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BACKGROUND: Anti-programmed cell death-1(anti-PD1) treatment has shown promising antitumor efficacy in patients with advanced hepatocellular carcinoma (HCC). This study sought to explore the functional significance of programmed death ligand-1 (PD-L1) expression in tumor cells in the tumor microenvironment. METHODS: The mouse liver cancer cell line BNL-MEA was transfected with PD-L1 plasmids and stable clones expressing PD-L1 were selected. An orthotopic HCC model was generated by implanting the cells into the subcapsular space of BALB/c mice. Cell growth features were measured by proliferation assay, colony formation, flow cytometry (in vitro), ultrasonography, and animal survival (in vivo). The changes in T-cell function were examined by cytokine assay, expression of T-cell related genes, and flow cytometry. The efficacy of anti-PD1 therapy was compared between the parental and PD-L1-expressing tumors. RESULTS: PD-L1 expression did not affect growth characteristics of BNL-MEA cells but downregulated the expression of genes related to T-cell activation in the tumor microenvironment. Co-culture of PD-L1-expressing BNL-MEA cells with CD8+ T cells reduced T-cell proliferation and expression of cytokines IFNγ and TNFα. Tumors with PD-L1 expression showed better response to anti-PD1 therapy and depletion of CD8+ T cells abolished the antitumor effect. The difference in treatment response between parental and PD-L1-expressing tumors disappeared when a combination of anti-PD1 and sorafenib was given. CONCLUSIONS: PD-L1 expression in HCC cells may inhibit T-cell function in the liver tumor microenvironment. Anti-PD1 therapy appeared more effective in PD-L1-expressing than nonexpressing tumors, but the difference was diminished by the addition of sorafenib.
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Exosomes are nano-vesicles transporting bioactive material between cells. This study explored the prognostic association of exosomal TGF-ß1 with lymph node (LN) metastasis of gastric cancer (GC). TGF-ß1 expressions in the exosomes isolated from the gastroepiploic veins of 61 GC patients analyzed by ELISA. The regulatory T (Treg) cells in celiac LNs of gastric cancer analyzed by immunohistochemistry. Exosomal TGF-ß1 expression and the ratio of Treg cells in draining LNs were both significantly associated with pathological stages and LN metastasis of gastric cancer. Besides, the exosomal TGF-ß1 expression and Treg proportion in LN were also significantly correlated in gastric cancer patients. Recombinant TGF-ß1 and exosomes isolated from GC patients were used to induce FOXP3+ Treg cells from naïve T cells in vitro. Compared to the control, recombinant TGF-ß1 induced more CD25 (41%), FOXP3 (19%) and CTLA-4 (47%), while reduced CD45RA expression by 38% in primary naïve T cell cultures (p<0.01). Exosomes treatment induced more CD25 and 45% higher CTLA-4 expression, and increased 29% higher of CD45RA-negative cells than recombinant TGF-ß1 did (p<0.01). Adding TGF-ß1 neutralizing antibody partially abrogated the effects of exosomes on Treg induction. Our study showed exosomal TGF-ß1 related to lymph node metastasis and the ratio of Treg cells in lymph nodes of gastric cancers. Exosomes from gastric cancer patients could induce Treg cells formation through the effect of TGF-ß1.
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c-Maf belongs to the large Maf family of transcription factors and plays a key role in the regulation of cytokine production and differentiation of TH2, TH17, TFH, and Tr1 cells. Invariant natural killer T (iNKT) cells can rapidly produce large quantity of TH-related cytokines such as IFN-γ, IL-4, and IL-17A upon stimulation by glycolipid antigens, such as α-galactosylceramide (α-GalCer). However, the role of c-Maf in iNKT cells and iNKT cells-mediated diseases remains poorly understood. In this study, we demonstrate that α-GalCer-stimulated iNKT cells express c-Maf transcript and protein. By using c-Maf-deficient fetal liver cell-reconstituted mice, we further show that c-Maf-deficient iNKT cells produce less IL-17A than their wild-type counterparts after α-GalCer stimulation. While c-Maf deficiency does not affect the development and activation of iNKT cells, c-Maf is essential for the induction of IL-17-producing iNKT (iNKT17) cells by IL-6, TGF-ß, and IL-1ß, and the optimal expression of RORγt. Accordingly, c-Maf-deficient iNKT17 cells lose the ability to recruit neutrophils into the lungs. Taken together, c-Maf is a positive regulator for the expression of IL-17A and RORγt in iNKT17 cells. It is a potential therapeutic target in iNKT17 cell-mediated inflammatory disease.
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Galectin-3 is a ß-galactoside-binding animal lectin with diverse functions, including regulation of T helper (Th) 1 and Th2 responses. Current data indicate that galectin-3 expressed in dendritic cells (DCs) may be contributory. Th17 cells have emerged as critical inducers of tissue inflammation in autoimmune disease and important mediators of host defense against fungal pathogens, although little is known about galectin-3 involvement in Th17 development. We investigated the role of galectin-3 in the induction of Th17 immunity in galectin-3-deficient (gal3(-/-)) and gal3(+/+) mouse bone marrow-derived DCs. We demonstrate that intracellular galectin-3 negatively regulates Th17 polarization in response to the dectin-1 agonist curdlan (a ß-glucan present on the cell wall of fungal species) and lipopolysaccharide, agents that prime DCs for Th17 differentiation. On activation of dectin-1, gal3(-/-) DCs secreted higher levels of the Th17-axis cytokine IL-23 compared with gal3(+/+) DCs and contained higher levels of activated c-Rel, an NF-κB subunit that promotes IL-23 expression. Levels of active Raf-1, a kinase that participates in downstream inhibition of c-Rel binding to the IL23A promoter, were impaired in gal3(-/-) DCs. Modulation of Th17 by galectin-3 in DCs also occurred in vivo because adoptive transfer of gal3(-/-) DCs exposed to Candida albicans conferred higher Th17 responses and protection against fungal infection. We conclude that galectin-3 suppresses Th17 responses by regulating DC cytokine production.
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Citocinas/metabolismo , Células Dendríticas/metabolismo , Galectina 3/metabolismo , Células Th17/inmunología , Traslado Adoptivo , Animales , Candida albicans/inmunología , Candida albicans/fisiología , Candidiasis/inmunología , Candidiasis/microbiología , Candidiasis/patología , Polaridad Celular/efectos de los fármacos , Pollos , Células Dendríticas/efectos de los fármacos , Células Dendríticas/enzimología , Células Dendríticas/microbiología , Activación Enzimática/efectos de los fármacos , Quinasas MAP Reguladas por Señal Extracelular/metabolismo , Galectina 3/deficiencia , Inmunidad/efectos de los fármacos , Interleucina-23/biosíntesis , Lectinas Tipo C/agonistas , Lipopolisacáridos/farmacología , Ratones , Ratones Endogámicos C57BL , Modelos Biológicos , Proteínas Proto-Oncogénicas c-raf/metabolismo , Proteínas Proto-Oncogénicas c-rel/metabolismo , Transducción de Señal/efectos de los fármacos , Células Th17/efectos de los fármacos , beta-Glucanos/farmacologíaRESUMEN
Cytokine storm has been postulated as one of the major causes of mortality in patients with severe respiratory viral infections such as influenza. With the help of an influenza Ag- specific mouse experimental system, we report that CD4(+) T cells contribute effector cytokines leading to lung inflammation in acute influenza. Although virus can no longer be detected from tissues 14 d postinfection, virus-derived Ag continues to drive a CD4(+) T cell response after viral clearance. Ag-specific CD4(+) T cells proliferate and evolve into memory CD4(+) T cells efficiently, but the production of effector cytokines is seriously hampered during this phase. This decoupling of proliferation and effector cytokine production doesn't appear in conjunction with increased suppression by regulatory T cells or decreased induction of transcription factors. Rather, GATA-3 and ROR-γt levels are elevated when compared with cells that have effector cytokine production. T-bet dominance over GATA-3 and ROR-γt decreases with the disarmament of effector cytokine production. Importantly, upon reinfection, these decoupled cells produce elevated levels of IFN-γ and were effective in virus eradication. These results provide a mechanism through altered T-bet dominance to dampen the cytokine storm without impeding the generation of memory T cells in influenza virus infection.
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Linfocitos T CD4-Positivos/inmunología , Citocinas/biosíntesis , Interferón gamma/metabolismo , Memoria/fisiología , Infecciones por Orthomyxoviridae/inmunología , Infecciones por Orthomyxoviridae/metabolismo , Proteínas de Dominio T Box/antagonistas & inhibidores , Proteínas de Dominio T Box/biosíntesis , Animales , Linfocitos T CD4-Positivos/patología , Linfocitos T CD4-Positivos/virología , Embrión de Pollo , Citocinas/fisiología , Subtipo H1N1 del Virus de la Influenza A/patogenicidad , Ratones , Ratones Endogámicos C57BL , Ratones Transgénicos , Infecciones por Orthomyxoviridae/patología , Índice de Severidad de la Enfermedad , Proteínas de Dominio T Box/fisiologíaRESUMEN
Galectin-3 (gal3) is known for its immunoregulatory functions in infectious, autoimmune, and inflammatory diseases. However, little is known about its regulatory role in the host's IL-17A response to infection. Using a mouse model of histoplasmosis in which both Th1 and Th17 responses contribute to fungal clearance, we investigated how gal3 regulates IL-17A responses. Our study showed that Histoplasma infection induced gal3(-/-) dendritic cells to produce significantly higher levels of IL-23, TGF-ß1, and IL-1ß than did gal3(+/+) cells. Infected by the same inoculum of Histoplasma, gal3(-/-) mice had lower fungal burden and produced higher levels of IL-23/IL-17-axis cytokines and lower levels of IL-12 and IFN-γ. Additionally, there was an increase in Th17 cells and a reduction in Th1 cells in infected gal3(-/-) mice. In vitro Th1/Th17-skewing experiments excluded the intrinsic effect of gal3 on Th cell differentiation. Although neutrophils from both gal3(+/+) and gal3(-/-) mice produced IL-17A upon IL-23 stimulation, their contribution to IL-17A production was greater in gal3(-/-) mice than in gal3(+/+) mice. Compared with gal3(+/+) dendritic cells, adoptive transfer of gal3(-/-) dendritic cells resulted in production of significantly higher levels of IL-17-axis cytokines and reduced fungal burden. It appears that reduced fungal burden and preferential IL-17A response in gal3(-/-) mice by both Th17 cells and neutrophils were the result of preferential production of IL-23/IL-17-axis cytokines by dendritic cells. Our study showed that gal3 negatively regulates IL-17A responses through inhibition of IL-23/IL-17-axis cytokine production by dendritic cells.
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Citocinas/biosíntesis , Células Dendríticas/inmunología , Células Dendríticas/metabolismo , Galectina 3/metabolismo , Histoplasma/inmunología , Histoplasmosis/inmunología , Histoplasmosis/metabolismo , Animales , Diferenciación Celular/inmunología , Galectina 3/genética , Histoplasmosis/genética , Interacciones Huésped-Patógeno/inmunología , Interleucina-17/biosíntesis , Interleucina-17/farmacología , Interleucina-23/biosíntesis , Interleucina-23/farmacología , Activación de Macrófagos/inmunología , Macrófagos/inmunología , Macrófagos/microbiología , Ratones , Ratones Noqueados , Neutrófilos/inmunología , Neutrófilos/metabolismo , Células Th17/citología , Células Th17/inmunologíaRESUMEN
Exposure to protein allergen epicutaneously, inducing a Th2-dominant immune response, sensitizes the host to the development of atopic disease. Antigen-driven bystander effect demonstrates that polarized T cells could instruct naïve T cells to differentiate into T cells with similar phenotype. In this study, we aimed to determine the contribution of antigen-driven bystander effect on epicutaneous sensitization with a newly introduced protein allergen. BALB/c mice were immunized intraperitoneally with BSA emulsified in alum, known to induce a Th2 response, three weeks before given BSA and OVA epicutaneously. Lymph node cells from these mice restimulated with OVA secreted higher levels IL-4, IL-5 and IL-13 as compared with cells from mice without BSA immunization. In addition, BALB/c mice immunized subcutaneously with BSA emulsified in complete Freund's adjuvant, known to induce a Th1-predominant response, also induced higher Th1 as well as Th2 cytokine response when restimulated with OVA as compared with mice without immunization. We demonstrated that subcutaneous immunization with BSA in CFA induced Th2 as well as Th1 response. The threshold of epicutaneous sensitization to OVA was also reduced, possibly due to increased expressions of IL-4 and IL-10 in the draining lymph nodes during the early phase of sensitization. In conclusion, antigen-driven bystander effect, whether it is of Th1- or Th2-predominant nature, can accelerate epicutaneous sensitization by a newly introduced protein allergen. These results provide a possible explanation for mono- to poly-sensitization spread commonly observed in atopic children.