RESUMO
Resolvin E1 (RvE1) is a specialized pro-resolving lipid mediator derived from eicosapentaenoic acid and plays a critical role in resolving inflammation and tissue homeostasis. Th17 cells are a distinct group of T helper (Th) cells with tissue-destructive functions in autoimmune and chronic inflammatory diseases via the secretion of IL-17. Dendritic cell (DC)-mediated antigen presentation regulates the Th17-induced progression of inflammation and tissue destruction. In this study, we hypothesized that the RvE1 would restore homeostatic balance and inflammation by targeting the Th17 function. We designed three experiments to investigate the impact of RvE1 on different phases of Th17 response and the potential role of DCs: First CD4+ T cells were induced by IL-6/TGFß to measure the effect of RvE1 on Th17 differentiation in an inflammatory milieu. Second, we measured the impact of RvE1 on DC-stimulated Th17 differentiation in a co-culture model. Third, we measured the effect of RvE1 on DC maturation. RvE1 blocked the CD25, CCR6 and IL-17 expression; IL-17, IL-21, IL-10, and IL-2 production, suggesting inhibition of T cell activation, Th17 stimulation and chemoattraction. RvE1 also suppressed the activation of DCs by limiting their pro-inflammatory cytokine production. Our findings collectively demonstrated that the RvE1 targeted the Th17 activation and the DC function as a potential mechanism for inflammatory resolution and acquired immune response.
Assuntos
Células Dendríticas/imunologia , Ácido Eicosapentaenoico/análogos & derivados , Interleucina-17/metabolismo , Ativação Linfocitária/efeitos dos fármacos , Células Th17/imunologia , Animais , Apresentação de Antígeno/imunologia , Diferenciação Celular/efeitos dos fármacos , Diferenciação Celular/imunologia , Proliferação de Células/efeitos dos fármacos , Células Dendríticas/citologia , Ácido Eicosapentaenoico/farmacologia , Feminino , Inflamação/imunologia , Subunidade alfa de Receptor de Interleucina-2/biossíntese , Interleucina-6/biossíntese , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Células Th17/citologia , Células Th17/efeitos dos fármacos , Fator de Crescimento Transformador beta/metabolismoRESUMO
γδ T cells are a numerically significant subset of immune cells in ruminants, where they may comprise up to 70 % of all peripheral blood mononuclear cells (PBMCs) in young animals and 25 % in adults. These cells can be activated through traditional TCR-dependent mechanisms, or alternatively in a TCR-independent manner by pattern recognition receptors and have been shown to uptake antigen, as well as process and present it to αß T cells. We have identified a novel CD11b+ subset of γδ T cells in normal sheep peripheral blood. An increase in the frequency of these cells in sheep peripheral blood in response to immunization with an experimental recombinant subunit Rift Valley fever (RVF) vaccine was observed. However, injection of the vaccine adjuvant ISA-25VG alone without the recombinant RVF virus antigens demonstrated the same effect, pointing to an antigen-independent innate immune function of CD11b+ γδ T cells in response to the adjuvant. In vitro studies showed repeatable increases of CD11b-, CD14-, CD86-, CD40-, CD72-, and IFNγ- expressing γδ T cells in PBMCs after 24 h of incubation in the absence of a mitogen. Moreover, the majority of these myeloid-like γδ T cells were demonstrated to process exogenous antigen even in the absence of mitogen. ConA activation increased CD25- and MHCII- expression in γδ T cells, but not the myeloid associated receptors CD14 or CD11b or co-stimulatory molecules such as CD86 and CD40. Considering the role of CD11b and CD14 in the activation of innate immunity, we hypothesize that this subpopulation of sheep γδ T cells may function as innate antigen presenting and pro-inflammatory cells during immune responses. The results presented here also suggest that stress molecules and/or damage-associated molecular patterns may be involved in triggering antigen presenting and pro-inflammatory functions of γδ T cells, given their appearance in vitro in the absence of specific stimulation. Taken together, these data suggest that the early appearance of γδ T cells following adjuvant administration and their possible role in early activation of αß T cell subsets may non-specifically contribute to augmented innate immunity and may promote strong initiation of the adaptive immune response to vaccines in general.
Assuntos
Linfócitos Intraepiteliais/imunologia , Febre do Vale de Rift/imunologia , Vacinas Virais/imunologia , Animais , Apresentação de Antígeno , Antígenos de Diferenciação/biossíntese , Antígeno CD11b/biossíntese , Proliferação de Células , Células Cultivadas , Feminino , Imunidade Inata , Imunogenicidade da Vacina , Imunofenotipagem , Subunidade alfa de Receptor de Interleucina-2/biossíntese , Células Mieloides/imunologia , Febre do Vale de Rift/prevenção & controle , OvinosRESUMO
BACKGROUND: Melanoma patients who have detectable serum soluble NKG2D ligands either at the baseline or post-treatment of PD1/PDL1 blockade exhibit poor overall survival. Among families of soluble human NKG2D ligands, the soluble human MHC I chain-related molecule (sMIC) was found to be elevated in melanoma patients and mostly associated with poor response to PD1/PDL1 blockade therapy. METHODS: In this study, we aim to investigate whether co-targeting tumor-released sMIC enhances the therapeutic outcome of PD1/PDL1 blockade therapy for melanoma. We implanted sMIC-expressing B16F10 melanoma tumors into syngeneic host and evaluated therapeutic efficacy of anti-sMIC antibody and anti-PDL1 antibody combination therapy in comparison with monotherapy. We analyzed associated effector mechanism. We also assessed sMIC/MIC prevalence in metastatic human melanoma tumors. RESULTS: We found that the combination therapy of the anti-PDL1 antibody with an antibody targeting sMIC significantly improved animal survival as compared to monotherapies and that the effect of combination therapy depends significantly on NK cells. We show that combination therapy significantly increased IL-2Rα (CD25) on NK cells which sensitizes NK cells to low dose IL-2 for survival. We demonstrate that sMIC negatively reprograms gene expression related to NK cell homeostatic survival and proliferation and that antibody clearing sMIC reverses the effect of sMIC and reprograms NK cell for survival. We further show that sMIC/MIC is abundantly present in metastatic human melanoma tumors. CONCLUSIONS: Our findings provide a pre-clinical proof-of-concept and a new mechanistic understanding to underscore the significance of antibody targeting sMIC to improve therapeutic efficacy of anti-PD1/PDL1 antibody for MIC/sMIC+ metastatic melanoma patients.
Assuntos
Antineoplásicos Imunológicos/uso terapêutico , Antígeno B7-H1/antagonistas & inibidores , Antígenos de Histocompatibilidade Classe I/imunologia , Inibidores de Checkpoint Imunológico/uso terapêutico , Células Matadoras Naturais/citologia , Melanoma Experimental/tratamento farmacológico , Subfamília K de Receptores Semelhantes a Lectina de Células NK/agonistas , Proteínas de Neoplasias/antagonistas & inibidores , Receptor de Morte Celular Programada 1/antagonistas & inibidores , Animais , Antineoplásicos Imunológicos/farmacologia , Linfócitos T CD8-Positivos/imunologia , Sobrevivência Celular , Sinergismo Farmacológico , Antígenos de Histocompatibilidade Classe I/fisiologia , Homeostase , Humanos , Inibidores de Checkpoint Imunológico/farmacologia , Subunidade alfa de Receptor de Interleucina-2/biossíntese , Subunidade alfa de Receptor de Interleucina-2/genética , Ligantes , Neoplasias Pulmonares/prevenção & controle , Neoplasias Pulmonares/secundário , Masculino , Melanoma Experimental/imunologia , Melanoma Experimental/secundário , Camundongos , Camundongos Transgênicos , Proteínas de Neoplasias/fisiologia , Solubilidade , Organismos Livres de Patógenos Específicos , Transplante Isogênico , Microambiente Tumoral , Regulação para CimaRESUMO
In the present study, we investigated the effects of immunophenotyping on prognosis of diffuse large B cell lymphoma (DLBCL) with central nervous system (CNS) relapse treated with rituximab-CHOP (R-CHOP). CNS relapse occurred in 9.5% of DLBCL patients. At the diagnosis of DLBCL, CD25 was detected in 14.3% of cases. CD25 positivity correlated with an advanced stage, higher R-IPI, higher CNS-IPI, the presence of B symptoms, the presence of extranodal involvement >1, and bone involvement. Moreover CNS relapse was more frequently observed in patients with CD25+ than in those with CD25-. The univariate analysis showed that an advanced stage, high-risk R-IPI, high-risk CNS-IPI, bone involvement, and CD25+ were associated with shorter overall survival (OS). The multivariate analysis confirmed that CD25+ and high-risk CNS-IPI were independent adverse prognostic factors for shorter OS. A Kaplan-Meier analysis revealed the potential of CD25+ as a prognostic factor in patients with CNS relapse and that it correlated with shorter survival. The present results showed that the expression of CD25 in DLBCL patients with CNS relapse was associated with the patient prognosis independent other prognostic factors. The establishment of a treatment strategy for CNS relapse patients with CD25+ DLBCL cells is needed to improve poor outcomes.
Assuntos
Biomarcadores Tumorais/imunologia , Neoplasias do Sistema Nervoso Central/imunologia , Subunidade alfa de Receptor de Interleucina-2/biossíntese , Linfoma Difuso de Grandes Células B/patologia , Recidiva Local de Neoplasia/imunologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Protocolos de Quimioterapia Combinada Antineoplásica/uso terapêutico , Neoplasias do Sistema Nervoso Central/patologia , Ciclofosfamida/uso terapêutico , Doxorrubicina/uso terapêutico , Feminino , Humanos , Imunofenotipagem , Linfoma Difuso de Grandes Células B/tratamento farmacológico , Linfoma Difuso de Grandes Células B/imunologia , Masculino , Pessoa de Meia-Idade , Recidiva Local de Neoplasia/patologia , Prednisona/uso terapêutico , Prognóstico , Estudos Retrospectivos , Rituximab/uso terapêutico , Vincristina/uso terapêutico , Adulto JovemRESUMO
Invariant NKT (iNKT) cells have the unique ability to shape immunity during antitumor immune responses and other forms of sterile and nonsterile inflammation. Recent studies have highlighted a variety of classes of endogenous and pathogen-derived lipid antigens that can trigger iNKT cell activation under sterile and nonsterile conditions. However, the context and mechanisms that drive the presentation of self-lipid antigens in sterile inflammation remain unclear. Here we report that endoplasmic reticulum (ER)-stressed myeloid cells, via signaling events modulated by the protein kinase RNA-like ER kinase (PERK) pathway, increase CD1d-mediated presentation of immunogenic endogenous lipid species, which results in enhanced iNKT cell activation both in vitro and in vivo. In addition, we demonstrate that actin cytoskeletal reorganization during ER stress results in an altered distribution of CD1d on the cell surface, which contributes to enhanced iNKT cell activation. These results define a previously unidentified mechanism that controls iNKT cell activation during sterile inflammation.
Assuntos
Células Apresentadoras de Antígenos/imunologia , Células Dendríticas/imunologia , Estresse do Retículo Endoplasmático/imunologia , Ativação Linfocitária , Células T Matadoras Naturais/imunologia , Animais , Apresentação de Antígeno , Antígenos CD1d/biossíntese , Antígenos CD1d/imunologia , Autoantígenos/imunologia , Carcinoma Pulmonar de Lewis/patologia , Linhagem Celular Tumoral , Técnicas de Cocultura , Citoesqueleto/ultraestrutura , Endossomos/imunologia , Glicoesfingolipídeos/imunologia , Glicoesfingolipídeos/metabolismo , Humanos , Subunidade alfa de Receptor de Interleucina-2/biossíntese , Lipídeos/imunologia , Lisossomos/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Células THP-1 , Tapsigargina/farmacologia , Resposta a Proteínas não Dobradas/imunologia , eIF-2 Quinase/deficiência , eIF-2 Quinase/fisiologiaRESUMO
Invariant NKT (iNKT) cells express markers of both T and NK cells and may produce various cytokines to regulate liver immunity. However, the role of iNKT cells in the progression of HBV-relative liver cirrhosis (HBV-LC) is incompletely understood. Here, we investigated the impact of peripheral iNKT cells on a cohort of patients with HBV-LC. The frequency, number, activation status, apoptosis and proliferation ability of peripheral iNKT cells were detected with flow cytometry. The impact of peripheral iNKT cells on the proliferation of hepatocyte cell line (MIHA) and activation of hepatic stellate cell line (LX-2) was detected with flow cytometry and PCR. In HBV-LC patients, the frequency and absolute number of peripheral iNKT cells significantly reduced, but the expression levels of CD25, interleukin (IL)-4, IL-13 and interferon (IFN)-γ increased. No difference was observed in the proliferation and apoptosis of circulating iNKT cells between patients and healthy controls (HCs). CXCR6 (CD186), known to be closely associated with iNKT cells migration from the periphery to the liver, was highly expressed on peripheral iNKT cells in HBV-LC patients. Furthermore, peripheral iNKT cells had a profound impact on MIHA cell proliferation and LX-2 cell activation through IL-4 or IL-13. Our data suggest that in HBV-LC patients, highly activated peripheral iNKT cells may migrate to the liver and affect hepatocyte cell line (MIHA) proliferation and hepatic stellate cell line (LX-2) activation through the expression of type 2 cytokines, which may result in excessive healing and contributing to the progression of fibrosis toward cirrhosis in liver.
Assuntos
Células Estreladas do Fígado/metabolismo , Hepatite B Crônica/patologia , Hepatócitos/metabolismo , Cirrose Hepática/patologia , Fígado/patologia , Células T Matadoras Naturais/imunologia , Adulto , Idoso , Apoptose/fisiologia , Linhagem Celular , Movimento Celular/fisiologia , Proliferação de Células/fisiologia , Progressão da Doença , Feminino , Vírus da Hepatite B/imunologia , Humanos , Interferon gama/biossíntese , Subunidade alfa de Receptor de Interleucina-2/biossíntese , Interleucina-4/biossíntese , Fígado/citologia , Fígado/virologia , Cirrose Hepática/virologia , Ativação Linfocitária/imunologia , Masculino , Pessoa de Meia-Idade , Receptores CXCR6/metabolismoRESUMO
BACKGROUND: Chronic lymphocytic leukemia (CLL) is a condition characterized by the accumulation of morphologically mature monoclonal lymphocytes B with the CD19+/CD5+/CD23+ phenotype in lymphoid tissue, peripheral blood and bone marrow. The clinical course of patients with CLL is heterogeneous, ranging from indolent to aggressive. The role of lymphocyte activation in the natural history of CLL is still a matter of discussion. OBJECTIVES: The aim of this study was to determine the percentages and absolute numbers of lymphocytes B and T in peripheral blood and bone marrow of CLL patients. Moreover, we analyzed the relationship between the number of CD25-positive and CD69-positive lymphocytes and the established prognostic factors in CLL. MATERIAL AND METHODS: The study included 80 untreated patients with CLL and 20 healthy subjects. The immunophenotype of peripheral blood mononuclear cells (in both groups) and bone marrow cells (solely in the CLL group) was determined by means of flow cytometry. RESULTS: Patients with CLL showed a higher absolute number of activated lymphocytes B with phenotypes CD19+CD25+ and CD19+CD69+, as well as a higher absolute number of CD3+CD25+ lymphocytes T than the controls. The enhanced activation of peripheral blood and bone marrow lymphocytes was associated with higher Rai stages, an increased concentration of lactate dehydrogenase and beta-2 microglobulin and the progression of the disease. The number of lymphocytes B CD19+ZAP-70+ correlated positively with the number of CD19+CD25+ B cells and CD3+CD69+ T cells. CONCLUSIONS: The study confirmed the association between an unfavorable prognosis and a high expression of activation markers in CLL patients. The determination of CD25+ and CD69+ lymphocytes T and B constitutes a valuable diagnostic tool, completing the cytometric evaluation of CLL.
Assuntos
Linfócitos B/patologia , Leucemia Linfocítica Crônica de Células B/sangue , Subpopulações de Linfócitos/imunologia , Linfócitos T/patologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Antígenos CD/biossíntese , Antígenos de Diferenciação de Linfócitos T/biossíntese , Linfócitos B/imunologia , Feminino , Humanos , Subunidade alfa de Receptor de Interleucina-2/biossíntese , Lectinas Tipo C/biossíntese , Leucemia Linfocítica Crônica de Células B/imunologia , Leucemia Linfocítica Crônica de Células B/patologia , Ativação Linfocitária/imunologia , Contagem de Linfócitos , Masculino , Pessoa de Meia-Idade , Prognóstico , Linfócitos T/imunologiaRESUMO
Objectives: γδ T cells, a non-conventional innate lymphocyte subset containing cells that can be activated by lipids and phosphoantigens, are abnormally regulated in systemic sclerosis (SSc). To further evaluate the significance of this dysregulation, we compared how exposure to an autoantigenic lipid, cardiolipin (CL), during co-stimulation with an amino-bisphosphonate (zoledronate, zol), affects the activation and cytokine production of SSc and healthy control (HC) γδ T cells. Methods: Expression of CD25 on Vγ9+, Vδ1+, and total CD3+ T cells in cultured peripheral blood mononuclear cells (PBMCs), their binding of CD1d tetramers, and the effect of monoclonal antibody (mAb) blockade of CD1d were monitored by flow cytometry after 4 days of in vitro culture. Intracellular production of IFNγ and IL-4 was assessed after overnight culture. Results: Percentages of CD25+ among CD3+ and Vδ1+ T cells were elevated significantly in short-term cultured SSc PBMC compared to HC. In SSc but not HC, CL and zol, respectively, suppressed %CD25+ Vγ9+ and Vδ1+ T cells but, when combined, CL + zol significantly activated both subsets in HC and partially reversed inhibition by the individual reagents in SSc. Importantly, Vδ1+ T cells in both SSc and HC were highly reactive with lipid presenting CD1d tetramers, and a CD1d-blocking mAb decreased CL-induced enhancement of %SSc CD25+ Vδ1+ T cells in the presence of zol. %IFNγ+ cells among Vγ9+ T cells of SSc was lower than HC cultured in medium, CL, zol, or CL + zol, whereas %IFNγ+ Vδ1+ T cells was lower only in the presence of CL or CL + zol. %IL-4+ T cells were similar in SSc and HC in all conditions, with the exception of being increased in SSc Vγ9+ T cells in the presence of CL. Conclusion: Abnormal functional responses of γδ T cell subsets to stimulation by CL and phosphoantigens in SSc may contribute to fibrosis and immunosuppression, characteristics of this disease.
Assuntos
Cardiolipinas/farmacologia , Ativação Linfocitária/imunologia , Escleroderma Sistêmico/imunologia , Subpopulações de Linfócitos T/imunologia , Ácido Zoledrônico/farmacologia , Adulto , Idoso , Células Cultivadas , Citocinas/biossíntese , Citocinas/imunologia , Difosfonatos/farmacologia , Feminino , Humanos , Subunidade alfa de Receptor de Interleucina-2/biossíntese , Subunidade alfa de Receptor de Interleucina-2/imunologia , Ativação Linfocitária/efeitos dos fármacos , Masculino , Pessoa de Meia-Idade , Receptores de Antígenos de Linfócitos T gama-delta , Subpopulações de Linfócitos T/efeitos dos fármacosRESUMO
BACKGROUND: Loss of neuroprotective role of CD4+ helper T cells, regulatory T cells, and M2 microglia constitutively results in the rapid neural death in the "rapidly progressive phase" of amyotrophic lateral sclerosis (ALS). AIM: We aimed to investigate relative count of CD4+ and regulatory T lymphocytes and expression level of IL2Ra and FOXP3 genes in peripheral blood mononuclear cells (PBMCs) from patients with ALS. METHOD: We performed a flow cytometric analysis on PBMC from 38 patients with ALS and 32 controls to determine the count of CD4+ and CD4+CD25+ cells. Quantitative real-time PCR analyses were implemented to determine the level of expression of FOXP3 and IL-2Rα (CD25) genes in the peripheral blood mononucleated cells. RESULTS: We found a significant higher proportion of CD4+ T cells (p value < 0.001), along with a significantly reduced proportion of CD4+CD25+ Treg cells (p value = 0.001, p value = 0.02), in the peripheral blood of patient's with ALS. CONCLUSION: The results of our study are in line with the hypothesis that in the early phase of ALS, neuroprotective helper T cells infiltrate in the affected areas in the lumbar spinal cord. This was reflected in higher peripheral percentage of CD4+ helper T cells and higher expression of FOXP3 and IL-2Rα. The observed demise in the number of active CD4+CD25+ regulatory T cells might indicate early signs of progression to later stages of ALS in our study group. Interestingly, disease duration was the sole independent significant determining factor that predicted CD4+CD25+ regulatory T cell counts in the peripheral blood of patients at various stages of ALS, according to a logistic regression model.
Assuntos
Esclerose Lateral Amiotrófica/imunologia , Fatores de Transcrição Forkhead/imunologia , Subunidade alfa de Receptor de Interleucina-2/imunologia , Leucócitos Mononucleares/imunologia , Linfócitos T Reguladores/imunologia , Adulto , Idoso , Esclerose Lateral Amiotrófica/sangue , Esclerose Lateral Amiotrófica/genética , Progressão da Doença , Feminino , Fatores de Transcrição Forkhead/biossíntese , Fatores de Transcrição Forkhead/sangue , Fatores de Transcrição Forkhead/genética , Expressão Gênica , Humanos , Subunidade alfa de Receptor de Interleucina-2/biossíntese , Subunidade alfa de Receptor de Interleucina-2/sangue , Subunidade alfa de Receptor de Interleucina-2/genética , Leucócitos Mononucleares/metabolismo , Masculino , Pessoa de Meia-Idade , Linfócitos T Reguladores/metabolismoRESUMO
Johne's disease (JD) is a chronic wasting disease of ruminants caused by infection with Mycobacterium avium subspecies paratuberculosis (MAP). JD is particularly problematic on US dairy farms: estimates show that over 50% of farms are MAP-contaminated and as many as 91% of dairy herds could be infected. Although estimates vary widely, JD may cost the dairy industry between $200 million and $1.5 billion every year. One major obstacle to JD management is that JD is difficult to detect in many animals, in part due to the variable immunity against MAP demonstrated by JD+ cattle. To characterize the diversity of immune responses against MAP, peripheral blood mononuclear cells from 154 JD test negative and 96 JD test positive cows from the same dairy herds were stimulated with MAP in vitro. The activation of CD4+, CD8+ and γδ T cells and surface IgM+ B cells was measured using flow cytometry. CD4+CD45R0+ T cells, γδ+MHCII+ and γδ+MHCII- T cells and SIgM+ B cells from JD test positive cows all exhibited increased proportions expressing CD25 after MAP stimulation, while CD8+ T cells did not demonstrate increased CD25 expression in response to MAP.
Assuntos
Linfócitos B/imunologia , Doenças dos Bovinos/imunologia , Ativação Linfocitária , Mycobacterium avium subsp. paratuberculosis/imunologia , Paratuberculose/imunologia , Linfócitos T/imunologia , Animais , Linfócitos T CD4-Positivos/imunologia , Bovinos , Doenças dos Bovinos/microbiologia , Células Cultivadas , Resistência à Doença/genética , Feminino , Predisposição Genética para Doença , Imunofenotipagem , Subunidade alfa de Receptor de Interleucina-2/biossíntese , Linfócitos Intraepiteliais/imunologiaRESUMO
The majority of genetic variants associated with common human diseases map to enhancers, non-coding elements that shape cell-type-specific transcriptional programs and responses to extracellular cues. Systematic mapping of functional enhancers and their biological contexts is required to understand the mechanisms by which variation in non-coding genetic sequences contributes to disease. Functional enhancers can be mapped by genomic sequence disruption, but this approach is limited to the subset of enhancers that are necessary in the particular cellular context being studied. We hypothesized that recruitment of a strong transcriptional activator to an enhancer would be sufficient to drive target gene expression, even if that enhancer was not currently active in the assayed cells. Here we describe a discovery platform that can identify stimulus-responsive enhancers for a target gene independent of stimulus exposure. We used tiled CRISPR activation (CRISPRa) to synthetically recruit a transcriptional activator to sites across large genomic regions (more than 100 kilobases) surrounding two key autoimmunity risk loci, CD69 and IL2RA. We identified several CRISPRa-responsive elements with chromatin features of stimulus-responsive enhancers, including an IL2RA enhancer that harbours an autoimmunity risk variant. Using engineered mouse models, we found that sequence perturbation of the disease-associated Il2ra enhancer did not entirely block Il2ra expression, but rather delayed the timing of gene activation in response to specific extracellular signals. Enhancer deletion skewed polarization of naive T cells towards a pro-inflammatory T helper (TH17) cell state and away from a regulatory T cell state. This integrated approach identifies functional enhancers and reveals how non-coding variation associated with human immune dysfunction alters context-specific gene programs.
Assuntos
Autoimunidade/genética , Sistemas CRISPR-Cas/genética , Repetições Palindrômicas Curtas Agrupadas e Regularmente Espaçadas/genética , Elementos Facilitadores Genéticos/genética , Animais , Antígenos CD/biossíntese , Antígenos CD/genética , Antígenos CD/imunologia , Antígenos de Diferenciação de Linfócitos T/biossíntese , Antígenos de Diferenciação de Linfócitos T/genética , Antígenos de Diferenciação de Linfócitos T/imunologia , Diferenciação Celular , Linhagem Celular , Cromatina/genética , Feminino , Regulação da Expressão Gênica/genética , Humanos , Subunidade alfa de Receptor de Interleucina-2/biossíntese , Subunidade alfa de Receptor de Interleucina-2/genética , Subunidade alfa de Receptor de Interleucina-2/imunologia , Lectinas Tipo C/biossíntese , Lectinas Tipo C/genética , Lectinas Tipo C/imunologia , Camundongos , Receptores de Antígenos de Linfócitos T/genética , Receptores de Antígenos de Linfócitos T/imunologia , Células Th17/citologia , Células Th17/imunologiaRESUMO
T-follicular helper (Tfh) cells differentiate through a multistep process, culminating in germinal center (GC) localized GC-Tfh cells that provide support to GC-B cells. T-follicular regulatory (Tfr) cells have critical roles in the control of Tfh cells and GC formation. Although Tfh-cell differentiation is inhibited by IL-2, regulatory T (Treg) cell differentiation and survival depend on it. Here, we describe a CD25- subpopulation within both murine and human PD1+CXCR5+Foxp3+ Tfr cells. It is preferentially located in the GC and can be clearly differentiated from CD25+ non-GC-Tfr, Tfh, and effector Treg (eTreg) cells by the expression of a wide range of molecules. In comparison to CD25+ Tfr and eTreg cells, CD25- Tfr cells partially down-regulate IL-2-dependent canonical Treg features, but retain suppressive function, while simultaneously up-regulating genes associated with Tfh and GC-Tfh cells. We suggest that, similar to Tfh cells, Tfr cells follow a differentiation pathway generating a mature GC-localized subpopulation, CD25- Tfr cells.
Assuntos
Centro Germinativo/citologia , Centro Germinativo/imunologia , Subunidade alfa de Receptor de Interleucina-2/genética , Linfócitos T Auxiliares-Indutores/imunologia , Linfócitos T Reguladores/imunologia , Animais , Regulação para Baixo/imunologia , Fatores de Transcrição Forkhead/biossíntese , Humanos , Subunidade alfa de Receptor de Interleucina-2/biossíntese , Tecido Linfoide/citologia , Tecido Linfoide/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fator 1 de Ligação ao Domínio I Regulador Positivo/biossíntese , Proteínas Proto-Oncogênicas/biossíntese , Receptores CXCR5/biossíntese , Proteínas Repressoras/biossínteseRESUMO
The synergistic effect of IL-18/IL-15/IL-12 stimulation potently activates NK cells, inducing high levels of IFN-γ production. As a result of this potent stimulatory effect, NK cell pre-activation with IL-18/IL-15/IL-12 is being developed as a cancer immunotherapy. Ex vivo expansion of NK cells enables the efficient generation of large numbers of NK cells for wide-scale and repeated therapeutic use, and is thus an important source of NK cells for clinical application. However, the effects of IL-18/IL-15/IL-12 stimulation on ex vivo expanded NK cells have not yet been assessed. Thus, the present study assessed the effects of IL-18/IL-15/IL-12 stimulation on NK cells expanded ex vivo using K562-based artificial antigen presenting cells expressing membrane-bound IL-21. We report that ex vivo expanded NK cells stimulated with IL-18/IL-15/IL-12 produce high levels of IFN-γ and TNFα, have potent cytotoxicity, and maintain prolonged IFN-γ production following removal of stimulation. IL-18/IL-15/IL-12 stimulation induces a phenotypically unique IFN-γ-producing population with reduced CD16 expression and greater CD25 expression as compared to stimulated IFN-γ- NK cells and unstimulated NK cells. We elucidate that the mechanism of synergy for induction and maintenance of IFN-γ production is not due to a further enhancement of STAT4 activation compared to stimulation with IL-12 alone. Furthermore, we demonstrate that the synergistic increase in IFN-γ is not solely under translational regulation, as elevated levels of IFN-γ mRNA contribute to the synergistic increase in IFN-γ. Overall, this study characterizes the response of ex vivo expanded NK cells to IL-18/IL-15/IL-12 stimulation and supports the use of ex vivo expanded NK cells as a feasible and efficient source of IL-18/IL-15/IL-12 pre-activated NK cells for adoptive transfer in cancer immunotherapies.
Assuntos
Interferon gama/biossíntese , Interleucina-12/farmacologia , Interleucina-15/farmacologia , Interleucina-18/farmacologia , Células Matadoras Naturais/imunologia , Ativação Linfocitária/imunologia , Fator de Transcrição STAT4/metabolismo , Transferência Adotiva/métodos , Células Cultivadas , Humanos , Imunoterapia/métodos , Subunidade alfa de Receptor de Interleucina-2/biossíntese , Interleucinas/metabolismo , Neoplasias/terapia , Receptores de IgG/biossíntese , Fator de Necrose Tumoral alfaRESUMO
Genome wide association studies and fine mapping has established a firm link between the IL2RA gene, encoding the interleukin-2 receptor α-chain CD25, and susceptibility to multiple sclerosis (MS). We hypothesized that gene expression in peripheral blood mononuclear cells (PBMCs) from healthy controls (HCs) and MS patients are associated with IL2RA SNP rs2104286 and that gene expression levels correlate with soluble CD25 (sCD25) concentrations - that are affected by rs2104286. We used the Affymetrix Human Gene ST 1.0 microarray to analyze gene expression levels in PBMCs from 18 HCs and 51MS patients. Plasma concentrations of sCD25 were measured by ELISA in all individuals. In HCs 266 genes correlated with sCD25 with Spearman's rho≥0.707; 70 of these genes had a false discovery rate (FDR) value of q<0.05. These genes were highest expressed in cells belonging to the innate immune system. Gene-networks were focused around NFKB1, TNF, BCL6 and STAT1. Eighteen genes correlated with sCD25 with rho≥0.707 in relapsing remitting MS versus 33 in secondary progressive and 34 in primary progressive MS. None had a FDR<0.05. Thirty-eight and 23 genes were differentially expressed between rs2104286 genotype-groups in MS patients and HCs respectively, however they were not significant after FDR correction. Our study indicates that rs2104286 influences gene expression in PBMCs in HCs as shown by the high correlations with the rs2104286-affected sCD25 protein. Correlations were strongest in HCs suggesting that immunological alterations may obscure the role of the IL2RA SNP rs2104286 in established MS.
Assuntos
Regulação da Expressão Gênica , Subunidade alfa de Receptor de Interleucina-2 , Leucócitos Mononucleares/metabolismo , Esclerose Múltipla , Polimorfismo de Nucleotídeo Único , Adulto , Feminino , Perfilação da Expressão Gênica , Estudo de Associação Genômica Ampla , Humanos , Subunidade alfa de Receptor de Interleucina-2/biossíntese , Subunidade alfa de Receptor de Interleucina-2/genética , Masculino , Esclerose Múltipla/sangue , Esclerose Múltipla/genética , Análise de Sequência com Séries de OligonucleotídeosRESUMO
Chronic myeloid leukemia (CML) is a stem cell-derived leukemia in which neoplastic cells exhibit the Philadelphia chromosome and the related oncoprotein BCR-ABL1. The disease is characterized by an accumulation of myeloid precursor cells in the peripheral blood and bone marrow (BM). A small fraction of neoplastic cells in the CML clone supposedly exhibits self-renewal and thus long-term disease-propagating ability. However, so far, little is known about the phenotype, function, and target expression profiles of these leukemic stem cells (LSCs). Recent data suggest that CML LSCs aberrantly express the interleukin-2 receptor alpha chain CD25. Whereas normal CD34+/CD38- BM stem cells display only low amounts of CD25 or lack CD25 altogether, CD34+/CD38- LSCs express CD25 strongly in more than 90% of all patients with untreated CML. As a result, CD25 can be used to identify and quantify CML LSCs. In addition, it has been shown that CD25 serves as a negative growth regulator of CML LSCs. Here, we review the value of CD25 as a novel marker and potential drug target in CML LSCs.
Assuntos
Biomarcadores Tumorais/biossíntese , Proteínas de Fusão bcr-abl/metabolismo , Regulação Leucêmica da Expressão Gênica , Subunidade alfa de Receptor de Interleucina-2/biossíntese , Leucemia Mielogênica Crônica BCR-ABL Positiva/diagnóstico , Leucemia Mielogênica Crônica BCR-ABL Positiva/metabolismo , Proteínas de Neoplasias/metabolismo , Células-Tronco Neoplásicas/metabolismo , ADP-Ribosil Ciclase 1/metabolismo , Antígenos CD34/metabolismo , Células da Medula Óssea/metabolismo , Humanos , Glicoproteínas de Membrana/metabolismo , Células-Tronco Neoplásicas/patologiaRESUMO
T-cell population consists of two major subsets, CD4+ T cells and CD8+ T cells, which can be distinguished by the expression of CD4 or CD8 molecules, respectively. Although they play quite different roles in the immune system, many of their basic cellular processes such as proliferation following stimulation are presumably common. In this study, we have carefully analyzed time-course of G0/1 transition as well as cell cycle progression in the two subsets of quiescent T-cell population following in vitro growth stimulation. We found that CD8+ T cells promote G0/1 transition more rapidly and drive their cell cycle progression faster compared to CD4+ T cells. In addition, expression of CD25 and effects of its blockade revealed that IL-2 is implicated in the rapid progression, but not the earlier G0/1 transition, of CD8+ T cells.
Assuntos
Linfócitos T CD4-Positivos/citologia , Linfócitos T CD8-Positivos/citologia , Fase G1/genética , Ativação Linfocitária/imunologia , Fase de Repouso do Ciclo Celular/genética , Animais , Proliferação de Células , Células Cultivadas , Interleucina-2/metabolismo , Subunidade alfa de Receptor de Interleucina-2/biossíntese , CamundongosRESUMO
CD4 T cell activation and differentiation mechanisms constitute a complex and intricate signaling network involving several regulatory proteins. IRF2BP2 is a transcriptional repressor that is involved in gene-expression regulation in very diverse biologic contexts. Information regarding the IRF2BP2 regulatory function in CD4 T lymphocytes is very limited and suggests a role for this protein in repressing the expression of different cytokine genes. Here, we showed that Irf2bp2 gene expression was decreased in CD4 T cells upon activation. To investigate the possible regulatory roles for IRF2BP2 in CD4 T cell functions, this protein was ectopically expressed in murine primary-activated CD4 T lymphocytes through retroviral transduction. Interestingly, ectopic expression of IRF2BP2 led to a reduction in CD25 expression and STAT5 phosphorylation, along with an impaired proliferative capacity. The CD69 expression was also diminished in IRF2BP2-overexpressing cells, whereas CD44 and CD62L levels were not altered. In vivo, transferred, IRF2BP2-overexpressing, transduced cells displayed an impaired expansion capacity compared with controls. Furthermore, overexpression of IRF2BP2 in differentiated Th cells resulted in slightly reduced IL-4 and pro-TGF-ß production in Th2 and iTregs but had no effect on IFN-γ or IL-17 expression in Th1 and Th17 cells, respectively. Taken together, our data suggest a role for IRF2BP2 in regulating CD4 T cell activation by repressing proliferation and the expression of CD25 and CD69 induced by TCR stimuli.
Assuntos
Linfócitos T CD4-Positivos/imunologia , Ativação Linfocitária/imunologia , Receptores de Antígenos de Linfócitos T/imunologia , Fatores de Transcrição/imunologia , Animais , Antígenos CD/biossíntese , Antígenos CD/genética , Antígenos de Diferenciação de Linfócitos T/biossíntese , Antígenos de Diferenciação de Linfócitos T/genética , Apoptose/imunologia , Células Cultivadas , Citocinas/biossíntese , Citocinas/genética , Feminino , Regulação da Expressão Gênica/imunologia , Humanos , Subunidade alfa de Receptor de Interleucina-2/biossíntese , Subunidade alfa de Receptor de Interleucina-2/genética , Lectinas Tipo C/biossíntese , Lectinas Tipo C/genética , Ativação Linfocitária/genética , Masculino , Camundongos , Camundongos Endogâmicos C57BL , RNA Mensageiro/biossíntese , RNA Mensageiro/genética , Quimera por Radiação , Proteínas Recombinantes de Fusão/metabolismo , Fator de Transcrição STAT5/metabolismo , Transdução GenéticaRESUMO
BACKGROUND: Head and neck squamous cell carcinomas (HNSCC) are known to evade the host immune response. How premalignant oral lesions modulate the immune response, however, has yet to be elucidated. MATERIALS AND METHODS: A mouse model of oral carcinogenesis was used to determine how mediators from premalignant oral lesion cells vs. HNSCC cells impact on immune cytokine production and activation. RESULTS: Media conditioned by premalignant lesion cells elicited an increased production of T cell-associated cytokines and proinflammatory mediators from cervical lymph node cells compared to media conditioned by HNSCC cells or media alone. In the presence of premalignant lesion cell-conditioned media, CD4(+) T cell expression of the IL-2 receptor CD25 and CD8(+) T cell expression of the activation marker CD69 was greater, compared to what was induced in HNSCC cell-conditioned media or media alone. CONCLUSION: Premalignant lesion cells promote a proinflammatory environment and induce immune changes before HNSCC tumors are established.
Assuntos
Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Carcinoma de Células Escamosas/imunologia , Citocinas/biossíntese , Neoplasias de Cabeça e Pescoço/imunologia , Neoplasias Bucais/imunologia , Lesões Pré-Cancerosas/imunologia , Animais , Citocinas/imunologia , Modelos Animais de Doenças , Feminino , Interferon gama/biossíntese , Interferon gama/imunologia , Subunidade alfa de Receptor de Interleucina-2/biossíntese , Subunidade alfa de Receptor de Interleucina-2/imunologia , Ativação Linfocitária , Camundongos , Camundongos Endogâmicos C57BL , Carcinoma de Células Escamosas de Cabeça e Pescoço , Células Tumorais CultivadasRESUMO
T follicular helper (Tfh) cells are a subset of T cells carrying the CD4 antigen; they are important in supporting plasma cell and germinal centre responses. The initial induction of Tfh cell properties occurs within the first few days after activation by antigen recognition on dendritic cells, although how dendritic cells promote this cell-fate decision is not fully understood. Moreover, although Tfh cells are uniquely defined by expression of the follicle-homing receptor CXCR5 (refs 1, 2), the guidance receptor promoting the earlier localization of activated T cells at the interface of the B-cell follicle and T zone has been unclear. Here we show that the G-protein-coupled receptor EBI2 (GPR183) and its ligand 7α,25-dihydroxycholesterol mediate positioning of activated CD4 T cells at the interface of the follicle and T zone. In this location they interact with activated dendritic cells and are exposed to Tfh-cell-promoting inducible co-stimulator (ICOS) ligand. Interleukin-2 (IL-2) is a cytokine that has multiple influences on T-cell fate, including negative regulation of Tfh cell differentiation. We demonstrate that activated dendritic cells in the outer T zone further augment Tfh cell differentiation by producing membrane and soluble forms of CD25, the IL-2 receptor α-chain, and quenching T-cell-derived IL-2. Mice lacking EBI2 in T cells or CD25 in dendritic cells have reduced Tfh cells and mount defective T-cell-dependent plasma cell and germinal centre responses. These findings demonstrate that distinct niches within the lymphoid organ T zone support distinct cell fate decisions, and they establish a function for dendritic-cell-derived CD25 in controlling IL-2 availability and T-cell differentiation.
Assuntos
Diferenciação Celular , Células Dendríticas/imunologia , Interleucina-2/imunologia , Receptores Acoplados a Proteínas G/metabolismo , Linfócitos T Auxiliares-Indutores/citologia , Linfócitos T Auxiliares-Indutores/imunologia , Animais , Membrana Celular/metabolismo , Células Dendríticas/citologia , Feminino , Centro Germinativo/imunologia , Hidroxicolesteróis/metabolismo , Proteína Coestimuladora de Linfócitos T Induzíveis/metabolismo , Subunidade alfa de Receptor de Interleucina-2/biossíntese , Subunidade alfa de Receptor de Interleucina-2/química , Subunidade alfa de Receptor de Interleucina-2/deficiência , Subunidade alfa de Receptor de Interleucina-2/metabolismo , Ativação Linfocitária , Masculino , Camundongos , Plasmócitos/imunologia , Receptores Acoplados a Proteínas G/deficiência , Receptores Acoplados a Proteínas G/genética , SolubilidadeRESUMO
BACKGROUND: Recently, identification of CD25 (interleukin-2 receptor alpha) expression on leukemic blasts was correlated to early treatment failure and unfavorable outcome in acute myeloid leukemia (AML) patients. Here we wished to determine whether quantification of CD25 on peripheral blood CD4+ T cells could improve prognostication in newly diagnosed AML patients. METHODS: The mean fluorescence intensity (MFI) of CD25 expression and frequencies of peripheral blood CD4+ T cells with varying levels of CD25 and CD127 expression were assessed by flow cytometry in all studied individuals. RESULTS: Using univariate (unadjusted) and multivariate (adjusted) analyses we demonstrated that detection of high pretreatment CD25 expression on circulating CD4+ T cells was associated with significantly decreased survival rate of AML patients subjected to standard induction chemotherapy. These associations held true for both entire group of analyzed AML patients and different subgroups of patients identified by presence or absence of favorable and adverse molecular prognostic factors. CONCLUSIONS: Our data indicate that quantification of CD25 expression on peripheral blood CD4+ T cells could become a novel, easily accessible method of shortened survival prognostication of AML patients subjected to standard cytotoxic therapy.