RESUMO
The pathogenesis of allergic contact dermatitis (ACD) requires the activation of Ag-specific T cells, including effector and regulatory T cells. The differentiation and function of these T cells is epigenetically regulated through DNA methylation and histone modifications. However, the roles of altered histone H3K27 methylation in T cells in the development of ACD remain unknown. Two types of histone H3K27 demethylases, Utx and Jmjd3, have been reported in mammals. To determine the role of the histone H3K27 demethylase expression of T cells in the development of ACD, we generated T cell-specific, Utx-deficient (Utx KO) mice or Jmjd3-deficient (Jmjd3 KO) mice. Unlike control mice, Utx KO mice had severer symptoms of ACD, whereas Jmjd3 KO mice showed symptoms identical to those in control mice. In Utx KO mice with ACD, the massive infiltration of myeloid cells, including neutrophils and dendritic cells, has been observed. In addition, the expression of proinflammatory cytokines in CD4+ T cells of the draining lymph nodes (LNs) and in CD8+ T cells of the skin was increased in Utx KO mice, whereas the ratio of Foxp3+ regulatory CD4+ T cells to Foxp3- conventional CD4+ T cells was decreased in both the draining LNs and the skin of Utx KO mice with ACD. Furthermore, Foxp3+ regulatory CD4+ T cells of Utx KO mice with ACD expressed a decreased level of CCR4 (a skin-tropic chemokine receptor) in comparison with control. Thus, in CD4+ T cells, Utx could potentially be involved in the regulation of the pathogenesis of ACD.
Assuntos
Dermatite de Contato/imunologia , Histona Desmetilases/metabolismo , Histonas/metabolismo , Histona Desmetilases com o Domínio Jumonji/metabolismo , Pele/imunologia , Subpopulações de Linfócitos T/imunologia , Linfócitos T Reguladores/imunologia , Animais , Citocinas/metabolismo , Modelos Animais de Doenças , Fatores de Transcrição Forkhead/metabolismo , Histona Desmetilases/genética , Histonas/genética , Humanos , Mediadores da Inflamação/metabolismo , Histona Desmetilases com o Domínio Jumonji/genética , Camundongos , Camundongos Knockout , Receptores CCR4/metabolismoRESUMO
Thymic stromal lymphopoietin (TSLP) acts on dendritic cells (DCs), which prime helper T (Th) cells to become type 2 cytokine producing cells. Recently, a different set of populations of TSLP-responsive DCs has been discovered. Here, we identified two populations of CD103loEpCAMhi migratory DCs (fraction I and fraction II) that accumulated in skin-draining lymph nodes in response to TSLP expressed in the mouse skin. Fraction I DCs with CD11b+PDL2hi expression primed naïve Th cells to differentiate into cells secreting IFN-γ, IL-17A and IL-22, while fraction II DCs with CD11bloPDL2+ expression primed naïve Th cells to differentiate into cells secreting IL-4, IL-5, IL-9, IL-13 and IL-10. Fraction I DCs migrated from the skin via IL-4Rα signaling pathway, whereas fraction II DCs migrated partially via TSLPR signaling pathway. All suggest that at least two populations of CD103loEpCAMhi DCs with distinct functions and pathways could migrate in response to TSLP expression in the skin.
Assuntos
Citocinas/metabolismo , Células Dendríticas/metabolismo , Animais , Antígenos CD/imunologia , Antígenos CD/metabolismo , Citocinas/fisiologia , Células Dendríticas/fisiologia , Feminino , Cadeias alfa de Integrinas/imunologia , Cadeias alfa de Integrinas/metabolismo , Interleucina-17/metabolismo , Linfonodos/metabolismo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Transgênicos , Pele/metabolismo , Linfócitos T Auxiliares-Indutores/metabolismo , Células Th2/metabolismo , Linfopoietina do Estroma do TimoRESUMO
General anesthesia is thought to suppress the immune system and negatively affect postoperative infection and the long-term prognosis of cancer. However, the mechanism underlying immunosuppression induced by general anesthetics remains unclear. In this study, we focused on propofol, which is widely used for sedation under general anesthesia and intensive care and examined its effects on the T cell function and T cell-dependent immune responses. We found that propofol suppressed T cell glycolytic metabolism, differentiation into effector T cells, and cytokine production by effector T cells. CD8 T cells activated and differentiated into effector cells in the presence of propofol in vitro showed reduced antitumor activity. Furthermore, propofol treatment suppressed the increase in the number of antigen-specific CD8 T cells during Listeria infection. In contrast, the administration of propofol improved inflammatory conditions in mouse models of inflammatory diseases, such as OVA-induced allergic airway inflammation, hapten-induced contact dermatitis, and experimental allergic encephalomyelitis. These results suggest that propofol may reduce tumor and infectious immunity by suppressing the T cell function and T cell-dependent immune responses while improving the pathogenesis and prognosis of chronic inflammatory diseases by suppressing inflammation.
Assuntos
Linfócitos T CD8-Positivos , Propofol , Propofol/farmacologia , Animais , Camundongos , Linfócitos T CD8-Positivos/imunologia , Linfócitos T CD8-Positivos/efeitos dos fármacos , Camundongos Endogâmicos C57BL , Linfócitos T/imunologia , Linfócitos T/efeitos dos fármacos , Ativação Linfocitária/efeitos dos fármacos , Inflamação/imunologia , Diferenciação Celular/efeitos dos fármacos , Citocinas/metabolismo , Listeriose/imunologia , Modelos Animais de Doenças , Encefalomielite Autoimune Experimental/imunologia , Encefalomielite Autoimune Experimental/tratamento farmacológico , FemininoRESUMO
The cytokine thymic stromal lymphopoietin (TSLP) functions as a regulator of bone marrow B-cell development and a key initiator of allergic inflammation. In the current study, we show that mature B cells, derived from transgenic mice with systemically elevated levels of TSLP (K5-TSLP mice), exhibit markedly enhanced mitogenic responses in vitro and that this enhanced responsiveness leads to polyclonal B-cell activation and development of autoimmune hemolytic anemia in vivo. In contrast, B cells derived from K5-TSLP mice lacking CD4(+) T cells failed to show polyclonal activation. Furthermore, neither mature B-cell activation nor hemolytic anemia occurred in IL-4-deficient K5-TSLP mice. Consistent with these findings, activation of mature B cells occurred independently of B-cell intrinsic TSLP signals. Taken together, our results demonstrate that systemic alterations in TSLP, through induction of IL-4 from CD4(+) T cells and other cell types, functions as an important factor in peripheral B-cell homeostasis and promotion of humoral autoimmunity.
Assuntos
Linfócitos B/imunologia , Linfócitos T CD4-Positivos/imunologia , Citocinas/imunologia , Interleucina-4/imunologia , Ativação Linfocitária/imunologia , Anemia Hemolítica Autoimune/imunologia , Animais , Autoimunidade/imunologia , Separação Celular , Modelos Animais de Doenças , Ensaio de Imunoadsorção Enzimática , Citometria de Fluxo , Camundongos , Camundongos Knockout , Linfopoietina do Estroma do TimoRESUMO
Allergic contact dermatitis (ACD) and atopic dermatitis develop through delayed-type hypersensitivity reactions mediated by T cells. The development of immunomodulatory drugs, such as Jak inhibitors, would be useful for the long-term management of these diseases owing to their profile of favorable adverse effects. However, the efficacy of Jak inhibitors for ACD treatment has not been fully determined under a variety of settings. Therefore, we evaluated the effects of ruxolitinib, a Jak inhibitor for Jak1 and Jak2, using a mouse ACD model. As a result, the lower numbers of immune cells, including CD4+ T cells, CD8+ T cells, neutrophils, and possibly macrophages, as well as milder pathophysiological aspects have been observed in the inflamed skin of ACD with the administration of ruxolitinib. In addition, the treatment of differentiating T cells with ruxolitinib downregulated the level of IL-2-mediated glycolysis in vitro. Furthermore, symptoms of ACD did not develop in T-cell-specific Pgam1-deficient mice whose T cells had no glycolytic capacity. Taken together, our data suggest that the downregulation of glycolysis in T cells by ruxolitinib could be an important factor in the suppression of ACD development in mice.
Assuntos
Dermatite Alérgica de Contato , Inibidores de Janus Quinases , Camundongos , Animais , Inibidores de Janus Quinases/farmacologia , Inibidores de Janus Quinases/uso terapêutico , Linfócitos T CD8-Positivos , Pirazóis/farmacologia , Pirazóis/uso terapêutico , Modelos Animais de DoençasRESUMO
The cytokine TSLP was originally identified in a murine thymic stromal cell line as a lymphoid growth factor. After the discovery of TSLP, extensive molecular genetic analyses and gene targeting experiments have demonstrated that TSLP plays an essential role in allergic diseases. In this review, we discuss the current status of TSLP and its functional role in allergic diseases particularly by focusing on effects of TSLP on haematopoietic cells in mouse models. It is our conclusion that a number of research areas, i.e., a new source of TSLP, effects of TSLP on non-haematopoietic and haematopoietic cells, synergistic interactions of cytokines including IL-25 and IL-33 and a regulation of TSLP expression and its function, are critically needed to understand the whole picture of TSLP involvement in allergic diseases. The mouse models will thus contribute further to our understanding of TSLP involvement in allergic diseases and development of therapeutic measures for human allergic diseases.
Assuntos
Citocinas/fisiologia , Modelos Animais de Doenças , Hipersensibilidade/imunologia , Camundongos , Animais , Citocinas/metabolismo , Humanos , Hipersensibilidade/metabolismo , Leucócitos/imunologia , Leucócitos/metabolismo , Linfopoietina do Estroma do TimoRESUMO
NKT cells are characterized by their production of both T(h)1 and T(h)2 cytokines immediately after stimulation with alpha-galactosylceramide (alpha-GalCer), which is composed of alpha-galactopyranose linked to ceramide (itself composed of sphingosine and fatty-acyl chains); the chain length of the ceramide varies and this affects the ability of alpha-GalCer to stimulate cytokine production. However, the contribution of its galactopyranose sugar moiety remains unclear. We synthesized alpha-carba-GalCer, which has an alpha-linked carba-galactosyl moiety; here, the 5a'-oxygen atom of the D-galactopyranose ring of alpha-GalCer is replaced by a methylene group. The alpha-carba-GalCer was more stable and showed higher affinity to the NKT receptor. It thus enhanced and prolonged production of IL-12 and IFN-gamma compared with alpha-GalCer, resulting in augmented NKT cell-mediated adjuvant effects in vivo. The alpha-carba-GalCer, which has an ether linkage, was more resistant to degradation by liver microsomes than was alpha-GalCer, which has an acetal bond. Modulation of the sugar moiety in glycolipids might therefore provide optimal therapeutic reagents for protective immune responses against tumor or pathogens.
Assuntos
Adjuvantes Imunológicos/farmacologia , Cicloexanóis/farmacologia , Citocinas/biossíntese , Galactosilceramidas/farmacologia , Células T Matadoras Naturais/efeitos dos fármacos , Células Th1/imunologia , Adjuvantes Imunológicos/síntese química , Adjuvantes Imunológicos/química , Animais , Linhagem Celular , Cicloexanóis/síntese química , Cicloexanóis/química , Citocinas/análise , Galactosilceramidas/síntese química , Galactosilceramidas/química , Glicolipídeos/metabolismo , Humanos , Injeções Intravenosas , Ligantes , Camundongos , Células T Matadoras Naturais/imunologiaRESUMO
To elucidate whether leukocyte cell-derived chemotaxin 2 (LECT2) controls the progression of staphylococcal enterotoxin A (SEA)-induced toxicity, we examined the role of LECT2 in a mouse model. Almost all the C57BL/6J (B6) mice survived for 72 h after the injection of 0.1 µg of SEA and 20 mg of d-galactosamine (d-GalN). However, the same treatment protocol in LECT2(-/-) mice produced a high lethality (~90%), severe hepatic apoptosis, and massive hepatic and pulmonary hemorrhage, similar to the situation observed in B6 mice treated with 1.0 µg SEA/d-GalN. The plasma LECT2 levels in B6 mice treated with 1.0 µg SEA/d-GalN were inversely correlated with the plasma cytokine levels and were associated with prognosis. LECT2 administration increased the survival of B6 mice and down-regulated TNF-α and IL-6. These results suggest the involvement of LECT2 in the regulation of fatal SEA-induced toxicity in d-GalN-sensitized mice.
Assuntos
Doença Hepática Induzida por Substâncias e Drogas/patologia , Enterotoxinas , Galactosamina/imunologia , Peptídeos e Proteínas de Sinalização Intercelular/imunologia , Fígado/patologia , Pulmão/patologia , Choque Séptico/imunologia , Linfócitos T/imunologia , Animais , Apoptose , Doença Hepática Induzida por Substâncias e Drogas/imunologia , Modelos Animais de Doenças , Regulação para Baixo/efeitos dos fármacos , Regulação para Baixo/imunologia , Enterotoxinas/imunologia , Enterotoxinas/toxicidade , Feminino , Citometria de Fluxo , Hemorragia/induzido quimicamente , Marcação In Situ das Extremidades Cortadas , Peptídeos e Proteínas de Sinalização Intercelular/administração & dosagem , Peptídeos e Proteínas de Sinalização Intercelular/farmacologia , Interleucina-6/metabolismo , Fígado/efeitos dos fármacos , Fígado/imunologia , Pulmão/efeitos dos fármacos , Pulmão/imunologia , Camundongos , Camundongos Endogâmicos C57BL , Choque Séptico/induzido quimicamente , Choque Séptico/patologia , Linfócitos T/efeitos dos fármacos , Fatores de Tempo , Fator de Necrose Tumoral alfa/metabolismoRESUMO
Primary tumours establish metastases by interfering with distinct organs. In pre-metastatic organs, a tumour-friendly microenvironment supports metastatic cells and is prepared by many factors including tissue resident cells, bone marrow-derived cells and abundant fibrinogen depositions. However, other components are unclear. Here, we show that a third organ, originally regarded as a bystander, plays an important role in metastasis by directly affecting the pre-metastatic soil. In our model system, the liver participated in lung metastasis as a leucocyte supplier. These liver-derived leucocytes displayed liver-like characteristics and, thus, were designated hepato-entrained leucocytes (HepELs). HepELs had high expression levels of coagulation factor X (FX) and vitronectin (Vtn) and relocated to fibrinogen-rich hyperpermeable regions in pre-metastatic lungs; the cells then switched their expression from Vtn to thrombospondin, both of which were fibrinogen-binding proteins. Cell surface marker analysis revealed that HepELs contained B220+CD11c+NK1.1+ cells. In addition, an injection of B220+CD11c+NK1.1+ cells successfully eliminated fibrinogen depositions in pre-metastatic lungs via FX Moreover, B220+CD11c+NK1.1+ cells demonstrated anti-metastatic tumour ability with IFNγ induction. These findings indicate that liver-primed B220+CD11c+NK1.1+ cells suppress lung metastasis.
Assuntos
Células Matadoras Naturais/imunologia , Fígado/patologia , Neoplasias Pulmonares/patologia , Pulmão/patologia , Metástase Neoplásica , Lesões Pré-Cancerosas , Animais , Antígenos CD11 , Feminino , Fibrinogênio/metabolismo , Citometria de Fluxo , Humanos , Interferon gama/imunologia , Antígenos Comuns de Leucócito , Fígado/imunologia , Pulmão/imunologia , Neoplasias Pulmonares/imunologia , Neoplasias Pulmonares/secundário , Masculino , CamundongosRESUMO
In the pathogenesis of atopic dermatitis (AD), skin barrier dysfunction and T-helper (Th) type 2 immune reactions play important roles. Alterations in the stratum spinosum of AD have not been studied in much detail. In this study, we investigated the changes of structural proteins and adhesion molecules residing in the stratum spinosum of AD lesional skin, and whether Th2 cytokines including interleukin (IL)-4 and IL-13 alter the expression of these proteins. Skin samples were collected from patients with AD and from healthy controls. Normal human epidermal keratinocytes were cultured and differentiated in the presence or absence of either IL-4 or IL-13 in vitro. The expression of keratin 1, keratin 10, desmoglein 1 and desmocollin 1 was examined by immunofluorescence and western blotting. The expression of these proteins was downregulated in AD lesional skin, and IL-4 and IL-13 were shown to suppress their expression. These results suggest that the stratum spinosum is impaired in AD lesional skin, possibly by Th2 cytokines, which may be involved in the pathogenesis of AD.
Assuntos
Dermatite Atópica/imunologia , Desmogleína 1/imunologia , Epiderme/imunologia , Interleucina-13/imunologia , Interleucina-4/imunologia , Queratina-10/imunologia , Queratina-1/imunologia , Adulto , Células Cultivadas , Desmogleína 1/metabolismo , Regulação para Baixo , Epiderme/metabolismo , Feminino , Humanos , Interleucina-13/sangue , Interleucina-4/sangue , Queratina-1/metabolismo , Queratina-10/metabolismo , Masculino , Pessoa de Meia-Idade , Adulto JovemRESUMO
T helper type 2 (Th2) cytokines, IL-4 and IL-13, attenuate the expression of genes that regulate epidermal cellular structures and the barrier function at the terminal stage of keratinocyte differentiation. However, whether these Th2 cytokines act at earlier stages remains unknown. We investigated the roles of cytokines in expression levels of mRNAs and/or proteins in primary mouse keratinocytes and human keratinocyte HaCaT cells at earlier stages. We showed that IL-4 downregulated the expression levels of Krt1, Krt10, Dsg1, and Dsc1 via IL-4Rα- and signal transducer and activator of transcription factor 6 (STAT6)-dependent mechanisms in differentiating mouse keratinocytes at early stages. As the expression levels of keratin-1 and -10 in the keratinocytes transiently expressing an active form of STAT6 were not downregulated, STAT6 and other IL-4-induced molecules may synergistically regulate this expression. The restoration of the downregulated expression levels of Krt1 and Krt10 induced by IL-4 with the MEK (mitogen-activated protein kinase (MAPK)/extracellular signal-regulated kinase kinase) inhibitor U0126 indicated the involvement of the p44/42 MAPK signaling pathway in the attenuated expression. IL-13 also downregulated the expression of the four genes. Furthermore, IL-4 or IL-13 caused the downregulation of these genes in HaCaT cells and promoted the fragmentation of cell sheets with mechanical stress. Our results showed that IL-4 or IL-13 acted on differentiating keratinocytes in vitro at early stages to attenuate the gene expression.
Assuntos
Interleucina-13/metabolismo , Interleucina-3/metabolismo , Interleucina-4/metabolismo , Queratinócitos/fisiologia , Células Th2/fisiologia , Animais , Linhagem Celular , Dermatite Atópica/genética , Dermatite Atópica/metabolismo , Dermatite Atópica/patologia , Desmocolinas/genética , Desmocolinas/metabolismo , Desmogleína 1/genética , Desmogleína 1/metabolismo , Células Epidérmicas , Epiderme/fisiologia , Humanos , Queratina-1/genética , Queratina-1/metabolismo , Queratina-10/genética , Queratina-10/metabolismo , Queratinócitos/citologia , Queratinas Específicas do Cabelo/genética , Queratinas Específicas do Cabelo/metabolismo , Sistema de Sinalização das MAP Quinases/fisiologia , Glicoproteínas de Membrana/genética , Glicoproteínas de Membrana/metabolismo , Camundongos , Camundongos Endogâmicos BALB C , Camundongos Knockout , Cultura Primária de Células , Células Th2/citologiaRESUMO
This protocol describes methods to identify, purify and culture CD1d restricted invariant natural killer T (iNKT) cells from mouse tissue or human blood samples. The methods for identification and purification of iNKT cells are based on the interaction between iNKT cell receptor and its ligand. The iNKT cell receptor is composed of the invariant V alpha 14 J alpha 18/V beta 8.2 in mice or V alpha 24 J alpha 18/V beta 11 in humans and is expressed only on iNKT cells but not on conventional T cells. The iNKT cell antigen receptor in both species recognizes alpha-galactosylceramide (alpha-GalCer) presented by the MHC class I-like CD1d. Thus, alpha-GalCer-loaded CD1d dimer can be used for analysis and purification by fluorescence-activated cell sorting (FACS). Isolation of 1 x 10(6) purified iNKT cells from mouse thymus, spleen or liver requires 5-6 mice and takes 1-2 h for mononuclear cell preparation from mouse tissues, 1.5 h for enrichment by magnetic beads and 4 h for detection and purification of the iNKT cells by FACS. In the case of isolation of human peripheral blood mononuclear cells (PBMCs) from whole blood, it takes 2 h and requires 5 ml of blood to obtain 5 x 10(6) PBMCs, which contain 500-25,000 iNKT cells.