflammation, defined by decreased hepatic infiltration and activation of proinflammatory MoMFs, as assessed by immunohistochemistry, mRNA expression, and flow cytometry. Likewise, mass cytometry by time-of-flight on intrahepatic leukocytes showed that ITGß1Ab reduced levels of infiltrating proinflammatory monocytes. Furthermore, ITGß1Ab treatment significantly ameliorated liver injury and fibrosis. CONCLUSIONS: Lipotoxic EVs mediate monocyte adhesion to LSECs mainly through an ITGß1-dependent mechanism. ITGß1Ab ameliorates diet-induced NASH in mice by reducing MoMF-driven inflammation, suggesting that blocking ITGß1 is a potential anti-inflammatory therapeutic strategy in human NASH. LAY SUMMARY: Herein, we report that a cell adhesion molecule termed integrin ß1 (ITGß1) plays a key role in the progression of non-alcoholic steatohepatitis (NASH). ITGß1 is released from hepatocytes under lipotoxic stress as a cargo of extracellular vesicles, and mediates monocyte adhesion to liver sinusoidal endothelial cells, which is an essential step in hepatic inflammation. In a mouse model of NASH, blocking ITGß1 reduces liver inflammation, injury and fibrosis. Hence, ITGß1 inhibition may serve as a new therapeutic strategy for NASH.


Asunto(s)
Anticuerpos Neutralizantes , Adhesión Celular/inmunología , Hepatocitos/inmunología , Integrina beta1/inmunología , Lisofosfatidilcolinas/farmacología , Macrófagos/inmunología , Enfermedad del Hígado Graso no Alcohólico/inmunología , Animales , Anticuerpos Neutralizantes/administración & dosificación , Anticuerpos Neutralizantes/inmunología , Dieta Alta en Grasa , Modelos Animales de Enfermedad , Vesículas Extracelulares/inmunología , Hepatocitos/metabolismo , Humanos , Cirrosis Hepática/prevención & control , Ratones , Monocitos/inmunología , Enfermedad del Hígado Graso no Alcohólico/terapia
10.
Eur J Immunol ; 49(11): 2030-2043, 2019 11.
Artículo en Inglés | MEDLINE | ID: mdl-31318439

RESUMEN

Plasmacytoid dendritic cells (pDCs) are found in the CNS during neuroinflammation and have been reported to exert regulatory functions in multiple sclerosis (MS) and its animal model experimental autoimmune encephalomyelitis (EAE). However, the mechanisms of entry of pDCs into the CNS as well as their phenotype and innate functional properties, once recruited into the CNS, have not been thoroughly examined. Herein, we show that pDCs rapidly accumulate into the brain and spinal cord during the acute phase of EAE, and maintain the expression of numerous phenotypic markers typical of peripheral pDCs. Functionally, CNS-pDCs constitutively expressed IRF7 and were able to rapidly produce type I IFNs and IL-12p40 upon ex vivo TLR-9 stimulation. Using adoptive transfer experiments, we provide evidence that CNS-pDC are recruited from the blood and accumulate into the CNS during the acute phase of EAE. Accumulation of pDCs into the CNS was strongly inhibited in the absence of CD29, but not CD18, suggesting a major role for ß1 but not ß2 integrins. Indeed, blocking the CD49d α4-integrins during acute EAE drastically diminished CNS-pDC numbers. Together, our results demonstrate that circulating pDCs are actively recruited into the CNS during acute EAE through a mechanism largely dependent on CD49d/CD29-integrins.


Asunto(s)
Encéfalo/inmunología , Células Dendríticas/inmunología , Encefalomielitis Autoinmune Experimental/inmunología , Integrina alfa4/inmunología , Integrina beta1/inmunología , Médula Espinal/inmunología , Traslado Adoptivo , Animales , Encéfalo/efectos de los fármacos , Encéfalo/patología , Movimiento Celular/inmunología , Células Dendríticas/efectos de los fármacos , Células Dendríticas/patología , Células Dendríticas/trasplante , Encefalomielitis Autoinmune Experimental/inducido químicamente , Encefalomielitis Autoinmune Experimental/genética , Encefalomielitis Autoinmune Experimental/patología , Femenino , Regulación de la Expresión Génica , Inmunidad Innata , Integrina alfa4/genética , Integrina beta1/genética , Factor 7 Regulador del Interferón/genética , Factor 7 Regulador del Interferón/inmunología , Interferón Tipo I/genética , Interferón Tipo I/inmunología , Subunidad p40 de la Interleucina-12/genética , Subunidad p40 de la Interleucina-12/inmunología , Ratones , Ratones Endogámicos C57BL , Glicoproteína Mielina-Oligodendrócito/administración & dosificación , Fragmentos de Péptidos/administración & dosificación , Toxina del Pertussis/administración & dosificación , Transducción de Señal , Médula Espinal/efectos de los fármacos , Médula Espinal/patología , Receptor Toll-Like 9/genética , Receptor Toll-Like 9/inmunología
11.
J Immunol ; 202(9): 2782-2794, 2019 05 01.
Artículo en Inglés | MEDLINE | ID: mdl-30894426

RESUMEN

Abs exert several of their effector functions by binding to cell surface receptors. For murine IgG3 (mIgG3), the identity of its receptors (and the very existence of a receptor) is still under debate, as not all mIgG3 functions can be explained by interaction with FcγRI. This implies the existence of an alternate receptor, whose identity we sought to pinpoint. We found that blockage of integrin ß1 selectively hampered binding of mIgG3 to macrophages and mIgG3-mediated phagocytosis. Manganese, an integrin activator, increased mIgG3 binding to macrophages. Blockage of FcγRI or Itgb1 inhibited binding of different mIgG3 Abs to variable extents. Our results are consistent with the notion that Itgb1 functions as part of an IgG receptor complex. Given the more ancient origin of integrins in comparison with FcγR, this observation could have far-ranging implications for our understanding of the evolution of Ab-mediated immunity as well as in immunity to microorganisms, pathogenesis of autoimmune diseases, and Ab engineering.


Asunto(s)
Inmunoglobulina G/inmunología , Integrina beta1/inmunología , Macrófagos/inmunología , Fagocitosis , Receptores de IgG/inmunología , Animales , Inmunoglobulina G/genética , Integrina beta1/genética , Ratones , Ratones Noqueados , Receptores de IgG/genética
12.
Mol Cancer Ther ; 18(6): 1127-1136, 2019 06.
Artículo en Inglés | MEDLINE | ID: mdl-30926634

RESUMEN

Integrin ß1 receptor, expressed on the surface of tumor cells and macrophages in the tumor microenvironment (TME), has been implicated in both tumor progression and resistance to multiple modalities of therapy. OS2966 is the first clinical-ready humanized monoclonal antibody to block integrin ß1 and was recently orphan designated by the FDA Office of Orphan Products Development. Here, we tested therapeutic potential of OS2966-mediated integrin ß1 blockade to enhance the efficacy of oncolytic herpes simplex virus-1 (oHSV) through evaluation of virus replication, tumor cell killing efficiency, effect on the antiviral signaling pathway, co-culture assays of oHSV-infected cells with macrophages, and in vivo bioluminescence imaging on mammary fat pad triple-negative breast cancer xenograft and subcutaneous and intracranial glioma xenografts. OS2966 treatment decreased interferon signaling and proinflammatory cytokine induction in oHSV-treated tumor cells and inhibited migration of macrophages, resulting in enhanced oHSV replication and cytotoxicity. OS2966 treatment also significantly enhanced oHSV replication and oHSV-mediated antitumor efficacy in orthotopic xenograft models, including triple-negative breast cancer and glioblastoma. The results demonstrated the synergistic potential of the combinatory treatment approach with OS2966 to improve antitumor efficacy of conventional oHSV therapy.


Asunto(s)
Anticuerpos Bloqueadores/uso terapéutico , Herpesvirus Humano 1/fisiología , Integrina beta1/inmunología , Viroterapia Oncolítica/métodos , Virus Oncolíticos/fisiología , Animales , Neoplasias Encefálicas/metabolismo , Neoplasias Encefálicas/patología , Neoplasias Encefálicas/terapia , Neoplasias de la Mama/metabolismo , Neoplasias de la Mama/patología , Neoplasias de la Mama/terapia , Línea Celular Tumoral , Movimiento Celular/inmunología , Técnicas de Cocultivo , Terapia Combinada/métodos , Femenino , Glioma/metabolismo , Glioma/patología , Glioma/terapia , Humanos , Macrófagos/metabolismo , Ratones , Ratones Desnudos , Células RAW 264.7 , Replicación Viral/inmunología , Ensayos Antitumor por Modelo de Xenoinjerto
13.
Sci Rep ; 9(1): 1283, 2019 02 04.
Artículo en Inglés | MEDLINE | ID: mdl-30718722

RESUMEN

The importance of neutrophils in the pathogenesis of autoimmune rheumatic diseases, such as systemic lupus erythematosus (SLE) and rheumatoid arthritis (RA), is increasingly recognised. Generation of reactive oxygen species (ROS) and release of neutrophil extracellular traps (NETs) by activated neutrophils are both thought to contribute to pathology; although the underlying mechanisms, particularly the effects of IgG autoantibodies upon neutrophil function, are not fully understood. Therefore, we determined whether purified IgG from patients with SLE or RA have differential effects upon neutrophil activation and function. We found that SLE- and RA-IgG both bound human neutrophils but differentially regulated neutrophil function. RA- and SLE-IgG both increased PMA-induced ß1 integrin-mediated adhesion to fibronectin, whilst only SLE-IgG enhanced αMß2 integrin-mediated adhesion to fibrinogen. Interestingly, only SLE-IgG modulated neutrophil adhesion to endothelial cells. Both SLE- and RA-IgG increased ROS generation and DNA externalisation by unstimulated neutrophils. Only SLE-IgG however, drove DNA externalisation following neutrophil activation. Co-culture of neutrophils with resting endothelium prevented IgG-mediated increase of extracellular DNA, but this inhibition was overcome for SLE-IgG when the endothelium was stimulated with TNF-α. This differential pattern of neutrophil activation has implications for understanding SLE and RA pathogenesis and may highlight avenues for development of novel therapeutic strategies.


Asunto(s)
Artritis Reumatoide/inmunología , Autoanticuerpos/inmunología , Células Endoteliales de la Vena Umbilical Humana/inmunología , Inmunoglobulina G/inmunología , Integrina beta1/inmunología , Lupus Eritematoso Sistémico/inmunología , Activación Neutrófila , Neutrófilos/inmunología , Artritis Reumatoide/patología , Células Endoteliales de la Vena Umbilical Humana/patología , Humanos , Lupus Eritematoso Sistémico/patología , Neutrófilos/patología
14.
Mol Med Rep ; 19(3): 1739-1746, 2019 Mar.
Artículo en Inglés | MEDLINE | ID: mdl-30628700

RESUMEN

The present study aimed to evaluate the effects of Krüppel­like factor 2 (KLF2) on the differentiation of endothelial progenitor cells (EPCs) to endothelial cells (ECs) induced by shear stress, and to investigate the corresponding mechanisms. Cultured rat late EPCs were exposed to shear stress (12 dyn/cm2) for different lengths of time. Reverse transcription­quantitative polymerase chain reaction (RT­qPCR) was used to measure the initial KLF2 mRNA levels in each group. Subsequently, the EPCs were treated with anti­integrin ß1 or ß3 antibodies to block integrin ß1 and ß3, respectively, or cytochalasin D to destroy F­actin, and the subsequent expression levels of KLF2 in EPCs were measured. Then, KLF2 small interfering RNAs (siRNAs) were transfected into EPCs, and RT­qPCR was used to measure the mRNA expression level of KLF2. Additionally, flow cytometry was applied to evaluate the protein levels of cluster of differentiation 31 (CD31) and the von Willebrand factor (vWF), and the regulatory effects of KLF2 in the promoter region of vWF were determined via a luciferase assay. High shear stress upregulated KLF2 expression, while blocking integrin ß1/ß3 or destroying F­actin resulted in a corresponding decrease in KLF2 expression. Downregulation of KLF2 expression by siKLF2 inhibited the differentiation of EPCs to ECs under shear stress conditions, while the expression of EC­specific markers decreased, including CD31 and vWF. Various lengths of the vWF promoter region induced vWF expression, and EPCs co­transfected with KLF2 significantly increased the vWF expression levels compared with the group treated with vWF alone (P<0.01). In conclusion, shear stress may upregulate KLF2 expression, which may be associated with the integrin­actin cytoskeleton system. Most importantly, the shear stress­induced differentiation of EPCs may be mediated by KLF2.


Asunto(s)
Diferenciación Celular/genética , Células Progenitoras Endoteliales/metabolismo , Endotelio Vascular/crecimiento & desarrollo , Factores de Transcripción de Tipo Kruppel/genética , Estrés Mecánico , Actinas/genética , Animales , Anticuerpos Antiidiotipos/administración & dosificación , Citocalasina D/metabolismo , Células Endoteliales/citología , Células Endoteliales/metabolismo , Células Progenitoras Endoteliales/citología , Regulación del Desarrollo de la Expresión Génica/genética , Integrina beta1/inmunología , Molécula-1 de Adhesión Celular Endotelial de Plaqueta/genética , Ratas , Factor de von Willebrand/genética
15.
Cell Physiol Biochem ; 51(4): 1815-1829, 2018.
Artículo en Inglés | MEDLINE | ID: mdl-30504725

RESUMEN

BACKGROUND/AIMS: Mycobacteria-induced diseases, especially tuberculosis, cause more than 1 million deaths each year, which is higher than any other single bacterial pathogen. Neutral sphingomyelinase 2 (Nsm2) has been implied in many physiological processes and diseases, but the role of Nsm2 in pathogen-host interactions and mycobacterial infections has barely been studied. METHODS: We investigated the role of the Nsm2/ceramide system in systemic infection of mice and murine macrophages with Mycobacterium bovis Bacillus Calmette-Guérin (BCG) as a model for mycobacterial infection. For in vitro assays we isolated bone marrow-derived macrophages from Wildtype mice or Nsm2-heterozygous and investigated the role of Nsm2 for macrophage migration/clustering as well as the involvement of p38 mitogen-activated protein kinases (p38K), c-Jun N-terminal kinase (JNK), ß1-integrin and Rac1 activity by Western blot and microscopic studies. For in vivo assays we injected mice intravenously with BCG and analyzed infected tissues for the role of Nsm2-mediated activation of ß1-integrin in granuloma formation and bacterial burden. RESULTS: Our results reveal that BCG infection of macrophages results in rapid stimulation of Nsm2. Genetic and pharmacological studies demonstrate that Nsm2 stimulates a signaling cascade via p38K and JNK to an activation of surface ß1-integrin and Rac1 that leads to the formation of granuloma-like macrophages clusters in vitro and granuloma in vivo. Heterozygosity of Nsm2 in macrophages or antibody-mediated neutralization of active b1-integrin reduced macrophage clusters in vitro and granuloma formation in vivo. Most importantly, Nsm2 heterozygosity or treatment with neutralizing antibodies against ß1-integrin protected mice from systemic BCG infections and chronic infections of the liver and spleen. CONCLUSION: The findings indicate that the Nsm2/ ceramide system plays an important role in systemic infection of mice with mycobacteria by regulating a signaling cascade via p38K, JNK, b1-integrin and Rac1.


Asunto(s)
Integrina beta1/inmunología , Mycobacterium bovis/inmunología , Transducción de Señal , Esfingomielina Fosfodiesterasa/inmunología , Tuberculosis/veterinaria , Animales , Ceramidas/inmunología , Granuloma/inmunología , Granuloma/microbiología , Granuloma/patología , Granuloma/veterinaria , Macrófagos/inmunología , Macrófagos/microbiología , Macrófagos/patología , Ratones , Tuberculosis/inmunología , Tuberculosis/microbiología , Tuberculosis/patología
16.
Cell Physiol Biochem ; 50(3): 1015-1028, 2018.
Artículo en Inglés | MEDLINE | ID: mdl-30355914

RESUMEN

BACKGROUND/AIMS: The mitogenic effects of periodic mechanical stress on chondrocytes have been studied extensively, but the mechanisms whereby chondrocytes sense and respond to mechanical stimuli remain to be determined. We explored the question and verified the key role of G protein coupled receptor kinase interacting protein 1 (GIT1) signaling in periodic mechanical stress-induced chondrocyte proliferation. METHODS: Two steps were undertaken in the experiment. In the first step, the cells were maintained under non-pressure conditions or periodic mechanical stress for 1 h prior to Western blot analysis. In the second step, the cells were pretreated with short hairpin RNA (shRNA) targeted to GIT1 or Src or control scrambled shRNA, or transfected with GIT1 wild-type or GIT1 mutant Y321F, or focal adhesion kinase (FAK) wild-type or FAK mutants Y397F or Y576F/Y577, respectively. Moreover, the cells were pretreated with blocking antibody against integrin ß1 or PP2. Then the cells were maintained under non-pressure conditions or periodic mechanical stress for 1 h prior to Western blot analysis, and for 3 days, 8 h per day, prior to direct cell counting and CCK-8 assay, respectively. RESULTS: Periodic mechanical stress significantly induced sustained phosphorylation of GIT1 at Tyr321. Reduction of GIT1 with shRNA targeted to GIT1 and GIT1 mutant Y321F inhibited periodic mechanical stress-promoted chondrocyte proliferation, accompanied by attenuated extracellular signal-regulated kinase (ERK)1/2 and FAK phosphorylation at Tyr576/577. However, activation of Src and FAK-Tyr397 was not prevented upon GIT1 suppression. Furthermore, pretreatment with blocking antibody against integrin ß1, Src-selective inhibitor, PP2, and shRNA targeted to Src blocked GIT1 activation under periodic mechanical stress. In addition, GIT1 phosphorylation at Tyr321 was not reduced upon pretreatment with FAK mutants Y397F or Y576F/Y577 under conditions of periodic mechanical stress. CONCLUSION: These findings collectively suggested that periodic mechanical stress promoted chondrocyte proliferation through at least two separate pathways, integrin ß1-Src-GIT1-FAK(Tyr576/577)-ERK1/2, and the other parallel GIT1-independent integrin ß1-FAK(Tyr397)-ERK1/2.


Asunto(s)
Proteínas de Ciclo Celular/metabolismo , Proteína Quinasa 1 Activada por Mitógenos/metabolismo , Proteína Quinasa 3 Activada por Mitógenos/metabolismo , Fosfoproteínas/metabolismo , Estrés Mecánico , Animales , Proteínas de Ciclo Celular/antagonistas & inhibidores , Proteínas de Ciclo Celular/genética , Proliferación Celular , Células Cultivadas , Condrocitos/citología , Condrocitos/metabolismo , Quinasa 1 de Adhesión Focal/genética , Quinasa 1 de Adhesión Focal/metabolismo , Integrina beta1/inmunología , Integrina beta1/metabolismo , Mutagénesis Sitio-Dirigida , Fosfoproteínas/antagonistas & inhibidores , Fosfoproteínas/genética , Fosforilación , Interferencia de ARN , ARN Interferente Pequeño/metabolismo , Ratas , Ratas Sprague-Dawley , Familia-src Quinasas/metabolismo
17.
J Biomol Tech ; 29(3): 71-78, 2018 09.
Artículo en Inglés | MEDLINE | ID: mdl-30174558

RESUMEN

Efficient extraction of proteins is a great challenge for numerous downstream proteomic analyses. During the protein extraction procedure, it is critical to maintain the conformational stability, integrity, as well as higher yield of the protein. To do so, 5-different lysis buffers of Tris and HEPES have been used as the primary buffering reagents with variable compositions at different concentrations and pH using human cancer cells. In this study, different protein lysates of human breast cancer cells T47D and MDA-MB-231 and ovarian cancer cell PA-1 were subjected to run SDS-PAGE for separation of proteins based on their molecular size, followed by Coomassie blue, silver staining, and immunoblot assays to compare the extraction yield of total cytoplasmic proteins, glyceraldehyde-3-phosphate dehydrogenase (GAPDH) and the integral membrane protein, integrin ß-1. Our results revealed that Tris-based lysis buffer with 50 mM concentration, pH 7.5, is relatively the efficient and reliable protein extraction method for a wide range of MW subcellular markers, cytoplasmic GAPDH and transmembrane integrin ß-1 proteins. We anticipate that this simple and cost-effective protein extraction protocol might be extremely useful across a broad range of subcellular proteins in different biologic samples.


Asunto(s)
Neoplasias de la Mama/patología , Citosol/química , Biología Molecular/métodos , Proteínas de Neoplasias/aislamiento & purificación , Neoplasias Ováricas/patología , Tampones (Química) , Línea Celular Tumoral , Electroforesis en Gel de Poliacrilamida , Femenino , Gliceraldehído-3-Fosfato Deshidrogenasa (Fosforilante)/inmunología , Gliceraldehído-3-Fosfato Deshidrogenasa (Fosforilante)/aislamiento & purificación , Humanos , Immunoblotting , Integrina beta1/inmunología , Integrina beta1/aislamiento & purificación , Proteínas de la Membrana/inmunología , Proteínas de la Membrana/aislamiento & purificación , Proteínas de Neoplasias/inmunología
18.
PLoS One ; 12(10): e0186628, 2017.
Artículo en Inglés | MEDLINE | ID: mdl-29040328

RESUMEN

SHARPIN (Shank-Associated RH Domain-Interacting Protein) is a component of the linear ubiquitin chain assembly complex (LUBAC), which enhances TNF-induced NF-κB activity. SHARPIN-deficient (Sharpincpdm/cpdm) mice display multi-organ inflammation and chronic proliferative dermatitis (cpdm) due to TNF-induced keratinocyte apoptosis. In cells, SHARPIN also inhibits integrins independently of LUBAC, but it has remained enigmatic whether elevated integrin activity levels in the dermis of Sharpincpdm/cpdm mice is due to increased integrin activity or is secondary to inflammation. In addition, the functional contribution of increased integrin activation to the Sharpincpdm/cpdm phenotype has not been investigated. Here, we find increased integrin activity in keratinocytes from Tnfr1-/- Sharpincpdm/cpdm double knockout mice, which do not display chronic inflammation or proliferative dermatitis, thus suggesting that SHARPIN indeed acts as an integrin inhibitor in vivo. In addition, we present evidence for a functional contribution of integrin activity to the Sharpincpdm/cpdm skin phenotype. Treatment with an integrin beta 1 function blocking antibody reduced epidermal hyperproliferation and epidermal thickness in Sharpincpdm/cpdm mice. Our data indicate that, while TNF-induced cell death triggers the chronic inflammation and proliferative dermatitis, absence of SHARPIN-dependent integrin inhibition exacerbates the epidermal hyperproliferation in Sharpincpdm/cpdm mice.


Asunto(s)
Proteínas Portadoras/genética , Dermatitis/tratamiento farmacológico , Epidermis/efectos de los fármacos , Integrina beta1/genética , Queratinocitos/efectos de los fármacos , Receptores Tipo I de Factores de Necrosis Tumoral/genética , Animales , Anticuerpos Neutralizantes/farmacología , Apoptosis , Proteínas Portadoras/inmunología , Proliferación Celular , Enfermedad Crónica , Dermatitis/genética , Dermatitis/inmunología , Dermatitis/patología , Epidermis/inmunología , Epidermis/patología , Femenino , Eliminación de Gen , Regulación de la Expresión Génica , Inflamación , Integrina beta1/inmunología , Péptidos y Proteínas de Señalización Intracelular , Queratinocitos/inmunología , Queratinocitos/patología , Masculino , Ratones , Ratones Noqueados , FN-kappa B/genética , FN-kappa B/inmunología , Fenotipo , Receptores Tipo I de Factores de Necrosis Tumoral/deficiencia , Receptores Tipo I de Factores de Necrosis Tumoral/inmunología , Transducción de Señal , Ubiquitina/genética , Ubiquitina/inmunología
19.
Clin Exp Immunol ; 190(3): 315-327, 2017 12.
Artículo en Inglés | MEDLINE | ID: mdl-28862740

RESUMEN

The role of macrophage infiltrates in oral mucosal acute graft-versus-host disease (AGVHD) remains unclear, although clinical studies suggest that macrophage infiltration correlates directly with the severity of AGVHD. In this study, we investigated the role of M1 macrophage infiltration in the oral mucosa of rats with AGVHD. Lewis rat spleen cells were injected into (Lewis × Brown Norway) F1 rats to induce systemic GVHD. Tongue samples were evaluated using histology, immunohistochemistry, dual immunofluorescence, real-time reverse transcription-polymerase chain reaction, Transwell migration assays and Stamper-Woodruff binding assays. At the onset of oral mucosal AGVHD, dual immunofluorescence and migration assays revealed that M1 macrophages had accumulated in the basement membrane (BM) region via the laminin/CD29 ß1 integrin pathway. Macrophage-secreted matrix metalloproteinase-2 was related to BM degradation. The adhesion of macrophages to the oral epithelium could be inhibited by pretreating macrophages with a CC chemokine receptor 2 (CCR2) antibody and/or pretreating lesion sections with monocyte chemoattractant protein-1 (MCP-1) antibody. Our data show that the migration and adhesion of M1 macrophages are associated with oral mucosal AGVHD, which is mediated in part by both laminin/CD29 ß 1 intern and MCP-1/CCR2 pathways. Therefore, our study provides additional support for the contribution of macrophage infiltrate to the development of oral mucosal AGVHD.


Asunto(s)
Movimiento Celular/inmunología , Enfermedad Injerto contra Huésped/inmunología , Macrófagos/microbiología , Mucosa Bucal/inmunología , Enfermedad Aguda , Animales , Quimiocina CCL2/inmunología , Femenino , Enfermedad Injerto contra Huésped/patología , Integrina beta1/inmunología , Laminina/inmunología , Macrófagos/patología , Masculino , Metaloproteinasa 2 de la Matriz/inmunología , Mucosa Bucal/patología , Ratas , Ratas Endogámicas Lew , Receptores CCR2/inmunología
20.
Biochem Biophys Res Commun ; 491(3): 629-635, 2017 09 23.
Artículo en Inglés | MEDLINE | ID: mdl-28760342

RESUMEN

Acupuncture therapy is performed by applying the needle insertion at discrete cutaneous locations and used for the treatments of diverse symptoms and disorders. In order to elucidate mechanistic basis on how acupuncture stimulation (AS) produces therapeutic effects, it is primarily important to understand tissue responses locally at the acupuncture site (acupoint). Here, we investigated integrin protein as molecular target responding to and integrating AS. Signals of α6 and ß1 integrins were clearly induced at zusanli acupoint 24 h after AS in areas of nuclear clusters around the needle track. Induction levels of integrin were largely reduced by needle insertion at non-acupuncture point or without needle rotation. Phospho-Erk1/2 was initially decreased below the basal level after AS but increased 24 h later. Induction pattern of phospho-Erk1/2 was as similar as that of α6 integrin in its selectivity to needling procedure and tissue distribution. We further found that mRNA expression of P2X3 purinergic receptor was upregulated in the dorsal root ganglion (DRG) after AS, but decreased by the inhibition of Erk1/2 activity at the acupuncture area. Moreover, AS-mediated integrin activation was required for Erk1/2 activation at the acupuncture site and regulation of pain sensitivity in the hind paw. The present results provide a new evidence on acupuncture-specific tissue response in terms of integrin induction, and further suggest that integrin activation may be involved in transmitting mechanosensory signals from the acupoint to afferent nerve fiber.


Asunto(s)
Terapia por Acupuntura/métodos , Integrina alfa6/inmunología , Integrina beta1/inmunología , Sistema de Señalización de MAP Quinasas/inmunología , Neuralgia/inmunología , Neuralgia/terapia , Puntos de Acupuntura , Animales , Masculino , Mecanotransducción Celular/inmunología , Neuralgia/diagnóstico , Estimulación Física/métodos , Ratas , Ratas Sprague-Dawley , Resultado del Tratamiento
SELECCIÓN DE REFERENCIAS
DETALLE DE LA BÚSQUEDA
...