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1.
Antioxidants (Basel) ; 13(4)2024 Apr 22.
Artigo em Inglês | MEDLINE | ID: mdl-38671946

RESUMO

The positional cloning of single nucleotide polymorphisms (SNPs) of the neutrophil cytosolic factor 1 (Ncf1) gene, advocating that a low oxidative burst drives autoimmune disease, demands an understanding of the underlying molecular causes. A cellular target could be T cells, which have been shown to be regulated by reactive oxygen species (ROS). However, the pathways by which ROS mediate T cell signaling remain unclear. The adaptor molecule linker for activation of T cells (LAT) is essential for coupling T cell receptor-mediated antigen recognition to downstream responses, and it contains several cysteine residues that have previously been suggested to be involved in redox regulation. To address the possibility that ROS regulate T cell-dependent inflammation through LAT, we established a mouse strain with cysteine-to-serine mutations at positions 120 and 172 (LATSS). We found that redox regulation of LAT through C120 and C172 mediate its localization and phosphorylation. LATSS mice had reduced numbers of double-positive thymocytes and naïve peripheral T cells. Importantly, redox insensitivity of LAT enhanced T cell-dependent autoimmune inflammation in collagen-induced arthritis (CIA), a mouse model of rheumatoid arthritis (RA). This effect was reversed on an NCF1-mutated (NCF1m1j), ROS-deficient, background. Overall, our data show that LAT is redox-regulated, acts to repress T cell activation, and is targeted by ROS induced by NCF1 in antigen-presenting cells (APCs).

2.
Elife ; 112022 05 19.
Artigo em Inglês | MEDLINE | ID: mdl-35587260

RESUMO

Chronic autoimmune diseases are associated with mutations in PTPN22, a modifier of T cell receptor (TCR) signaling. As with all protein tyrosine phosphatases, the activity of PTPN22 is redox regulated, but if or how such regulation can modulate inflammatory pathways in vivo is not known. To determine this, we created a mouse with a cysteine-to-serine mutation at position 129 in PTPN22 (C129S), a residue proposed to alter the redox regulatory properties of PTPN22 by forming a disulfide with the catalytic C227 residue. The C129S mutant mouse showed a stronger T-cell-dependent inflammatory response and development of T-cell-dependent autoimmune arthritis due to enhanced TCR signaling and activation of T cells, an effect neutralized by a mutation in Ncf1, a component of the NOX2 complex. Activity assays with purified proteins suggest that the functional results can be explained by an increased sensitivity to oxidation of the C129S mutated PTPN22 protein. We also observed that the disulfide of native PTPN22 can be directly reduced by the thioredoxin system, while the C129S mutant lacking this disulfide was less amenable to reductive reactivation. In conclusion, we show that PTPN22 functionally interacts with Ncf1 and is regulated by oxidation via the noncatalytic C129 residue and oxidation-prone PTPN22 leads to increased severity in the development of T-cell-dependent autoimmunity.


Assuntos
Doenças Autoimunes , Linfócitos T , Animais , Dissulfetos/metabolismo , Inflamação/metabolismo , Camundongos , Oxirredução , Proteína Tirosina Fosfatase não Receptora Tipo 22/genética , Receptores de Antígenos de Linfócitos T/metabolismo , Linfócitos T/metabolismo
4.
Nat Commun ; 12(1): 610, 2021 01 27.
Artigo em Inglês | MEDLINE | ID: mdl-33504785

RESUMO

The introduction of the CTLA-4 recombinant fusion protein has demonstrated therapeutic effects by selectively modulating T-cell activation in rheumatoid arthritis. Here we show, using a forward genetic approach, that a mutation in the SH3gl1 gene encoding the endocytic protein Endophilin A2 is associated with the development of arthritis in rodents. Defective expression of SH3gl1 affects T cell effector functions and alters the activation threshold of autoreactive T cells, thereby leading to complete protection from chronic autoimmune inflammatory disease in both mice and rats. We further show that SH3GL1 regulates human T cell signaling and T cell receptor internalization, and its expression is upregulated in rheumatoid arthritis patients. Collectively our data identify SH3GL1 as a key regulator of T cell activation, and as a potential target for treatment of autoimmune diseases.


Assuntos
Aciltransferases/deficiência , Artrite Reumatoide/enzimologia , Artrite Reumatoide/imunologia , Doenças Autoimunes/enzimologia , Doenças Autoimunes/imunologia , Ativação Linfocitária/imunologia , Linfócitos T/imunologia , Aciltransferases/genética , Aciltransferases/metabolismo , Animais , Artrite Reumatoide/prevenção & controle , Autoimunidade , Endocitose , Feminino , Humanos , Células Jurkat , Linfonodos/metabolismo , Linfonodos/patologia , Masculino , Camundongos , Mutação/genética , Ratos , Receptores de Antígenos de Linfócitos T/metabolismo , Transdução de Sinais , Regulação para Cima/genética
5.
Proc Natl Acad Sci U S A ; 117(40): 24986-24997, 2020 10 06.
Artigo em Inglês | MEDLINE | ID: mdl-32958661

RESUMO

It has proven difficult to identify the underlying genes in complex autoimmune diseases. Here, we use forward genetics to identify polymorphisms in the vitamin D receptor gene (Vdr) promoter, controlling Vdr expression and T cell activation. We isolated these polymorphisms in a congenic mouse line, allowing us to study the immunomodulatory properties of VDR in a physiological context. Congenic mice overexpressed VDR selectively in T cells, and thus did not suffer from calcemic effects. VDR overexpression resulted in an enhanced antigen-specific T cell response and more severe autoimmune phenotypes. In contrast, vitamin D3-deficiency inhibited T cell responses and protected mice from developing autoimmune arthritis. Our observations are likely translatable to humans, as Vdr is overexpressed in rheumatic joints. Genetic control of VDR availability codetermines the proinflammatory behavior of T cells, suggesting that increased presence of VDR at the site of inflammation might limit the antiinflammatory properties of its ligand.


Assuntos
Inflamação/genética , Receptores de Calcitriol/genética , Linfócitos T/imunologia , Animais , Regulação da Expressão Gênica/genética , Humanos , Inflamação/imunologia , Camundongos , Polimorfismo Genético , Linfócitos T/metabolismo , Vitamina D/genética , Deficiência de Vitamina D/genética , Deficiência de Vitamina D/imunologia
6.
J Clin Invest ; 128(9): 4115-4131, 2018 08 31.
Artigo em Inglês | MEDLINE | ID: mdl-30124469

RESUMO

Congenital neutropenia is characterized by low absolute neutrophil numbers in blood, leading to recurrent bacterial infections, and patients often require life-long granulocyte CSF (G-CSF) support. X-linked neutropenia (XLN) is caused by gain-of-function mutations in the actin regulator Wiskott-Aldrich syndrome protein (WASp). To understand the pathophysiology in XLN and the role of WASp in neutrophils, we here examined XLN patients and 2 XLN mouse models. XLN patients had reduced myelopoiesis and extremely low blood neutrophil number. However, their neutrophils had a hyperactive phenotype and were present in normal numbers in XLN patient saliva. Murine XLN neutrophils were hyperactivated, with increased actin dynamics and migration into tissues. We provide molecular evidence that the hyperactivity of XLN neutrophils is caused by WASp in a constitutively open conformation due to contingent phosphorylation of the critical tyrosine-293 and plasma membrane localization. This renders WASp activity less dependent on regulation by PI3K. Our data show that the amplitude of WASp activity inside a cell could be enhanced by cell-surface receptor signaling even in the context in which WASp is already in an active conformation. Moreover, these data categorize XLN as an atypical congenital neutropenia in which constitutive activation of WASp in tissue neutrophils compensates for reduced myelopoiesis.


Assuntos
Doenças Genéticas Ligadas ao Cromossomo X/genética , Doenças Genéticas Ligadas ao Cromossomo X/metabolismo , Neutropenia/genética , Neutropenia/metabolismo , Neutrófilos/metabolismo , Proteína da Síndrome de Wiskott-Aldrich/genética , Proteína da Síndrome de Wiskott-Aldrich/metabolismo , Animais , Síndrome Congênita de Insuficiência da Medula Óssea , Feminino , Mutação com Ganho de Função , Técnicas de Introdução de Genes , Humanos , Masculino , Camundongos , Camundongos da Linhagem 129 , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Neutropenia/congênito , Neutrófilos/ultraestrutura , Fagocitose , Fosforilação , Conformação Proteica , Proteína da Síndrome de Wiskott-Aldrich/química
7.
Infect Immun ; 85(10)2017 10.
Artigo em Inglês | MEDLINE | ID: mdl-28760930

RESUMO

The obligate intracellular parasite Toxoplasma gondii can actively infect any nucleated cell type, including cells from the immune system. The rapid transfer of T. gondii from infected dendritic cells to effector natural killer (NK) cells may contribute to the parasite's sequestration and shielding from immune recognition shortly after infection. However, subversion of NK cell functions, such as cytotoxicity or production of proinflammatory cytokines, such as gamma interferon (IFN-γ), upon parasite infection might also be beneficial to the parasite. In the present study, we investigated the effects of T. gondii infection on NK cells. In vitro, infected NK cells were found to be poor at killing target cells and had reduced levels of IFN-γ production. This could be attributed in part to the inability of infected cells to form conjugates with their target cells. However, even upon NK1.1 cross-linking of NK cells, the infected NK cells also exhibited poor degranulation and IFN-γ production. Similarly, NK cells infected in vivo were also poor at killing target cells and producing IFN-γ. Increased levels of transforming growth factor ß production, as well as increased levels of expression of SHP-1 in the cytosol of infected NK cells upon infection, were observed in infected NK cells. However, the phosphorylation of STAT4 was not altered in infected NK cells, suggesting that transcriptional regulation mediates the reduced IFN-γ production, which was confirmed by quantitative PCR. These data suggest that infection of NK cells by T. gondii impairs NK cell recognition of target cells and cytokine release, two mechanisms that independently could enhance T. gondii survival.


Assuntos
Imunomodulação , Células Matadoras Naturais/microbiologia , Células Matadoras Naturais/fisiologia , Toxoplasma/imunologia , Toxoplasmose Animal/imunologia , Animais , Citotoxicidade Imunológica , Células Dendríticas/imunologia , Células Dendríticas/microbiologia , Interações Hospedeiro-Parasita , Interferon gama/biossíntese , Interferon gama/imunologia , Células Matadoras Naturais/imunologia , Camundongos , Fosforilação , Proteína Tirosina Fosfatase não Receptora Tipo 6/biossíntese , Proteína Tirosina Fosfatase não Receptora Tipo 6/genética , Fator de Transcrição STAT4/metabolismo , Toxoplasma/fisiologia , Fator de Crescimento Transformador beta/biossíntese
10.
Lab Anim (NY) ; 37(4): 161-3, 2008 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-18356915

RESUMO

At the authors' facility, housing arrangements for Xenopus laevis were cumbersome and labor-intensive, requiring technicians to wash frog tanks by hand several times a week. The authors describe an alternative housing solution they implemented by modifying a rack system that was originally used to maintain zebrafish. The rack's self-contained water circulation and filtration system saved technicians time and labor, and a commercial chiller attached to the mechanism efficiently controlled frogs' environmental temperature.


Assuntos
Criação de Animais Domésticos , Abrigo para Animais , Xenopus laevis/fisiologia , Animais , Meio Ambiente
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