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1.
PLoS One ; 18(1): e0280590, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-36662882

RESUMO

C-C motif chemokine receptor-like 2 (CCRL2) is a non-signaling 7 transmembrane receptor that binds chemotactic ligands to shape leukocyte recruitment to sites of inflammation. However, there is a lack of consensus on the ligands that directly bind CCRL2 or their functional impact. Studies with CCRL2 knockout mice have demonstrated that neutrophils have impaired degranulation and migration in response to CXCL8, where the underlying molecular mechanism is proposed to be due to the formation of CCRL2 heterodimers with the chemokine receptor CXCR2. Herein, we characterized the ligands that bind directly to CCRL2 and interrogated the impact of CCRL2 neutralization on CXCL8 signaling in neutrophils using pharmacological antibody tools. Using flow cytometry and Surface Plasmon Resonance microscopy (SPRm) cell binding experiments, we confirmed that chemerin, but not previously reported C-C chemokines, binds CCRL2. Furthermore, we identified human and mouse CCRL2 antibodies that neutralized chemerin binding to CCRL2. Unexpectedly, we found that neutralization of CCRL2 with these antibodies did not attenuate CXCL8-induced human neutrophil degranulation nor CXCL8-induced murine neutrophil recruitment to the peritoneum. Based on the observed differences in modulating CCRL2 function with neutralizing antibodies compared to the reported CCRL2 deficient murine models, we hypothesize that the ligand binding function of CCRL2 is dispensable for CXCL8 signaling in neutrophils. Finally, extensive profiling of CCRL2 expression on peripheral blood leukocytes revealed monocytes, dendritic cells (DC), and subpopulations of natural killer T (NKT) cells as additional targets, highlighting potential roles for CCRL2 in human cell types beyond neutrophils that warrants future investigation.


Assuntos
Ativação de Neutrófilo , Receptores CCR , Humanos , Animais , Camundongos , Receptores CCR/metabolismo , Anticorpos Neutralizantes/farmacologia , Ligantes , Transdução de Sinais , Interleucina-8 , Neutrófilos/metabolismo
2.
J Exp Med ; 202(6): 841-51, 2005 Sep 19.
Artigo em Inglês | MEDLINE | ID: mdl-16172261

RESUMO

Experimental autoimmune encephalomyelitis (EAE), a Th1-mediated inflammatory disease of the central nervous system (CNS), is a model of human multiple sclerosis. Cytosolic phospholipase A2alpha (cPLA2alpha), which initiates production of prostaglandins, leukotrienes, and platelet-activating factor, is present in EAE lesions. Using myelin oligodendrocyte glycoprotein (MOG) immunization, as well as an adoptive transfer model, we showed that cPLA2alpha-/- mice are resistant to EAE. Histologic examination of the CNS from MOG-immunized mice revealed extensive inflammatory lesions in the cPLA2alpha+/- mice, whereas the lesions in cPLA2alpha-/- mice were reduced greatly or completely absent. MOG-specific T cells generated from WT mice induced less severe EAE in cPLA2alpha-/- mice compared with cPLA2alpha+/- mice, which indicates that cPLA2alpha plays a role in the effector phase of EAE. Additionally, MOG-specific T cells from cPLA2alpha-/- mice, transferred into WT mice, induced EAE with delayed onset and lower severity compared with EAE that was induced by control cells; this indicates that cPLA2alpha also plays a role in the induction phase of EAE. MOG-specific T cells from cPLA2alpha-/- mice were deficient in production of Th1-type cytokines. Consistent with this deficiency, in vivo administration of IL-12 rendered cPLA2alpha-/- mice susceptible to EAE. Our data indicate that cPLA2alpha plays an important role in EAE development and facilitates differentiation of T cells toward the Th1 phenotype.


Assuntos
Diferenciação Celular/imunologia , Citosol/enzimologia , Encefalomielite Autoimune Experimental/enzimologia , Encefalomielite Autoimune Experimental/genética , Fosfolipases A/deficiência , Células Th1/imunologia , Animais , Encefalomielite Autoimune Experimental/imunologia , Feminino , Fosfolipases A2 do Grupo IV , Imunidade Inata/genética , Imunofenotipagem , Interleucina-12/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas da Mielina , Glicoproteína Associada a Mielina/imunologia , Glicoproteína Mielina-Oligodendrócito , Fosfolipases A/genética , Fosfolipases A/metabolismo , Fosfolipases A2 , Medula Espinal/imunologia , Medula Espinal/patologia , Células Th1/citologia
3.
Bioorg Med Chem Lett ; 19(13): 3485-8, 2009 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-19464884

RESUMO

Tpl2 (cot/MAP3K8) is an upstream kinase of MEK in the ERK pathway. It plays an important role in Tumor Necrosis Factor-alpha (TNF-alpha) production and signaling. We have discovered that 8-halo-4-(3-chloro-4-fluoro-phenylamino)-6-[(1H-[1,2,3]triazol-4-ylmethyl)-amino]-quinoline-3-carbonitriles (4) are potent inhibitors of this enzyme. In order to improve the inhibition of TNF-alpha production in LPS-stimulated human blood, a series of analogs with a variety of substitutions around the triazole moiety were studied. We found that a cyclic amine group appended to the triazole ring could considerably enhance potency, aqueous solubility, and cell membrane permeability. Optimization of these cyclic amine groups led to the identification of 8-chloro-4-(3-chloro-4-fluorophenylamino)-6-((1-(1-ethylpiperidin-4-yl)-1H-1,2,3-triazol-4-yl)methylamino)quinoline-3-carbonitrile (34). In a LPS-stimulated rat inflammation model, compound 34 showed good efficacy in inhibiting TNF-alpha production.


Assuntos
Anti-Inflamatórios/química , MAP Quinase Quinase Quinases/antagonistas & inibidores , Nitrilas/química , Inibidores de Proteínas Quinases/química , Proteínas Proto-Oncogênicas/antagonistas & inibidores , Quinolinas/química , Fator de Necrose Tumoral alfa/sangue , Animais , Anti-Inflamatórios/síntese química , Anti-Inflamatórios/farmacocinética , Feminino , Humanos , Lipopolissacarídeos/farmacologia , MAP Quinase Quinase Quinases/metabolismo , Monócitos/efeitos dos fármacos , Monócitos/imunologia , Nitrilas/síntese química , Nitrilas/farmacocinética , Inibidores de Proteínas Quinases/síntese química , Inibidores de Proteínas Quinases/farmacocinética , Proteínas Proto-Oncogênicas/metabolismo , Quinolinas/síntese química , Quinolinas/farmacocinética , Ratos , Ratos Sprague-Dawley , Fator de Necrose Tumoral alfa/biossíntese
4.
J Neuroimmunol ; 204(1-2): 29-37, 2008 Nov 15.
Artigo em Inglês | MEDLINE | ID: mdl-18829119

RESUMO

Cytosolic phospholipase A2 alpha (cPLA2 alpha) is the rate-limiting enzyme for release of arachidonic acid, which is converted primarily to prostaglandins via the cyclooxygenase (COX) 1/2 pathways, and leukotrienes via the 5-lipoxygenase (LO) pathway. We utilized inhibitors of cPLA2 alpha, COX-1/2 and 5-LO to determine the potential roles of these enzymes in development of experimental autoimmune encephalomyelitis (EAE), an animal model of multiple sclerosis (MS). Blocking cPLA2 alpha prevented EAE development and greatly reduced antigen-induced production of Th1-type cytokines and IL-17. Blocking COX-1/2 delayed onset and reduced severity of EAE, and reduced production of Th1-type cytokines, but not IL-17. Blocking 5-LO delayed onset and reduced cumulative severity of EAE, but did not reduce production of Th1-type cytokines or IL-17. Finally, blockade of cPLA2 alpha from the onset of clinical EAE reduced duration of EAE relapses. Therefore, cPLA2 alpha represents a potential therapeutic target for treatment of MS.


Assuntos
Encefalomielite Autoimune Experimental/prevenção & controle , Encefalomielite Autoimune Experimental/fisiopatologia , Fosfolipases A2 do Grupo IV/antagonistas & inibidores , Células Th1/fisiologia , Análise de Variância , Animais , Benzoatos/farmacologia , Benzoatos/uso terapêutico , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Inibidores de Ciclo-Oxigenase/farmacologia , Inibidores de Ciclo-Oxigenase/uso terapêutico , Citocinas/metabolismo , Modelos Animais de Doenças , Encefalomielite Autoimune Experimental/induzido quimicamente , Encefalomielite Autoimune Experimental/enzimologia , Inibidores Enzimáticos/farmacologia , Inibidores Enzimáticos/uso terapêutico , Feminino , Glicoproteínas , Hidroxiureia/análogos & derivados , Hidroxiureia/farmacologia , Hidroxiureia/uso terapêutico , Inibidores de Lipoxigenase/farmacologia , Inibidores de Lipoxigenase/uso terapêutico , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Endogâmicos NOD , Glicoproteína Mielina-Oligodendrócito , Naproxeno/farmacologia , Naproxeno/uso terapêutico , Fragmentos de Peptídeos , Índice de Gravidade de Doença , Vazamento Acidental em Seveso , Sulfonamidas/farmacologia , Sulfonamidas/uso terapêutico , Células Th1/efeitos dos fármacos , Fatores de Tempo
5.
J Neuroimmunol ; 182(1-2): 124-34, 2007 Jan.
Artigo em Inglês | MEDLINE | ID: mdl-17182110

RESUMO

Interactions between PD-1 and its two differentially expressed ligands, PD-L1 and PD-L2, attenuate T cell activation and effector function. To determine the role of these molecules in autoimmune disease of the CNS, PD-1-/-, PD-L1-/- and PD-L2-/- mice were generated and immunized to induce experimental autoimmune encephalomyelitis (EAE). PD-1-/- and PD-L1-/- mice developed more severe EAE than wild type and PD-L2-/- mice. Consistent with this, PD-1-/- and PD-L1-/- cells produced elevated levels of the pro-inflammatory cytokines IFN-gamma, TNF, IL-6 and IL-17. These results demonstrate that interactions between PD-1/PD-L1, but not PD-1/PDL-2, are crucial in attenuating T cell responses in EAE.


Assuntos
Antígenos de Diferenciação/metabolismo , Antígeno B7-1/metabolismo , Encefalomielite Autoimune Experimental/fisiopatologia , Glicoproteínas de Membrana/metabolismo , Peptídeos/metabolismo , Animais , Antígeno B7-H1 , Encefalomielite Autoimune Experimental/imunologia , Encefalomielite Autoimune Experimental/metabolismo , Encefalomielite Autoimune Experimental/patologia , Glicoproteínas/imunologia , Humanos , Interferon gama/biossíntese , Interleucina-17/biossíntese , Interleucina-6/biossíntese , Linfonodos/metabolismo , Linfonodos/patologia , Ativação Linfocitária , Glicoproteínas de Membrana/deficiência , Camundongos , Camundongos Knockout , Camundongos Transgênicos , Glicoproteína Mielina-Oligodendrócito , Fragmentos de Peptídeos/imunologia , Peptídeos/deficiência , Proteína 2 Ligante de Morte Celular Programada 1 , Receptor de Morte Celular Programada 1 , Índice de Gravidade de Doença , Linfócitos T/imunologia , Fator de Necrose Tumoral alfa/biossíntese
6.
Neurosci Lett ; 332(3): 185-9, 2002 Nov 08.
Artigo em Inglês | MEDLINE | ID: mdl-12399011

RESUMO

Experimental autoimmune encephalomyelitis (EAE) is an inflammatory disease of the central nervous system (CNS) that can be induced in susceptible mice by the transfer of autoreactive T cells that recognize myelin basic protein (MBP). The onset and subsequent recovery from disease are associated with distinct patterns of cytokine and chemokine expression within the inflammatory lesions of the CNS. Given the likely importance of the local cytokine milieu in regulating the disease process, it would be preferable to administer cytokines locally to the CNS and reduce systemic delivery in order to evaluate their immunoregulatory roles in EAE. For this purpose, we have used retrovirally transduced T cells from MBP-specific T cell receptor transgenic mice in an attempt to target cytokine delivery to the CNS where MBP is primarily expressed. We have found that T cells expressing granulocyte macrophage colony-stimulating factor (GM-CSF) induce severe, chronic EAE from which mice fail to recover. Our results indicate that increased local GM-CSF expression could play an important role in inducing chronic EAE.


Assuntos
Encefalomielite Autoimune Experimental/patologia , Fator Estimulador de Colônias de Granulócitos e Macrófagos/fisiologia , Retroviridae/genética , Linfócitos T/fisiologia , Animais , Antígenos/imunologia , Fator Estimulador de Colônias de Granulócitos e Macrófagos/biossíntese , Fator Estimulador de Colônias de Granulócitos e Macrófagos/genética , Camundongos , Camundongos Transgênicos , Proteína Básica da Mielina/imunologia , Proteína Básica da Mielina/fisiologia , RNA Mensageiro/biossíntese , Transdução Genética
7.
J Allergy Clin Immunol ; 115(2): 309-15, 2005 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-15696086

RESUMO

BACKGROUND: T(H)2-mediated allergic asthma is characterized by eosinophilia, mucus overproduction, and airway hyperresponsiveness (AHR). Although it is clear that T(H)2 cells and their cytokines play an important role in AHR, the roles of T(H)1 cells and neutrophils in AHR are controversial. OBJECTIVE: We sought to determine the roles of T(H)1 cells and neutrophils in AHR. METHODS: Ovalbumin-specific CD4(+) T cells were purified from DO11.10 mice, differentiated into T(H)1 cells, and injected into naive BALB/c, IL-4RalphaKO, or IL-8RKO mice. After ovalbumin antigen challenge, cytokine mRNA levels in lung samples, as well as inflammatory cell types and numbers in bronchoalveolar lavage fluid (BALF), were determined. AHR was assessed by measuring resistance in tracheostomized mice and enhanced pause in freely moving mice. RESULTS: T(H)1 cells induced AHR as robust as T(H)2 cells. They also induced lung inflammation dominated by neutrophils. Neither AHR nor inflammation were reduced when T(H)1 cells were transferred into IL-4RalphaKO mice. When IL-8RKO mice were used as recipients of T(H)1 cells, neutrophilia was greatly reduced, but the AHR was as strong as that seen in wild-type mice. On the other hand, dexamethasone treatment had no effect on neutrophilia but has significantly reduced AHR. Reduction in AHR was accompanied by a reduction in the numbers of lymphocytes and macrophages in BALF. CONCLUSIONS: T(H)1 cells can induce strong AHR independent of IL-4 and IL-13. The AHR is associated with the presence of lymphocytes and macrophages, but not neutrophils, in BALF. Our results point to a pathway whereby T(H)1 cells mediate AHR independent of neutrophilic inflammation.


Assuntos
Hiper-Reatividade Brônquica/fisiopatologia , Bronquite/patologia , Neutrófilos/patologia , Células Th1/imunologia , Transferência Adotiva , Animais , Hiper-Reatividade Brônquica/imunologia , Hiper-Reatividade Brônquica/metabolismo , Citocinas/genética , Dexametasona/farmacologia , Glucocorticoides/farmacologia , Interleucina-13/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Muco/metabolismo , Ovalbumina/farmacologia , RNA Mensageiro/metabolismo , Receptores de Interleucina-8A/deficiência , Índice de Gravidade de Doença , Células Th2/imunologia
8.
Biochem Biophys Res Commun ; 324(2): 534-46, 2004 Nov 12.
Artigo em Inglês | MEDLINE | ID: mdl-15474460

RESUMO

Matrix metalloproteinases (MMPs) degrade extracellular matrix proteins, which may be responsible for enlargement of alveoli in chronic obstructive pulmonary disease (COPD) and remodeling of pulmonary tissue associated with chronic asthma. Here, we provide novel evidence that MMP-12 is expressed and secreted by normal human bronchial epithelial cell cultures (NHBECs) and reveal the regulation of MMP-12 gene expression by tumor necrosis factor-alpha (TNF-alpha), epidermal growth factor (EGF), and interferon gamma (IFN-gamma). Reverse transcription-polymerase chain reaction analyses demonstrated MMP-12 mRNA presence in unstimulated differentiated NHBEC cultures. Cultures stimulated independently with EGF or IFN-gamma failed to alter MMP-12 mRNA abundance, while TNF-alpha, TNF-alpha+EGF, or TNF-alpha+IFN-gamma elicited relatively early (6 h) peak increases in MMP-12 mRNA levels. Western blot analyses specifically indicated the presence of MMP-12 in differentiated NHBEC-conditioned media. These findings indicate that the bronchial epithelium may be an important source of elastolytic activity in COPD and tissue remodeling in chronic asthma.


Assuntos
Asma/metabolismo , Brônquios/citologia , Células Epiteliais/citologia , Metaloendopeptidases/metabolismo , Ar , Western Blotting , Caseínas/química , Células Cultivadas , Meios de Cultivo Condicionados/farmacologia , Células Epiteliais/metabolismo , Proteínas de Fluorescência Verde/metabolismo , Humanos , Interferon gama/metabolismo , Metaloproteinase 12 da Matriz , Mucinas/metabolismo , Estrutura Terciária de Proteína , Doença Pulmonar Obstrutiva Crônica/metabolismo , RNA Mensageiro/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Fatores de Tempo
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