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1.
Nat Immunol ; 13(9): 832-42, 2012 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-22842346

RESUMO

The mechanisms by which tumor microenvironments modulate nucleic acid-mediated innate immunity remain unknown. Here we identify the receptor TIM-3 as key in circumventing the stimulatory effects of nucleic acids in tumor immunity. Tumor-associated dendritic cells (DCs) in mouse tumors and patients with cancer had high expression of TIM-3. DC-derived TIM-3 suppressed innate immune responses through the recognition of nucleic acids by Toll-like receptors and cytosolic sensors via a galectin-9-independent mechanism. In contrast, TIM-3 interacted with the alarmin HMGB1 to interfere with the recruitment of nucleic acids into DC endosomes and attenuated the therapeutic efficacy of DNA vaccination and chemotherapy by diminishing the immunogenicity of nucleic acids released from dying tumor cells. Our findings define a mechanism whereby tumor microenvironments suppress antitumor immunity mediated by nucleic acids.


Assuntos
Células Dendríticas/imunologia , Proteína HMGB1/imunologia , Imunidade Inata , Neoplasias/imunologia , Ácidos Nucleicos/imunologia , Receptores Virais/imunologia , Microambiente Tumoral/imunologia , Animais , Células Dendríticas/metabolismo , Ensaio de Imunoadsorção Enzimática , Citometria de Fluxo , Proteína HMGB1/metabolismo , Receptor Celular 2 do Vírus da Hepatite A , Humanos , Immunoblotting , Vigilância Imunológica/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Microscopia Confocal , Microscopia de Fluorescência , Neoplasias/metabolismo , Receptores de Reconhecimento de Padrão/imunologia , Receptores Virais/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa
2.
FASEB J ; 36(1): e22065, 2022 01.
Artigo em Inglês | MEDLINE | ID: mdl-34847625

RESUMO

Neutrophil trafficking is a key component of the inflammatory response. Here, we have investigated the role of the immunomodulatory lectin Galectin-9 (Gal-9) on neutrophil recruitment. Our data indicate that Gal-9 is upregulated in the inflamed vasculature of RA synovial biopsies and report the release of Gal-9 into the extracellular environment following endothelial cell activation. siRNA knockdown of endothelial Gal-9 resulted in reduced neutrophil adhesion and neutrophil recruitment was significantly reduced in Gal-9 knockout mice in a model of zymosan-induced peritonitis. We also provide evidence for Gal-9 binding sites on human neutrophils; Gal-9 binding induced neutrophil activation (increased expression of ß2 integrins and reduced expression of CD62L). Intra-vital microscopy confirmed a pro-recruitment role for Gal-9, with increased numbers of transmigrated neutrophils following Gal-9 administration. We studied the role of both soluble and immobilized Gal-9 on human neutrophil recruitment. Soluble Gal-9 significantly strengthened the interaction between neutrophils and the endothelium and inhibited neutrophil crawling on ICAM-1. When immobilized, Gal-9 functioned as an adhesion molecule and captured neutrophils from the flow. Neutrophils adherent to Gal-9 exhibited a spread/activated phenotype that was inhibited by CD18 and CD44 neutralizing antibodies, suggesting a role for these molecules in the pro-adhesive effects of Gal-9. Our data indicate that Gal-9 is expressed and released by the activated endothelium and functions both in soluble form and when immobilized as a neutrophil adhesion molecule. This study paves the way for further investigation of the role of Gal-9 in leukocyte recruitment in different inflammatory settings.


Assuntos
Antígenos CD18/metabolismo , Galectinas/metabolismo , Células Endoteliais da Veia Umbilical Humana/metabolismo , Receptores de Hialuronatos/metabolismo , Neutrófilos/metabolismo , Migração Transendotelial e Transepitelial , Animais , Adesão Celular , Humanos , Camundongos
3.
Immunity ; 41(2): 270-82, 2014 Aug 21.
Artigo em Inglês | MEDLINE | ID: mdl-25065622

RESUMO

The ß-galactoside-binding protein galectin-9 is critical in regulating the immune response, but the mechanism by which it functions remains unclear. We have demonstrated that galectin-9 is highly expressed by induced regulatory T cells (iTreg) and was crucial for the generation and function of iTreg cells, but not natural regulatory T (nTreg) cells. Galectin-9 expression within iTreg cells was driven by the transcription factor Smad3, forming a feed-forward loop, which further promoted Foxp3 expression. Galectin-9 increased iTreg cell stability and function by directly binding to its receptor CD44, which formed a complex with transforming growth factor-ß (TGF-ß) receptor I (TGF-ßRI), and activated Smad3. Galectin-9 signaling was further found to regulate iTreg cell induction by dominantly acting through the CNS1 region of the Foxp3 locus. Our data suggest that exogenous galectin-9, in addition to being an effector molecule for Treg cells, acts synergistically with TGF-ß to enforce iTreg cell differentiation and maintenance.


Assuntos
Fatores de Transcrição Forkhead/biossíntese , Galectinas/imunologia , Receptores de Hialuronatos/imunologia , Linfócitos T Reguladores/imunologia , Animais , Diferenciação Celular/imunologia , Colite/genética , Colite/imunologia , Galectinas/genética , Receptor Celular 2 do Vírus da Hepatite A , Ativação Linfocitária/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas Serina-Treonina Quinases/imunologia , Receptor do Fator de Crescimento Transformador beta Tipo I , Receptores de Fatores de Crescimento Transformadores beta/imunologia , Receptores Virais/imunologia , Transdução de Sinais/imunologia , Proteína Smad3/imunologia , Fator de Crescimento Transformador beta/imunologia
4.
J Immunol ; 199(8): 2721-2728, 2017 10 15.
Artigo em Inglês | MEDLINE | ID: mdl-28877989

RESUMO

Stimulation of several TNF receptor family proteins has been shown to dampen inflammatory disease in murine models through augmenting the number and/or activity of regulatory T cells (Tregs). We recently found that one molecule, 4-1BB, used binding to Galectin-9 to exert its immunosuppressive effects and drive expansion of CD8+Foxp3- Tregs. We now show that ligation of another TNFR family molecule, DR3, which has previously been found to strongly expand CD4+Foxp3+ Tregs and suppress inflammation, also requires Galectin-9. We found that the extracellular region of DR3 directly binds to Galectin-9, and that Galectin-9 associates with DR3 in Tregs. From studies in vitro with Galectin-9-/- CD4+ T cells and Tregs, we found that stimulatory activity induced by ligating DR3 was in part dependent on Galectin-9. In vivo, in a model of experimental autoimmune encephalomyelitis, we show that an agonist of DR3 suppressed disease, correlating with expansion of CD4+Foxp3+ Tregs, and this protective effect was lost in Galectin-9-/- mice. Similar results were seen in an allergic lung inflammation model. Thus, we demonstrate a novel function of Galectin-9 in facilitating activity of DR3 related to Treg-mediated suppression.


Assuntos
Encefalomielite Autoimune Experimental/imunologia , Galectinas/metabolismo , Inflamação/imunologia , Esclerose Múltipla/imunologia , Subpopulações de Linfócitos T/imunologia , Linfócitos T Reguladores/imunologia , Membro 9 da Superfamília de Receptores de Fatores de Necrose Tumoral/metabolismo , Animais , Proliferação de Células , Células Cultivadas , Fatores de Transcrição Forkhead/metabolismo , Galectinas/genética , Humanos , Tolerância Imunológica , Ativação Linfocitária , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Ligação Proteica , Membro 25 de Receptores de Fatores de Necrose Tumoral/metabolismo , Transdução de Sinais
5.
Int J Mol Sci ; 20(11)2019 May 29.
Artigo em Inglês | MEDLINE | ID: mdl-31146370

RESUMO

Galectin-9 (Gal-9) enhances tumor immunity mediated by T cells, macrophages, and dendritic cells. Its expression level in various cancers correlates with prognosis. Furthermore, Gal-9 directly induces apoptosis in various cancers; however, its mechanism of action and bioactivity has not been clarified. We evaluated Gal-9 antitumor effect against esophageal squamous cell carcinoma (ESCC) to analyze the dynamics of apoptosis-related molecules, elucidate its mechanism of action, and identify relevant changes in miRNA expressions. KYSE-150 and KYSE-180 cells were treated with Gal-9 and their proliferation was evaluated. Gal-9 inhibited cell proliferation in a concentration-dependent manner. The xenograft mouse model established with KYSE-150 cells was administered with Gal-9 and significant suppression in the tumor growth observed. Gal-9 treatment of KYSE-150 cells increased the number of Annexin V-positive cells, activation of caspase-3, and collapse of mitochondrial potential, indicating apoptosis induction. c-Jun NH2-terminal kinase (JNK) and p38 mitogen-activated protein kinase (p38) phosphorylation were activated and could be involved in apoptosis. Therefore, Gal-9 induces mitochondria-mediated apoptosis of ESCC and inhibits cell proliferation in vitro and in vivo with JNK and p38 activation.


Assuntos
Antineoplásicos/farmacologia , Apoptose/efeitos dos fármacos , Carcinoma de Células Escamosas/metabolismo , Neoplasias Esofágicas/metabolismo , Galectinas/farmacologia , Animais , Antineoplásicos/uso terapêutico , Carcinoma de Células Escamosas/tratamento farmacológico , Linhagem Celular Tumoral , Neoplasias Esofágicas/tratamento farmacológico , Galectinas/uso terapêutico , Humanos , Sistema de Sinalização das MAP Quinases , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Mitocôndrias/metabolismo
6.
Cell Physiol Biochem ; 50(5): 1856-1868, 2018.
Artigo em Inglês | MEDLINE | ID: mdl-30396167

RESUMO

BACKGROUND/AIMS: Galectin-9 is a soluble immune modulator with versatile functions, including a role as an immune checkpoint molecule. Therefore, the amount of galectin-9 in the blood may reflect an individual's immunological balance. Many studies have conducted galectin-9 measurements; however, the reported galectin-9 concentration in the blood varies greatly, even within healthy controls. This study investigates the variation between the reported and actual concentrations of galectin-9. METHODS: A GalPharma ELISA and an R&D Systems ELISA kit were directly compared using the same set of plasma and a series of recombinant galectins, including degraded galectin-9. Furthermore, galectin-9 in plasma was concentrated using anti-galectin-9 antibody-conjugated beads, and subjected to western blotting to estimate the quantity and integrity of galectin-9 and assess the consistency of ELISA measurements. RESULTS: The R&D Systems' ELISA indicated a 50-fold higher median concentration of plasma galectin-9 than that indicated by the GalPharma ELISA. This variation is due to aberrantly enhanced reactivity of the R&D Systems' ELISA to degraded galectin-9 present in small quantities in the plasma. The GalPharma ELISA could detect only intact galectin-9 and its results correlated well with the plasma galectin-9 level obtained by western blotting. CONCLUSION: ELISA kits from R&D Systems reacts aberrantly higher against degraded galectin-9 than the intact galectin-9. Therefore, the existence of a small amount of degraded galectin-9 in a test sample hinders the quantification. As galectin-9 is a fragile protein, this is a serious concern when using this kit. Based on quantifications from the GalPharma ELISA, the median (25th-75th percentiles) galectin-9 concentration in healthy subjects in the current study cohort was calculated as 110 pg/mL (67 -154 pg/mL).


Assuntos
Galectinas/sangue , Falência Hepática Aguda/sangue , Ensaio de Imunoadsorção Enzimática , Galectinas/metabolismo , Humanos , Limite de Detecção , Falência Hepática Aguda/diagnóstico , Kit de Reagentes para Diagnóstico
7.
PLoS Pathog ; 12(6): e1005677, 2016 06.
Artigo em Inglês | MEDLINE | ID: mdl-27253379

RESUMO

Identifying host immune determinants governing HIV transcription, latency and infectivity in vivo is critical to developing an HIV cure. Based on our recent finding that the host factor p21 regulates HIV transcription during antiretroviral therapy (ART), and published data demonstrating that the human carbohydrate-binding immunomodulatory protein galectin-9 regulates p21, we hypothesized that galectin-9 modulates HIV transcription. We report that the administration of a recombinant, stable form of galectin-9 (rGal-9) potently reverses HIV latency in vitro in the J-Lat HIV latency model. Furthermore, rGal-9 reverses HIV latency ex vivo in primary CD4+ T cells from HIV-infected, ART-suppressed individuals (p = 0.002), more potently than vorinostat (p = 0.02). rGal-9 co-administration with the latency reversal agent "JQ1", a bromodomain inhibitor, exhibits synergistic activity (p<0.05). rGal-9 signals through N-linked oligosaccharides and O-linked hexasaccharides on the T cell surface, modulating the gene expression levels of key transcription initiation, promoter proximal-pausing, and chromatin remodeling factors that regulate HIV latency. Beyond latent viral reactivation, rGal-9 induces robust expression of the host antiviral deaminase APOBEC3G in vitro and ex vivo (FDR<0.006) and significantly reduces infectivity of progeny virus, decreasing the probability that the HIV reservoir will be replenished when latency is reversed therapeutically. Lastly, endogenous levels of soluble galectin-9 in the plasma of 72 HIV-infected ART-suppressed individuals were associated with levels of HIV RNA in CD4+ T cells (p<0.02) and with the quantity and binding avidity of circulating anti-HIV antibodies (p<0.009), suggesting a role of galectin-9 in regulating HIV transcription and viral production in vivo during therapy. Our data suggest that galectin-9 and the host glycosylation machinery should be explored as foundations for novel HIV cure strategies.


Assuntos
Linfócitos T CD4-Positivos/virologia , Galectinas/metabolismo , Infecções por HIV/metabolismo , Ativação Viral/fisiologia , Latência Viral/fisiologia , Fármacos Anti-HIV/uso terapêutico , Western Blotting , Ensaio de Imunoadsorção Enzimática , Citometria de Fluxo , Perfilação da Expressão Gênica , Infecções por HIV/tratamento farmacológico , Infecções por HIV/virologia , HIV-1/fisiologia , Humanos , Reação em Cadeia da Polimerase , Transcrição Gênica/fisiologia , Transcriptoma
8.
J Gastroenterol Hepatol ; 33(2): 492-499, 2018 Feb.
Artigo em Inglês | MEDLINE | ID: mdl-28618039

RESUMO

BACKGROUND AND AIM: Chronic liver diseases progress from chronic inflammation to fibrosis to tumorigenesis. Galectin-9, a ß-galactoside-specific animal lectin, is indicated to contribute to all three steps of progression. The aim of this study was to determine which of the three steps was most dominant in elevating the serum galectin-9 concentration and to test the possibility of galectin-9 as a serum biomarker. METHODS: Japanese patients with chronic hepatitis, liver cirrhosis, hepatocellular carcinoma (HCC), non-alcoholic fatty liver disease, or alcoholic liver disease who provided informed consent were enrolled in this study. Serum galectin-9 levels were measured using a sandwich ELISA. Multiple regression analyses were performed using ezr to identify factors that determined serum galectin-9 concentration. RESULTS: One hundred one patients with 50 of chronic hepatitis and 51 of liver cirrhosis were enrolled; the cohort included 45 cases of hepatitis C virus infection, 13 cases of hepatitis B virus infection, and 46 cases with HCC-related complications. The median serum galectin-9 concentration was 77.54 pg/mL (interquartile range: 18.89-241.9 pg/mL). Multiple linear regression analyses proved Fibrosis-4 index and aspartate aminotransferase to platelet ratio index, indexes of liver fibrosis, were able to predict the serum galectin-9 levels with statistical significance. A multiple logistic regression analysis determined 10 pg/mL increase in the serum galectin-9 concentration presented an odds ratio of 3.90 for liver fibrosis progression. CONCLUSIONS: The serum galectin-9 concentration represents a potential biomarker of liver fibrosis in patients with chronic liver diseases, regardless of chronic inflammation or the presence of HCC complications. Furthermore, higher serum galectin-9 levels are a predictor for liver fibrosis progression.


Assuntos
Galectinas/sangue , Cirrose Hepática/diagnóstico , Idoso , Biomarcadores/sangue , Doença Crônica , Progressão da Doença , Ensaio de Imunoadsorção Enzimática , Humanos , Cirrose Hepática/etiologia , Hepatopatias/complicações , Pessoa de Meia-Idade , Análise de Regressão
9.
Biochem Biophys Res Commun ; 490(4): 1287-1293, 2017 09 02.
Artigo em Inglês | MEDLINE | ID: mdl-28687490

RESUMO

Galectin-9 (G9) is a tandem-repeat type ß-galactoside-specific animal lectin having N-terminal and C-terminal carbohydrate recognition domains (N-CRD and C-CRD, respectively) joined by a linker peptide that is involved in the immune system. G9 is divalent in glycan binding, and structural information about the spatial arrangement of the two CRDs is very important for elucidating its biological functions. As G9 is protease sensitive due to the long linker, the protease-resistant mutant form of G9 (G9Null) was developed by modification of the linker peptide, while retaining its biological functions. The X-ray structure of a mutant form of G9Null with the replacement of Arg221 by Ser (G9Null_R221S) having two CRDs was determined. The structure of G9Null_R221S was compact to associate the two CRDs in the back-to-back orientation with a large interface area, including hydrogen bonds and hydrophobic interactions. A metal ion was newly found in the galectin structure, possibly contributing to the stable structure of protein. The presented X-ray structure was thought to be one of the stable structures of G9, which likely occurs in solution. This was supported by structural comparisons with other tandem-repeated galectins and the analyses of protein thermostability by CD spectra measurements.


Assuntos
Galactosídeos/química , Galectinas/química , Metais/química , Mutação , Adenoviridae/química , Sequência de Aminoácidos , Animais , Sítios de Ligação , Clonagem Molecular , Cristalografia por Raios X , Escherichia coli/genética , Escherichia coli/metabolismo , Galectinas/genética , Galectinas/metabolismo , Expressão Gênica , Humanos , Ligação de Hidrogênio , Interações Hidrofóbicas e Hidrofílicas , Modelos Moleculares , Ligação Proteica , Conformação Proteica em alfa-Hélice , Conformação Proteica em Folha beta , Domínios e Motivos de Interação entre Proteínas , Proteínas Recombinantes/química , Proteínas Recombinantes/genética , Proteínas Recombinantes/metabolismo , Alinhamento de Sequência , Homologia de Sequência de Aminoácidos , Toxascaris/química
10.
Int J Mol Sci ; 18(1)2017 Jan 01.
Artigo em Inglês | MEDLINE | ID: mdl-28045432

RESUMO

Dysregulation of apoptosis is a major hallmark in cancer biology that might equip tumors with a higher malignant potential and chemoresistance. The anti-cancer activities of lectin, defined as a carbohydrate-binding protein that is not an enzyme or antibody, have been investigated for over a century. Recently, galectin-9, which has two distinct carbohydrate recognition domains connected by a linker peptide, was noted to induce apoptosis in thymocytes and immune cells. The apoptosis of these cells contributes to the development and regulation of acquired immunity. Furthermore, human recombinant galectin-9, hG9NC (null), which lacks an entire region of the linker peptide, was designed to resist proteolysis. The hG9NC (null) has demonstrated anti-cancer activities, including inducing apoptosis in hematological, dermatological and gastrointestinal malignancies. In this review, the molecular characteristics, history and apoptosis-inducing potential of galectin-9 are described.


Assuntos
Antineoplásicos/uso terapêutico , Apoptose/efeitos dos fármacos , Galectinas/uso terapêutico , Neoplasias/tratamento farmacológico , Animais , Antineoplásicos/análise , Antineoplásicos/metabolismo , Galectinas/análise , Galectinas/metabolismo , Humanos , Neoplasias/metabolismo , Neoplasias/patologia , Proteínas Recombinantes/análise , Proteínas Recombinantes/metabolismo , Proteínas Recombinantes/uso terapêutico
11.
J Biol Chem ; 290(27): 16797-811, 2015 Jul 03.
Artigo em Inglês | MEDLINE | ID: mdl-25947381

RESUMO

Galectin-9 (gal-9) is a multifunctional ß-galactoside-binding lectin, frequently released in the extracellular medium, where it acts as a pleiotropic immune modulator. Despite its overall immunosuppressive effects, a recent study has reported bimodal action of gal-9 on human resting blood T cells with apoptosis occurring in the majority of them, followed by a wave of activation and expansion of Th1 cells in the surviving population. Our knowledge of the signaling events triggered by exogenous gal-9 in T cells remains limited. One of these events is cytosolic calcium (Ca(2+)) release reported in some murine and human T cells. The aim of this study was to investigate the contribution of Ca(2+) mobilization to apoptotic and nonapoptotic effects of exogenous gal-9 in human T cells. We found that the T cell receptor (TCR)-CD3 complex and the Lck kinase were required for Ca(2+) mobilization but not for apoptosis induction in Jurkat cells. These data were confirmed in human CD4(+) T cells from peripheral blood as follows: a specific Lck chemical inhibitor abrogated Ca(2+) mobilization but not apoptosis induction. Moreover, Lck activity was also required for the production of Th1-type cytokines, i.e. interleukin-2 and interferon-γ, which resulted from gal-9 stimulation in peripheral CD4(+) T cells. These findings indicate that gal-9 acts on T cells by two distinct pathways as follows: one mimicking antigen-specific activation of the TCR with a mandatory contribution of proximal elements of the TCR complex, especially Lck, and another resulting in apoptosis that is independent of this complex.


Assuntos
Apoptose , Complexo CD3/metabolismo , Galectinas/metabolismo , Proteína Tirosina Quinase p56(lck) Linfócito-Específica/metabolismo , Receptores de Antígenos de Linfócitos T/metabolismo , Linfócitos T/metabolismo , Complexo CD3/genética , Cálcio/metabolismo , Citocinas/genética , Citocinas/metabolismo , Galectinas/genética , Humanos , Proteína Tirosina Quinase p56(lck) Linfócito-Específica/genética , Receptores de Antígenos de Linfócitos T/genética , Linfócitos T/citologia
12.
Clin Gastroenterol Hepatol ; 14(4): 606-612.e3, 2016 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-26499927

RESUMO

BACKGROUND & AIMS: Fewer than 50% of patients with acute liver failure (ALF) recover spontaneously, and ALF has high mortality without liver transplantation. Kupffer cells have been reported to mediate liver inflammation during drug-induced injury. Galectin-9 is produced by Kupffer cells and has diverse roles in regulating immunity. We investigated whether plasma levels of galectin-9 are associated with outcomes of patients with ALF. METHODS: We analyzed plasma samples (collected at time of hospital admission) and clinical data from 149 patients included in the Acute Liver Failure Study Group from July 2006 through November 2010 (110 had acetaminophen-induced hepatotoxicity and 39 had nonacetaminophen drug-induced liver injury). We compared data with those from all patients enrolled in the study (from July 1, 2006 through October 30, 2013), and from healthy individuals of similar ages with no evidence of liver disease (control subjects). Plasma levels of galectin-9 were measured using a polyclonal antibody and colorimetric assay. RESULTS: Patients with ALF had statistically higher plasma levels of galectin-9 than control subjects, but levels did not differ significantly between patients with acetaminophen-induced liver injury and drug-induced liver injury. A level of galectin-9 above 690 pg/mL was associated with a statistically significant increase in risk for mortality or liver transplantation caused by ALF. Competing risk analyses associated level of galectin-9 with transplant-free survival, independently of Model For End-Stage Liver Disease score or systemic inflammatory response syndrome. CONCLUSIONS: A one-time measurement of plasma galectin-9 level can be used to assign patients with ALF to high-, intermediate-, and low-risk groups. The combination of galectin-9 level and Model For End-Stage Liver Disease score was more closely associated with patient outcome than either value alone. These data might be used to determine patient prognoses and prioritize patients for liver transplantation. ClinicalTrials.gov ID NCT00518440.


Assuntos
Galectinas/sangue , Falência Hepática Aguda/induzido quimicamente , Falência Hepática Aguda/mortalidade , Plasma/química , Acetaminofen/efeitos adversos , Adulto , Antipiréticos/efeitos adversos , Estudos de Coortes , Colorimetria , Feminino , Humanos , Imunoensaio , Falência Hepática Aguda/diagnóstico , Masculino , Pessoa de Meia-Idade , Prognóstico , Análise de Sobrevida
13.
J Immunol ; 192(5): 2418-31, 2014 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-24477913

RESUMO

The T cell Ig and mucin domain-containing molecule (TIM) family of receptors have emerged as potential therapeutic targets to correct abnormal immune function in chronic inflammatory conditions. TIM-3 serves as a functional receptor in structural cells of the airways and via the ligand galectin-9 (Gal-9) can modulate the inflammatory response. The aim of this study was to investigate TIM-3 expression and function in neutrophils, focusing on its potential role in cystic fibrosis (CF) lung disease. Results revealed that TIM-3 mRNA and protein expression values of circulating neutrophils were equal between healthy controls (n = 20) and people with CF (n = 26). TIM-3 was detected on resting neutrophil membranes by FACS analysis, and expression levels significantly increased post IL-8 or TNF-α exposure (p < 0.05). Our data suggest a novel role for TIM-3/Gal-9 signaling involving modulation of cytosolic calcium levels. Via TIM-3 interaction, Gal-9 induced neutrophil degranulation and primed the cell for enhanced NADPH oxidase activity. Killing of Pseudomonas aeruginosa was significantly increased upon bacterial opsonization with Gal-9 (p < 0.05), an effect abrogated by blockade of TIM-3 receptors. This mechanism appeared to be Gram-negative bacteria specific and mediated via Gal-9/ LPS binding. Additionally, we have demonstrated that neutrophil TIM-3/Gal-9 signaling is perturbed in the CF airways due to proteolytic degradation of the receptor. In conclusion, results suggest a novel neutrophil defect potentially contributing to the defective bacterial clearance observed in the CF airways and suggest that manipulation of the TIM-3 signaling pathway may be of therapeutic value in CF, preferably in conjunction with antiprotease treatment.


Assuntos
Fibrose Cística/imunologia , Galectinas/imunologia , Pulmão/imunologia , Proteínas de Membrana/imunologia , Neutrófilos/imunologia , Pseudomonas aeruginosa/imunologia , Fibrose Cística/microbiologia , Fibrose Cística/patologia , Feminino , Receptor Celular 2 do Vírus da Hepatite A , Humanos , Lipopolissacarídeos/imunologia , Pulmão/microbiologia , Pulmão/patologia , Masculino , Neutrófilos/microbiologia , Transdução de Sinais/imunologia
14.
Am J Physiol Renal Physiol ; 306(8): F822-32, 2014 Apr 15.
Artigo em Inglês | MEDLINE | ID: mdl-24477688

RESUMO

Antiglomerular basement membrane glomerulonephritis (anti-GBM GN) is a Th1- and Th17-predominant autoimmune disease. Galectin-9 (Gal-9), identified as the ligand of Tim-3, functions in diverse biological processes and leads to the apoptosis of CD4(+)Tim-3(+) T cells. It is still unclear how Gal-9 regulates the functions of Th1 and Th17 cells and prevents renal injury in anti-GBM GN. In this study, Gal-9 was administered to anti-GBM GN mice for 7 days. We found that Gal-9 retarded the increase of Scr, ameliorated renal tubular injury, and reduced the formation of crescents. The infiltration of Th1 and Th17 cells into the spleen and kidneys significantly decreased in Gal-9-treated nephritic mice. The reduced infiltration of Th1 and Th17 cells might be associated with the downregulation of CCL-20, CXCL-9, and CXCL-10 mRNAs in the kidney. In parallel, the blood levels of IFN-γ and IL-17A declined in Gal-9-treated nephritic mice at days 21 and 28. In addition, an enhanced Th2 cell-mediated immune response was observed in the kidneys of nephritic mice after a 7-day injection of Gal-9. In conclusion, the protective role of Gal-9 in anti-GBM GN is associated with the inhibition of Th1 and Th17 cell-mediated immune responses and enhanced Th2 immunity in the kidney.


Assuntos
Doença Antimembrana Basal Glomerular/prevenção & controle , Galectinas/farmacologia , Células Th1/efeitos dos fármacos , Células Th1/imunologia , Células Th17/efeitos dos fármacos , Células Th17/imunologia , Animais , Doença Antimembrana Basal Glomerular/imunologia , Autoanticorpos/imunologia , Linfócitos T CD4-Positivos/imunologia , Receptor Celular 2 do Vírus da Hepatite A , Rim/imunologia , Camundongos , Receptores Virais/biossíntese
15.
J Virol ; 87(9): 4835-45, 2013 May.
Artigo em Inglês | MEDLINE | ID: mdl-23408620

RESUMO

Galectin-9 is a pleiotropic immune modulator affecting numerous cell types of innate and adaptive immunity. Patients with chronic infection with either hepatitis C virus (HCV) or HIV have elevated circulating levels. Limited data exist on the regulation of natural killer (NK) cell function through interaction with galectin-9. We found that galectin-9 ligation downregulates multiple immune-activating genes, including eight involved in the NK cell-mediated cytotoxicity pathway, impairs lymphokine-activated killing, and decreases the proportion of gamma interferon (IFN-γ)-producing NK cells that had been stimulated with interleukin-12 (IL-12)/IL-15. We demonstrate that the transcriptional and functional changes induced by galectin-9 are independent of Tim-3. Consistent with these results for humans, we find that the genetic absence of galectin-9 in mice is associated with greater IFN-γ production by NK cells and enhanced degranulation. We also show that in the setting of a short-term (4-day) murine cytomegalovirus infection, terminally differentiated NKs accumulate in the livers of galectin-9 knockout mice, and that hepatic NKs spontaneously produce significantly more IFN-γ in this setting. Taken together, our results indicate that galectin-9 engagement impairs the function of NK cells, including cytotoxicity and cytokine production.


Assuntos
Galectinas/imunologia , Infecções por Herpesviridae/imunologia , Células Matadoras Naturais/imunologia , Animais , Células Cultivadas , Citotoxicidade Imunológica , Galectinas/genética , Receptor Celular 2 do Vírus da Hepatite A , Infecções por Herpesviridae/genética , Infecções por Herpesviridae/virologia , Humanos , Interleucina-12/imunologia , Interleucina-15/imunologia , Proteínas de Membrana/genética , Proteínas de Membrana/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Muromegalovirus/fisiologia , Receptores Virais/genética , Receptores Virais/imunologia
16.
Blood ; 119(18): 4192-204, 2012 May 03.
Artigo em Inglês | MEDLINE | ID: mdl-22438246

RESUMO

Galectin-9 (Gal-9) is a tandem repeat-type member of the galectin family and is a ligand for T-cell immunoglobulin mucin domain 3 (Tim-3), a type-I glycoprotein that is persistently expressed on dysfunctional T cells during chronic infection. Studies in autoimmune diseases and chronic viral infections show that Tim-3 is a regulatory molecule that inhibits Th1 type immune responses. Here we show that soluble Gal-9 interacts with Tim-3 expressed on the surface of activated CD4(+) T cells and renders them less susceptible to HIV-1 infection and replication. The Gal-9/Tim-3 interaction on activated CD4(+) T cells, leads to down-regulation of HIV-1 coreceptors and up-regulation of the cyclin-dependent kinase inhibitor p21 (also known as cip-1 and waf-1). We suggest that higher expression of Tim-3 during chronic infection has evolved to limit persistent immune activation and associated tissue damage. These data demonstrate a novel mechanism for Gal-9/Tim-3 interactions to induce resistance of activated CD4(+) T cells to HIV-1 infection and suggest that Gal-9 may play a role in HIV-1 pathogenesis and could be used as a novel microbicide to prevent HIV-1 infection.


Assuntos
Linfócitos T CD4-Positivos/metabolismo , Galectinas/metabolismo , Infecções por HIV/imunologia , HIV-1/fisiologia , Proteínas de Membrana/metabolismo , Viremia/imunologia , Apoptose , Linfócitos T CD4-Positivos/virologia , Células Cultivadas/virologia , Inibidor de Quinase Dependente de Ciclina p21/biossíntese , Inibidor de Quinase Dependente de Ciclina p21/genética , Regulação da Expressão Gênica , Genes p53 , Infecções por HIV/sangue , Infecções por HIV/virologia , Receptor Celular 2 do Vírus da Hepatite A , Humanos , Integrinas/biossíntese , Integrinas/genética , Ativação Linfocitária , Ligação Proteica , Mapeamento de Interação de Proteínas , RNA Interferente Pequeno/farmacologia , Receptores CXCR4/biossíntese , Receptores CXCR4/genética , Receptores CXCR5/biossíntese , Receptores CXCR5/genética , Proteína Supressora de Tumor p53/biossíntese , Carga Viral , Ligação Viral , Replicação Viral
17.
Blood ; 119(13): 3064-72, 2012 Mar 29.
Artigo em Inglês | MEDLINE | ID: mdl-22323453

RESUMO

NK-cell function is regulated by the integration of signals received from activating and inhibitory receptors. Here we show that a novel immune receptor, T-cell Ig and mucin-containing domain-3 (Tim-3), is expressed on resting human NK cells and is up-regulated on activation. The NK92 NK-cell line engineered to overexpress Tim-3 showed a marked increase in IFN-γ production in the presence of soluble rhGal-9 or Raji tumor cells engineered to express Gal-9. The Tim-3(+) population of low-dose IL-12/IL-18-activated primary NK cells significantly increased IFN-γ production in response to soluble rhGal-9, Gal-9 presented by cell lines, and primary acute myelogenous leukemia (AML) targets that endogenously express Gal-9. This effect is highly specific as Tim-3 Ab blockade significantly decreased IFN-γ production, and Tim-3 cross-linking induced ERK activation and degradation of IκBα. Exposure to Gal-9-expressing target cells had little effect on CD107a degranulation. Reconstituted NK cells obtained from patients after hematopoietic cell transplantation had diminished expression of Tim-3 compared with paired donors. This observation correlates with the known IFN-γ defect seen early posttransplantation. In conclusion, we show that Tim-3 functions as a human NK-cell coreceptor to enhance IFN-γ production, which has important implications for control of infectious disease and cancer.


Assuntos
Galectinas/farmacologia , Interferon gama/metabolismo , Células Matadoras Naturais/efeitos dos fármacos , Células Matadoras Naturais/metabolismo , Proteínas de Membrana/genética , Adulto , Células Cultivadas , Galectinas/genética , Galectinas/metabolismo , Regulação da Expressão Gênica/efeitos dos fármacos , Regulação da Expressão Gênica/fisiologia , Células HEK293 , Transplante de Células-Tronco Hematopoéticas , Receptor Celular 2 do Vírus da Hepatite A , Humanos , Interferon gama/sangue , Células Jurkat , Leucemia/sangue , Leucemia/genética , Leucemia/imunologia , Leucemia/terapia , Proteínas de Membrana/antagonistas & inibidores , Proteínas de Membrana/metabolismo , Proteínas de Membrana/fisiologia , Receptores de Células Matadoras Naturais/genética , Receptores de Células Matadoras Naturais/metabolismo , Receptores de Células Matadoras Naturais/fisiologia , Proteínas Recombinantes/farmacologia
18.
Blood ; 120(3): 682-90, 2012 Jul 19.
Artigo em Inglês | MEDLINE | ID: mdl-22677125

RESUMO

T-cell immunoglobulin mucin-3 (Tim-3) is expressed on pathogenic T cells, and its ligand galectin-9 (gal-9) is up-regulated in inflamed tissues. When Tim-3(+) T cells encounter high gal-9 levels, they are deleted. Tim-3 is up-regulated on activated T cells during GVHD. Inhibition of Tim-3/gal-9 binding by infusion of a Tim-3-Ig fusion protein or Tim-3(-/-) donor T cells increased T-cell proliferation and GVHD lethality. When the Tim-3/gal-9 pathway engagement was augmented using gal-9 transgenic recipients, GVHD lethality was slowed. Together, these data indicate a potential for modulating this pathway to reduce disease by increasing Tim-3 or gal-9 engagement. Paradoxically, when Tim-3/gal-9 was inhibited in the absence of donor T-regulatory cells (Tregs), GVHD was inhibited. GVHD reduction was associated with decreased colonic inflammatory cytokines as well as epithelial barrier destruction. CD25-depleted Tim-3(-/-) donor T cells underwent increased activation-induced cell death because of increased IFN-γ production. To our knowledge, these studies are the first to show that although the absence of Tim-3/gal-9 pathway interactions augments systemic GVHD, concurrent donor Treg depletion paradoxically and surprisingly inhibits GVHD. Thus, although donor Tregs typically inhibit GVHD, under some conditions, such Tregs actually may contribute to GVHD by reducing activation-induced T-cell death.


Assuntos
Transplante de Medula Óssea/efeitos adversos , Galectinas/metabolismo , Doença Enxerto-Hospedeiro/imunologia , Receptores Virais/metabolismo , Transdução de Sinais/imunologia , Linfócitos T Reguladores/imunologia , Doença Aguda , Animais , Morte Celular/imunologia , Divisão Celular/imunologia , Galectinas/genética , Galectinas/imunologia , Doença Enxerto-Hospedeiro/metabolismo , Doença Enxerto-Hospedeiro/mortalidade , Receptor Celular 2 do Vírus da Hepatite A , Interferon gama/antagonistas & inibidores , Interferon gama/sangue , Intestinos/imunologia , Intestinos/patologia , Depleção Linfocítica/métodos , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Transgênicos , Receptores Virais/genética , Receptores Virais/imunologia , Linfócitos T Reguladores/metabolismo , Regulação para Cima/imunologia
19.
J Immunol ; 188(9): 4631-43, 2012 May 01.
Artigo em Inglês | MEDLINE | ID: mdl-22467659

RESUMO

Stromal keratitis is a chronic immunopathological lesion of the eye caused by HSV-1 infection that can result in blindness. Because the inflammatory lesions are primarily orchestrated by Th1 cells, and to a lesser extent by Th17 cells, inhibiting their activity represents a useful form of therapy. In this study we evaluated the therapeutic potential of galectin-1 (gal-1), an endogenous lectin that in some autoimmune diseases was shown to suppress the functions of Th1 and Th17 cells. Treatment was begun at different times after ocular infection with HSV and the outcome was assessed clinically as well as for effects on various immune parameters. Treatment with recombinant gal-1 significantly diminished stromal keratitis lesion severity and the extent of corneal neovascularization. Treated mice had reduced numbers of IFN-γ- and IL-17-producing CD4(+) T cells, as well as neutrophil infiltration in the cornea. Furthermore, disease severity was greater in gal-1 knockout mice compared with their wild-type counterparts. The many effects of gal-1 treatment include reduction in the production of proinflammatory cytokines and chemokines, increased production of IL-10, and inhibitory effects on molecules involved in neovascularization. To our knowledge, our findings are the first to show that gal-1 treatment represents a useful approach to control lesion severity in a virally induced immunopathological disease.


Assuntos
Neovascularização da Córnea/imunologia , Galectina 1/imunologia , Herpesvirus Humano 1/imunologia , Ceratite Herpética/imunologia , Animais , Cegueira/genética , Cegueira/imunologia , Cegueira/metabolismo , Cegueira/patologia , Cegueira/virologia , Neovascularização da Córnea/genética , Neovascularização da Córnea/metabolismo , Neovascularização da Córnea/patologia , Neovascularização da Córnea/virologia , Galectina 1/genética , Galectina 1/metabolismo , Herpesvirus Humano 1/genética , Herpesvirus Humano 1/metabolismo , Interleucina-10/genética , Interleucina-10/imunologia , Interleucina-10/metabolismo , Ceratite Herpética/genética , Ceratite Herpética/metabolismo , Ceratite Herpética/patologia , Camundongos , Camundongos Knockout , Células Th1/imunologia , Células Th1/metabolismo , Células Th1/patologia , Células Th17/imunologia , Células Th17/metabolismo , Células Th17/patologia
20.
J Virol ; 86(19): 10606-20, 2012 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-22811539

RESUMO

Ocular infection with herpes simplex virus 1 (HSV-1) results in a chronic immunoinflamammtory reaction in the cornea, which is primarily orchestrated by CD4(+) T cells. Hence, targeting proinflammatory CD4(+) T cells or increasing the representation of cells that regulate their function is a relevant therapeutic strategy. In this report, we demonstrate that effective therapeutic control can be achieved using a combination of approaches under circumstances where monotherapy is ineffective. We use a convenient and highly effective monoclonal antibody (MAb) approach with MAbT25 to expand cells that express the tumor necrosis factor receptor superfamily member 25 (TNFRSF25). In naïve animals, these are predominantly cells that are Foxp3-positive regulatory T cells. MAbT25 treatment before or at the time of initial HSV infection was an effective means of reducing the severity of subsequent stromal keratitis lesions. However, MAbT25 treatment was not effective if given 6 days after infection since it expanded proinflammatory effector T cells, which also express TNFRSF25. Therefore, the MAbT25 procedure was combined with galectin-9 (Gal-9), an approach that compromises the activity of T cells involved in tissue damage. The combination therapy provided highly effective lesion control over that achieved by treatment with one of them. The beneficial outcome of the combination therapy was attributed to the expansion of the regulatory T cell population that additionally expressed activation markers such as CD103 needed to access inflammatory sites. Additionally, there was a marked reduction of CD4(+) gamma interferon-producing effector T cells responsible for orchestrating the tissue damage. The approach that we describe has potential application to control a wide range of inflammatory diseases, in addition to stromal keratitis, an important cause of human blindness.


Assuntos
Anticorpos/química , Galectinas/metabolismo , Herpes Simples/metabolismo , Membro 25 de Receptores de Fatores de Necrose Tumoral/imunologia , Membro 25 de Receptores de Fatores de Necrose Tumoral/metabolismo , Simplexvirus/metabolismo , Animais , Anticorpos Monoclonais/química , Antígenos CD/biossíntese , Apoptose , Linfócitos T CD4-Positivos/metabolismo , Proliferação de Células , Córnea/virologia , Cricetinae , Feminino , Fatores de Transcrição Forkhead/metabolismo , Proteínas de Fluorescência Verde/metabolismo , Inflamação , Cadeias alfa de Integrinas/biossíntese , Ceratite Herpética/imunologia , Ceratite Herpética/virologia , Cinética , Camundongos , Camundongos Endogâmicos C57BL
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