RESUMEN

Desmogleins are involved in cell adhesion conferring structural skin integrity. However, their role in inflammation has been barely studied, and whether desmoglein-4 modulates psoriasis lesions is completely unknown. In this study, we assessed the impact of desmoglein-4 deficiency on the severity of imiquimod (IMQ)-induced skin inflammation and psoriasiform lesions. To this end, desmoglein-4-/- Oncins France Colony A (OFA) with Sprague-Dawley (SD) genetic background were used. Additionally, human RNA-Seq datasets from psoriasis (PSO), atopic dermatitis (AD), and a healthy cohort were analyzed to obtain a desmosome gene expression overview. OFA rats displayed an intense skin inflammation while SD showed only mild inflammatory changes after IMQ treatment. We found that IMQ treatment increased CD3+ T cells in skin from both OFA and SD, being higher in desmoglein-4-deficient rats. In-depth transcriptomic analysis determined that PSO displayed twofold less DSG4 expression than healthy samples while both, PSO and AD showed more than three-fold change expression of DSG3 and DSC2 genes. Although underlying mechanisms are still unknown, these results suggest that the lack of desmoglein-4 may contribute to immune-mediated skin disease progression, promoting leukocyte recruitment to skin. Although further research is needed, targeting desmoglein-4 could have a potential impact on designing new biomarkers for skin diseases.

Asunto(s)

Desmogleínas/deficiencia , Psoriasis/metabolismo , Piel/metabolismo , Animales , Complejo CD3/metabolismo , Estudios de Casos y Controles , Quimiotaxis de Leucocito , Desmogleínas/genética , Modelos Animales de Enfermedad , Regulación hacia Abajo , Femenino , Humanos , Imiquimod , Mediadores de Inflamación/metabolismo , Psoriasis/inducido químicamente , Psoriasis/inmunología , Psoriasis/patología , Ratas Sprague-Dawley , Ratas Transgénicas , Piel/inmunología , Piel/patología , Linfocitos T/inmunología , Linfocitos T/metabolismo

RESUMEN

BACKGROUND: The study examines the function of hypoxia-mediated down-regulation of microRNAs (miRNAs) (mir-30c, mir-135a, and mir-27a) in the process of bladder cancer immune escape. METHODS: Quantitative Real-time PCR (qRT-PCR) was carried out to determine gene expression levels of Drosha and Dicer under hypoxia treatment, while western blotting and flow cytometry were used to determine protein expression. Seven reported miRNAs were identified via qRT-PCR assay. Flow cytometry detection of CD3/CD4/CD8-positive expression and statistics. Enzyme-linked immunosorbent assay (ELISA) detected cellular immune factors content. Cell apoptosis was checked via flow cytometry assay. Luciferase report assay and western blot assays were both used to verify the relationship between miRNAs and Casitas B-lineage lymphoma proto-oncogene b (Cbl-b). The animal model was established and Hematoxylin-eosin (HE) staining, TdT-mediated dUTP Nick-End Labeling (TUNEL) staining, and immunohistochemistry (IHC) assays were separately used to verify the conclusions. RESULTS: The CD3 + /CD4 + expression was increased in the hypoxia group, while CD3 + /CD8 + expression, the cellular immune factors content Interleukin-2 (IL-2) and Tumor Necrosis Factor-α (TNFα) along with the cell apoptosis were suppressed. The protein expression of Cbl-b was found to be up-regulated in the hypoxia group. After constructing the overexpression/ knockdown of Cbl-b in peripheral blood mononuclear cell (PBMC), Cbl-b has been found to promote tumor immune escape in bladder cancer. Furthermore, Cbl-b had been identified as the co-targets of mir-30c, mir-135a, and mir-27a and down-regulation of miRNA biogenesis promotes Cbl-b expression and deactivating T cells in vitro/in vivo. CONCLUSION: Hypoxia-mediated down-regulation of miRNAs' biogenesis promotes tumor immune escape in bladder cancer, which could bring much more advance to the medical research on tumors.

Asunto(s)

Regulación hacia Abajo/inmunología , MicroARNs/metabolismo , Escape del Tumor/inmunología , Hipoxia Tumoral/inmunología , Neoplasias de la Vejiga Urinaria/inmunología , Proteínas Adaptadoras Transductoras de Señales/genética , Animales , Apoptosis , Complejo CD3/metabolismo , Antígenos CD4/metabolismo , Antígenos CD8/metabolismo , ARN Helicasas DEAD-box/genética , Modelos Animales de Enfermedad , Regulación Neoplásica de la Expresión Génica , Humanos , Interleucina-2/metabolismo , Leucocitos Mononucleares/metabolismo , Masculino , Ratones Endogámicos C57BL , MicroARNs/biosíntesis , Estudios Prospectivos , Proto-Oncogenes Mas , Proteínas Proto-Oncogénicas c-cbl/genética , Distribución Aleatoria , Ribonucleasa III/genética , Factor de Necrosis Tumoral alfa/metabolismo , Regulación hacia Arriba , Neoplasias de la Vejiga Urinaria/metabolismo

RESUMEN

In mycobacterial infections, the number of cells from two newly discovered subpopulations of CD3+ myeloid cells are increased at the infection site; one type expresses the T cell receptor (CD3+TCRαß+) and the other does not (CD3+TCRαß-). The role of Mycobacterium tuberculosis (Mtb) virulence in generating these subpopulations and the ability of these cells to migrate remains unclear. In this study, monocyte-derived macrophages (MDMs) infected in vitro with either a virulent (H37Rv) or an avirulent (H37Ra) Mtb strain were phenotypically characterized based on three MDM phenotypes (CD3-, CD3+TCRαß+, and CD3+TCRαß-); then, their migration ability upon Mtb infection was evaluated. We found no differences in the frequency of CD3+ MDMs at 24 h of infection with either Mtb strain. However, H37Rv infection increased the frequency of CD3+TCRαß+ MDMs at a multiplicity of infection of 1 and altered the expression of CD1b, CD1c, and TNF on the surface of cells from both the CD3+ MDM subpopulations; it also modified the expression of CCR2, CXCR1, and CCR7, thus affecting CCL2 and IL-8 levels. Moreover, H37Rv infection decreased the migration ability of the CD3- MDMs, but not CD3+ MDMs. These results confirm that the CD3+ macrophage subpopulations express chemokine receptors that respond to chemoattractants, facilitating cell migration. Together, these data suggest that CD3+ MDMs are a functional subpopulation involved in the immune response against Mtb.

Asunto(s)

Complejo CD3/metabolismo , Movimiento Celular , Macrófagos/citología , Macrófagos/metabolismo , Mycobacterium tuberculosis/fisiología , Receptores de Antígenos de Linfocitos T/metabolismo , Microambiente Celular , Humanos , Inflamación/patología , Ligandos , Modelos Biológicos , Monocitos/metabolismo , Mycobacterium tuberculosis/patogenicidad , Fenotipo , Receptores de Quimiocina/metabolismo , Receptores del Factor de Necrosis Tumoral/genética , Receptores del Factor de Necrosis Tumoral/metabolismo , Tuberculosis/inmunología , Tuberculosis/microbiología , Tuberculosis/patología , Factor de Necrosis Tumoral alfa/genética , Factor de Necrosis Tumoral alfa/metabolismo , Virulencia

RESUMEN

The immunomodulatory activity of plant lectins has been evaluated because of their high selectivity for glycans linked to receptors on innate and adaptative immune cells. ArtinM is a mannosyl-binding lectin, obtained from the seeds of Artocarpus heterophyllus, that induces the differentiation of CD4+ T cells and macrophages by interacting with CD3 and TLR2/CD14, respectively. This ArtinM property ultimately favors the combat of intracellular pathogens, opening new perspectives on the lectins application as immunomodulatory agents. The current section describes protocols for purification and evaluation of ArtinM biological activity. The purification is based on the ArtinM-D-mannose affinity. The effect of inducing IL-12 production by murine macrophages cell line is adopted to evaluate the ArtinM biological activity.

Asunto(s)

Artocarpus/metabolismo , Linfocitos T CD4-Positivos/citología , Factores Inmunológicos/farmacología , Macrófagos/citología , Lectinas de Plantas/farmacología , Animales , Artocarpus/química , Complejo CD3/metabolismo , Linfocitos T CD4-Positivos/efectos de los fármacos , Linfocitos T CD4-Positivos/metabolismo , Diferenciación Celular/efectos de los fármacos , Línea Celular , Factores Inmunológicos/aislamiento & purificación , Interleucina-12/metabolismo , Receptores de Lipopolisacáridos/metabolismo , Macrófagos/efectos de los fármacos , Macrófagos/metabolismo , Manosa/metabolismo , Ratones , Lectinas de Plantas/aislamiento & purificación , Células RAW 264.7 , Semillas/química , Semillas/metabolismo , Receptor Toll-Like 2/metabolismo

RESUMEN

The pathogenesis of cutaneous leishmaniasis (CL) caused by Leishmania (Viannia) braziliensis is dictated mainly by the immune-mediated-tissue inflammation developed. The understanding of the immunological mechanisms that generate tissue damage or resolution of lesions is the key to the development of effective vaccine protocols and proper therapeutic schemes. It is clear that the specific immune response mediated by T cells is responsible for the beneficial outcome of the disease, however, the roles of CD4+ T, CD8+ T, NK and NKT cell subpopulations in immunopathogenesis of CL need to be elucidated. Peripheral blood cells from patients before, during and after the antimonial therapy, as well as healthy individuals (HI) were cultured with (LbAgS) or without (NS) L. braziliensis antigens (LbAg). Afterwards, the frequencies of LbAg-specific-cytotoxic CD8+ T, CD4+ T, NK and CD3+CD56+ NKT cells, as well as their activation and exhaustion profiles, were defined by flow cytometry. We observed higher frequencies of CD8+ T, NK and CD3+CD56+ NKT cells and lower frequencies of CD4+ T lymphocytes in LbAgS cell cultures from patients before treatment. The specific response to LbAg resulted in an expansion of cytotoxic-activated CD4+ T, CD8+ T, and NK cells, before and during treatment, indicating specificity in the response by these cells against L. braziliensis. Furthermore, comparing the differences of frequencies of cytotoxic-activated CD4+T, CD8+T, and NK cells, among before and during treatment patients and HI groups, we conclude that these cell populations are in charge of immune response elicited by antimonial therapy. Interestingly, we also observed that NK cells were induced by LbAg to an exhaustion profile during all clinical stages of the disease. The increased antigen-specific activation and cytotoxic activity are in line with the strong inflammatory response described in this disease, a likely cause of tissue damage. These findings reinforce the involvement of these distinct cytotoxic-activated cell populations in the immunopathogenesis of CL, showing a character of specificity in this immune response.

Asunto(s)

Antígenos de Protozoos/inmunología , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD8-positivos/inmunología , Células Asesinas Naturales/inmunología , Leishmania braziliensis/inmunología , Leishmaniasis Cutánea/inmunología , Leishmaniasis Cutánea/patología , Adulto , Anciano , Complejo CD3/metabolismo , Antígeno CD56/metabolismo , Citocinas/metabolismo , Femenino , Humanos , Leishmaniasis Cutánea/parasitología , Masculino , Persona de Mediana Edad , Linfocitos T Citotóxicos , Adulto Joven

RESUMEN

Glucose and nutrient uptake is essential in supporting T cell activation and is increased upon CD3/CD28 stimulation. As T cells from pleural effusions secondary to lung cancer show impaired function, we hypothesized that these cells might have altered expression of nutrient transporters. Here, we analysed by flow cytometry the expression of the transferrin receptor CD71, amino acid transporter CD98 and glucose transporter Glut1 and glucose uptake in pleural effusion-derived T cells from lung cancer patients, after stimulation via CD3/CD28 under normoxia or hypoxia (2% O2 ). We compared the response of T cells from pleural effusions secondary to lung cancer with that of T cells from nonmalignant effusions. In memory T cells from both groups, anti-CD3/CD28-stimulation under normoxia upregulated CD98 and CD71 expression (measured as median fluorescence intensity, MFI) in comparison with anti-CD3-stimulation. Costimulation under hypoxia tended to increase CD98 expression compared to CD3-stimulation in memory T cells from both groups. Remarkably, in the cancer group, memory T cells stimulated via CD3/CD28 under hypoxia failed to increase CD71 and Glut1 expression levels compared to the cells receiving anti-CD3 stimulation, a phenomenon that contrasted with the behaviour of memory T cells from nonmalignant effusions. Consequently, glucose uptake by memory T cells from the cancer group was not increased after CD3/CD28 stimulation under hypoxia, implying that their glycolytic metabolism is defective. As this process is required for inducing an antitumoural response, our study suggests that memory T cells are rendered dysfunctional and are unable to eliminate lung tumour cells.

Asunto(s)

Antígenos CD28/metabolismo , Complejo CD3/metabolismo , Transportador de Glucosa de Tipo 1/metabolismo , Glucosa/metabolismo , Memoria Inmunológica/inmunología , Neoplasias Pulmonares/metabolismo , Derrame Pleural/metabolismo , Linfocitos T/inmunología , Adulto , Anciano , Anciano de 80 o más Años , Femenino , Humanos , Neoplasias Pulmonares/inmunología , Activación de Linfocitos/inmunología , Masculino , Persona de Mediana Edad , Derrame Pleural/inmunología , Linfocitos T/metabolismo

RESUMEN

BACKGROUND: Systemic lupus erythematosus (SLE) is the prototype of systemic autoimmune disease, characterized by loss of immune tolerance against self-antigens where autoantibody production is the hallmark of disease. B-cell-activating factor (BAFF) and A proliferation-inducing ligand (APRIL) are cytokines that promote autoreactive cell survival, immunoglobulin-class switching and autoantibody responses in human and mouse SLE models. BAFF and APRIL exert their functions through interactions with their receptors BAFF-R and TACI that are differentially expressed in B lymphocyte subsets, monocytes, dendritic cells and T lymphocytes. BAFF stimulation favors T lymphocyte activation and cytokine production through BAFF-R, which could contribute to the Th1, Th17 and/or Th2 response dysregulation observed in SLE patients. OBJECTIVE: To evaluate the expression of the cytokines BAFF and APRIL and their association with the receptors BAFF-R and TACI on CD3+ T cells and to evaluate Th1/Th2/Th17 cytokine profile in patients with SLE. METHODS: Fifteen healthy controls (HC) and 36 SLE patients were included, and their demographic and clinical data were assessed. The disease activity index (Mex-SLEDAI) and damage index (SLICC) were applied to the SLE patients. BAFF-R and TACI expression on CD3+ T cells were evaluated by flow cytometry. Serum BAFF and APRIL concentrations were measured by enzyme-linked immunosorbent assays (ELISA). Cytokine levels of Th1 (IL-12, IL-2, IFN-γ, TNF-α), Th2 (IL-4, IL-6, IL-10, IL-13) and Th17 (IL-1ß e IL-17) were quantified with a multiplex assay (MAGPIX). Statistical analysis was performed using PASW Statistics v.20 and GraphPad Prism v.6 software. RESULTS: No differences in BAFF-R or TACI expression on the CD3+ T cells of SLE and HC were observed. BAFF-R expression correlates inversely with disease activity (r = -0.538, p < 0.01), while TACI correlates with disease activity (r = 0.530, p < 0.05). Serum BAFF and APRIL levels were high in SLE patients and correlated with the disease activity index Mex-SLEDAI (r = 0.621, p < 0.01 and r = 0.416, p < 0.05). SLE patients were found to have significantly higher levels of IL-12, IFN-γ, TNF-α, IL-6, IL-10, IL-13, IL-1ß and IL-17 compared to HC (p < 0.05). Cytokines IL-17 (r = 0.526) and TNF-α (r = 0.410) correlate with disease activity (p < 0.05), while APRIL (r = 0.477), IL-10 (r = 0.426) and IFN-γ (r = 0.440) levels were associated with organ damage (p < 0.01). Serum BAFF expression levels correlate with IL-4 (r = 0.424; p < 0.05), IL-6 (r = 0.420; p < 0.05) and IL-10 (r = 0.459; p < 0.01), whereas APRIL levels correlate with IL-2 (r = 0.666; p < 0.01), IL-12 (r = 0.611; p < 0.01) and TNF-α (r = 0.471; p < 0.05) cytokines. A subgroup of SLE patients with high serum BAFF levels (>2 ng/mL) also showed increased APRIL, IL-2, IL-6 and IL-10 levels (p < 0.05). Finally, BAFF, IL-4 and TNF-α serum levels were associated with high titers of antinuclear antibodies. CONCLUSIONS: The study demonstrates an imbalance in the Th1/Th2 cytokine profile, with increased proinflammatory cytokines, as well as BAFF and APRIL serum levels. Associations of BAFF with Th2 profile cytokines and disease activity, as well as APRIL with Th1 profile cytokines and organ damage, suggest that BAFF and APRIL generated in the autoimmunity context could through still unknown mechanisms, modulate the microenvironment, and perpetuate the inflammatory response, autoantibody production and organ damage observed in SLE patients.

Asunto(s)

Factor Activador de Células B/metabolismo , Receptor del Factor Activador de Células B/metabolismo , Citocinas/sangre , Lupus Eritematoso Sistémico/inmunología , Linfocitos T Colaboradores-Inductores/inmunología , Proteína Activadora Transmembrana y Interactiva del CAML/metabolismo , Miembro 13 de la Superfamilia de Ligandos de Factores de Necrosis Tumoral/metabolismo , Adulto , Autoanticuerpos/sangre , Complejo CD3/metabolismo , Femenino , Humanos , Lupus Eritematoso Sistémico/sangre , Persona de Mediana Edad , Adulto Joven

RESUMEN

BACKGROUND AIMS: Although mesenchymal stromal cells (MSCs) have shown therapeutic potential in intestinal tissue repair, controversy concerning their short survival and poor biodistribution in recipient tissues still remains. Therefore, we investigated the paracrine role of MSC in three-dimensional culture of colon with experimental colitis. METHODS: Colitis was induced in mice by oral administration of dextran sulfate sodium (DSS) for 7 days. Inflammatory responses were assessed on the basis of clinical signs, morphological, and histopathological parameters. On days 2 and 5, colonic explants were removed, and a three-dimensional culture was performed. The structural integrity of the intestinal mucosa was tested by treating the cultures with MSC or conditioned medium (CM) for 24 h, and then the colons were analyzed for histology/immunohistochemistry and interleukin (IL)-6 production. RESULTS: Histological analysis demonstrated that both MSC and CM treatment reduced colon damage in organ culture. An increase in cell proliferation (Ki-67 staining) was observed after CM treatment. Additionally, MSC treatment was able to reduce CD3+ cells. The therapeutic effect of MSC and CM was mediated by the downregulation of IL-6. DISCUSSION: The intestinal in vitro model has shown to be potentially useful for studying cellular interactions in a three-dimensional cell arrangement. Moreover, our results provide strong evidence that both MSC and CM treatments can alleviate colonic damage in organ culture. Importantly, these results suggest that MSC-secreted factors are able to protect the colon from inflammation caused by DSS-induced colitis independent of cell transplantation.

Asunto(s)

Colitis/tratamiento farmacológico , Colon/patología , Células Madre Mesenquimatosas/metabolismo , Técnicas de Cultivo de Órganos/métodos , Animales , Complejo CD3/metabolismo , Proliferación Celular , Colitis/inducido químicamente , Medios de Cultivo Condicionados/farmacología , Sulfato de Dextran/toxicidad , Modelos Animales de Enfermedad , Femenino , Humanos , Interleucina-6/metabolismo , Mucosa Intestinal/citología , Mucosa Intestinal/efectos de los fármacos , Masculino , Ratones Endogámicos C57BL , Placenta/citología , Embarazo

RESUMEN

PURPOSE: To evaluate the acute response of natural killer (NK) cell subsets of chronic kidney disease patients submitted to intradialytic exercise in a randomized crossover study. METHODS: Nine patients were submitted to a single bout of 20-min intradialytic exercise and a control hemodialysis (HD) session with an interval of 7 days between them. Peripheral blood sample was collected at baseline, during HD and immediately after HD in each trial to evaluate the peripheral frequency of NK cells and their subsets (CD3-CD56bright and CD3-CD56dim), systemic cortisol concentrations, C-reactive protein (CRP), creatine kinase activity (CK), and urea and creatinine levels. RESULTS: HD therapy induced a significant decrease in NK cells frequency (p = 0.039), NK CD3-CD56bright cells (p = 0.04), and CD3-CD56dim cells (p = 0.036). On the other hand, no significant alterations were observed in NK cells and NK subsets during and after intradialytic exercise trial (p > 0.05). Neither trial altered CRP levels or serum CK activity during and after HD therapy (p > 0.05). However, HD therapy increased cortisol concentrations after HD therapy (p = 0.034). CONCLUSIONS: This study suggests the potential role of intradialytic exercise to prevent the decrease in peripheral frequency of NK cell subsets during HD therapy. Moreover, moderate intensity intradialytic exercise did not exacerbate the systemic inflammation or induce muscle damage during HD therapy.

Asunto(s)

Ejercicio Físico/fisiología , Células Asesinas Naturales , Esfuerzo Físico/fisiología , Insuficiencia Renal Crónica/sangre , Insuficiencia Renal Crónica/terapia , Anciano , Proteína C-Reactiva/metabolismo , Complejo CD3/metabolismo , Antígeno CD56/metabolismo , Creatina Quinasa/sangre , Creatinina/sangre , Estudios Cruzados , Femenino , Humanos , Hidrocortisona/sangre , Células Asesinas Naturales/metabolismo , Recuento de Linfocitos , Masculino , Persona de Mediana Edad , Proyectos Piloto , Diálisis Renal , Urea/sangre

RESUMEN

Glutaric acidemia type I (GA I) is an inherited neurometabolic disorder caused by a severe deficiency of the mitochondrial glutaryl-CoA dehydrogenase (GCDH) activity. Patients usually present progressive cortical leukodystrophy and commonly develop acute bilateral striatal degeneration mainly during infections that markedly worse their prognosis. A role for quinolinic acid (QA), a key metabolite of the kynurenine pathway, which is activated during inflammatory processes, on the pathogenesis of the acute striatum degeneration occurring in GA I was proposed but so far has not yet been evaluated. Therefore, we investigated whether an acute intrastriatal administration of quinolinic acid (QA) could induce histopathological alterations in the striatum of 30-day-old wild-type (WT) and GCDH knockout (Gcdh-/-) mice. Striatum morphology was evaluated by hematoxylin and eosin, T lymphocyte presence (CD3), and glial activation (GFAP and S100ß) by immunohistochemistry and 3-nitrotyrosine (YNO2) by immunofluorescence. QA provoked extensive vacuolation, edema, and especially lymphocyte infiltration in the striatum of Gcdh-/-. QA also enhanced CD3 staining and the number of YNO2 positive cells in Gcdh-/- mice, relatively to WT, indicating T lymphocyte infiltration and nitrosative stress, respectively. QA-treated WT mice also showed an increase of GFAP and S100ß staining, which is indicative of reactive astrogliosis, whereas the levels of these astrocytic proteins were not changed in Gcdh-/- QA-injected mice. The present data indicate that QA significantly contributes to the histopathological changes observed in the striatum of Gcdh-/- mice.

Asunto(s)

Errores Innatos del Metabolismo de los Aminoácidos/patología , Encefalopatías Metabólicas/patología , Cuerpo Estriado/efectos de los fármacos , Regulación de la Expresión Génica/efectos de los fármacos , Glutaril-CoA Deshidrogenasa/deficiencia , Inflamación/inducido químicamente , Inflamación/genética , Ácido Quinolínico/toxicidad , Errores Innatos del Metabolismo de los Aminoácidos/dietoterapia , Errores Innatos del Metabolismo de los Aminoácidos/genética , Animales , Encefalopatías Metabólicas/dietoterapia , Encefalopatías Metabólicas/genética , Complejo CD3/metabolismo , Cuerpo Estriado/metabolismo , Modelos Animales de Enfermedad , Relación Dosis-Respuesta a Droga , Regulación de la Expresión Génica/genética , Proteína Ácida Fibrilar de la Glía/metabolismo , Glutaril-CoA Deshidrogenasa/genética , Lisina/administración & dosificación , Ratones , Ratones Endogámicos C57BL , Ratones Noqueados , Oxidación-Reducción/efectos de los fármacos , Subunidad beta de la Proteína de Unión al Calcio S100/metabolismo , Estadísticas no Paramétricas , Factores de Tiempo , Tirosina/análogos & derivados , Tirosina/metabolismo

RESUMEN

BACKGROUND: Anti-CD3 therapy can induce immunosuppression by several non mutually exclusive mechanisms that have been proposed to explain the therapeutic effect the administration anti-CD3 mAb, but its immunoregulatory mechanism is still not completely clear. In T cells, microRNAs (miRNAs) regulate several pathways, including those associated with immune tolerance. Here, we report changes in miRNA expression in T cells following treatment with anti-human CD3 antibodies. Peripheral blood mononuclear cells were cultured in the presence of the monoclonal antibody OKT3 or a recombinant fragment of humanized anti-CD3. Following these treatments, the expression profiles of 31 miRNA species were assessed in T cells using TaqMan arrays. RESULTS: Eight of the tested miRNAs (miR-155, miR-21, miR-146a, miR-210, miR-17, miR-590-5p, miR-106b and miR-301a) were statistically significantly up- or down-regulated relative to untreated cells. CONCLUSIONS: Stimulation of T cells with anti-human CD3 antibodies alters miRNA expression patterns, including of miRNA species associated with immune regulatory pathways.

Asunto(s)

Anticuerpos Monoclonales/inmunología , Complejo CD3/metabolismo , Perfilación de la Expresión Génica , MicroARNs/genética , Linfocitos T/metabolismo , Complejo CD3/inmunología , Humanos

RESUMEN

Mechanisms leading to decreased platelet count in immune thrombocytopenia (ITP) are heterogeneous. This study describes increased platelet apoptosis involving loss of mitochondrial membrane potential (ΔΨm), caspase 3 activation (aCasp3) and phosphatidylserine (PS) externalization in a cohort of adult ITP patients. Apoptosis was not related to platelet activation, as PAC-1 binding, P-selectin exposure and GPIb-IX internalization were not increased. Besides, ITP platelets were more sensitive to apoptotic stimulus in terms of aCasp3. Incubation of normal platelets with ITP plasma induced loss of ΔΨm, while PS exposure and aCasp3 remained unaltered. The increase in PS exposure observed in ITP platelets could be reproduced in normal platelets incubated with ITP plasma by adding normal CD3+ lymphocytes to the system as effector cells. Addition of leupeptin -a cathepsin B inhibitor- to this system protected platelets from apoptosis. Increased PS exposure was also observed when normal platelets and CD3+ lymphocytes were incubated with purified IgG from ITP patients and was absent when ITP plasma was depleted of auto-antibodies, pointing to the latter as responsible for platelet damage. Apoptosis was present in platelets from all patients carrying anti-GPIIb-IIIa and anti-GPIb auto-antibodies but was absent in the patient with anti-GPIa-IIa auto-antibodies. Platelet damage inversely correlated with platelet count and decreased during treatment with a thrombopoietin receptor agonist. These results point to a key role for auto-antibodies in platelet apoptosis and suggest that antibody-dependent cell cytotoxicity is the mechanism underlying this phenomenon.

Asunto(s)

Autoanticuerpos/inmunología , Plaquetas/patología , Púrpura Trombocitopénica Idiopática/inmunología , Adulto , Anciano , Anciano de 80 o más Años , Plaquetas/efectos de los fármacos , Plaquetas/inmunología , Complejo CD3/metabolismo , Calcimicina/farmacología , Caspasa 3/metabolismo , Humanos , Linfocitos/inmunología , Linfocitos/metabolismo , Persona de Mediana Edad , Fosfatidilserinas/metabolismo , Plasma , Activación Plaquetaria , Púrpura Trombocitopénica Idiopática/sangre , Adulto Joven

RESUMEN

There is an ongoing need to develop strategic combinations of therapeutic agents to prevent type 1 diabetes (T1D) or to preserve islet ß-cell mass in new-onset disease. Although clinical trials using candidate therapeutics are commonly based on preclinical studies, concern is growing regarding the reproducibility as well as the potential clinical translation of reported results using animal models of human disorders. In response, the National Institutes of Health Immune Tolerance Network and JDRF established a multicenter consortium of academic institutions designed to assess the efficacy and intergroup reproducibility of clinically applicable immunotherapies for reversing new-onset disease in the NOD mouse model of T1D. Predicated on prior studies, this consortium conducted coordinated, prospective studies, using joint standard operating procedures, fixed criteria for study entry, and common reagents, to optimize combined anti-CD3 treatment plus interleukin-1 (IL-1) blockade to reverse new-onset disease in NOD mice. We did not find that IL-1 blockade with anti-IL-1ß monoclonal antibody or IL-1trap provided additional benefit for reversing new-onset disease compared with anti-CD3 treatment alone. These results demonstrate the value of larger, multicenter preclinical studies for vetting and prioritizing therapeutics for future clinical use.

Asunto(s)

Anticuerpos Monoclonales/administración & dosificación , Enfermedades Autoinmunes/tratamiento farmacológico , Complejo CD3/química , Diabetes Mellitus Tipo 1/tratamiento farmacológico , Células Secretoras de Insulina/efectos de los fármacos , Insulina/metabolismo , Interleucina-1beta/antagonistas & inhibidores , Animales , Anticuerpos Monoclonales/química , Anticuerpos Monoclonales/uso terapéutico , Enfermedades Autoinmunes/inmunología , Enfermedades Autoinmunes/metabolismo , Investigación Biomédica/métodos , Complejo CD3/metabolismo , Diabetes Mellitus Tipo 1/inmunología , Diabetes Mellitus Tipo 1/metabolismo , Esquema de Medicación , Quimioterapia Combinada , Femenino , Fragmentos Fab de Inmunoglobulinas/administración & dosificación , Fragmentos Fab de Inmunoglobulinas/química , Fragmentos Fab de Inmunoglobulinas/uso terapéutico , Inmunoterapia/métodos , Secreción de Insulina , Células Secretoras de Insulina/inmunología , Células Secretoras de Insulina/metabolismo , Proteína Accesoria del Receptor de Interleucina-1/antagonistas & inhibidores , Proteína Accesoria del Receptor de Interleucina-1/metabolismo , Interleucina-1beta/metabolismo , Ratones Endogámicos NOD , Estudios Multicéntricos como Asunto , Proyectos Piloto , Receptores Tipo I de Interleucina-1/antagonistas & inhibidores , Receptores Tipo I de Interleucina-1/metabolismo , Proteínas Recombinantes de Fusión/uso terapéutico , Reproducibilidad de los Resultados , Proyectos de Investigación , Organismos Libres de Patógenos Específicos , Estados Unidos

RESUMEN

We described a 21-year-old woman with a diagnosis of Sjögren syndrome that came for consultation with a localized mass over her left arm of fast growth. Lab results were normal; a Doppler ultrasound showed the presence of partial thrombosis in the left axillary vein; a magnetic resonance imaging showed edema on the biceps muscle and the biopsy of the mass disclosed the presence of severe lymphocyte infiltrate within the connective tissue and scarce muscle fibers. Immunostaining showed positive results for antígeno comun leucocitario in spanish (leukocyte common antigen) (ACL) and CD3. Those results are consistent with the diagnosis of focal myositis. The patient was treated with low doses of prednisone and methotrexate, with good response.

Asunto(s)

Miositis/diagnóstico , Trombosis/diagnóstico , Antígenos CD20/metabolismo , Brazo/patología , Vena Axilar/patología , Biopsia , Complejo CD3/metabolismo , Antígenos CD8/metabolismo , Diagnóstico Diferencial , Edema/diagnóstico , Femenino , Humanos , Linfocitos/citología , Imagen por Resonancia Magnética , Metotrexato/uso terapéutico , Músculo Esquelético/patología , Miositis/diagnóstico por imagen , Prednisona/uso terapéutico , Trombosis/diagnóstico por imagen , Ultrasonografía Doppler , Adulto Joven

RESUMEN

Evidence indicates that more than 90 % of infected individuals never develop active tuberculosis. This fact highlights the relevance of the immune response in tuberculosis control. The inducible co-stimulator (ICOS) is a regulator of the function, differentiation, proliferation, and activation of T cells. Moreover, T cells synthesise nitric oxide (NO), interferon gamma (IFN-γ), and interleukin (IL)-10, which help regulate the immune response to tuberculosis. Therefore, we assessed the synthesis of NO, IFN-γ, and IL-10 in CD3+ICOS+ T cells from healthy individuals, household contacts (HHC), and patients with active pulmonary tuberculosis (PTB), previously stimulated with the antigen H37Rv. Our results indicated a significant increase in both the percentage of ICOS+ cells and CD3+ICOS+ T cells producing NO, IFN-γ, and IL-10 in cells obtained from patients with PTB (p < 0.01). In addition, a high mitochondrial membrane potential (ΔΨ m) in CD3+ICOS+ T cells was observed in the cells from HHC and from PTB patients, and is associated with the activation of T cells. In conclusion, results show that the CD3+ICOS+ T cells obtained from PTB patients are the main producers of NO, IFN-γ, and IL-10. In addition, our results imply that NO is a modulator of ICOS expression of T cells from PTB patients.

Asunto(s)

Interferón gamma/metabolismo , Interleucina-10/metabolismo , Óxido Nítrico/metabolismo , Linfocitos T/inmunología , Tuberculosis Pulmonar/inmunología , Adulto , Complejo CD3/metabolismo , Células Cultivadas , Familia , Femenino , Voluntarios Sanos , Humanos , Hidrolasas/inmunología , Proteína Coestimuladora de Linfocitos T Inducibles/metabolismo , Activación de Linfocitos , Masculino , Linfocitos T/metabolismo , Tuberculosis Pulmonar/metabolismo

RESUMEN

Human T-lymphotropic virus type 1 (HTLV-1) is a human retrovirus related to the chronic neuroinflammatory disease HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). CD4(+) T cells activation appears to play a key role on HTLV-1 infection. Here we investigated the expression of genes associated to T cell activation CD3e molecule, epsilon (CD3ɛ), lymphocyte-specific protein tyrosine kinase (LCK), vav 1 guanine nucleotide exchange factor (VAV1), and zeta-chain (TCR) associated protein kinase 70kDa (ZAP70) on T lymphocytes of HTLV-1-infected individuals and compared to healthy uninfected individuals (CT). We observed that CD3ɛ, LCK, ZAP70, and VAV1 gene expression were increased in CD4(+) T cells from HAM/TSP group compared to HTLV-1 asymptomatic patients (HAC). Moreover, ZAP70 and VAV1 were also upregulated in HAM/TSP compared to CT group. We detected a positive correlation among all these genes. We also observed that CD3ɛ, LCK, and VAV1 genes had a positive correlation with the proviral load (PVL) and Tax expression. These results suggest that PVL and Tax protein could drive CD3ɛ, LCK, and VAV1 gene expression in CD4(+) T cells, and these genes function on a synchronized way on the CD4(+) T cell activation. The elucidation of the mechanisms underlying T cell receptor signaling pathway is of considerable interest and might lead to new insights into the mechanism of HAM/TSP.

Asunto(s)

Linfocitos T CD4-Positivos/inmunología , Perfilación de la Expresión Génica , Paraparesia Espástica Tropical/inmunología , Receptores de Antígenos de Linfocitos T/inmunología , Transducción de Señal/inmunología , Adulto , Anciano , Complejo CD3/metabolismo , Linfocitos T CD4-Positivos/enzimología , Linfocitos T CD4-Positivos/virología , Estudios de Casos y Controles , Femenino , Expresión Génica , Humanos , Masculino , Persona de Mediana Edad , Paraparesia Espástica Tropical/genética , Proteínas Tirosina Quinasas/metabolismo , Proteínas Proto-Oncogénicas c-vav/metabolismo , Reacción en Cadena en Tiempo Real de la Polimerasa , Linfocitos T/metabolismo , Carga Viral , Proteína Tirosina Quinasa ZAP-70/metabolismo

RESUMEN

The aim of this study was to explore the immune protective mechanism of rMOMP protein vaccine in intraocular hypertension and retinal optic nerve injury in rats. The rMOMP protein ophthalmic vaccine was prepared and quality-controlled. Sixty normal adult SD rats were randomly divided into two groups to establish a chronic ocular hypertension model and an optic nerve injury model. The model rats were vaccinated with rMOMP-CS ophthalmic vaccine. Fluorogold retrograde tracing was used to observe retinal ganglion cells, and an immunofluorescence method to determine the expression of retinal GAP43, CD3, BDNF, and GDNF. rMOMP protein ophthalmic vaccine met the requirements for medicinal use. The number of retinal ganglion cells (RGCs) of the rMOMP-CS group in the chronic ocular hypertension model was significantly higher than that of the CS group (P < 0.05). The count of RGCs of the rMOMP-CS group in the optic nerve clamping injury model was significantly higher than that of the CS group (P < 0.01). Thus, rMOMP protein ophthalmic vaccine can induce an increase in the expression of retinal neurotrophic factors, thereby exerting a protective effect on damaged retinal optic nerve.

Asunto(s)

Proteínas de la Membrana Bacteriana Externa/inmunología , Sistema Inmunológico/inmunología , Hipertensión Ocular/inmunología , Traumatismos del Nervio Óptico/inmunología , Vacunas/inmunología , Animales , Proteínas de la Membrana Bacteriana Externa/genética , Factor Neurotrófico Derivado del Encéfalo/inmunología , Factor Neurotrófico Derivado del Encéfalo/metabolismo , Complejo CD3/inmunología , Complejo CD3/metabolismo , Enfermedad Crónica , Femenino , Proteína GAP-43/inmunología , Proteína GAP-43/metabolismo , Factor Neurotrófico Derivado de la Línea Celular Glial/inmunología , Factor Neurotrófico Derivado de la Línea Celular Glial/metabolismo , Sistema Inmunológico/efectos de los fármacos , Masculino , Microscopía Confocal , Hipertensión Ocular/metabolismo , Hipertensión Ocular/prevención & control , Traumatismos del Nervio Óptico/metabolismo , Traumatismos del Nervio Óptico/prevención & control , Sustancias Protectoras/administración & dosificación , Distribución Aleatoria , Ratas Sprague-Dawley , Proteínas Recombinantes/inmunología , Retina/inmunología , Retina/metabolismo , Células Ganglionares de la Retina/inmunología , Células Ganglionares de la Retina/metabolismo , Estilbamidinas/metabolismo , Vacunación/métodos , Vacunas/administración & dosificación

RESUMEN

Many RNA virus CNS infections cause neurological disease. Because Piry virus has a limited human pathogenicity and exercise reduces activation of microglia in aged mice, possible influences of environment and aging on microglial morphology and behavior in mice sublethal encephalitis were investigated. Female albino Swiss mice were raised either in standard (S) or in enriched (EE) cages from age 2 to 6 months (young - Y), or from 2 to 16 months (aged - A). After behavioral tests, mice nostrils were instilled with Piry-virus-infected or with normal brain homogenates. Brain sections were immunolabeled for virus antigens or microglia at 8 days post-infection (dpi), when behavioral changes became apparent, and at 20 and 40 dpi, after additional behavioral testing. Young infected mice from standard (SYPy) and enriched (EYPy) groups showed similar transient impairment in burrowing activity and olfactory discrimination, whereas aged infected mice from both environments (EAPy, SAPy) showed permanent reduction in both tasks. The beneficial effects of an enriched environment were smaller in aged than in young mice. Six-hundred and forty microglial cells, 80 from each group were reconstructed. An unbiased, stereological sampling approach and multivariate statistical analysis were used to search for microglial morphological families. This procedure allowed distinguishing between microglial morphology of infected and control subjects. More severe virus-associated microglial changes were observed in young than in aged mice, and EYPy seem to recover microglial homeostatic morphology earlier than SYPy . Because Piry-virus encephalitis outcomes were more severe in aged mice, it is suggested that the reduced inflammatory response in those individuals may aggravate encephalitis outcomes.

Asunto(s)

Envejecimiento , Encéfalo/patología , Encefalitis Viral/patología , Encefalitis Viral/terapia , Ambiente , Microglía/patología , Análisis de Varianza , Animales , Complejo CD3/metabolismo , Proteínas de Unión al Calcio/metabolismo , Modelos Animales de Enfermedad , Encefalitis Viral/fisiopatología , Conducta Exploratoria , Femenino , Imagenología Tridimensional , Memoria/fisiología , Ratones , Proteínas de Microfilamentos/metabolismo , Rhabdoviridae/patogenicidad , Olfato/fisiología , Factores de Tiempo

RESUMEN

NY-ESO-1 is a cancer-testis antigen aberrantly expressed in melanomas, which may serve as a robust and specific target in immunotherapy. NY-ESO-1 antigen expression, tumor features, and the immune profile of tumor infiltrating lymphocytes were assessed in primary cutaneous melanoma. NY-ESO-1 protein was detected in 20% of invasive melanomas (16/79), rarely in in situ melanoma (1/10) and not in benign nevi (0/20). Marked intratumoral heterogeneity of NY-ESO-1 protein expression was observed. NY-ESO-1 expression was associated with increased primary tumor thickness (P = 0.007) and inversely correlated with superficial spreading melanoma (P < 0.02). NY-ESO-1 expression was also associated with reduced numbers and density of CD3+ tumor infiltrating lymphocytes (P = 0.017). When NY-ESO-1 protein was expressed, CD3+ T cells were less diffusely infiltrating the tumor and were more often arranged in small clusters (P = 0.010) or as isolated cells (P = 0.002) than in large clusters of more than five lymphocytes. No correlation of NY-ESO-1 expression with gender, age, tumor site, ulceration, lymph node sentinel status, or survival was observed. NY-ESO-1 expression in melanoma was associated with tumor progression, including increased tumor thickness, and with reduced tumor infiltrating lymphocytes.

Asunto(s)

Antígenos de Neoplasias/metabolismo , Linfocitos T CD8-positivos/inmunología , Linfocitos Infiltrantes de Tumor/inmunología , Melanoma/inmunología , Proteínas de la Membrana/metabolismo , Adulto , Anciano , Anciano de 80 o más Años , Antígenos de Neoplasias/biosíntesis , Brasil , Complejo CD3/metabolismo , Antígenos CD8/metabolismo , Vacunas contra el Cáncer/inmunología , Progresión de la Enfermedad , Femenino , Factores de Transcripción Forkhead/metabolismo , Humanos , Inmunoterapia , Masculino , Melanoma/patología , Proteínas de la Membrana/biosíntesis , Persona de Mediana Edad , Biopsia del Ganglio Linfático Centinela , Neoplasias Cutáneas , Adulto Joven , Melanoma Cutáneo Maligno

RESUMEN

We investigated the roles of CD3McAb and rhIL-2 activated bone marrow in the killing and purging of leukemia cells. Cytotoxicity of activated bone marrow was detected with MTT assay. CFU-GM level in activated bone marrow and the destruction of leukemia cells were measured using the semi-solid cell culture. Immune activation markers in activated bone marrow were detected by indirect immunofluorescence assay. Bone marrow activated by CD3McAb and rhIL-2 displayed significantly upregulated the killing and purging abilities on the leukemia cell line K562 and HL-60. Such effects were superior to that of bone marrow activated by rhIL-2 or CD3McAb alone (P < 0.05, P < 0.01). Activation by rhIL-2 and (or) CD3McAb exerted no obvious influence on CFU-GM level in bone marrow. Compared with bone marrow activated by rhIL-2 or CD3McAb alone, the synergistic effect of both CD3McAb+ and hIL-2 caused significant increase of CD3(+), CD8(+), CD19(+), CD25(+), CD38(+), and CD56(+) levels. Our study indicates that CD3McAb enhanced the killing and purging effects of rhIL-2 activated bone marrow on leukemia cells.

Asunto(s)

Anticuerpos Monoclonales/farmacología , Células de la Médula Ósea/efectos de los fármacos , Purgación de la Médula Ósea , Interleucina-2/farmacología , Adolescente , Adulto , Anticuerpos Monoclonales/inmunología , Células de la Médula Ósea/metabolismo , Células de la Médula Ósea/patología , Purgación de la Médula Ósea/métodos , Trasplante de Médula Ósea , Complejo CD3/inmunología , Complejo CD3/metabolismo , Línea Celular Tumoral , Supervivencia Celular/efectos de los fármacos , Ensayo de Unidades Formadoras de Colonias , Femenino , Células Progenitoras de Granulocitos y Macrófagos , Células HL-60 , Humanos , Inmunofenotipificación , Células K562 , Leucemia/metabolismo , Leucemia/patología , Leucemia/terapia , Masculino , Persona de Mediana Edad , Fenotipo , Adulto Joven