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1.
J Med Chem ; 63(23): 15050-15071, 2020 Dec 10.
Artigo em Inglês | MEDLINE | ID: mdl-33261314

RESUMO

Scaffold hopping and structure-based drug design were employed to identify substituted 4-aminoquinolines and 4-aminonaphthyridines as potent, small molecule inhibitors of tumor necrosis factor alpha (TNFα). Structure-activity relationships in both the quinoline and naphthyridine series leading to the identification of compound 42 with excellent potency and pharmacokinetic profile are discussed. X-ray co-crystal structure analysis and ultracentrifugation experiments clearly demonstrate that these inhibitors distort the TNFα trimer upon binding, leading to aberrant signaling when the trimer binds to TNF receptor 1 (TNFR1). Pharmacokinetic-pharmacodynamic activity of compound 42 in a TNF-induced IL-6 mouse model and in vivo activity in a collagen antibody-induced arthritis model, where it showed biologic-like in vivo efficacy, will be discussed.

2.
ACS Med Chem Lett ; 11(3): 266-271, 2020 Mar 12.
Artigo em Inglês | MEDLINE | ID: mdl-32184955

RESUMO

Necroptosis has been implicated in a variety of disease states, and RIPK3 is one of the kinases identified to play a critical role in this signaling pathway. In an effort to identify RIPK3 kinase inhibitors with a novel profile, mechanistic studies were incorporated at the hit triage stage. Utilization of these assays enabled identification of a Type II DFG-out inhibitor for RIPK3, which was confirmed by protein crystallography. Structure-based drug design on the inhibitors targeting this previously unreported conformation enabled an enhancement in selectivity against key off-target kinases.

3.
Immunity ; 44(6): 1350-64, 2016 06 21.
Artigo em Inglês | MEDLINE | ID: mdl-27261277

RESUMO

T follicular helper (Tfh) cells promote affinity maturation of B cells in germinal centers (GCs), whereas T follicular regulatory (Tfr) cells limit the GC reaction. Store-operated Ca(2+) entry (SOCE) through Ca(2+) release-activated Ca(2+) (CRAC) channels mediated by STIM and ORAI proteins is a fundamental signaling pathway in T lymphocytes. Conditional deletion of Stim1 and Stim2 genes in T cells abolished SOCE and strongly reduced antibody-mediated immune responses following viral infection caused by impaired differentiation and function of Tfh cells. Conversely, aging Stim1Stim2-deficient mice developed humoral autoimmunity with spontaneous autoantibody production due to abolished Tfr cell differentiation in the presence of residual Tfh cells. Mechanistically, SOCE controlled Tfr and Tfh cell differentiation through NFAT-mediated IRF4, BATF, and Bcl-6 transcription-factor expression. SOCE had a dual role in controlling the GC reaction by regulating both Tfh and Tfr cell differentiation, thus enabling protective B cell responses and preventing humoral autoimmunity.


Assuntos
Autoimunidade , Linfócitos B/imunologia , Centro Germinativo/imunologia , Imunidade Humoral , Molécula 1 de Interação Estromal/metabolismo , Molécula 2 de Interação Estromal/metabolismo , Linfócitos T/imunologia , Animais , Fatores de Transcrição de Zíper de Leucina Básica/genética , Fatores de Transcrição de Zíper de Leucina Básica/metabolismo , Canais de Cálcio Ativados pela Liberação de Cálcio/metabolismo , Sinalização do Cálcio , Células Cultivadas , Fatores Reguladores de Interferon/genética , Fatores Reguladores de Interferon/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Fatores de Transcrição NFATC/metabolismo , Proteína ORAI1/metabolismo , Proteínas Proto-Oncogênicas c-bcl-6/genética , Proteínas Proto-Oncogênicas c-bcl-6/metabolismo , Molécula 1 de Interação Estromal/genética , Molécula 2 de Interação Estromal/genética
4.
J Immunol ; 196(2): 573-85, 2016 Jan 15.
Artigo em Inglês | MEDLINE | ID: mdl-26673135

RESUMO

The function of CD4(+) T cells is dependent on Ca(2+) influx through Ca(2+) release-activated Ca(2+) (CRAC) channels formed by ORAI proteins. To investigate the role of ORAI1 in proinflammatory Th1 and Th17 cells and autoimmune diseases, we genetically and pharmacologically modulated ORAI1 function. Immunization of mice lacking Orai1 in T cells with MOG peptide resulted in attenuated severity of experimental autoimmune encephalomyelitis (EAE). The numbers of T cells and innate immune cells in the CNS of ORAI1-deficient animals were strongly reduced along with almost completely abolished production of IL-17A, IFN-γ, and GM-CSF despite only partially reduced Ca(2+) influx. In Th1 and Th17 cells differentiated in vitro, ORAI1 was required for cytokine production but not the expression of Th1- and Th17-specific transcription factors T-bet and RORγt. The differentiation and function of induced regulatory T cells, by contrast, was independent of ORAI1. Importantly, induced genetic deletion of Orai1 in adoptively transferred, MOG-specific T cells was able to halt EAE progression after disease onset. Likewise, treatment of wild-type mice with a selective CRAC channel inhibitor after EAE onset ameliorated disease. Genetic deletion of Orai1 and pharmacological ORAI1 inhibition reduced the leukocyte numbers in the CNS and attenuated Th1/Th17 cell-mediated cytokine production. In human CD4(+) T cells, CRAC channel inhibition reduced the expression of IL-17A, IFN-γ, and other cytokines in a dose-dependent manner. Taken together, these findings support the conclusion that Th1 and Th17 cell function is particularly dependent on CRAC channels, which could be exploited as a therapeutic approach to T cell-mediated autoimmune diseases.


Assuntos
Canais de Cálcio/imunologia , Encefalomielite Autoimune Experimental/imunologia , Subpopulações de Linfócitos T/imunologia , Células Th1/imunologia , Células Th17/imunologia , Animais , Separação Celular , Cromatografia Líquida , Encefalomielite Autoimune Experimental/patologia , Citometria de Fluxo , Humanos , Camundongos , Camundongos Transgênicos , Proteína ORAI1 , Reação em Cadeia da Polimerase em Tempo Real , Medula Espinal/imunologia , Medula Espinal/patologia , Linfócitos T Reguladores/imunologia , Espectrometria de Massas em Tandem
5.
Immunity ; 42(4): 654-64, 2015 Apr 21.
Artigo em Inglês | MEDLINE | ID: mdl-25888258

RESUMO

Missense mutations in the nucleotide-binding oligomerization domain (NOD)-like receptor pyrin domain containing family of gene 12 (Nlrp12) are associated with periodic fever syndromes and atopic dermatitis in humans. Here, we have demonstrated a crucial role for NLRP12 in negatively regulating pathogenic T cell responses. Nlrp12(-/-) mice responded to antigen immunization with hyperinflammatory T cell responses. Furthermore, transfer of CD4(+)CD45RB(hi)Nlrp12(-/-) T cells into immunodeficient mice led to more severe colitis and atopic dermatitis. NLRP12 deficiency did not, however, cause exacerbated ascending paralysis during experimental autoimmune encephalomyelitis (EAE); instead, Nlrp12(-/-) mice developed atypical neuroinflammatory symptoms that were characterized by ataxia and loss of balance. Enhanced T-cell-mediated interleukin-4 (IL-4) production promotes the development of atypical EAE disease in Nlrp12(-/-) mice. These results define an unexpected role for NLRP12 as an intrinsic negative regulator of T-cell-mediated immunity and identify altered NF-κB regulation and IL-4 production as key mediators of NLRP12-associated disease.


Assuntos
Ataxia/imunologia , Colite/imunologia , Dermatite Atópica/imunologia , Encefalomielite Autoimune Experimental/imunologia , Imunidade Celular , Interleucina-4/imunologia , Peptídeos e Proteínas de Sinalização Intracelular/imunologia , Animais , Ataxia/genética , Ataxia/patologia , Autoimunidade , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/patologia , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/patologia , Colite/genética , Colite/patologia , Células Dendríticas/imunologia , Células Dendríticas/patologia , Dermatite Atópica/genética , Dermatite Atópica/patologia , Encefalomielite Autoimune Experimental/genética , Encefalomielite Autoimune Experimental/patologia , Regulação da Expressão Gênica , Interleucina-4/genética , Peptídeos e Proteínas de Sinalização Intracelular/genética , Macrófagos/imunologia , Macrófagos/patologia , Camundongos , Camundongos Knockout , NF-kappa B/genética , NF-kappa B/imunologia , Transdução de Sinais
6.
J Clin Invest ; 124(10): 4549-63, 2014 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-25157823

RESUMO

Calcium signaling is critical for lymphocyte function, and intracellular Ca2+ concentrations are regulated by store-operated Ca2+ entry (SOCE) through Ca2+ release-activated Ca2+ (CRAC) channels. In patients, loss-of-function mutations in CRAC channel components ORAI1 and STIM1 abolish SOCE and are associated with recurrent and chronic viral infections. Here, using mice with conditional deletion of Stim1 and its homolog Stim2 in T cells, we determined that both components are required for the maintenance of virus-specific memory CD8+ T cells and recall responses following secondary infection. In the absence of STIM1 and STIM2, acute viral infections became chronic. Early during infection, STIM1 and STIM2 were required for the differentiation of naive CD8+ T cells into fully functional cytolytic effector cells and mediated the production of cytokines and prevented cellular exhaustion in viral-specific CD8+ effector T cells. Importantly, memory and recall responses by CD8+ T cells required expression of STIM1 and STIM2 in CD4+ T cells. CD4+ T cells lacking STIM1 and STIM2 were unable to provide "help" to CD8+ T cells due to aberrant regulation of CD40L expression. Together, our data indicate that STIM1, STIM2, and CRAC channel function play distinct but synergistic roles in CD4+ and CD8+ T cells during antiviral immunity.


Assuntos
Infecções por Arenaviridae/imunologia , Linfócitos T CD4-Positivos/citologia , Linfócitos T CD8-Positivos/citologia , Canais de Cálcio/metabolismo , Glicoproteínas de Membrana/metabolismo , Animais , Células da Medula Óssea/citologia , Células da Medula Óssea/virologia , Linfócitos T CD4-Positivos/virologia , Ligante de CD40/metabolismo , Linfócitos T CD8-Positivos/virologia , Canais de Cálcio/genética , Sinalização do Cálcio , Moléculas de Adesão Celular/genética , Moléculas de Adesão Celular/metabolismo , Homozigoto , Humanos , Memória Imunológica , Vírus da Coriomeningite Linfocítica , Glicoproteínas de Membrana/genética , Proteínas de Membrana/genética , Proteínas de Membrana/metabolismo , Camundongos , Camundongos Knockout , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/metabolismo , Molécula 1 de Interação Estromal , Molécula 2 de Interação Estromal
7.
Eur J Immunol ; 43(12): 3343-54, 2013 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-23939929

RESUMO

Entry of lymphocytes into secondary lymphoid organs (SLOs) involves intravascular arrest and intracellular calcium ion ([Ca(2+)]i) elevation. TCR activation triggers increased [Ca(2+)]i and can arrest T-cell motility in vitro. However, the requirement for [Ca(2+)]i elevation in arresting T cells in vivo has not been tested. Here, we have manipulated the Ca(2+) release-activated Ca(2+) (CRAC) channel pathway required for [Ca(2+)]i elevation in T cells through genetic deletion of stromal interaction molecule (STIM) 1 or by expression of a dominant-negative ORAI1 channel subunit (ORAI1-DN). Interestingly, the absence of CRAC did not interfere with homing of naïve CD4(+) T cells to SLOs and only moderately reduced crawling speeds in vivo. T cells expressing ORAI1-DN lacked TCR activation induced [Ca(2+)]i elevation, yet arrested motility similar to control T cells in vitro. In contrast, antigen-specific ORAI1-DN T cells had a twofold delayed onset of arrest following injection of OVA peptide in vivo. CRAC channel function is not required for homing to SLOs, but enhances spatiotemporal coordination of TCR signaling and motility arrest.


Assuntos
Linfócitos T CD4-Positivos/imunologia , Canais de Cálcio/imunologia , Cálcio/imunologia , Glicoproteínas de Membrana/imunologia , Animais , Canais de Cálcio/genética , Glicoproteínas de Membrana/genética , Camundongos , Camundongos Knockout , Proteína ORAI1 , Peptídeos/imunologia , Peptídeos/farmacologia , Receptores de Antígenos de Linfócitos T/genética , Receptores de Antígenos de Linfócitos T/imunologia , Molécula 1 de Interação Estromal
8.
EMBO Mol Med ; 5(9): 1311-21, 2013 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-23922331

RESUMO

Store-operated calcium entry (SOCE) through Ca(2+) release-activated Ca(2+) (CRAC) channels regulates the function of many immune cells. Patients with loss-of-function mutations in the CRAC channel genes ORAI1 or STIM1 are immunodeficient and are prone to develop virus-associated tumours. This and the reported role of Ca(2+) signals in cytotoxic lymphocyte function suggest that SOCE may be critical for tumour immune surveillance. Using conditional knock out mice lacking STIM1 and its homologue STIM2, we find that SOCE in CD8(+) T cells is required to prevent the engraftment of melanoma and colon carcinoma cells and to control tumour growth. SOCE is essential for the cytotoxic function of CTLs both in vivo and in vitro by regulating the degranulation of CTLs, their expression of Fas ligand and production of TNF-α and IFN-γ. Our results emphasize an important role of SOCE in antitumour immunity, which is significant given recent reports arguing in favour of CRAC channel inhibition for cancer therapy.


Assuntos
Linfócitos T CD8-Positivos/imunologia , Cálcio/metabolismo , Glicoproteínas de Membrana/metabolismo , Neoplasias/imunologia , Transdução de Sinais , Linfócitos T Citotóxicos/imunologia , Animais , Canais de Cálcio , Glicoproteínas de Membrana/deficiência , Camundongos , Camundongos Knockout , Dados de Sequência Molecular , Análise de Sequência de DNA , Molécula 1 de Interação Estromal , Molécula 2 de Interação Estromal
9.
Cell Mol Life Sci ; 70(15): 2637-56, 2013 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-23052215

RESUMO

Calcium (Ca(2+)) influx is required for the activation and function of all cells in the immune system. It is mediated mainly by store-operated Ca(2+) entry (SOCE) through Ca(2+) release-activated Ca(2+) (CRAC) channels located in the plasma membrane. CRAC channels are composed of ORAI proteins that form the channel pore and are activated by stromal interaction molecules (STIM) 1 and 2. Located in the membrane of the endoplasmic reticulum, STIM1 and STIM2 have the dual function of sensing the intraluminal Ca(2+) concentration in the ER and to activate CRAC channels. A decrease in the ER's Ca(2+) concentration induces STIM multimerization and translocation into puncta close to the plasma membrane where they bind to and activate ORAI channels. Since the identification of ORAI and STIM genes as the principal mediators of CRAC channel function, substantial advances have been achieved in understanding the molecular regulation and physiological role of CRAC channels in cells of the immune system and other organs. In this review, we discuss the mechanisms that regulate CRAC channel function and SOCE, the role of recently identified proteins and mechanisms that modulate the activation of ORAI/STIM proteins and the consequences of CRAC channel dysregulation for lymphocyte function and immunity.


Assuntos
Canais de Cálcio/metabolismo , Cálcio/metabolismo , Moléculas de Adesão Celular/metabolismo , Linfócitos/fisiologia , Proteínas de Membrana/metabolismo , Modelos Moleculares , Proteínas de Neoplasias/metabolismo , Conformação Proteica , Canais de Cálcio/genética , Moléculas de Adesão Celular/genética , Retículo Endoplasmático/metabolismo , Proteínas de Membrana/genética , Proteínas de Neoplasias/genética , Proteína ORAI1 , Oxirredução , Fosforilação , Polimerização , Processamento de Proteína Pós-Traducional/fisiologia , Molécula 1 de Interação Estromal , Molécula 2 de Interação Estromal , Ubiquitinação
11.
J Physiol ; 590(17): 4157-67, 2012 Sep 01.
Artigo em Inglês | MEDLINE | ID: mdl-22615435

RESUMO

Store-operated Ca(2+) entry (SOCE) in cells of the immune system is mediated by Ca(2+) release-activated Ca(2+) (CRAC) channels that are formed by ORAI1 and its homologues ORAI2 and ORAI3. They are activated by stromal interaction molecules (STIM) 1 and 2 in response to depletion of endoplasmic reticulum Ca(2+) stores. Loss-of-function mutations in the human ORAI1 and STIM1 genes abolish CRAC channel function and SOCE in a variety of non-excitable cells including lymphocytes and other immune cells, resulting in a unique clinical syndrome termed CRAC channelopathy. It is dominated by severe immunodeficiency and autoimmunity due to impaired SOCE and defects in the function of several lymphocyte subsets. These include CD8(+) T cells, CD4(+) effector and regulatory T cells, natural killer (NK) cells and B cells. This review provides a concise discussion of the role of CRAC channels in these lymphocyte populations and the regulation of adaptive immune responses to infection, in autoimmunity and inflammation.


Assuntos
Autoimunidade , Canais de Cálcio/imunologia , Canais de Cálcio/metabolismo , Infecções/imunologia , Infecções/metabolismo , Linfócitos/imunologia , Linfócitos/metabolismo , Glicoproteínas de Membrana/imunologia , Glicoproteínas de Membrana/metabolismo , Animais , Canais de Cálcio/genética , Sinalização do Cálcio , Humanos , Síndromes de Imunodeficiência/genética , Síndromes de Imunodeficiência/imunologia , Síndromes de Imunodeficiência/metabolismo , Glicoproteínas de Membrana/genética , Modelos Biológicos , Mutação
12.
Front Biosci (Elite Ed) ; 4: 2253-68, 2012 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-22202035

RESUMO

Calcium signals play a critical role in many cell-type specific effector functions during innate and adaptive immune responses. The predominant mechanism to raise intracellular (Ca²âº) used by most immune cells is store-operated Ca²âº entry (SOCE), whereby the depletion of endoplasmic reticulum (ER) Ca²âº stores triggers the influx of extracellular Ca²âº. SOCE in immune cells is mediated by the highly Ca²âº selective Ca²âº-release-activated Ca²âº (CRAC) channel, encoded by ORAI1, ORAI2 and ORAI3 genes. ORAI proteins are activated by stromal interaction molecules (STIM) 1 and 2, which act as sensors of ER Ca²âº store depletion. The importance of SOCE mediated by STIM and ORAI proteins for immune function is evident from the immunodeficiency and autoimmunity in patients with mutations in STIM1 and ORAI1 genes. These patients and studies in gene-targeted mice have revealed an essential role for ORAI/STIM proteins in the function of several immune cells. This review focuses on recent advances made towards understanding the role of SOCE in immune cells with an emphasis on the immune dysregulation that results from defects in SOCE in human patients and transgenic mice.


Assuntos
Cálcio/metabolismo , Sistema Imunitário/fisiopatologia , Animais , Canais de Cálcio/genética , Canais de Cálcio/fisiologia , Humanos , Sistema Imunitário/citologia , Sistema Imunitário/patologia , Transporte de Íons , Proteínas de Membrana/genética , Proteínas de Membrana/fisiologia , Mutação , Proteínas de Neoplasias/genética , Proteínas de Neoplasias/fisiologia , Proteína ORAI1 , Molécula 1 de Interação Estromal
13.
Nat Immunol ; 12(10): 1010-6, 2011 Sep 04.
Artigo em Inglês | MEDLINE | ID: mdl-21892172

RESUMO

The adaptor ASC contributes to innate immunity through the assembly of inflammasome complexes that activate the cysteine protease caspase-1. Here we demonstrate that ASC has an inflammasome-independent, cell-intrinsic role in cells of the adaptive immune response. ASC-deficient mice showed defective antigen presentation by dendritic cells (DCs) and lymphocyte migration due to impaired actin polymerization mediated by the small GTPase Rac. Genome-wide analysis showed that ASC, but not the cytoplasmic receptor NLRP3 or caspase-1, controlled the mRNA stability and expression of Dock2, a guanine nucleotide-exchange factor that mediates Rac-dependent signaling in cells of the immune response. Dock2-deficient DCs showed defective antigen uptake similar to that of ASC-deficient cells. Ectopic expression of Dock2 in ASC-deficient cells restored Rac-mediated actin polymerization, antigen uptake and chemotaxis. Thus, ASC shapes adaptive immunity independently of inflammasomes by modulating Dock2-dependent Rac activation and actin polymerization in DCs and lymphocytes.


Assuntos
Actinas/química , Proteínas do Citoesqueleto/fisiologia , Proteínas Ativadoras de GTPase/fisiologia , Inflamassomos/fisiologia , Proteínas rac de Ligação ao GTP/metabolismo , Actinas/metabolismo , Imunidade Adaptativa , Animais , Apresentação do Antígeno , Proteínas Reguladoras de Apoptose , Proteínas Adaptadoras de Sinalização CARD , Movimento Celular , Quimiotaxia de Leucócito , Células Dendríticas/imunologia , Proteínas Ativadoras de GTPase/genética , Fatores de Troca do Nucleotídeo Guanina , Linfócitos/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Polimerização , Estabilidade de RNA
14.
Anal Chim Acta ; 701(1): 112-8, 2011 Sep 02.
Artigo em Inglês | MEDLINE | ID: mdl-21763816

RESUMO

A new rapid method for the determination of actinides in emergency concrete and brick samples has been developed at the Savannah River Site Environmental Lab (Aiken, SC, USA) that can be used in emergency response situations or for routine analysis. If a radiological dispersive device (RDD), Improvised Nuclear Device (IND) or nuclear accident occurs, there will be a urgent need for rapid analyses of many different environmental matrices, including building materials such as concrete and brick, to support dose mitigation and environmental clean-up. The new method for actinides in concrete and brick method utilizes a rapid sodium hydroxide fusion method, a lanthanum fluoride matrix removal step, and a column separation process with stacked TEVA, TRU and DGA Resin cartridges. Alpha emitters are prepared using rare earth microprecipitation for counting by alpha spectrometry. The method showed high chemical recoveries and effective removal of interferences. The determination of actinides in concrete and brick sample analysis can be performed in less than 8h with excellent quality for emergency samples. The rapid fusion technique is a rugged sample digestion method that ensures that any refractory actinide particles are effectively digested.

15.
Eur J Immunol ; 41(8): 2260-8, 2011 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-21681738

RESUMO

The Nlrp3 inflammasome has been proposed to play an important role in antifungal host defense. However, studies exploring the role of the inflammasome in antifungal host defense have been limited to the direct effects on IL-1ß processing. Although IL-1ß has important direct effects on the innate immune response, important effects of the caspase-1-dependent cytokines IL-1ß and IL-18 are exerted on the initiation of the adaptive Th1 and Th17 cellular responses. No studies have been employed to assess the impact of the inflammasome on the Th1/Th17 defense mechanisms in vivo during candidiasis. In the present study, we demonstrate an essential role for caspase-1 and ASC (apoptosis-associated speck-like protein containing a caspase recruitment domain) in disseminated candidiasis through regulating antifungal Th1 and Th17 responses. Caspase-1(-/-) and ASC(-/-) mice display diminished Th1/Th17 responses, followed by increased fungal outgrowth and lower survival. These observations identify a critical role for the inflammasome in controlling protective adaptive immune responses during invasive fungal infection.


Assuntos
Candidíase/imunologia , Inflamassomos/imunologia , Células Th1/imunologia , Células Th17/imunologia , Animais , Proteínas Reguladoras de Apoptose , Células da Medula Óssea/imunologia , Células da Medula Óssea/metabolismo , Proteínas Adaptadoras de Sinalização CARD , Candida albicans/imunologia , Candida albicans/fisiologia , Candidíase/microbiologia , Caspase 1/genética , Caspase 1/imunologia , Caspase 1/metabolismo , Proteínas do Citoesqueleto/genética , Proteínas do Citoesqueleto/imunologia , Proteínas do Citoesqueleto/metabolismo , Células Dendríticas/imunologia , Células Dendríticas/metabolismo , Ensaio de Imunoadsorção Enzimática , Feminino , Citometria de Fluxo , Predisposição Genética para Doença , Interações Hospedeiro-Patógeno/imunologia , Imunidade Celular/imunologia , Inflamassomos/genética , Inflamassomos/metabolismo , Interleucina-18/genética , Interleucina-18/imunologia , Interleucina-18/metabolismo , Interleucina-1beta/genética , Interleucina-1beta/imunologia , Interleucina-1beta/metabolismo , Rim/imunologia , Rim/microbiologia , Rim/patologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Células Th1/metabolismo , Células Th17/metabolismo , Fator de Necrose Tumoral alfa/imunologia , Fator de Necrose Tumoral alfa/metabolismo
16.
Toxicol Appl Pharmacol ; 252(3): 289-97, 2011 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-21396389

RESUMO

Acetaminophen (APAP) overdose is the leading cause of acute liver failure in the US and UK. Recent studies implied that APAP-induced injury is partially mediated by interleukin-1ß (IL-1ß), which can activate and recruit neutrophils, exacerbating injury. Mature IL-1ß is formed by caspase-1, dependent on inflammasome activation. The objective of this invetstigation was to evaluate the role of the Nalp3 inflammasome on release of damage associated molecular patterns (DAMPs), hepatic neutrophil accumulation and liver injury (ALT, necrosis) after APAP overdose. Mice deficient for each component of the Nalp3 inflammasome (caspase-1, ASC and Nalp3) were treated with 300mg/kg APAP for 24h; these mice had similar neutrophil recruitment and liver injury as APAP-treated C57Bl/6 wildtype animals. In addition, plasma levels of DAMPs (DNA fragments, keratin-18, hypo- and hyper-acetylated forms of high mobility group box-1 protein) were similarly elevated with no significant difference between wildtype and gene knockout mice. In addition, aspirin treatment, which has been postulated to attenuate cytokine formation and the activation of the Nalp3 inflammasome after APAP, had no effect on release of DAMPs, hepatic neutrophil accumulation or liver injury. Together, these data confirm the release of DAMPs and a sterile inflammatory response after APAP overdose. However, as previously reported minor endogenous formation of IL-1ß and the activation of the Nalp3 inflammasome have little impact on APAP hepatotoxicity. It appears that the Nalp3 inflammasome is not a promising therapeutic target to treat APAP overdose.


Assuntos
Acetaminofen/toxicidade , Proteínas de Transporte/imunologia , Doença Hepática Induzida por Substâncias e Drogas/imunologia , Inflamassomos/imunologia , Neutrófilos/imunologia , Alanina Transaminase/sangue , Animais , Caspase 1/sangue , Glutationa/sangue , Proteína HMGB1/sangue , Inflamação/induzido quimicamente , Inflamação/imunologia , Queratina-18/sangue , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteína 3 que Contém Domínio de Pirina da Família NLR , Estatísticas não Paramétricas
17.
Immunity ; 34(1): 75-84, 2011 Jan 28.
Artigo em Inglês | MEDLINE | ID: mdl-21236705

RESUMO

Peripheral peptidolgycan (PGN) is present within antigen-presenting cells in the central nervous system (CNS) of multiple sclerosis (MS) patients, possibly playing a role in neuroinflammation. Accordingly, PGN is linked with disease progression in the animal model of MS, experimental autoimmune encephalomyelitis (EAE), but the role of specific PGN-sensing proteins is unknown. Here we report that the progression of EAE was dependent on the intracellular PGN sensors NOD1 and NOD2 and their common downstream adaptor molecule, receptor interacting protein 2 (RIP2; also known as RIPK2 and RICK). We found that RIP2, but not toll-like receptor 2 (TLR2), played a critical role in the activation of CNS-infiltrating dendritic cells. Our results suggest that PGN in the CNS is involved in the pathogenesis of EAE through the activation of infiltrating dendritic cells via NOD1-, NOD2-, and RIP2-mediated pathways.


Assuntos
Sistema Nervoso Central/imunologia , Células Dendríticas/metabolismo , Encefalomielite Autoimune Experimental/imunologia , Esclerose Múltipla/imunologia , Proteína Serina-Treonina Quinases de Interação com Receptores/metabolismo , Animais , Movimento Celular/genética , Células Cultivadas , Células Dendríticas/imunologia , Células Dendríticas/patologia , Modelos Animais de Doenças , Progressão da Doença , Encefalomielite Autoimune Experimental/fisiopatologia , Humanos , Inflamação , Camundongos , Camundongos Knockout , Proteína Adaptadora de Sinalização NOD1/genética , Proteína Adaptadora de Sinalização NOD1/imunologia , Proteína Adaptadora de Sinalização NOD1/metabolismo , Proteína Adaptadora de Sinalização NOD2/genética , Proteína Adaptadora de Sinalização NOD2/imunologia , Proteína Adaptadora de Sinalização NOD2/metabolismo , Prostaglandinas/imunologia , Prostaglandinas/metabolismo , Proteína Serina-Treonina Quinase 2 de Interação com Receptor , Proteína Serina-Treonina Quinases de Interação com Receptores/genética , Proteína Serina-Treonina Quinases de Interação com Receptores/imunologia , Transdução de Sinais/genética , Transdução de Sinais/imunologia
18.
Trends Mol Med ; 17(2): 57-64, 2011 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-21163704

RESUMO

The NOD-like receptor (NLR) family members are cytosolic sensors of microbial components and danger signals. A subset of NLRs control inflammasome assembly that results in caspase-1 activation and, in turn, IL-1ß and IL-18 production. Excessive inflammasome activation can cause autoinflammatory disorders, including the hereditary periodic fevers. Autoinflammatory and autoimmune diseases form a disease spectrum of aberrant, immune-mediated inflammation against self, through innate and adaptive immunity. However, the role of inflammasomes in autoimmune disease is less clear than in autoinflammation, despite the numerous effects IL-1ß and IL-18 can have on shaping adaptive immunity. We summarize the role of inflammasomes in autoimmune disorders, highlight the need for a better understanding of inflammasomes in these conditions and offer suggestions for future research directions.


Assuntos
Autoimunidade/imunologia , Inflamassomos/imunologia , Animais , Doenças Autoimunes/imunologia , Autoimunidade/genética , Humanos
19.
Toxicol Sci ; 118(1): 7-18, 2010 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-20538741

RESUMO

Idiosyncratic adverse drug reactions (IADRs) occur in a minority of patients yet account for the majority of postmarketing use restrictions by the Food and Drug Administration. Despite the impact of these toxicities, the underlying mechanisms are still poorly understood. Animal models of IADRs would be beneficial in understanding mechanisms and in developing assays with predictive potential. Recent work exploring the interactions between inflammatory stress and drugs associated with human idiosyncratic drug-induced liver injury (IDILI) has led to the development of the first animal models that apply to a range of drugs. Here, we discuss hypotheses for the mechanisms of IDILI and focus on a murine model of trovafloxacin-induced hepatotoxicity as an example related to the inflammatory stress hypothesis.


Assuntos
Anti-Infecciosos/toxicidade , Doença Hepática Induzida por Substâncias e Drogas/etiologia , Modelos Animais de Doenças , Efeitos Colaterais e Reações Adversas Relacionados a Medicamentos/etiologia , Fluoroquinolonas/toxicidade , Inflamação/metabolismo , Naftiridinas/toxicidade , Animais , Doença Hepática Induzida por Substâncias e Drogas/metabolismo , Efeitos Colaterais e Reações Adversas Relacionados a Medicamentos/metabolismo , Humanos , Estresse Fisiológico/fisiologia
20.
J Immunol ; 184(9): 4610-4, 2010 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-20368281

RESUMO

Multiple sclerosis is an autoimmune disease in which self-reactive T cells attack oligodendrocytes that myelinate axons in the CNS. Experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis, is dependent on caspase-1; however, the role of Nod-like receptors upstream of caspase-1 is unknown. Danger- and pathogen-associated molecular patterns activate Nod-like receptor 3, which activates caspase-1 through the adaptor protein, apoptosis-associated speck-like protein containing CARD (ASC). We report that the progression of EAE is dependent on ASC and caspase-1 but not Nod-like receptor 3. ASC(-/-) mice were even more protected from the progression of EAE than were caspase-1(-/-) mice, suggesting that an inflammasome-independent function of ASC contributes to the progression of EAE. We found that CD4(+) T cells deficient in ASC exhibited impaired survival; accordingly, ASC(-/-) mice had fewer myelin oligodendrocyte glycoprotein-specific T cells in the draining lymph nodes and CNS.


Assuntos
Apoptose/imunologia , Proteínas Adaptadoras de Sinalização CARD/fisiologia , Encefalomielite Autoimune Experimental/imunologia , Animais , Apoptose/genética , Proteínas Adaptadoras de Sinalização CARD/deficiência , Proteínas Adaptadoras de Sinalização CARD/genética , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD4-Positivos/patologia , Proteínas de Transporte/fisiologia , Caspase 1/fisiologia , Proliferação de Células , Sobrevivência Celular/genética , Sobrevivência Celular/imunologia , Células Cultivadas , Sistema Nervoso Central/imunologia , Sistema Nervoso Central/patologia , Citocinas/biossíntese , Progressão da Doença , Encefalomielite Autoimune Experimental/enzimologia , Encefalomielite Autoimune Experimental/genética , Encefalomielite Autoimune Experimental/prevenção & controle , Linfonodos/imunologia , Linfonodos/patologia , Linfopenia/genética , Linfopenia/imunologia , Linfopenia/patologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas da Mielina , Glicoproteína Associada a Mielina/toxicidade , Glicoproteína Mielina-Oligodendrócito , Proteína 3 que Contém Domínio de Pirina da Família NLR
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