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1.
Nat Commun ; 10(1): 4768, 2019 10 18.
Artigo em Inglês | MEDLINE | ID: mdl-31628339

RESUMO

B-1a cells are long-lived, self-renewing innate-like B cells that predominantly inhabit the peritoneal and pleural cavities. In contrast to conventional B-2 cells, B-1a cells have a receptor repertoire that is biased towards bacterial and self-antigens, promoting a rapid response to infection and clearing of apoptotic cells. Although B-1a cells are known to primarily originate from fetal tissues, the mechanisms by which they arise has been a topic of debate for many years. Here we show that in the fetal liver versus bone marrow environment, reduced IL-7R/STAT5 levels promote immunoglobulin kappa gene recombination at the early pro-B cell stage. As a result, differentiating B cells can directly generate a mature B cell receptor (BCR) and bypass the requirement for a pre-BCR and pairing with surrogate light chain. This 'alternate pathway' of development enables the production of B cells with self-reactive, skewed specificity receptors that are peculiar to the B-1a compartment. Together our findings connect seemingly opposing lineage and selection models of B-1a cell development and explain how these cells acquire their unique properties.


Assuntos
Subpopulações de Linfócitos B/imunologia , Linfócitos B/imunologia , Diferenciação Celular/imunologia , Receptores de Células Precursoras de Linfócitos B/imunologia , Receptores de Antígenos de Linfócitos B/imunologia , Animais , Subpopulações de Linfócitos B/metabolismo , Linfócitos B/metabolismo , Medula Óssea/imunologia , Medula Óssea/metabolismo , Diferenciação Celular/genética , Cadeias Leves Substitutas da Imunoglobulina/genética , Cadeias Leves Substitutas da Imunoglobulina/imunologia , Cadeias Leves Substitutas da Imunoglobulina/metabolismo , Fígado/embriologia , Fígado/imunologia , Fígado/metabolismo , Ativação Linfocitária/genética , Ativação Linfocitária/imunologia , Camundongos Endogâmicos C57BL , Camundongos Knockout , Receptores de Células Precursoras de Linfócitos B/genética , Receptores de Células Precursoras de Linfócitos B/metabolismo , Receptores de Antígenos de Linfócitos B/genética , Receptores de Antígenos de Linfócitos B/metabolismo , Receptores de Interleucina-7/genética , Receptores de Interleucina-7/imunologia , Receptores de Interleucina-7/metabolismo , Fator de Transcrição STAT5/genética , Fator de Transcrição STAT5/imunologia , Fator de Transcrição STAT5/metabolismo
2.
Nat Commun ; 10(1): 4415, 2019 09 27.
Artigo em Inglês | MEDLINE | ID: mdl-31562329

RESUMO

Many autoimmune diseases are characterized by the production of autoantibodies. The current view is that CD4+ T follicular helper (Tfh) cells are the main subset regulating autoreactive B cells. Here we report a CXCR5+PD1+ Tfh subset of CD8+ T cells whose development and function are negatively modulated by Stat5. These CD8+ Tfh cells regulate the germinal center B cell response and control autoantibody production, as deficiency of Stat5 in CD8 T cells leads to an increase of CD8+ Tfh cells, resulting in the breakdown of B cell tolerance and concomitant autoantibody production. CD8+ Tfh cells share similar gene signatures with CD4+ Tfh, and require CD40L/CD40 and TCR/MHCI interactions to deliver help to B cells. Our study thus highlights the diversity of follicular T cell subsets that contribute to the breakdown of B-cell tolerance.


Assuntos
Linfócitos B/imunologia , Linfócitos T CD8-Positivos/imunologia , Tolerância Imunológica/imunologia , Receptor de Morte Celular Programada 1/imunologia , Receptores CXCR5/imunologia , Linfócitos T Auxiliares-Indutores/imunologia , Autoanticorpos/imunologia , Autoanticorpos/metabolismo , Doenças Autoimunes/genética , Doenças Autoimunes/imunologia , Doenças Autoimunes/metabolismo , Linfócitos B/metabolismo , Antígenos CD40/genética , Antígenos CD40/imunologia , Antígenos CD40/metabolismo , Ligante de CD40/genética , Ligante de CD40/imunologia , Ligante de CD40/metabolismo , Linfócitos T CD8-Positivos/metabolismo , Perfilação da Expressão Gênica/métodos , Humanos , Tolerância Imunológica/genética , Receptor de Morte Celular Programada 1/genética , Receptor de Morte Celular Programada 1/metabolismo , Receptores CXCR5/genética , Receptores CXCR5/metabolismo , Fator de Transcrição STAT5/genética , Fator de Transcrição STAT5/imunologia , Fator de Transcrição STAT5/metabolismo , Subpopulações de Linfócitos T/imunologia , Subpopulações de Linfócitos T/metabolismo , Linfócitos T Auxiliares-Indutores/metabolismo
3.
Nat Commun ; 10(1): 2042, 2019 05 03.
Artigo em Inglês | MEDLINE | ID: mdl-31053703

RESUMO

Metabolic pathways that regulate T-cell function show promise as therapeutic targets in diverse diseases. Here, we show that at rest cultured human effector memory and central memory CD4+ T-cells have elevated levels of glycolysis and oxidative phosphorylation (OXPHOS), in comparison to naïve T-cells. Despite having low resting metabolic rates, naive T-cells respond to TCR stimulation with robust and rapid increases in glycolysis and OXPHOS. This early metabolic switch requires Akt activity to support increased rates of glycolysis and STAT5 activity for amino acid biosynthesis and TCA cycle anaplerosis. Importantly, both STAT5 inhibition and disruption of TCA cycle anaplerosis are associated with reduced IL-2 production, demonstrating the functional importance of this early metabolic program. Our results define STAT5 as a key node in modulating the early metabolic program following activation in naive CD4+ T-cells and in turn provide greater understanding of how cellular metabolism shapes T-cell responses.


Assuntos
Linfócitos T CD4-Positivos/imunologia , Proteínas Proto-Oncogênicas c-akt/metabolismo , Receptores de Antígenos de Linfócitos T/metabolismo , Fator de Transcrição STAT5/metabolismo , Linfócitos T CD4-Positivos/metabolismo , Células Cultivadas , Ciclo do Ácido Cítrico/imunologia , Glicólise/imunologia , Voluntários Saudáveis , Humanos , Memória Imunológica , Ativação Linfocitária , Fosforilação Oxidativa , Cultura Primária de Células , Proteínas Proto-Oncogênicas c-akt/imunologia , Receptores de Antígenos de Linfócitos T/imunologia , Fator de Transcrição STAT5/imunologia
4.
J Immunol ; 202(10): 2924-2944, 2019 05 15.
Artigo em Inglês | MEDLINE | ID: mdl-30988120

RESUMO

Clonal expansion of B cell chronic lymphocytic leukemia (B-CLL) occurs within lymphoid tissue pseudofollicles. IL-15, a stromal cell-associated cytokine found within spleens and lymph nodes of B-CLL patients, significantly boosts in vitro cycling of blood-derived B-CLL cells following CpG DNA priming. Both IL-15 and CpG DNA are elevated in microbe-draining lymphatic tissues, and unraveling the basis for IL-15-driven B-CLL growth could illuminate new therapeutic targets. Using CpG DNA-primed human B-CLL clones and approaches involving both immunofluorescent staining and pharmacologic inhibitors, we show that both PI3K/AKT and JAK/STAT5 pathways are activated and functionally important for IL-15→CD122/ɣc signaling in ODN-primed cells expressing activated pSTAT3. Furthermore, STAT5 activity must be sustained for continued cycling of CFSE-labeled B-CLL cells. Quantitative RT-PCR experiments with inhibitors of PI3K and STAT5 show that both contribute to IL-15-driven upregulation of mRNA for cyclin D2 and suppression of mRNA for DNA damage response mediators ATM, 53BP1, and MDC1. Furthermore, protein levels of these DNA damage response molecules are reduced by IL-15, as indicated by Western blotting and immunofluorescent staining. Bioinformatics analysis of ENCODE chromatin immunoprecipitation sequencing data from cell lines provides insight into possible mechanisms for STAT5-mediated repression. Finally, pharmacologic inhibitors of JAKs and STAT5 significantly curtailed B-CLL cycling when added either early or late in a growth response. We discuss how the IL-15-induced changes in gene expression lead to rapid cycling and possibly enhanced mutagenesis. STAT5 inhibitors might be an effective modality for blocking B-CLL growth in patients.


Assuntos
Ciclina D2/imunologia , Dano ao DNA/imunologia , Interleucina-15/imunologia , Leucemia Linfocítica Crônica de Células B/imunologia , Proteínas Proto-Oncogênicas c-akt/imunologia , Fator de Transcrição STAT5/imunologia , Transdução de Sinais/imunologia , Proteínas Adaptadoras de Transdução de Sinal/imunologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Proteínas Mutadas de Ataxia Telangiectasia/imunologia , Proteínas de Ciclo Celular/imunologia , Linhagem Celular Tumoral , Feminino , Regulação Neoplásica da Expressão Gênica/imunologia , Humanos , Leucemia Linfocítica Crônica de Células B/patologia , Masculino , Pessoa de Meia-Idade , Proteína 1 de Ligação à Proteína Supressora de Tumor p53/imunologia , Regulação para Cima/imunologia
5.
Nat Commun ; 10(1): 354, 2019 01 21.
Artigo em Inglês | MEDLINE | ID: mdl-30664665

RESUMO

Regulatory T (Treg) cells control self-tolerance, inflammatory responses and tissue homeostasis. In mature Treg cells, continued expression of FOXP3 maintains lineage identity, while T cell receptor (TCR) signaling and interleukin-2 (IL-2)/STAT5 activation support the suppressive effector function of Treg cells, but how these regulators synergize to control Treg cell homeostasis and function remains unclear. Here we show that TCR-activated posttranslational modification by O-linked N-Acetylglucosamine (O-GlcNAc) stabilizes FOXP3 and activates STAT5, thus integrating these critical signaling pathways. O-GlcNAc-deficient Treg cells develop normally but display modestly reduced FOXP3 expression, strongly impaired lineage stability and effector function, and ultimately fatal autoimmunity in mice. Moreover, deficiency in protein O-GlcNAcylation attenuates IL-2/STAT5 signaling, while overexpression of a constitutively active form of STAT5 partially ameliorates Treg cell dysfunction and systemic inflammation in O-GlcNAc deficient mice. Collectively, our data demonstrate that protein O-GlcNAcylation is essential for lineage stability and effector function in Treg cells.


Assuntos
Acetilglucosamina/metabolismo , Linhagem da Célula/imunologia , Fatores de Transcrição Forkhead/imunologia , Processamento de Proteína Pós-Traducional , Receptores de Antígenos de Linfócitos T/imunologia , Fator de Transcrição STAT5/imunologia , Linfócitos T Reguladores/imunologia , Acetilglucosamina/imunologia , Animais , Autoimunidade , Linhagem da Célula/genética , Feminino , Fatores de Transcrição Forkhead/genética , Genes Reporter , Humanos , Interleucina-2/genética , Interleucina-2/imunologia , Masculino , Camundongos , Camundongos Transgênicos , Cultura Primária de Células , Receptores de Antígenos de Linfócitos T/genética , Fator de Transcrição STAT5/genética , Tolerância a Antígenos Próprios , Transdução de Sinais , Linfócitos T Reguladores/citologia
7.
Fish Shellfish Immunol ; 84: 962-969, 2019 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-30399402

RESUMO

STAT plays important roles in innate immunity during JAK/STAT signaling pathway, and STAT5 is particularly focused due to the existence of duplicated forms in fish and mammal. In Chinese tongue sole, stat5bl was suggested to be a candidate related to Vibrio harveyi resistance based on previous QTL screening. In this study, the full length of stat5bl cDNA was cloned and its expression patterns were analyzed. stat5bl was predominantly expressed in immune tissues, where the highest level was observed in liver, followed by skin and gill. Time course expression patterns were examined in six tissues (liver, skin, gill, kidney, intestine, spleen) after V. harveyi infection. stat5bl could be up-regulated by V. harveyi infection in all tissues except liver, despite the timepoints of peak were different. In contrast, stat5bl was significantly downregulated in liver. To elucidate the role of stat5bl in liver, in vitro RNAi were performed using primary liver cell culture. Knockdown of stat5bl could regulate the expression of genes closely related to JAK/STAT pathway. This study would enlarge our understanding of stat5bl in fish immunity.


Assuntos
Doenças dos Peixes/imunologia , Linguados/genética , Linguados/imunologia , Regulação da Expressão Gênica/imunologia , Imunidade Inata/genética , Fator de Transcrição STAT5/genética , Fator de Transcrição STAT5/imunologia , Sequência de Aminoácidos , Animais , Sequência de Bases , Proteínas de Peixes/química , Proteínas de Peixes/genética , Proteínas de Peixes/imunologia , Perfilação da Expressão Gênica/veterinária , Filogenia , Fator de Transcrição STAT5/química , Alinhamento de Sequência/veterinária , Vibrio , Vibrioses
8.
J Autoimmun ; 95: 1-14, 2018 12.
Artigo em Inglês | MEDLINE | ID: mdl-30446251

RESUMO

Susceptibility to multiple autoimmune diseases is associated with common gene polymorphisms influencing IL-2 signaling and Treg function, making Treg-specific expansion by IL-2 a compelling therapeutic approach to treatment. As an in vivo IL-2 half-life enhancer we used a non-targeted, effector-function-silent human IgG1 as a fusion protein. An IL-2 mutein (N88D) with reduced binding to the intermediate affinity IL-2Rßγ receptor was engineered with a stoichiometry of two IL-2N88D molecules per IgG, i.e. IgG-(IL-2N88D)2. The reduced affinity of IgG-(IL-2N88D)2 for the IL-2Rßγ receptor resulted in a Treg-selective molecule in human whole blood pSTAT5 assays. Treatment of cynomolgus monkeys with single low doses of IgG-(IL-2N88D)2 induced sustained preferential activation of Tregs accompanied by a corresponding 10-14-fold increase in CD4+ and CD8+ CD25+FOXP3+ Tregs; conditions that had no effect on CD4+ or CD8+ memory effector T cells. The expanded cynomolgus Tregs had demethylated FOXP3 and CTLA4 epigenetic signatures characteristic of functionally suppressive cells. Humanized mice had similar selective in vivo responses; IgG-(IL-2N88D)2 increased Tregs while wild-type IgG-IL-2 increased NK cells in addition to Tregs. The expanded human Tregs had demethylated FOXP3 and CTLA4 signatures and were immunosuppressive. These results describe a next-generation immunotherapy using a long-lived and Treg-selective IL-2 that activates and expands functional Tregsin vivo. Patients should benefit from restored immune homeostasis in a personalized fashion to the extent that their autoimmune disease condition dictates opening up the possibility for remissions and cures.


Assuntos
Doenças Autoimunes/terapia , Imunoglobulina G/imunologia , Imunoterapia/métodos , Interleucina-2/imunologia , Linfotoxina-alfa/imunologia , Proteínas Recombinantes de Fusão/imunologia , Linfócitos T Reguladores/efeitos dos fármacos , Animais , Doenças Autoimunes/genética , Doenças Autoimunes/imunologia , Doenças Autoimunes/patologia , Sítios de Ligação , Linfócitos T CD8-Positivos/efeitos dos fármacos , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/patologia , Antígeno CTLA-4/genética , Antígeno CTLA-4/imunologia , Proliferação de Células , Metilação de DNA/efeitos dos fármacos , Modelos Animais de Doenças , Epigênese Genética , Fatores de Transcrição Forkhead/genética , Fatores de Transcrição Forkhead/imunologia , Humanos , Imunoglobulina G/administração & dosagem , Imunoglobulina G/química , Imunoglobulina G/genética , Interleucina-2/administração & dosagem , Interleucina-2/química , Interleucina-2/genética , Subunidade beta de Receptor de Interleucina-2/genética , Subunidade beta de Receptor de Interleucina-2/imunologia , Ativação Linfocitária/efeitos dos fármacos , Linfotoxina-alfa/administração & dosagem , Linfotoxina-alfa/química , Linfotoxina-alfa/genética , Macaca fascicularis , Masculino , Camundongos , Camundongos Transgênicos , Modelos Moleculares , Ligação Proteica , Isoformas de Proteínas/genética , Isoformas de Proteínas/imunologia , Estrutura Secundária de Proteína , Proteínas Recombinantes de Fusão/administração & dosagem , Proteínas Recombinantes de Fusão/química , Proteínas Recombinantes de Fusão/genética , Fator de Transcrição STAT5/genética , Fator de Transcrição STAT5/imunologia , Transdução de Sinais , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/patologia
9.
Nat Commun ; 9(1): 4874, 2018 11 19.
Artigo em Inglês | MEDLINE | ID: mdl-30451838

RESUMO

The metabolic checkpoint kinase mechanistic/mammalian target of rapamycin (mTOR) regulates natural killer (NK) cell development and function, but the exact underlying mechanisms remain unclear. Here, we show, via conditional deletion of Raptor (mTORC1) or Rictor (mTORC2), that mTORC1 and mTORC2 promote NK cell maturation in a cooperative and non-redundant manner, mainly by controlling the expression of Tbx21 and Eomes. Intriguingly, mTORC1 and mTORC2 regulate cytolytic function in an opposing way, exhibiting promoting and inhibitory effects on the anti-tumor ability and metabolism, respectively. mTORC1 sustains mTORC2 activity by maintaining CD122-mediated IL-15 signaling, whereas mTORC2 represses mTORC1-modulated NK cell effector functions by restraining STAT5-mediated SLC7A5 expression. These positive and negative crosstalks between mTORC1 and mTORC2 signaling thus variegate the magnitudes and kinetics of NK cell activation, and help define a paradigm for the modulation of NK maturation and effector functions.


Assuntos
Células Matadoras Naturais/imunologia , Proteína Companheira de mTOR Insensível à Rapamicina/genética , Proteína Regulatória Associada a mTOR/genética , Proteínas com Domínio T/genética , Animais , Diferenciação Celular , Regulação da Expressão Gênica , Humanos , Interleucina-15/genética , Interleucina-15/imunologia , Subunidade beta de Receptor de Interleucina-2/genética , Subunidade beta de Receptor de Interleucina-2/imunologia , Células Matadoras Naturais/citologia , Transportador 1 de Aminoácidos Neutros Grandes/genética , Transportador 1 de Aminoácidos Neutros Grandes/imunologia , Ativação Linfocitária , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Proteína Companheira de mTOR Insensível à Rapamicina/deficiência , Proteína Companheira de mTOR Insensível à Rapamicina/imunologia , Proteína Regulatória Associada a mTOR/deficiência , Proteína Regulatória Associada a mTOR/imunologia , Fator de Transcrição STAT5/genética , Fator de Transcrição STAT5/imunologia , Transdução de Sinais , Proteínas com Domínio T/imunologia
10.
Cell Death Dis ; 9(9): 905, 2018 09 05.
Artigo em Inglês | MEDLINE | ID: mdl-30185773

RESUMO

Foxp3+ regulatory T cells (Tregs) can inhibit immune responses and maintain immune tolerance by secreting immunosuppressive TGF-ß1 and IL-10. However, the efficiency of Tregs become the major obstacle to their use for immunotherapy. In this study, we investigated the relevance of the C-type lectin receptor CD69 to the suppressive function. Compared to CD4+Foxp3+CD69- Tregs (CD69- Tregs), CD4+Foxp3+CD69+ Tregs (CD69+ Tregs) displayed stronger ability to maintain immune tolerance. CD69+ Tregs expressed higher levels of suppression-associated markers such as CTLA-4, ICOS, CD38 and GITR, and secreted higher levels of IL-10 but not TGF-ß1. CD69+ Tregs from Il10+/+ rather than Il10-/- mice significantly inhibit the proliferation of CD4+ T cells. CD69 over-expression stimulated higher levels of IL-10 and c-Maf expression, which was compromised by silencing of STAT3 or STAT5. In addition, the direct interaction of STAT3 with the c-Maf promoter was detected in cells with CD69 over-expression. Moreover, adoptive transfer of CD69+ Tregs but not CD69-Tregs or CD69+ Tregs deficient in IL-10 dramatically prevented the development of inflammatory bowel disease (IBD) in mice. Taken together, CD69 is important to the suppressive function of Tregs by promoting IL-10 production. CD69+ Tregs have the potential to develop new therapeutic approach for autoimmune diseases like IBD.


Assuntos
Antígenos CD/imunologia , Antígenos de Diferenciação de Linfócitos T/imunologia , Colite/imunologia , Tolerância Imunológica/imunologia , Interleucina-10/imunologia , Lectinas Tipo C/imunologia , Linfócitos T Reguladores/imunologia , Transferência Adotiva/métodos , Animais , Linfócitos T CD4-Positivos/imunologia , Antígeno CTLA-4/imunologia , Proliferação de Células/fisiologia , Feminino , Doenças Inflamatórias Intestinais/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Fator de Transcrição STAT3/imunologia , Fator de Transcrição STAT5/imunologia , Fator de Crescimento Transformador beta1/imunologia
11.
Mucosal Immunol ; 11(5): 1398-1407, 2018 09.
Artigo em Inglês | MEDLINE | ID: mdl-29907868

RESUMO

Immune-mediated lung is considered the result of an exacerbated innate injury immune response, although a role for adaptive lymphocytes is emerging. αß T cells specific for S. aureus enterotoxin A orchestrate a Tγδ17 response during lung injury. However, the mechanism driving IL-17 production is unclear. Here, we show a role for IL-2 triggering IL-17 production by lung granular γδ T cells as IL-17 synthesis and neutrophil recruitment was reduced by IL-2 blocking mAbs in vitro and in vivo. Mass cytometry analysis revealed that lung γδ T cells responded directly to IL-2 as evident from STAT5 phosphorylation and RoRγt expression. IL-2 receptor blocking mAbs and JAK inhibition impaired STAT5 phosphorylation and IL-17 release. Moreover, inhalation of S. aureus enterotoxin A induced IL-2 secretion and caspase-1-dependent IL-1ß activation to drive IL-17 production. This T-cell-mediated inflammasome-dependent IL-17 response is maximum when lung Tγδ17 cells were sequentially stimulated first with IL-2 then IL-1ß. Interestingly, when IL-2 is given therapeutically to cancer patients it carries a known risk of lung injury that is largely indistinguishable from that seen in sepsis. Hence, this novel mechanism reveals therapeutic targets treating both acute lung injury and high-dose IL-2 toxicity in cancer.


Assuntos
Interleucina-17/imunologia , Interleucina-1beta/imunologia , Interleucina-2/imunologia , Pulmão/imunologia , Linfócitos T/imunologia , Animais , Anticorpos Monoclonais/imunologia , Caspase 1/imunologia , Janus Quinases/imunologia , Pulmão/microbiologia , Camundongos , Camundongos Endogâmicos C57BL , Neutrófilos/imunologia , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/imunologia , Fosforilação/imunologia , Fator de Transcrição STAT5/imunologia , Saccharomyces cerevisiae/imunologia
12.
J Immunol ; 201(1): 278-295, 2018 07 01.
Artigo em Inglês | MEDLINE | ID: mdl-29752311

RESUMO

Myeloid-derived suppressor cells (MDSCs) are known suppressors of antitumor immunity, affecting amino acid metabolism and T cell function in the tumor microenvironment. However, it is unknown whether MDSCs regulate B cell responses during tumor progression. Using a syngeneic mouse model of lung cancer, we show reduction in percentages and absolute numbers of B cell subsets including pro-, pre-, and mature B cells in the bone marrow (BM) of tumor-bearing mice. The kinetics of this impaired B cell response correlated with the progressive infiltration of MDSCs. We identified that IL-7 and downstream STAT5 signaling that play a critical role in B cell development and differentiation were also impaired during tumor progression. Global impairment of B cell function was indicated by reduced serum IgG levels. Importantly, we show that anti-Gr-1 Ab-mediated depletion of MDSCs not only rescued serum IgG and IL-7 levels but also reduced TGF-ß1, a known regulator of stromal IL-7, suggesting MDSC-mediated regulation of B cell responses. Furthermore, blockade of IL-7 resulted in reduced phosphorylation of downstream STAT5 and B cell differentiation in tumor-bearing mice and administration of TGF-ß-blocking Ab rescued these IL-7-dependent B cell responses. Adoptive transfer of BM-derived MDSCs from tumor-bearing mice into congenic recipients resulted in significant reductions of B cell subsets in the BM and in circulation. MDSCs also suppressed B cell proliferation in vitro in an arginase-dependent manner that required cell-to-cell contact. Our results indicate that tumor-infiltrating MDSCs may suppress humoral immune responses and promote tumor escape from immune surveillance.


Assuntos
Linfócitos B/imunologia , Interleucina-7/imunologia , Neoplasias Pulmonares/imunologia , Células Supressoras Mieloides/imunologia , Fator de Transcrição STAT5/imunologia , Evasão Tumoral/imunologia , Transferência Adotiva , Animais , Linfócitos B/citologia , Células da Medula Óssea/imunologia , Diferenciação Celular/imunologia , Linhagem Celular Tumoral , Proliferação de Células , Técnicas de Cocultura , Feminino , Imunoglobulina G/sangue , Interleucina-7/sangue , Camundongos , Camundongos Endogâmicos C57BL , Células Supressoras Mieloides/transplante , Fosforilação , Transdução de Sinais/imunologia , Fator de Crescimento Transformador beta/sangue , Microambiente Tumoral/imunologia
13.
Nat Commun ; 9(1): 1431, 2018 04 12.
Artigo em Inglês | MEDLINE | ID: mdl-29650953

RESUMO

Heat shock protein 27 (HSP27/HSPB1) is a stress-inducible chaperone that facilitates cancer development by its proliferative and anti-apoptotic functions. The OGX-427 antisense oligonucleotide against HSP27 has been reported to be beneficial against idiopathic pulmonary fibrosis. Here we show that OGX-427 is effective in two murine models of thrombopoietin- and JAKV617F-induced myelofibrosis. OGX-427 limits disease progression and is associated with a reduction in spleen weight, in megakaryocyte expansion and, for the JAKV617F model, in fibrosis. HSP27 regulates the proliferation of JAK2V617F-positive cells and interacts directly with JAK2/STAT5. We also show that its expression is increased in both CD34+ circulating progenitors and in the serum of patients with JAK2-dependent myeloproliferative neoplasms with fibrosis. Our data suggest that HSP27 plays a key role in the pathophysiology of myelofibrosis and represents a new potential therapeutic target for patients with myeloproliferative neoplasms.


Assuntos
Proteínas de Choque Térmico HSP27/genética , Janus Quinase 2/genética , Oligonucleotídeos/farmacologia , Mielofibrose Primária/tratamento farmacológico , Mielofibrose Primária/genética , Fator de Transcrição STAT5/genética , Animais , Células da Medula Óssea/imunologia , Células da Medula Óssea/patologia , Transplante de Medula Óssea , Linhagem Celular Tumoral , Modelos Animais de Doenças , Feminino , Células HEK293 , Proteínas de Choque Térmico HSP27/imunologia , Humanos , Janus Quinase 2/imunologia , Células K562 , Leucócitos/efeitos dos fármacos , Leucócitos/imunologia , Leucócitos/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Terapia de Alvo Molecular , Mutação , Mielofibrose Primária/imunologia , Mielofibrose Primária/patologia , Fator de Transcrição STAT5/imunologia , Trombopoetina/genética , Trombopoetina/imunologia , Transdução Genética , Irradiação Corporal Total
14.
Nat Commun ; 9(1): 1095, 2018 03 15.
Artigo em Inglês | MEDLINE | ID: mdl-29545616

RESUMO

Regulatory T (Treg) cells contribute to the anti-inflammatory response during atherogenesis. Here we show that during atherogenesis Treg cells lose Foxp3 expression and their immunosuppressive function, leading to the conversion of a fraction of these cells into T follicular helper (Tfh) cells. We show that Tfh cells are pro-atherogenic and that their depletion reduces atherosclerosis. Mechanistically, the conversion of Treg cells to Tfh cells correlates with reduced expression of IL-2Rα and pSTAT5 levels and increased expression of IL-6Rα. In vitro, incubation of naive T cells with oxLDL prevents their differentiation into Treg cells. Furthermore, injection of lipid-free Apolipoprotein AI (ApoAI) into ApoE-/- mice reduces intracellular cholesterol levels in Treg cells and prevents their conversion into Tfh cells. Together our results suggest that ApoAI, the main protein in high-density lipoprotein particles, modulates the cellular fate of Treg cells and thus influences the immune response during atherosclerosis.


Assuntos
Apolipoproteína A-I/imunologia , Aterosclerose/fisiopatologia , Diferenciação Celular , Linfócitos T Auxiliares-Indutores/citologia , Linfócitos T Reguladores/citologia , Animais , Apolipoproteína A-I/genética , Aterosclerose/genética , Aterosclerose/imunologia , Feminino , Humanos , Subunidade alfa de Receptor de Interleucina-2/genética , Subunidade alfa de Receptor de Interleucina-2/imunologia , Masculino , Camundongos , Camundongos Knockout , Receptores de Interleucina-6/genética , Receptores de Interleucina-6/imunologia , Fator de Transcrição STAT5/genética , Fator de Transcrição STAT5/imunologia , Linfócitos T Auxiliares-Indutores/imunologia , Linfócitos T Reguladores/imunologia
15.
Blood ; 131(14): 1587-1599, 2018 04 05.
Artigo em Inglês | MEDLINE | ID: mdl-29463562

RESUMO

Monocytes and macrophages play a key role in defending pathogens, removing the dead cells or cell debris, and wound healing. The mammalian target of rapamycin (mTOR) inhibitor rapamycin (RPM) is widely used in clinics to treat patients with organ transplantation or tumors. The role of mTOR in monocyte/macrophage development remains to be clarified. Here we found that mTOR intrinsically controls monocyte/macrophage development, as evidenced by the decreased percentages and cell numbers of CD11b+F4/80+ cells resulting from mTOR inhibition in SCID mice, mTOR-deficient mice, and mixed chimera mice, and the in vitro colony formation and monocyte/macrophage induction assays. However, Lyzs-mTOR knockout mice displayed normal levels of monocytes/macrophages, indicating that mTOR is not essential for the survival and maturation of monocytes/macrophages. Further studies showed that mTOR deficiency significantly reduced macrophage colony-stimulating factor receptor CD115 expression at the transcriptional and translational levels. The molecular mechanism studies indicate that the impaired monocyte/macrophage development caused by mTOR deficiency is mainly a result of the overactivated STAT5 and subsequent downregulation of IRF8, but not the altered cell metabolism and autophagy. Therefore, our work identifies that mTOR is an intrinsic master for monocyte/macrophage development at the early stages through regulating STAT5-IRF8-dependent CD115-expressing pathway. Long-term usage of RPM may cause a defect of myeloid progenitors in bone marrow.


Assuntos
Medula Óssea/imunologia , Fatores Reguladores de Interferon/imunologia , Macrófagos/imunologia , Monócitos/imunologia , Fator de Transcrição STAT5/imunologia , Serina-Treonina Quinases TOR/imunologia , Animais , Regulação da Expressão Gênica/imunologia , Humanos , Fatores Reguladores de Interferon/genética , Macrófagos/citologia , Camundongos , Camundongos Knockout , Camundongos SCID , Monócitos/citologia , Biossíntese de Proteínas/imunologia , Receptor de Fator Estimulador de Colônias de Macrófagos/genética , Receptor de Fator Estimulador de Colônias de Macrófagos/imunologia , Fator de Transcrição STAT5/genética , Transdução de Sinais/genética , Transdução de Sinais/imunologia , Serina-Treonina Quinases TOR/genética , Transcrição Genética/imunologia
16.
J Bone Miner Res ; 33(4): 732-742, 2018 04.
Artigo em Inglês | MEDLINE | ID: mdl-29206332

RESUMO

Osteoblasts provide a microenvironmental niche for B-cell commitment and maturation in the bone marrow (BM). Any abnormity of osteoblasts function may result in the defect of B lymphopoiesis. Signaling from mechanistic target of rapamycin complex 1 (mTORC1) has been implicated in regulating the expansion and differentiation of osteoblasts. Thus, we raise a hypothesis that mTORC1 signaling in osteoblasts plays a vital role in B-cell development. Inactivation of mTORC1 in osterix-expressing cells (mainly osteoblast lineage) through Osx-Cre-directed deletion of Raptor (an mTORC1-specific component) resulted in a reduction in the total B-cell population in the BM, which was due to a block in early B-cell development from the pro-B to pre-B cell stage. Further mechanistic studies revealed that this defect was the result of reduction of interleukin-7 (IL-7) expression in osterix-expressing immature osteoblasts, which caused the abnormality of IL-7/Stat5 signaling in early B lymphocytes, leading to an increased apoptosis of pre-B plus immature B cells. In vitro and in vivo studies demonstrated that the addition of exogenous IL-7 partially restored B lymphopoiesis in the BM of Raptor mutant mice. Furthermore, total BM cells cultured in conditioned media from Raptor null immature osteoblasts or media with anti-IL-7 neutralizing antibody failed to differentiate into pre-B and immature B cells, indicating that inactivation of mTORC1 in immature osteoblast cannot fully support normal B-cell development. Taken together, these findings demonstrate a novel role for mTORC1 in the regulation of bone marrow environments that support B-cell differentiation via regulating IL-7 expression. © 2017 American Society for Bone and Mineral Research.


Assuntos
Linfócitos B/imunologia , Diferenciação Celular/imunologia , Alvo Mecanístico do Complexo 1 de Rapamicina/imunologia , Transdução de Sinais/imunologia , Fator de Transcrição Sp7/imunologia , Animais , Apoptose/genética , Apoptose/imunologia , Linfócitos B/citologia , Diferenciação Celular/genética , Interleucina-7/genética , Interleucina-7/imunologia , Masculino , Alvo Mecanístico do Complexo 1 de Rapamicina/genética , Camundongos , Camundongos Knockout , Osteoblastos/citologia , Osteoblastos/imunologia , Fator de Transcrição STAT5/genética , Fator de Transcrição STAT5/imunologia , Transdução de Sinais/genética , Fator de Transcrição Sp7/genética
17.
Sci Signal ; 10(510)2017 Dec 19.
Artigo em Inglês | MEDLINE | ID: mdl-29259099

RESUMO

Interleukin-2 (IL-2) stimulates both activated CD4+ and CD8+ T cells to proliferate. IL-2 signals through an identical receptor complex and promotes the same dose-dependent phosphorylation of the canonical transcription factor STAT5 in both cell types. Despite this, CD8+ T cells enter the S phase earlier and proliferate to a greater extent than do CD4+ T cells in response to IL-2. We identified distinct IL-2 signaling dynamics in CD4+ and CD8+ T cells. In IL-2-stimulated CD8+ T cells, STAT5 phosphorylation increased rapidly and was sustained for 6 hours. In contrast, CD4+ T cells had a biphasic response, with maxima at 15 min and 2 to 4 hours after stimulation. Both cell types required vesicular trafficking, but only CD4+ T cells required new protein synthesis to maintain high phosphorylation of STAT5. Two subunits of the IL-2 receptor, IL-2Rß and IL-2Rγ, were twice as abundant in CD8+ T cells than in CD4+ T cells. Reduction of IL-2Rß abundance by 50% was sufficient to convert CD8+ T cells to a CD4+ T cell-like signaling pattern and delay S phase entry. These results suggest that the larger pool of IL-2Rß chains in CD8+ T cells is required to sustain IL-2 signaling and contributes to the quantitatively greater proliferative response to IL-2 relative to that of CD4+ T cells. This cell type-specific difference in IL-2Rß abundance appears to tune responses, potentially preventing extensive, autoimmune proliferation of CD4+ T cells, while still enabling sufficient proliferation of CD8+ T cells to control viral infections.


Assuntos
Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Subunidade beta de Receptor de Interleucina-2/imunologia , Interleucina-2/imunologia , Animais , Autoimunidade/imunologia , Proliferação de Células , Feminino , Imunidade Celular/imunologia , Subunidade gama Comum de Receptores de Interleucina/imunologia , Subunidade gama Comum de Receptores de Interleucina/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Fosforilação , Cultura Primária de Células , Proteínas Recombinantes/metabolismo , Fase S , Fator de Transcrição STAT5/imunologia , Fator de Transcrição STAT5/metabolismo
18.
Nat Commun ; 8(1): 1320, 2017 11 06.
Artigo em Inglês | MEDLINE | ID: mdl-29105654

RESUMO

Interleukin-15 (IL-15) is essential for the development and maintenance of natural killer (NK) cells. IL-15 activates STAT5 proteins, which can form dimers or tetramers. We previously found that NK cell numbers are decreased in Stat5a-Stat5b tetramer-deficient double knockin (DKI) mice, but the mechanism was not investigated. Here we show that STAT5 dimers are sufficient for NK cell development, whereas STAT5 tetramers mediate NK cell maturation and the expression of maturation-associated genes. Unlike the defective proliferation of Stat5 DKI CD8+ T cells, Stat5 DKI NK cells have normal proliferation to IL-15 but are susceptible to death upon cytokine withdrawal, with lower Bcl2 and increased active caspases. These findings underscore the importance of STAT5 tetramers in maintaining NK cell homoeostasis. Moreover, defective STAT5 tetramer formation could represent a cause of NK cell immunodeficiency, and interrupting STAT5 tetramer formation might serve to control NK leukaemia.


Assuntos
Células Matadoras Naturais/imunologia , Fator de Transcrição STAT5/imunologia , Animais , Diferenciação Celular/imunologia , Proliferação de Células , Sobrevivência Celular/imunologia , Citocinas/imunologia , Feminino , Expressão Gênica , Técnicas de Introdução de Genes , Homeostase , Células Matadoras Naturais/citologia , Masculino , Camundongos , Camundongos Transgênicos , Modelos Imunológicos , Estrutura Quaternária de Proteína , Proteínas Proto-Oncogênicas c-bcl-2/imunologia , Fator de Transcrição STAT5/química , Fator de Transcrição STAT5/genética
19.
JCI Insight ; 2(19)2017 10 05.
Artigo em Inglês | MEDLINE | ID: mdl-28978795

RESUMO

Foxp3+ Tregs possess potent immunosuppressive activity, which is critical for maintaining immune homeostasis and self-tolerance. Defects in Treg development or function result in inadvertent immune activation and autoimmunity. Despite recent advances in Treg biology, we still do not completely understand the molecular and cellular mechanisms governing the development and suppressive function of these cells. Here, we have demonstrated an essential role of the dedicator of cytokinesis 8 (DOCK8), guanine nucleotide exchange factors required for cytoskeleton rearrangement, cell migration, and immune cell survival in controlling Treg fitness and their function. Treg-specific DOCK8 deletion led to spontaneous multiorgan inflammation in mice due to uncontrolled T cell activation and production of proinflammatory cytokines. In addition, we show that DOCK8-deficient Tregs are defective in competitive fitness and in vivo suppressive function. Furthermore, DOCK8 controls IL-2 signaling, crucial for maintenance and competitive fitness of Tregs, via a STAT5-dependent manner. Our study provides potentially novel insights into the essential function of DOCK8 in Tregs and immune regulation, and it explains the autoimmune manifestations associated with DOCK8 deficiency.


Assuntos
Fatores de Troca do Nucleotídeo Guanina/imunologia , Interleucina-2/imunologia , Linfócitos T Reguladores/imunologia , Animais , Autoimunidade/imunologia , Fatores de Troca do Nucleotídeo Guanina/deficiência , Tolerância Imunológica/imunologia , Inflamação/imunologia , Mediadores da Inflamação/metabolismo , Ativação Linfocitária/imunologia , Camundongos Knockout , Fator de Transcrição STAT5/imunologia , Transdução de Sinais/imunologia
20.
Proc Natl Acad Sci U S A ; 114(46): 12111-12119, 2017 11 14.
Artigo em Inglês | MEDLINE | ID: mdl-29078395

RESUMO

Cytokines critically control immune responses, but how regulatory programs are altered to allow T cells to differentially respond to distinct cytokine stimuli remains poorly understood. Here, we have globally analyzed enhancer elements bound by IL-2-activated STAT5 and IL-21-activated STAT3 in T cells and identified Il2ra as the top-ranked gene regulated by an IL-2-activated STAT5-bound superenhancer and one of the top genes regulated by STAT3-bound superenhancers. Moreover, we found that STAT5 binding was rapidly superenriched at genes highly induced by IL-2 and that IL-2-activated STAT5 binding induces new and augmented chromatin interactions within superenhancer-containing genes. Based on chromatin interaction analysis by paired-end tag (ChIA-PET) sequencing data, we used CRISPR-Cas9 gene editing to target three of the STAT5 binding sites within the Il2ra superenhancer in mice. Each mutation decreased STAT5 binding and altered IL-2-induced Il2ra gene expression, revealing that individual elements within the superenhancer were not functionally redundant and that all were required for normal gene expression. Thus, we demonstrate cooperative utilization of superenhancer elements to optimize gene expression and show that STAT5 mediates IL-2-induced chromatin looping at superenhancers to preferentially regulate highly inducible genes, thereby providing new insights into the mechanisms underlying cytokine-dependent superenhancer function.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Elementos Facilitadores Genéticos , Interleucina-2/genética , Receptores de Interleucina-2/imunologia , Fator de Transcrição STAT5/imunologia , Animais , Sítios de Ligação , Linfócitos T CD8-Positivos/citologia , Sistemas CRISPR-Cas , Cromatina/química , Cromatina/imunologia , Edição de Genes , Regulação da Expressão Gênica , Genes Reporter , Loci Gênicos , Humanos , Interleucina-2/imunologia , Interleucinas/genética , Interleucinas/imunologia , Luciferases/genética , Luciferases/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Mutação , Ligação Proteica , Receptores de Interleucina-2/genética , Fator de Transcrição STAT5/genética , Transdução de Sinais , Transcrição Genética
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