RESUMEN
The receptor for advanced glycation endproducts (RAGE) is involved in multiple inflammatory processes. RAGE participates in adaptive and innate immune responses but its role in human immune cell responses has not been directly tested in vivo. We treated humanized mice (NSG) with the small molecule antagonist of RAGE, azeliragon, (AZ), and measured effects on xenogeneic (B6) skin graft rejection. AZ delayed the median time to xenograft rejection (22 vs 56â¯days, Pâ¯=â¯0.0001). PD-1 expression on CD4+ and CD8+ T cells was lower following AZ therapy. Transcriptome studies showed inhibition of pathways in splenocytes with AZ including IL-23, IL-17A and IL-1ß signaling. The serum levels of IL-1ß and IL-17A in AZ treated mice were reduced in mice that did not reject skin grafts. The RAGE antagonist prevented xenograft rejection by human immune cells in a murine model. A RAGE antagonist may be a useful inhibitor of adaptive human immune responses.
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Rechazo de Injerto , Interleucina-17 , Humanos , Ratones , Animales , Receptor para Productos Finales de Glicación Avanzada/metabolismo , Xenoinjertos , Trasplante Heterólogo , Ratones Endogámicos C57BLRESUMEN
Type 1 diabetes (T1D) is most likely caused by killing of ß cells by autoreactive CD8+ T cells. Methods to isolate and identify these cells are limited by their low frequency in the peripheral blood. We analyzed CD8+ T cells, reactive with diabetes Ags, with T cell libraries and further characterized their phenotype by CyTOF using class I MHC tetramers. In the libraries, the frequency of islet Ag-specific CD45RO+IFN-γ+CD8+ T cells was higher in patients with T1D compared with healthy control subjects. Ag-specific cells from the libraries of patients with T1D were reactive with ZnT8186-194, whereas those from healthy control recognized ZnT8186-194 and other Ags. ZnT8186-194-reactive CD8+ cells expressed an activation phenotype in T1D patients. We found TCR sequences that were used in multiple library wells from patients with T1D, but these sequences were private and not shared between individuals. These sequences could identify the Ag-specific T cells on a repeated draw, ex vivo in the IFN-γ+ CD8+ T cell subset. We conclude that CD8+ T cell libraries can identify Ag-specific T cells in patients with T1D. The T cell clonotypes can be tracked in vivo with identification of the TCR gene sequences.
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Linfocitos T CD8-positivos/inmunología , Diabetes Mellitus Tipo 1/genética , Diabetes Mellitus Tipo 1/inmunología , Células Secretoras de Insulina/inmunología , Receptores de Antígenos de Linfocitos T/genética , Receptores de Antígenos de Linfocitos T/inmunología , Linfocitos T CD8-positivos/patología , Diabetes Mellitus Tipo 1/patología , Femenino , Humanos , Células Secretoras de Insulina/patología , MasculinoRESUMEN
AIMS/HYPOTHESIS: The long-term effects of successful immune therapies for treatment of type 1 diabetes have not been well studied. The Autoimmunity-Blocking Antibody for Tolerance (AbATE) trial evaluated teplizumab, an Fc receptor non-binding humanised anti-CD3 monoclonal antibody in individuals with new-onset type 1 diabetes, and ended in 2011. Clinical drug-treated responders showed an increased frequency of 'partially exhausted' CD8+ T cells. We studied the clinical, immunological and metabolic status of participants after an average follow-up of 7 years. METHODS: Participants with detectable C-peptide at year 2 of AbATE returned for follow-up. C-peptide responses were assessed by 4 h mixed-meal tolerance test. Autoantibodies and HbA1c levels were measured and average daily insulin use was obtained from patient logs. Peripheral blood mononuclear cells were analysed by flow cytometry and cytokine release. RESULTS: Fifty-six per cent of the original participants returned. Three of the original control group who did not return had lost all detectable C-peptide by the end of the 2 year trial. The C-peptide responses to a mixed-meal tolerance test were similar overall in the drug vs control group of participants but were significantly improved, with less loss of C-peptide, in drug-treated responders identified at 1 year. However, the improvements in C-peptide response were not associated with lower HbA1c levels or insulin use. Drug-treated responders showed a significantly increased frequency of programmed cell death protein 1-positive central memory and anergic CD8+ T cells at follow-up. CONCLUSIONS/INTERPRETATION: These findings suggest there is reduced decline in C-peptide and persistent immunological responses up to 7 years after diagnosis of diabetes in individuals who respond to teplizumab. TRIAL REGISTRATION: ClinicalTrials.gov NCT02067923; the protocol is available at www.immunetolerance.org (ITN027AI).
Asunto(s)
Anticuerpos Monoclonales Humanizados/uso terapéutico , Diabetes Mellitus Tipo 1/sangre , Diabetes Mellitus Tipo 1/tratamiento farmacológico , Adolescente , Adulto , Área Bajo la Curva , Autoinmunidad , Péptido C/sangre , Complejo CD3/inmunología , Linfocitos T CD8-positivos/efectos de los fármacos , Niño , Citocinas/metabolismo , Femenino , Estudios de Seguimiento , Humanos , Hipoglucemiantes/uso terapéutico , Insulina/metabolismo , Islotes Pancreáticos/citología , Masculino , Ensayos Clínicos Controlados Aleatorios como Asunto , Inducción de Remisión , Resultado del Tratamiento , Adulto JovenRESUMEN
The ways in which environmental factors participate in the progression of autoimmune diseases are not known. After initiation, it takes years before hyperglycemia develops in patients at risk for type 1 diabetes (T1D). The receptor for advanced glycation endproducts (RAGE) is a scavenger receptor of the Ig family that binds damage-associated molecular patterns and advanced glycated endproducts and can trigger cell activation. We previously found constitutive intracellular RAGE expression in lymphocytes from patients with T1D. In this article, we show that there is increased RAGE expression in T cells from at-risk euglycemic relatives who progress to T1D compared with healthy control subjects, and in the CD8+ T cells in the at-risk relatives who do versus those who do not progress to T1D. Detectable levels of the RAGE ligand high mobility group box 1 were present in serum from at-risk subjects and patients with T1D. Transcriptome analysis of RAGE+ versus RAGE- T cells from patients with T1D showed differences in signaling pathways associated with increased cell activation and survival. Additional markers for effector memory cells and inflammatory function were elevated in the RAGE+ CD8+ cells of T1D patients and at-risk relatives of patients before disease onset. These studies suggest that expression of RAGE in T cells of subjects progressing to disease predates dysglycemia. These findings imply that RAGE expression enhances the inflammatory function of T cells, and its increased levels observed in T1D patients may account for the chronic autoimmune response when damage-associated molecular patterns are released after cell injury and killing.
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Linfocitos T CD8-positivos/inmunología , Diabetes Mellitus Tipo 1/inmunología , Inflamación/inmunología , Receptor para Productos Finales de Glicación Avanzada/metabolismo , Adolescente , Adulto , Enfermedades Asintomáticas , Supervivencia Celular , Células Cultivadas , Niño , Progresión de la Enfermedad , Femenino , Perfilación de la Expresión Génica , Humanos , Memoria Inmunológica , Activación de Linfocitos , Masculino , Riesgo , Transducción de Señal , Regulación hacia Arriba , Adulto JovenRESUMEN
Background: Herpes simplex virus type 2 (HSV-2; herpes) exacerbates human immunodeficiency virus type 1 (HIV) by unclear mechanisms. These studies tested the impact of HSV-2 on systemic T-cells and HIV reservoirs. Methods: Peripheral blood mononuclear cells from HIV-infected women on antiretroviral therapy who were HSV-2 seropositive or seronegative and HIV-uninfected controls were analyzed by flow cytometry. Cell-associated HIV DNA and RNA were quantified in the absence or presence of activating stimuli, recombinant interleukin 32γ (IL-32γ), and a RUNX1 inhibitor. RNA was assessed by nanostring. Results: CD4, but not CD8, T-cell phenotypes differed in HIV+/HSV-2+ versus HIV+/HSV-2- (overall P = .002) with increased frequency of CCR5+, CXCR4+, PD-1+, and CD69+ and decreased frequency of CCR10+ and CCR6+ T-cells. The changes were associated with higher HIV DNA. Paradoxically, IL-32, a proinflammatory cytokine, was lower in subpopulations of CD4+ T-cells in HSV-2+ versus HSV-2- women. Recombinant IL-32γ blocked HIV reactivation in CD4+ T-cells and was associated with an increase in RUNX1 expression; the blockade was overcome by a RUNX1 inhibitor. Conclusions: Herpes is associated with phenotypic changes in CD4+ T-cells, including a decrease in IL-32, which may contribute to increased HIV reservoirs. Blocking IL-32 may facilitate HIV reactivation to improve shock and kill strategies.
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Infecciones por VIH/inmunología , VIH-1/fisiología , Herpes Genital/inmunología , Herpesvirus Humano 2/fisiología , Interleucinas/inmunología , Adulto , Linfocitos T CD4-Positivos/inmunología , Linfocitos T CD8-positivos/inmunología , Coinfección/virología , Subunidad alfa 2 del Factor de Unión al Sitio Principal/antagonistas & inhibidores , Subunidad alfa 2 del Factor de Unión al Sitio Principal/metabolismo , Estudios Transversales , ADN Viral/aislamiento & purificación , Femenino , Humanos , Interleucinas/antagonistas & inhibidores , Leucocitos Mononucleares/inmunología , Persona de Mediana Edad , ARN Viral/aislamiento & purificación , Proteínas Recombinantes/metabolismo , Carga Viral , Adulto JovenRESUMEN
Oral administration of biologics may be a feasible approach for immune therapy that improves drug safety and potentiates mechanisms of tolerance at mucosal barriers. We tested the ability of a fully human non-FcR binding anti-CD3 mAb, foralumab, to prevent skin xenograft rejection in mice with human immune systems. At an intragastric dose of 15µg, the drug could transit through the small bowel. Serum absorption and binding of lymphoid cells was seen and proliferative responses of splenic CD8+ T cells to mitogen were reduced. Five consecutive daily doses, then weekly dosing led to indefinite graft acceptance without depletion of peripheral T cells. Proliferative and cytokine responses to activation of splenocytes with PHA were reduced. The serum levels of IL-10 but not TNF were increased 6days after application of the skin graft. Oral treatment with anti-CD3 mAb may represent a feasible approach for immune modulation.
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Anticuerpos Monoclonales/uso terapéutico , Complejo CD3/inmunología , Complejo CD3/metabolismo , Rechazo de Injerto/prevención & control , Trasplante de Piel , Administración Oral , Animales , Proliferación Celular , Humanos , Ratones , Linfocitos TRESUMEN
Immune-deficient mice, reconstituted with human stem cells, have been used to analyze human immune responses in vivo. Although they have been used to study immune responses to xenografts, allografts, and pathogens, there have not been models of autoimmune disease in which the mechanisms of the pathologic process can be analyzed. We have found that reconstituted "humanized" mice treated with anti-CTLA-4 Ab (ipilimumab) develop autoimmune disease characterized by hepatitis, adrenalitis, sialitis, anti-nuclear Abs, and weight loss. Induction of autoimmunity involved activation of T cells and cytokine production, and increased infiltration of APCs. When anti-CTLA-4 mAb-treated mice were cotreated with anti-CD3 mAb (teplizumab), hepatitis and anti-nuclear Abs were no longer seen and weight loss did not occur. The anti-CD3 blocked proliferation and activation of T cells, release of IFN-γ and TNF, macrophage infiltration, and release of IP-10 that was induced with anti-CTLA-4 mAb. We also found increased levels of T regulatory cells (CD25(+)CD127(-)) in the spleen and mesenteric lymph nodes in the mice treated with both Abs and greater constitutive phosphorylation of STAT5 in T regulatory cells in spleen cells compared with mice treated with anti-CTLA-4 mAb alone. We describe a model of human autoimmune disease in vivo. Humanized mice may be useful for understanding the mechanisms of biologics that are used in patients. Hepatitis, lymphadenopathy, and other inflammatory sequelae are adverse effects of ipilimumab treatment in humans, and this study may provide insights into this pathogenesis and the effects of immunologics on autoimmunity.
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Anticuerpos Monoclonales Humanizados/farmacología , Enfermedades Autoinmunes/terapia , Modelos Animales de Enfermedad , Trasplante de Células Madre/métodos , Linfocitos T/inmunología , Glándulas Suprarrenales/efectos de los fármacos , Glándulas Suprarrenales/inmunología , Glándulas Suprarrenales/metabolismo , Animales , Anticuerpos Monoclonales/toxicidad , Enfermedades Autoinmunes/inducido químicamente , Enfermedades Autoinmunes/inmunología , Citocinas/sangre , Citocinas/inmunología , Citocinas/metabolismo , Citometría de Flujo , Humanos , Subunidad gamma Común de Receptores de Interleucina/deficiencia , Subunidad gamma Común de Receptores de Interleucina/genética , Ipilimumab , Hígado/efectos de los fármacos , Hígado/inmunología , Hígado/metabolismo , Activación de Linfocitos/efectos de los fármacos , Activación de Linfocitos/inmunología , Macrófagos/efectos de los fármacos , Macrófagos/inmunología , Macrófagos/metabolismo , Ratones , Ratones Endogámicos NOD , Ratones Noqueados , Ratones SCID , Fosforilación/efectos de los fármacos , Fosforilación/inmunología , Factor de Transcripción STAT5/inmunología , Factor de Transcripción STAT5/metabolismo , Análisis de Supervivencia , Linfocitos T/efectos de los fármacos , Linfocitos T/metabolismo , Linfocitos T Reguladores/efectos de los fármacos , Linfocitos T Reguladores/inmunología , Linfocitos T Reguladores/metabolismo , Trasplante Heterólogo , Pérdida de Peso/efectos de los fármacos , Pérdida de Peso/inmunologíaRESUMEN
Intracellular (clade B) OVA-serpin protease inhibitors play an important role in tissue homeostasis by protecting cells from death in response to hypo-osmotic stress, heat shock, and other stimuli. It is not known whether these serpins influence immunological tolerance and the risk for autoimmune diseases. We found that a fraction of young autoimmune diabetes-prone NOD mice had elevated levels of autoantibodies against a member of clade B family known as serpinB13. High levels of anti-serpinB13 Abs were accompanied by low levels of anti-insulin autoantibodies, reduced numbers of islet-associated T cells, and delayed onset of diabetes. Exposure to anti-serpinB13 mAb alone also decreased islet inflammation, and coadministration of this reagent and a suboptimal dose of anti-CD3 mAb accelerated recovery from diabetes. In a fashion similar to that discovered in the NOD model, a deficiency in humoral activity against serpinB13 was associated with early onset of human type 1 diabetes. These findings suggest that, in addition to limiting exposure to proteases within the cell, clade B serpins help to maintain homeostasis by inducing protective humoral immunity.
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Autoanticuerpos/inmunología , Autoantígenos/inmunología , Diabetes Mellitus Tipo 1/inmunología , Serpinas/inmunología , Adolescente , Animales , Western Blotting , Niño , Preescolar , Humanos , Ratones , Ratones Endogámicos BALB C , Ratones Endogámicos C57BL , Ratones Endogámicos NOD , Ratones SCID , Transfección , Adulto JovenRESUMEN
BACKGROUND: Teplizumab, a FcR non-binding anti-CD3 mAb, is approved to delay progression of type 1 diabetes (T1D) at-risk patients. Previous investigations described the immediate effects of the 14-day treatment, but longer-term effects of the drug remain unknown. METHODS: With an extended analysis of study participants, we found that 36% were undiagnosed or remained clinical diabetes free after 5 years suggesting operational tolerance. Using single cell RNA-seq, we compared the phenotypes, transcriptome, and repertoire of peripheral blood CD8+ T cells including autoreactive T cells from study participants before and after teplizumab and features of responders and non-responders. RESULTS: At 3 months, there were transcriptional signatures of cell activation in CD4+ and CD8+ T cells including signaling that was reversed at 18 months. At that time, there was reduced expression of genes in T cell receptor and activation pathways in clinical responders. In CD8+ T cells, we found increased expression of genes associated with exhaustion and immune regulation with teplizumab treatment. These transcriptional features were further confirmed in an independent cohort. Pseudotime analysis showed differentiation of CD8+ exhausted and memory cells with teplizumab treatment. IL7R expression was reduced and patients with lower expression of CD127 had longer diabetes free intervals. In addition, the frequency of autoantigen reactive CD8+ T cells, that expanded in the placebo group over 18 months, did not increase in the teplizumab group. CONCLUSION: These findings indicate that teplizumab promotes operational tolerance in T1D, involving activation followed by exhaustion and regulation and prevents expansion of autoreactive T cells. CLINICALTRIALS: gov: NCT01030861. FUNDING: NIDDK/NIH, Juvenile Diabetes Research Foundation.
RESUMEN
Teplizumab has been approved for the delay of the onset of type 1 diabetes and may modulate new onset disease. We found that patients who were EBV positive at baseline had a more robust response to drug in two clinical trials and therefore postulated that latent virus has general effects in modifying immune responses. We compared the phenotypes, transcriptomes, and development of peripheral blood cells before and after teplizumab treatment. Higher number of Tregs and partially exhausted CD8 + T cells were found in EBV seropositive individuals at the baseline in the TN10 trial and AbATE trial. Single cell transcriptomics and functional assays identified downregulation of the T cell receptor and other signaling pathways before treatment. Impairments in function of adaptive immune cells were enhanced by teplizumab treatment in EBV seropositive individuals. Our data indicate that EBV can impair signaling pathways generally in immune cells, that broadly redirect cell differentiation.
RESUMEN
Herpes simplex virus type 2 (HSV-2) coinfection is associated with increased HIV-1 viral loads and expanded tissue reservoirs, but the mechanisms are not well defined. HSV-2 recurrences result in an influx of activated CD4+ T cells to sites of viral replication and an increase in activated CD4+ T cells in peripheral blood. We hypothesized that HSV-2 induces changes in these cells that facilitate HIV-1 reactivation and replication and tested this hypothesis in human CD4+ T cells and 2D10 cells, a model of HIV-1 latency. HSV-2 promoted latency reversal in HSV-2-infected and bystander 2D10 cells. Bulk and single-cell RNA-Seq studies of activated primary human CD4+ T cells identified decreased expression of HIV-1 restriction factors and increased expression of transcripts including MALAT1 that could drive HIV replication in both the HSV-2-infected and bystander cells. Transfection of 2D10 cells with VP16, an HSV-2 protein that regulates transcription, significantly upregulated MALAT1 expression, decreased trimethylation of lysine 27 on histone H3 protein, and triggered HIV latency reversal. Knockout of MALAT1 from 2D10 cells abrogated the response to VP16 and reduced the response to HSV-2 infection. These results demonstrate that HSV-2 contributes to HIV-1 reactivation through diverse mechanisms, including upregulation of MALAT1 to release epigenetic silencing.
Asunto(s)
Infecciones por VIH , ARN Largo no Codificante , Humanos , Herpesvirus Humano 2/genética , Linfocitos T CD4-Positivos , ARN Largo no Codificante/genética , Regulación hacia Arriba , Etopósido , Infecciones por VIH/genética , Latencia del VirusRESUMEN
Treatment with anti-CD3 mAb modulates immune responses that cause type 1 diabetes and other diseases. CD8+ Tregs can be induced in vitro and in vivo by mAb. However, 1/3 of patients do not respond to drug therapy and in an equal proportion, anti-CD3 mAb does not induce Tregs in vitro. The acquisition of CD8+ Treg activity is a function of the CD8+ cells and not the targets in the assay. To identify markers to differentiate responses of CD8+ Tregs, we analyzed genes differentially expressed in CD8+ T cells of non-responders compared with responders, and found that an inhibitory receptor NKG2A (CD159a) was highly expressed in cells from all non-responders tested. Application of a mAb agonistic to NKG2A during in vitro CD8+ Treg induction by anti-CD3 prevented induction of CD8+ Tregs. CD8+ T cells that are TNFR2+ but NKG2A- are the most potently induced Tregs. The level of NKG2A expression on resting CD8+ T cells inversely correlated with acquisition of regulatory function when activated. We suggest that the induction of human CD8+ Tregs by anti-CD3 mAb is controlled by a negative signaling through NKG2A, and that NKG2A may serve as a negative marker of human CD8+ Tregs.
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Anticuerpos Monoclonales/inmunología , Complejo CD3/inmunología , Linfocitos T CD8-positivos/inmunología , Linfocitos T CD8-positivos/metabolismo , Subfamília C de Receptores Similares a Lectina de Células NK/genética , Linfocitos T Reguladores/inmunología , Biomarcadores , Expresión Génica , Humanos , Subunidad alfa del Receptor de Interleucina-2/análisis , Activación de Linfocitos , Análisis por Micromatrices , Subfamília C de Receptores Similares a Lectina de Células NK/agonistas , Subfamília C de Receptores Similares a Lectina de Células NK/inmunología , Subfamília C de Receptores Similares a Lectina de Células NK/metabolismo , Reacción en Cadena de la Polimerasa , Isoformas de Proteínas , Receptores Tipo II del Factor de Necrosis Tumoral/análisisRESUMEN
Upon toll-like receptor 4 (TLR4) signaling in macrophages, the mammalian Swi/Snf-like BAF chromatin remodeling complex is recruited to many TLR4 target genes where it remodels their chromatin to promote transcription. Here, we show that, surprisingly, recruitment is not sufficient for chromatin remodeling; a second event, dependent on calcium/calmodulin (CaM), is additionally required. Calcium/CaM directly binds the HMG domain of the BAF57 subunit within the BAF complex. Calcium/CaM antagonists, including a CaM-binding peptide derived from BAF57, abolish BAF-dependent remodeling and gene expression without compromising BAF recruitment. BAF57 RNAi and BAF57 dominant negative mutants defective in CaM binding similarly impair the induction of BAF target genes. Our data implicate calcium/CaM in TLR4 signaling, and reveal a previously undescribed, recruitment-independent mode of regulation of the BAF complex that is probably achieved through a direct CaM-BAF interaction.
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Calcio/metabolismo , Calmodulina/metabolismo , Ensamble y Desensamble de Cromatina , Receptor Toll-Like 4/metabolismo , Proteínas Cromosómicas no Histona/química , Proteínas Cromosómicas no Histona/metabolismo , ADN Helicasas/metabolismo , Proteínas de Unión al ADN/química , Proteínas de Unión al ADN/metabolismo , Células HeLa , Humanos , Macrófagos/metabolismo , Proteínas Nucleares/metabolismo , Unión Proteica , Estructura Terciaria de Proteína , Factores de Transcripción/metabolismoRESUMEN
BACKGROUND: Our laboratory has shown that inhalational sensitization to new antigens is facilitated through an ongoing T(H)2-polarized inflammation of the lung, a phenomenon we call "collateral priming." OBJECTIVE: We were interested to analyze whether a T(H)1-polarized pulmonary inflammation also facilitates priming toward new antigens and which cytokine or cytokines are involved. METHODS: T(H)1-polarized T cells were generated in vitro and transferred into congenic mice. Mice were challenged initially with cognate antigen and an unrelated antigen; consecutively, they received cognate antigen or the secondary antigen. Airway inflammation, antigen-specific IgG2a levels, and airway hyperresponsiveness were assessed to determine the inflammatory phenotype, with antibody blocking studies used to determine cytokine requirements for T(H)1 collateral priming. RESULTS: Our experiments revealed that ongoing inflammation of the lung induced by the transfer of T(H)1-polarized cells also facilitates priming toward new antigens, which results in lymphocytic inflammation of the lung. Interestingly, blocking studies identified IL-17A as a major contributor to this pathology. Accordingly, we could demonstrate for the first time that T(H)17-polarized cells alone can facilitate priming toward new antigens, inducing lymphocytic airway inflammation and strong airway hyperresponsiveness. Flow cytometric analysis revealed priming of endogenous T cells for IL-17A secretion with a distinct memory/effector phenotype compared to T(H)1 cells, thus presenting an exciting model to further elucidate differentiation of T(H)17 cells. CONCLUSIONS: We show that airway inflammation mediated by T(H)17 cells facilitates sensitization to new antigens and confers increased airway responsiveness in a murine model of polysensitization, suggesting a mechanism involving IL-17A behind the increased risk for allergic sensitization in polysensitized subjects.
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Hiperreactividad Bronquial/inmunología , Activación de Linfocitos/inmunología , Neumonía/inmunología , Células Th17/inmunología , Traslado Adoptivo , Animales , Hiperreactividad Bronquial/metabolismo , Hiperreactividad Bronquial/patología , Separación Celular , Citometría de Flujo , Inhalación , Interleucina-17/inmunología , Interleucina-17/metabolismo , Pulmón/inmunología , Pulmón/metabolismo , Pulmón/patología , Ratones , Ratones Endogámicos BALB C , Ratones Transgénicos , Neumonía/metabolismo , Neumonía/patologíaRESUMEN
The receptor for advanced glycation endproducts (RAGE) is expressed in T cells after activation with antigen and is constitutively expressed in T cells from patients at-risk for and with type 1 diabetes mellitus (T1D). RAGE expression was associated with an activated T cell phenotype, leading us to examine whether RAGE is involved in T cell signaling. In primary CD4+ and CD8+ T cells from patients with T1D or healthy control subjects, RAGE- cells showed reduced phosphorylation of Erk. To study T cell receptor signaling in RAGE+ or-T cells, we compared signaling in RAGE+/+ Jurkat cells, Jurkat cells with RAGE eliminated by CRISPR/Cas9, or silenced with siRNA. In RAGE KO Jurkat cells, there was reduced phosphorylation of Zap70, Erk and MEK, but not Lck or CD3ξ. RAGE KO cells produced less IL-2 when activated with anti-CD3 +/- anti-CD28. Stimulation with PMA restored signaling and (with ionomycin) IL-2 production. Silencing RAGE with siRNA also decreased signaling. Our studies show that RAGE expression in human T cells is associated with an activated signaling cascade. These findings suggest a link between inflammatory products that are found in patients with diabetes, other autoimmune diseases, and inflammation that may enhance T cell reactivity.
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Linfocitos T CD4-Positivos/metabolismo , Linfocitos T CD8-positivos/metabolismo , Diabetes Mellitus Tipo 1/metabolismo , Receptor para Productos Finales de Glicación Avanzada/metabolismo , Transducción de Señal , Adolescente , Adulto , Linfocitos T CD4-Positivos/patología , Linfocitos T CD8-positivos/patología , Niño , Femenino , Productos Finales de Glicación Avanzada/metabolismo , Humanos , Inflamación/metabolismo , Células Jurkat , Masculino , Adulto JovenRESUMEN
Children and youth infected with severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) have milder disease than do adults, and even among those with the recently described multisystem inflammatory syndrome, mortality is rare. The reasons for the differences in clinical manifestations are unknown but suggest that age-dependent factors may modulate the antiviral immune response. We compared cytokine, humoral, and cellular immune responses in pediatric (children and youth, age <24 years) (n = 65) and adult (n = 60) patients with coronavirus disease 2019 (COVID-19) at a metropolitan hospital system in New York City. The pediatric patients had a shorter length of stay, decreased requirement for mechanical ventilation, and lower mortality compared to adults. The serum concentrations of interleukin-17A (IL-17A) and interferon-γ (IFN-γ), but not tumor necrosis factor-α (TNF-α) or IL-6, were inversely related to age. Adults mounted a more robust T cell response to the viral spike protein compared to pediatric patients as evidenced by increased expression of CD25+ on CD4+ T cells and the frequency of IFN-γ+ CD4+ T cells. Moreover, serum neutralizing antibody titers and antibody-dependent cellular phagocytosis were higher in adults compared to pediatric patients with COVID-19. The neutralizing antibody titer correlated positively with age and negatively with IL-17A and IFN-γ serum concentrations. There were no differences in anti-spike protein antibody titers to other human coronaviruses. Together, these findings demonstrate that the poor outcome in hospitalized adults with COVID-19 compared to children may not be attributable to a failure to generate adaptive immune responses.
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Betacoronavirus/fisiología , Infecciones por Coronavirus/inmunología , Infecciones por Coronavirus/virología , Hospitalización , Neumonía Viral/inmunología , Neumonía Viral/virología , Adolescente , Adulto , Anticuerpos Neutralizantes/inmunología , Anticuerpos Antivirales/sangre , COVID-19 , Niño , Infecciones por Coronavirus/sangre , Citocinas/sangre , Femenino , Humanos , Inmunoglobulina G/metabolismo , Masculino , Persona de Mediana Edad , Pandemias , Neumonía Viral/sangre , SARS-CoV-2 , Glicoproteína de la Espiga del Coronavirus/metabolismo , Resultado del TratamientoRESUMEN
The microbiome affects development and activity of the immune system, and may modulate immune therapies, but there is little direct information about this control in vivo. We studied how the microbiome affects regulation of human immune cells in humanized mice. When humanized mice were treated with a cocktail of 4 antibiotics, there was an increase in the frequency of effector T cells in the gut wall, circulating levels of IFN-γ, and appearance of anti-nuclear antibodies. Teplizumab, a non-FcR-binding anti-CD3ε antibody, no longer delayed xenograft rejection. An increase in CD8+ central memory cells and IL-10, markers of efficacy of teplizumab, were not induced. IL-10 levels were only decreased when the mice were treated with all 4 but not individual antibiotics. Antibiotic treatment affected CD11b+CD11c+ cells, which produced less IL-10 and IL-27, and showed increased expression of CD86 and activation of T cells when cocultured with T cells and teplizumab. Soluble products in the pellets appeared to be responsible for the reduced IL-27 expression in DCs. Similar changes in IL-10 induction were seen when human peripheral blood mononuclear cells were cultured with human stool samples. We conclude that changes in the microbiome may impact the efficacy of immunosuppressive medications by altering immune regulatory pathways.
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Anticuerpos Monoclonales Humanizados/inmunología , Microbioma Gastrointestinal/inmunología , Microbioma Gastrointestinal/fisiología , Tracto Gastrointestinal/inmunología , Leucocitos Mononucleares/inmunología , Leucocitos Mononucleares/metabolismo , Inmunidad Adaptativa/inmunología , Animales , Anticuerpos Antinucleares , Anticuerpos Monoclonales Humanizados/farmacología , Enfermedades Autoinmunes/inmunología , Enfermedades Autoinmunes/microbiología , Antígeno B7-2/metabolismo , Antígeno CD11b , Antígeno CD11c , Complejo CD3 , Linfocitos T CD8-positivos/inmunología , Citocinas/sangre , Citocinas/metabolismo , Modelos Animales de Enfermedad , Tracto Gastrointestinal/microbiología , Rechazo de Injerto/inmunología , Humanos , Inmunosupresores/farmacología , Inmunoterapia , Interferón gamma , Interleucina-10/metabolismo , Interleucina-27/metabolismo , Ratones , Ratones Noqueados , Membrana Mucosa/inmunología , Factor de Transcripción STAT5/metabolismo , Trasplante de Piel , Linfocitos T/inmunología , Trasplante HeterólogoRESUMEN
The receptor for glycation end products (RAGE) has been previously implicated in shaping the adaptive immune response. RAGE is expressed in T cells after activation and constitutively in T cells from patients with diabetes. The effects of RAGE on adaptive immune responses are not clear: Previous reports show that RAGE blockade affects Th1 responses. To clarify the role of RAGE in adaptive immune responses and the mechanisms of its effects, we examined whether RAGE plays a role in T cell activation in a Th2 response involving ovalbumin (OVA)-induced asthma in mice. WT and RAGE deficient wild-type and OT-II mice, expressing a T cell receptor specific for OVA, were immunized intranasally with OVA. Lung cellular infiltration and T cell responses were analyzed by immunostaining, FACS, and multiplex bead analyses for cytokines. RAGE deficient mice showed reduced cellular infiltration in the bronchial alveolar lavage fluid and impaired T cell activation in the mediastinal lymph nodes when compared with WT mice. In addition, RAGE deficiency resulted in reduced OT-II T cell infiltration of the lung and impaired IFNγ and IL-5 production when compared with WT mice and reduced infiltration when transferred into WT hosts. When cultured under conditions favoring the differentiation of T cells subsets, RAGE deficient T cells showed reduced production of IFNγ but increased production of IL-17. Our data show a stimulatory role for RAGE in T activation in OVA-induced asthma. This role is largely mediated by the effects of RAGE on T cell proliferation and differentiation. These findings suggest that RAGE may play a regulatory role in T cell responses following immune activation.
Asunto(s)
Asma/inducido químicamente , Asma/metabolismo , Ovalbúmina/toxicidad , Receptores Inmunológicos/metabolismo , Animales , Asma/genética , Diferenciación Celular/genética , Diferenciación Celular/fisiología , Femenino , Interferón gamma/metabolismo , Interleucina-17/metabolismo , Interleucina-5/metabolismo , Ratones , Receptor para Productos Finales de Glicación Avanzada , Receptores Inmunológicos/genética , Subgrupos de Linfocitos T/citología , Subgrupos de Linfocitos T/metabolismo , Células Th2/citología , Células Th2/metabolismoRESUMEN
Multiple sclerosis (MS) is a genetically mediated autoimmune disease of the central nervous system. B cells have recently emerged as major contributors to disease pathogenesis, but the mechanisms responsible for the loss of B cell tolerance in patients with MS are largely unknown. In healthy individuals, developing autoreactive B cells are removed from the repertoire at 2 tolerance checkpoints during early B cell development. Both of these central and peripheral B cell tolerance checkpoints are defective in patients with rheumatoid arthritis (RA) and type 1 diabetes (T1D). Here, we found that only the peripheral, but not the central, B cell tolerance checkpoint is defective in patients with MS. We show that this specific defect is accompanied by increased activation and homeostatic proliferation of mature naive B cells. Interestingly, all of these MS features parallel defects observed in FOXP3-deficient IPEX patients, who harbor nonfunctional Tregs. We demonstrate that in contrast to patients with RA or T1D, bone marrow central B cell selection in MS appears normal in most patients. In contrast, patients with MS suffer from a specific peripheral B cell tolerance defect that is potentially attributable to impaired Treg function and that leads to the accumulation of autoreactive B cell clones in their blood.