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1.
Immunity ; 53(3): 581-596.e5, 2020 09 15.
Artigo em Inglês | MEDLINE | ID: mdl-32707034

RESUMO

Glucocorticoids (GC) are the mainstay treatment option for inflammatory conditions. Despite the broad usage of GC, the mechanisms by which GC exerts its effects remain elusive. Here, utilizing murine autoimmune and allergic inflammation models, we report that Foxp3+ regulatory T (Treg) cells are irreplaceable GC target cells in vivo. Dexamethasone (Dex) administered in the absence of Treg cells completely lost its ability to control inflammation, and the lack of glucocorticoid receptor in Treg cells alone resulted in the loss of therapeutic ability of Dex. Mechanistically, Dex induced miR-342-3p specifically in Treg cells and miR-342-3p directly targeted the mTORC2 component, Rictor. Altering miRNA-342-3p or Rictor expression in Treg cells dysregulated metabolic programming in Treg cells, controlling their regulatory functions in vivo. Our results uncover a previously unknown contribution of Treg cells during glucocorticoid-mediated treatment of inflammation and the underlying mechanisms operated via the Dex-miR-342-Rictor axis.


Assuntos
Dexametasona/farmacologia , Glucocorticoides/farmacologia , Inflamação/tratamento farmacológico , MicroRNAs/genética , Proteína Companheira de mTOR Insensível à Rapamicina/metabolismo , Linfócitos T Reguladores/imunologia , Animais , Anti-Inflamatórios/farmacologia , Fatores de Transcrição Forkhead/metabolismo , Alvo Mecanístico do Complexo 2 de Rapamicina/metabolismo , Camundongos , Camundongos Endogâmicos C57BL , MicroRNAs/biossíntese , Receptores de Glucocorticoides/genética , Linfócitos T Reguladores/metabolismo
2.
Gut ; 73(7): 1110-1123, 2024 06 06.
Artigo em Inglês | MEDLINE | ID: mdl-38378253

RESUMO

OBJECTIVE: Intestinal fibrosis is considered an inevitable consequence of chronic IBD, leading to stricture formation and need for surgery. During the process of fibrogenesis, extracellular matrix (ECM) components critically regulate the function of mesenchymal cells. We characterised the composition and function of ECM in fibrostenosing Crohn's disease (CD) and control tissues. DESIGN: Decellularised full-thickness intestinal tissue platforms were tested using three different protocols, and ECM composition in different tissue phenotypes was explored by proteomics and validated by quantitative PCR (qPCR) and immunohistochemistry. Primary human intestinal myofibroblasts (HIMFs) treated with milk fat globule-epidermal growth factor 8 (MFGE8) were evaluated regarding the mechanism of their antifibrotic response, and the action of MFGE8 was tested in two experimental intestinal fibrosis models. RESULTS: We established and validated an optimal decellularisation protocol for intestinal IBD tissues. Matrisome analysis revealed elevated MFGE8 expression in CD strictured (CDs) tissue, which was confirmed at the mRNA and protein levels. Treatment with MFGE8 inhibited ECM production in normal control HIMF but not CDs HIMF. Next-generation sequencing uncovered functionally relevant integrin-mediated signalling pathways, and blockade of integrin αvß5 and focal adhesion kinase rendered HIMF non-responsive to MFGE8. MFGE8 prevented and reversed experimental intestinal fibrosis in vitro and in vivo. CONCLUSION: MFGE8 displays antifibrotic effects, and its administration may represent a future approach for prevention of IBD-induced intestinal strictures.


Assuntos
Antígenos de Superfície , Doença de Crohn , Matriz Extracelular , Fibrose , Proteínas do Leite , Humanos , Animais , Doença de Crohn/patologia , Doença de Crohn/metabolismo , Proteínas do Leite/metabolismo , Proteínas do Leite/farmacologia , Antígenos de Superfície/metabolismo , Matriz Extracelular/metabolismo , Miofibroblastos/metabolismo , Modelos Animais de Doenças , Camundongos , Ratos
3.
Gastroenterology ; 165(5): 1180-1196, 2023 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-37507073

RESUMO

BACKGROUND & AIMS: Fibroblasts play a key role in stricture formation in Crohn's disease (CD) but understanding its pathogenesis requires a systems-level investigation to uncover new treatment targets. We studied full-thickness CD tissues to characterize fibroblast heterogeneity and function by generating the first single-cell RNA sequencing (scRNAseq) atlas of strictured bowel and providing proof of principle for therapeutic target validation. METHODS: We performed scRNAseq of 13 fresh full-thickness CD resections containing noninvolved, inflamed nonstrictured, and strictured segments as well as 7 normal non-CD bowel segments. Each segment was separated into mucosa/submucosa or muscularis propria and analyzed separately for a total of 99 tissue samples and 409,001 cells. We validated cadherin-11 (CDH11) as a potential therapeutic target by using whole tissues, isolated intestinal cells, NanoString nCounter, next-generation sequencing, proteomics, and animal models. RESULTS: Our integrated dataset revealed fibroblast heterogeneity in strictured CD with the majority of stricture-selective changes detected in the mucosa/submucosa, but not the muscle layer. Cell-cell interaction modeling revealed CXCL14+ as well as MMP/WNT5A+ fibroblasts displaying a central signaling role in CD strictures. CDH11, a fibroblast cell-cell adhesion molecule, was broadly expressed and up-regulated, and its profibrotic function was validated using NanoString nCounter, RNA sequencing, tissue target expression, in vitro gain- and loss-of-function experiments, proteomics, and knock-out and antibody-mediated CDH11 blockade in experimental colitis. CONCLUSIONS: A full-thickness bowel scRNAseq atlas revealed previously unrecognized fibroblast heterogeneity and interactions in CD strictures and CDH11 was validated as a potential therapeutic target. These results provide a new resource for a better understanding of CD stricture formation and open potential therapeutic developments. This work has been posted as a preprint on Biorxiv under doi: 10.1101/2023.04.03.534781.


Assuntos
Colite , Doença de Crohn , Animais , Doença de Crohn/genética , Doença de Crohn/patologia , Constrição Patológica , Intestinos/patologia , Colite/patologia , Fibroblastos/patologia
4.
J Immunol ; 207(3): 765-770, 2021 08 01.
Artigo em Inglês | MEDLINE | ID: mdl-34301840

RESUMO

Glucocorticoids are a highly effective first-line treatment option for many inflammatory diseases, including asthma. Some patients develop a steroid-resistant condition, yet, the cellular and molecular mechanisms underlying steroid resistance remain largely unknown. In this study, we used a murine model of steroid-resistant airway inflammation and report that combining systemic dexamethasone and intranasal IL-27 is able to reverse the inflammation. Foxp3+ regulatory T cells (Tregs) were required during dexamethasone/IL-27 treatment of steroid-resistant allergic inflammation, and importantly, direct stimulation of Tregs via glucocorticoid or IL-27 receptors was essential. Mechanistically, IL-27 stimulation in Tregs enhanced expression of the agonistic glucocorticoid receptor-α isoform. Overexpression of inhibitory glucocorticoid receptor-ß isoform in Tregs alone was sufficient to elicit steroid resistance in a steroid-sensitive allergic inflammation model. Taken together, our results demonstrate for the first time, to our knowledge, that Tregs are instrumental during steroid resistance and that manipulating steroid responsiveness in Tregs may represent a novel strategy to treat steroid refractory asthma.


Assuntos
Asma/imunologia , Dexametasona/uso terapêutico , Interleucina-27/uso terapêutico , Hipersensibilidade Respiratória/imunologia , Linfócitos T Reguladores/imunologia , Alérgenos/imunologia , Animais , Asma/tratamento farmacológico , Células Cultivadas , Modelos Animais de Doenças , Resistência a Medicamentos , Fatores de Transcrição Forkhead/genética , Fatores de Transcrição Forkhead/metabolismo , Humanos , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Ovalbumina/imunologia , Isoformas de Proteínas/genética , Isoformas de Proteínas/metabolismo , Receptores de Glucocorticoides/genética , Receptores de Glucocorticoides/metabolismo , Hipersensibilidade Respiratória/tratamento farmacológico
5.
Proc Natl Acad Sci U S A ; 116(14): 6932-6937, 2019 04 02.
Artigo em Inglês | MEDLINE | ID: mdl-30894497

RESUMO

Clinical application of inhaled glucocorticoids (GCs) has been hampered in the case of steroid-resistant severe asthma. To overcome this limitation, we have developed a series of highly potent GCs, including VSGC12, VSG158, and VSG159 based on the structural insight into the glucocorticoid receptor (GR). Particularly, VSG158 exhibits a maximal repression of lung inflammation and is 10 times more potent than the currently most potent clinical GC, Fluticasone Furoate (FF), in a murine model of asthma. More importantly, VSG158 displays a unique property to reduce neutrophilic inflammation in a steroid-resistant airway inflammation model, which is refractory to clinically available GCs, including dexamethasone and FF. VSG158 and VSG159 are able to deliver effective treatments with reduced off-target and side effects. In addition, these GCs also display pharmacokinetic properties that are suitable for the inhalation delivery method for asthma treatment. Taken together, the excellent therapeutic and side-effect profile of these highly potent GCs holds promise for treating steroid-resistant severe asthma.


Assuntos
Antiasmáticos , Asma/tratamento farmacológico , Desenvolvimento de Medicamentos , Glucocorticoides , Animais , Antiasmáticos/química , Antiasmáticos/farmacologia , Asma/patologia , Modelos Animais de Doenças , Feminino , Glucocorticoides/química , Glucocorticoides/farmacologia , Masculino , Camundongos , Receptores de Glucocorticoides/agonistas , Índice de Gravidade de Doença
6.
J Immunol ; 202(6): 1680-1685, 2019 03 15.
Artigo em Inglês | MEDLINE | ID: mdl-30700587

RESUMO

IL-27 regulates immune responses in inflammation. The underlying mechanism of IL-27 functions has long been attributed to its ability to induce IL-10 production in activated CD4 T cells. In this study, we report that Foxp3+ regulatory T cells (Tregs) are the main target cells of IL-27, mediating its immunoregulatory functions in vivo. Systemically delivered IL-27 efficiently prevents the development of experimental autoimmune encephalomyelitis, an autoimmune inflammation in the CNS. However, it failed to do so upon Treg depletion. IL-27 signaling in Tregs was necessary, as transferring Tregs deficient in IL-27Rα or Lag3, a downstream molecule induced by IL-27, was unable to protect mice from experimental autoimmune encephalomyelitis. IL-27 efficiently induced IL-10 expression in CD4 T cells in vitro; however, we found no evidence supporting IL-27-induced IL-10 induction in CD4 T cells in vivo. Taken together, our results uncover an irreplaceable contribution of Tregs during IL-27-mediated control of inflammation.


Assuntos
Antígenos CD/imunologia , Autoimunidade/imunologia , Encefalomielite Autoimune Experimental/imunologia , Inflamação/imunologia , Interleucinas/imunologia , Linfócitos T Reguladores/imunologia , Animais , Interleucina-10/imunologia , Ativação Linfocitária/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteína do Gene 3 de Ativação de Linfócitos
7.
J Immunol ; 199(12): 3943-3951, 2017 12 15.
Artigo em Inglês | MEDLINE | ID: mdl-29093062

RESUMO

Understanding functions of Foxp3+ regulatory T cells (Tregs) during allergic airway inflammation remains incomplete. In this study, we report that, during cockroach Ag-induced allergic airway inflammation, Foxp3+ Tregs are rapidly mobilized into the inflamed lung tissues. However, the level of Treg accumulation in the lung was different depending on the type of inflammation. During eosinophilic airway inflammation, ∼30% of lung-infiltrating CD4 T cells express Foxp3, indicative of Tregs. On the contrary, only ∼10% of infiltrating CD4 T cells express Foxp3 during neutrophilic airway inflammation. Despite the different accumulation, the lung inflammation and inflammatory T cell responses were aggravated following Treg depletion, regardless of the type of inflammation, suggesting regulatory roles for Tregs. Interestingly, however, the extent to which inflammatory responses are aggravated by Treg depletion was significantly greater during eosinophilic airway inflammation. Indeed, lung-infiltrating Tregs exhibit phenotypic and functional features associated with potent suppression. Our results demonstrate that Tregs are essential regulators of inflammation, regardless of the type of inflammation, although the mechanisms used by Tregs to control inflammation may be shaped by environmental cues available to them.


Assuntos
Alveolite Alérgica Extrínseca/imunologia , Pulmão/imunologia , Infiltração de Neutrófilos/imunologia , Eosinofilia Pulmonar/imunologia , Linfócitos T Reguladores/imunologia , Adjuvantes Imunológicos/administração & dosagem , Adjuvantes Imunológicos/toxicidade , Alérgenos/administração & dosagem , Alérgenos/imunologia , Compostos de Alúmen/administração & dosagem , Compostos de Alúmen/toxicidade , Alveolite Alérgica Extrínseca/etiologia , Alveolite Alérgica Extrínseca/patologia , Animais , Baratas/imunologia , Modelos Animais de Doenças , Fatores de Transcrição Forkhead/análise , Adjuvante de Freund/administração & dosagem , Adjuvante de Freund/toxicidade , Perfilação da Expressão Gênica , Genes Reporter , Imunofenotipagem , Inflamação , Proteínas de Insetos/administração & dosagem , Proteínas de Insetos/imunologia , Pulmão/patologia , Camundongos Endogâmicos C57BL , Eosinofilia Pulmonar/etiologia , Eosinofilia Pulmonar/patologia , Organismos Livres de Patógenos Específicos
8.
J Immunol ; 193(7): 3792-802, 2014 Oct 01.
Artigo em Inglês | MEDLINE | ID: mdl-25143444

RESUMO

IL-33 has been implicated in the pathogenesis of asthma, atopic allergy, anaphylaxis, and other inflammatory diseases by promoting the production of proinflammatory cytokines and chemokines or Th2 immune responses. In this study, we analyzed the in vivo effect of IL-33 administration. IL-33 markedly promoted myelopoiesis in the bone marrow and myeloid cell emigration. Concomitantly, IL-33 induced hematopoietic stem and progenitor cell (HSPC) mobilization and extramedullary hematopoiesis. HSPC mobilization was mediated mainly through increased levels of CCL7 produced by vascular endothelial cells in response to IL-33. In vivo treatment of IL-33 rapidly induced phosphorylation of ERK, JNK, and p38, and inhibition of these signaling molecules completely blocked the production of CCL7 induced by IL-33. Consistently, inhibitor of CCR2 markedly reduced IL-33-mediated HSPC mobilization in vivo and migration of HSPCs in response to CCL7 in vitro. IL-33-mobilized HSPCs were capable of homing to, and of long-term reconstitution in, the bone marrow of irradiated recipients. Immune cells derived from these recipients had normal antifungal activity. The ability of IL-33 to promote migration of HSPCs and myeloid cells into the periphery and to regulate their antifungal activity represents a previously unrecognized role of IL-33 in innate immunity. These properties of IL-33 have clinical implications in hematopoietic stem cell transplantation.


Assuntos
Mobilização de Células-Tronco Hematopoéticas , Células-Tronco Hematopoéticas/imunologia , Interleucinas/farmacologia , Células Mieloides/imunologia , Mielopoese/imunologia , Receptores CCR2/imunologia , Animais , Autoenxertos , Transplante de Medula Óssea , Quimiocina CCL7/genética , Quimiocina CCL7/imunologia , MAP Quinases Reguladas por Sinal Extracelular/genética , MAP Quinases Reguladas por Sinal Extracelular/imunologia , Feminino , Imunidade Inata/efeitos dos fármacos , Imunidade Inata/genética , Interleucina-33 , Interleucinas/genética , Interleucinas/imunologia , Sistema de Sinalização das MAP Quinases/efeitos dos fármacos , Sistema de Sinalização das MAP Quinases/genética , Sistema de Sinalização das MAP Quinases/imunologia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Mielopoese/efeitos dos fármacos , Receptores CCR2/genética
9.
Antimicrob Agents Chemother ; 59(10): 6308-16, 2015 Oct.
Artigo em Inglês | MEDLINE | ID: mdl-26248370

RESUMO

Methicillin-resistant Staphylococcus aureus (MRSA) is a common pathogen associated with nosocomial pneumonia and is an increasing threat for severe community-acquired pneumonia. We have now investigated the role of interleukin-12 (IL-12) in protective immunity against lung infection with MRSA. The importance of IL-12 in protection from pulmonary MRSA infection was demonstrated by the finding that IL-12p35-deficient mice had a lower survival rate, higher bacterial burdens in lung and spleen, and decreased expression of interferon gamma (IFN-γ) in the lung compared to wild-type mice. These effects were completely reversed by replacement intranasal therapy with recombinant IL-12. Furthermore, exogenous IL-12 treatment of wild-type mice 24 h before pulmonary challenge with a lethal dose of MRSA significantly improved bacterial clearance and resulted in protection from death. The IL-12-treated mice had increased numbers of lung natural killer (NK) cells and neutrophils and higher levels of IFN-γ in the lung and serum compared to untreated mice. The major source of IL-12-driven IFN-γ expression in the lung was the NK cell, and the direct target of pulmonary IFN-γ was the lung macrophage, as shown using mice with a macrophage-specific defect in interferon gamma (IFN-γ) signaling (MIIG mice). Importantly, combination therapy with linezolid and IL-12 following intranasal MRSA infection significantly increased survival compared to that of mice receiving linezolid or IL-12 alone. These results indicate that IL-12-based immunotherapy may hold promise for treatment of MRSA pneumonia.


Assuntos
Antibacterianos/farmacologia , Subunidade p35 da Interleucina-12/farmacologia , Linezolida/farmacologia , Pulmão/efeitos dos fármacos , Pneumonia Bacteriana/tratamento farmacológico , Infecções Estafilocócicas/tratamento farmacológico , Animais , Quimioterapia Combinada , Feminino , Regulação da Expressão Gênica , Interferon gama/biossíntese , Interferon gama/genética , Interferon gama/imunologia , Subunidade p35 da Interleucina-12/genética , Subunidade p35 da Interleucina-12/imunologia , Pulmão/imunologia , Pulmão/microbiologia , Macrófagos Alveolares/efeitos dos fármacos , Macrófagos Alveolares/imunologia , Macrófagos Alveolares/microbiologia , Masculino , Staphylococcus aureus Resistente à Meticilina/efeitos dos fármacos , Staphylococcus aureus Resistente à Meticilina/crescimento & desenvolvimento , Staphylococcus aureus Resistente à Meticilina/patogenicidade , Camundongos , Camundongos Endogâmicos C57BL , Camundongos Knockout , Neutrófilos/efeitos dos fármacos , Neutrófilos/imunologia , Neutrófilos/microbiologia , Pneumonia Bacteriana/genética , Pneumonia Bacteriana/imunologia , Pneumonia Bacteriana/mortalidade , Transdução de Sinais , Infecções Estafilocócicas/genética , Infecções Estafilocócicas/imunologia , Infecções Estafilocócicas/mortalidade , Análise de Sobrevida
10.
Infect Immun ; 81(6): 2168-77, 2013 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-23545301

RESUMO

Severe sepsis and septic shock caused mainly by bacterial infections are life-threatening conditions that urge the development of novel therapies. However, host responses to and pathophysiology of sepsis have not been clearly understood, which remains a major obstacle for the development of effective therapeutics. Recently, we have shown that stimulation of a costimulatory molecule, CD137, enhanced survival of mice infected with the Gram-positive (G(+)) intracellular bacterium Listeria monocytogenes but decreased survival in a polymicrobial sepsis model. Herein, we report that CD137 deficiency or blocking of CD137 signaling decreased antibacterial responses of mice infected with G(+) bacteria (Staphylococcus aureus, Streptococcus pneumoniae, and Enterococcus faecalis) but increased these responses in mice infected with Gram-negative (G(-)) bacteria (Escherichia coli, Pseudomonas aeruginosa, and Salmonella enterica serovar Typhimurium). Consistent with these findings, stimulation of CD137 by administration of agonistic antibody enhanced responses against G(+) bacteria, whereas it decreased these responses against G(-) bacteria. Neutrophils were responsible for CD137-mediated opposite roles in control of G(+) and G(-) bacterial infections. Stimulation of CD137 enhanced activities of neutrophils against S. aureus but decreased these activities against E. coli, while CD137 blocking produced opposite results with the stimulation of CD137 in vivo and in vitro. Furthermore, we found that combined signaling of CD137 and Toll-like receptor 2 (TLR2) induced synergistic production of tumor necrosis factor alpha (TNF-α) and interleukin-6 (IL-6) by neutrophils, but combined signaling of CD137 and TLR4 did not. Our data strongly suggest that CD137 may play a dual role in sepsis in association with TLRs.


Assuntos
Infecções por Bactérias Gram-Negativas/imunologia , Infecções por Bactérias Gram-Positivas/imunologia , Neutrófilos/metabolismo , Membro 9 da Superfamília de Receptores de Fatores de Necrose Tumoral/metabolismo , Animais , Citocinas/genética , Citocinas/metabolismo , Regulação da Expressão Gênica/imunologia , Bactérias Gram-Negativas/imunologia , Bactérias Gram-Positivas/imunologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout , Transdução de Sinais , Organismos Livres de Patógenos Específicos , Receptor 2 Toll-Like/genética , Receptor 2 Toll-Like/metabolismo
11.
STAR Protoc ; 4(4): 102686, 2023 Dec 15.
Artigo em Inglês | MEDLINE | ID: mdl-37925636

RESUMO

Single-cell isolation techniques allow the investigation of physical and functional relationships between individual cells within a complex cell population. Here, we present a protocol for single-cell isolation from full-thickness intestinal tissue resections. We describe steps for pre-processing specimens, isolation of lamina propria and muscular layers, and red blood cell lysis. We then detail fixation of isolated cells and assessment of cell quality. The resulting cell suspension can be subjected to RNA sequencing on the 10× Chromium platform. For complete details on the use and execution of this protocol, please refer to Mukherjee et al.1.


Assuntos
Eritrócitos , Técnicas Histológicas , Humanos , Morte Celular , Separação Celular , Análise de Sequência de RNA
12.
bioRxiv ; 2023 Apr 04.
Artigo em Inglês | MEDLINE | ID: mdl-37066202

RESUMO

Background: Fibroblasts play a key role in stricture formation in Crohn's disease (CD) but understanding it's pathogenesis requires a systems-level investigation to uncover new treatment targets. We studied full thickness CD tissues to characterize fibroblast heterogeneity and function by generating the first single cell RNA sequencing (scRNAseq) atlas of strictured bowel and providing proof of principle for therapeutic target validation. Methods: We performed scRNAseq of 13 fresh full thickness CD resections containing non-involved, inflamed non-strictured, and strictured segments as well as 7 normal non-CD bowel segments. Each segment was separated into mucosa/submucosa or muscularis propria and analyzed separately for a total of 99 tissue samples and 409,001 cells. We validated cadherin-11 (CDH11) as a potential therapeutic target by using whole tissues, isolated intestinal cells, NanoString nCounter, next generation sequencing, proteomics and animal models. Results: Our integrated dataset revealed fibroblast heterogeneity in strictured CD with the majority of stricture-selective changes detected in the mucosa/submucosa, but not the muscle layer. Cell-cell interaction modeling revealed CXCL14+ as well as MMP/WNT5A+ fibroblasts displaying a central signaling role in CD strictures. CDH11, a fibroblast cell-cell adhesion molecule, was broadly expressed and upregulated, and its pro-fibrotic function was validated by NanoString nCounter, RNA sequencing, tissue target expression, in vitro gain- and loss-of-function experiments, proteomics, and two animal models of experimental colitis. Conclusion: A full-thickness bowel scRNAseq atlas revealed previously unrecognized fibroblast heterogeneity and interactions in CD strictures and CDH11 was validated as a potential therapeutic target. These results provide a new resource for a better understanding of CD stricture formation and opens potential therapeutic developments.

13.
Matrix Biol ; 113: 1-21, 2022 11.
Artigo em Inglês | MEDLINE | ID: mdl-36108990

RESUMO

OBJECTIVE: Inflammatory bowel diseases (IBD) cause chronic intestinal damage and extracellular matrix (ECM) remodeling. The ECM may play an active role in inflammation by modulating immune cell functions, including cell adhesion, but this hypothesis has not been tested in IBD. DESIGN: Primary human intestinal myofibroblast (HIMF)-derived ECM from IBD and controls, 3D decellularized colon or ECM molecule-coated scaffolds were tested for their adhesiveness for T cells. Matrisome was analysed via proteomics. Functional integrin blockade was used to investigate the underlying mechanism. Analysis of the pediatric Crohn's disease (CD) RISK inception cohort was used to explore an altered ECM gene expression as a potential predictor for a future complicated disease course. RESULTS: HIMF-derived ECM and 3D decellularized colonic ECM from IBD bound more T cells compared to control. Control HIMFs exposed to the pro-inflammatory cytokines Iinterleukin-1ß (IL-1ß) and tumor necrosis factor (TNF) increased, and to transforming growth factor-ß1 (TGF-ß1) decreased ECM adhesiveness to T cells. Matrisome analysis of the HIMF-derived ECM revealed collagen VI as a major culprit for differences in T cell adhesion. Collagen VI knockdown in HIMF reduced adhesion T cell as did the blockage of integrin αvß1. Elevated gene expression of collagen VI in biopsies of pediatric CD patients was linked to risk for future stricturing disease. CONCLUSION: HIMF-derived ECM in IBD binds a remarkably enhanced number of T cells, which is dependent on Collagen VI and integrin αvß1. Collagen VI expression is a risk factor for a future complicated CD course. Blocking immune cells retention may represent a novel approach to treatment in IBD.


Assuntos
Doenças Inflamatórias Intestinais , Miofibroblastos , Criança , Humanos , Miofibroblastos/metabolismo , Adesividade , Linfócitos T/patologia , Colágeno/metabolismo , Inflamação/metabolismo
14.
Oxid Med Cell Longev ; 2021: 5427153, 2021.
Artigo em Inglês | MEDLINE | ID: mdl-35003518

RESUMO

Sepsis is characterized by an initial net hyperinflammatory response, followed by a period of immunosuppression, termed immunoparalysis. During this immunosuppressive phase, patients may have difficulty eradicating invading pathogens and are susceptible to life-threatening secondary hospital-acquired infections. Due to progress in antimicrobial treatment and supportive care, most patients survive early sepsis. Mortality is more frequently attributed to subsequent secondary nosocomial infections and multiorgan system failure. 6-Gingerol is the major pharmacologically active component of ginger. Although it is known to exhibit a variety of biological activities, including anti-inflammation and antioxidation, the role of 6-gingerol in sepsis-induced immune dysfunction remains elusive. Thus, we investigated whether 6-gingerol improves septic host response to infections during sepsis. 6-Gingerol-treated mice showed significantly lower mortality in polymicrobial sepsis induced by cecal ligation and puncture LPS via enhanced bacterial clearance in the peritoneum, blood, and organs (liver, spleen, and kidney) and inhibited the production of TNF-α and IL-6 in TLR2 and/or TLR4-stimulated macrophages. In addition, we demonstrated that survival improvement of secondary infection following septic insult was associated with an initial response of enhanced neutrophil numbers and function at the infection site, reduced apoptosis of immune cells, and a shift from a T helper cell type 2 (Th2) to a T helper cell type 1 (Th1) cytokine balance in the hypoinflammation phase. Our overall findings suggest that 6-gingerol potentially restores sepsis-induced immune dysfunction by shifting the balance of Th1/Th2 and by regulating apoptosis of immune cells.


Assuntos
Catecóis/uso terapêutico , Citocinas/metabolismo , Álcoois Graxos/uso terapêutico , Doenças do Sistema Imunitário/tratamento farmacológico , Linfócitos/metabolismo , Sepse/complicações , Animais , Apoptose , Catecóis/farmacologia , Álcoois Graxos/farmacologia , Humanos , Doenças do Sistema Imunitário/fisiopatologia , Masculino , Camundongos
15.
Front Immunol ; 11: 181, 2020.
Artigo em Inglês | MEDLINE | ID: mdl-32117306

RESUMO

Graft-versus-host disease (GvHD) remains a significant complication of allogeneic hematopoietic cell transplantation (HCT), associated with significant morbidity and mortality. GvHD is characterized by dysregulated immune responses and resulting tissue damage of target organs. Recent investigations have focused on Foxp3+ regulatory T cells (Tregs) as a therapeutic tool, based on its regulatory functions in GvHD pathogenesis and their instrumental role in mitigating GvHD severity while preserving graft-versus-leukemia (GvL) activity. There are several challenges to its clinical application, including their paucity, impaired suppressive activity, and instability in vivo. Herein, we report that IL-27 pre-stimulation enhances suppressive functions of both mouse and human Tregs. In a complete MHC mismatched murine bone marrow transplant model, IL-27 pre-stimulated polyclonal iTregs diminish acute (a)GvHD lethality, while preserving the GvL effect. Allo-antigen specificity further improves suppressive functions when combined with IL-27 pre-stimulation. In a xenogeneic (human to mouse) GvHD model, IL-27 pre-stimulated human iTregs are superior in protecting recipients from GvHD. Lastly, we compared gene expression profiles of circulating Tregs isolated from HCT recipients with and without aGvHD and found that Tregs from aGvHD patients express distinct gene signatures enriched in immune activation and inflammation. Therefore, these results highlight a novel function of IL-27 in enforcing Treg functions to prevent aGvHD mediated lethality, proposing the hypothesis that dysregulated Treg functions may account for the potential mechanisms underlying GvHD development.


Assuntos
Transplante de Medula Óssea/efeitos adversos , Doença Enxerto-Hospedeiro/imunologia , Doença Enxerto-Hospedeiro/prevenção & controle , Transplante de Células-Tronco Hematopoéticas/efeitos adversos , Interleucinas/administração & dosagem , Linfócitos T Reguladores/imunologia , Transferência Adotiva/métodos , Aloenxertos/imunologia , Animais , Transplante de Medula Óssea/métodos , Doença Enxerto-Hospedeiro/sangue , Doença Enxerto-Hospedeiro/mortalidade , Efeito Enxerto vs Leucemia/imunologia , Transplante de Células-Tronco Hematopoéticas/métodos , Xenoenxertos/imunologia , Interleucinas/sangue , Leucemia/terapia , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Endogâmicos C57BL , Camundongos Knockout , Proteínas Recombinantes/administração & dosagem , Transcriptoma , Resultado do Tratamento
16.
Infect Immun ; 77(9): 3932-8, 2009 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-19564374

RESUMO

Sepsis, a leading cause of death worldwide, involves proinflammatory responses and inefficient bacterial clearance. Previously, we have shown that CD137 (4-1BB), a member of the tumor necrosis factor receptor superfamily, plays critical roles in eradicating infective Listeria monocytogenes, a gram-positive bacterium, and that stimulation of CD137 protects mice from sepsis-induced death. In this study, we unexpectedly found that CD137 activation aggravated polymicrobial sepsis due to mixed gram-positive and gram-negative bacterial infection induced by cecal ligation and puncture (CLP). CD137-deficient (CD137(-/-)) mice showed significantly lower mortality than CD137-sufficient (CD137(+/+)) mice in the CLP model. Administration of an agonistic anti-CD137 monoclonal antibody (MAb) to CD137(+/+) mice decreased their survival in this infection model, while administration of a blocking anti-CD137 ligand MAb (TKS-1) to such mice increased their survival. CD137(-/-) mice and TKS-1-treated CD137(+/+) mice had lower levels of chemokines/proinflammatory cytokines (monocyte chemoattractant protein 1, interleukin-6 [IL-6], tumor necrosis factor alpha, IL-12) and an anti-inflammatory cytokine (IL-10), exhibited improved bacterial clearance in the peritoneum, liver, and blood, and had greater numbers of infiltrated peritoneal neutrophils and macrophages in the CLP model than control mice. Our data suggest that CD137 activation aggravates polymicrobial sepsis induced by CLP.


Assuntos
Sepse/prevenção & controle , Transdução de Sinais/fisiologia , Membro 9 da Superfamília de Receptores de Fatores de Necrose Tumoral/antagonistas & inibidores , Animais , Anticorpos Monoclonais/uso terapêutico , Ceco , Movimento Celular , Leucócitos/fisiologia , Ligadura , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Punções , Sepse/microbiologia , Membro 9 da Superfamília de Receptores de Fatores de Necrose Tumoral/fisiologia
17.
Infect Immun ; 77(5): 2168-76, 2009 May.
Artigo em Inglês | MEDLINE | ID: mdl-19237524

RESUMO

The tumor necrosis factor receptor family molecule 4-1BB (CD137) has diverse roles in adaptive and innate immune responses. However, little is known of its role in bacterial infections. Previously, we showed that 4-1BB-deficient mice have enhanced susceptibility to Listeria monocytogenes infection, and mice pretreated with agonistic anti-4-1BB antibody (3E1) were much more resistant to L. monocytogenes infection than mice treated with control antibody. In this study, we report that stimulating 4-1BB by administering 3E1 in the early phase of L. monocytogenes infection is critical for promoting the survival of mice by inducing rapid infiltration of neutrophils and monocytes into L. monocytogenes-infected livers. The levels of tumor necrosis factor alpha, interleukin 6, and monocyte chemoattractant protein 1 in the livers of 3E1-treated mice increased as early as 30 min postinfection and peaked by 1 to 2 h, while those in mice treated with control antibody started to increase only at 16 h postinfection. Monocytes and neutrophils from the 3E1-treated mice had higher levels of activation markers, phagocytic activity, and reactive oxygen species than those from control mice. In vitro stimulation of 4-1BB induced the production of the inflammatory cytokines/chemokines of neutrophils, but not those of monocytes. These results suggest that 4-1BB stimulation of neutrophils in the early phase of L. monocytogenes infection causes rapid production of inflammatory cytokines/chemokines and that the subsequent infiltration of neutrophils and monocytes is crucial for eliminating the infecting L. monocytogenes.


Assuntos
Anticorpos Monoclonais/uso terapêutico , Listeria monocytogenes/imunologia , Listeriose/imunologia , Monócitos/imunologia , Neutrófilos/imunologia , Membro 9 da Superfamília de Receptores de Fatores de Necrose Tumoral/agonistas , Membro 9 da Superfamília de Receptores de Fatores de Necrose Tumoral/imunologia , Animais , Quimiocina CCL2/análise , Feminino , Interleucina-6/análise , Fígado/química , Fígado/imunologia , Fígado/patologia , Camundongos , Camundongos Endogâmicos BALB C , Fagocitose , Espécies Reativas de Oxigênio/metabolismo , Análise de Sobrevida , Fator de Necrose Tumoral alfa/análise
18.
JCI Insight ; 4(2)2019 Jan 24.
Artigo em Inglês | MEDLINE | ID: mdl-30674714

RESUMO

Foxp3+ CD4 Tregs are central regulators of inflammation, including allergic inflammation in the lung. There is increasing evidence that inflammatory factors undermine adequate Treg functions and homeostasis, resulting in prolonged and exacerbated inflammation. Therefore, identifying the factors is of the utmost important. IL-27 is an antiinflammatory cytokine implicated in immune regulation and tolerance. However, the cellular mechanisms underlying IL-27-mediated immune regulation in vivo remain largely unknown. Utilizing a cockroach antigen-induced allergic inflammation model in mice, we sought to test the roles of Tregs during IL-27-mediated regulation of allergic inflammation. Intranasally delivered IL-27 significantly reduced the development of airway inflammation. Unexpectedly, the IL-27-induced reduction occurred only in the presence of Tregs. Il27ra-/- and Treg-specific Il27ra-/- mice developed severe airway inflammation, and IL-27 treatment had little impact on diminishing the inflammatory responses. IL-27-induced treatment was restored following transfer of WT Tregs but not of Tregs deficient in Lag3, a molecule induced by IL-27 in Tregs. Finally, Tregs from asthmatic patients exhibited blunted STAT1 phosphorylation following IL-27 stimulation. Taken together, our results uncover that Tregs are the primary target cells of IL-27 in vivo to mediate its antiinflammatory functions, suggesting that altered IL-27 responsiveness in Tregs may underlie inadequate Treg functions and perpetuation of inflammation.

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