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1.
Exp Parasitol ; 207: 107789, 2019 Dec.
Artigo em Inglês | MEDLINE | ID: mdl-31669169

RESUMO

American visceral leishmaniasis is caused by the protozoan Leishmania infantum. The control of the disease depends on the magnitude of the Th1 cell response and IL-10 producing regulatory T cells. Administration of chemokine, such as CXCL10, has shown promising results in the leishmaniasis treatment. Previous studies from our group have shown that CXCL10 induces a reduction in parasite burden in the spleen and a decrease in IL-10 and TGF-ß production in L. infantum-infected BALB/c mice. This work investigated whether CXCL10-treatment reduces IL-10 + Treg cell populations (CD4+CD25+Foxp3+ and Tr1) and induces morphological changes in the spleen. BALB/c mice were infected and treated or not with CXCL10 on the 1st, 3rd and 7th days of infection. CXCL10-treatment was able to reduce the parasite load in the spleen in L. infantum-infected BALB/c mice and this decrease in the number of parasites correlated with the decrease in size of this organ in treated animals compared to untreated animals. 7, 23, and 45 days post-treatment (p.t.), the phenotype and frequency of IL-10 + Treg cells were evaluated by flow cytometry, and the morphological changes of the spleen were analyzed by optical microscopy. After 7 and 23 days p.t., CXCL10-treated animals showed a significant reduction of CD25-Foxp3-IL-10+ (Tr1) cells in the spleen when compared to untreated animals, whereas CD4+CD25+Foxp3+IL-10+ Treg cells reduced later at 23rd and 45th days p.t. Furthermore, while untreated animals showed a significant positive correlation between IL-10 production and Tr1 cells, in CXCL10-treated group this correlation was negative. Thus, these findings show that treatment with CXCL10 chemokine in L. infantum-infected BALB/c mice results in suppression of IL10+ Treg (Foxp3+ and Tr1) cells in the spleen, associated with a reduction in parasite load and splenomegaly.


Assuntos
Quimiocina CXCL10/uso terapêutico , Leishmaniose Visceral/tratamento farmacológico , Leishmaniose Visceral/imunologia , Baço/imunologia , Linfócitos T Reguladores/efeitos dos fármacos , Adjuvantes Imunológicos/uso terapêutico , Animais , Quimiocina CXCL10/administração & dosagem , Quimiocina CXCL10/farmacologia , Cricetinae , Citometria de Fluxo , Fatores de Transcrição Forkhead/imunologia , Humanos , Injeções Intraperitoneais , Leishmania infantum/efeitos dos fármacos , Leishmania infantum/imunologia , Leishmania infantum/patogenicidade , Masculino , Mesocricetus , Camundongos , Camundongos Endogâmicos BALB C , Tamanho do Órgão/efeitos dos fármacos , Tamanho do Órgão/imunologia , Carga Parasitária , Baço/parasitologia , Baço/patologia , Linfócitos T Reguladores/imunologia , Virulência
2.
Mol Immunol ; 116: 90-97, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31630080

RESUMO

BACKGROUND: A T cell subtype with the CD4+CD25-FoxP3+ phenotype was recently described. We aimed to investigate the frequency of these cells and their ability to produce cytokines in tumor-draining lymph nodes from patients with breast cancer (BC). MATERIALS AND METHODS: Mononuclear cells from lymph nodes of 20 patients with BC were activated and stained for appropriate markers. The cells were assayed with four-color flow cytometry. RESULTS: A very small fraction of CD4+CD25-FoxP3+ cells produced cytokines at levels that were significantly lower than in the regulatory (CD4+CD25+FoxP3+) and effector cell (CD4+CD25+FoxP3-) subpopulations. The expression of IFNγ and IL-2 in the CD4+CD25-FoxP3+ subset was significantly higher than in Treg cells, but lower than in the effector subset. Conversely, IL-22 expression in Treg cells was significantly higher than in the CD4+CD25-FoxP3+ subpopulation. The expression of IL-10 in the CD4+CD25-FoxP3+ subset was also significantly higher than in effector cells. CONCLUSION: We suggest that CD4+CD25-FoxP3+ cells in patients with BC are exhausted cells with an intermediate phenotype between effector and regulatory cells.


Assuntos
Neoplasias da Mama/imunologia , Linfócitos T CD4-Positivos/imunologia , Citocinas/imunologia , Fatores de Transcrição Forkhead/imunologia , Subunidade alfa de Receptor de Interleucina-2/imunologia , Linfonodos/imunologia , Adulto , Idoso , Feminino , Citometria de Fluxo/métodos , Humanos , Interferon gama/imunologia , Interleucina-10/imunologia , Interleucinas/imunologia , Pessoa de Meia-Idade , Linfócitos T Reguladores/imunologia
3.
Immunology ; 158(3): 161-170, 2019 11.
Artigo em Inglês | MEDLINE | ID: mdl-31517385

RESUMO

Regulatory T (Treg) cells are a subset of CD4+ T cells that are critical for the maintenance of self-tolerance. The forkhead box transcription factor Foxp3 is a master regulator for the Treg phenotype and function and its expression is essential in Treg cells, as the loss of Foxp3 results in lethal autoimmunity. Two major subsets of Treg cells have been described in vivo; thymus-derived Treg (tTreg) cells that develop in the thymus and peripherally induced Treg (pTreg) cells that are derived from conventional CD4+  Foxp3- T cells and are converted in peripheral tissues to cells that express Foxp3 and acquire suppressive ability. The transcription factor Helios, a member of the Ikaros transcription factor family, is expressed in 60-70% of Treg cells in both mouse and man, and is believed to be a marker of tTreg cells. In this review, we discuss the role and function of Helios in Treg cells, the controversy surrounding the use of Helios as a marker of tTreg cells, and how Helios controls specific aspects of the Treg cell program.


Assuntos
Antígenos de Diferenciação/imunologia , Proteínas de Ligação a DNA/imunologia , Fatores de Transcrição Forkhead/imunologia , Fator de Transcrição Ikaros/imunologia , Linfócitos T Reguladores/imunologia , Fatores de Transcrição/imunologia , Animais , Humanos , Camundongos
4.
Gastroenterology ; 157(6): 1584-1598, 2019 12.
Artigo em Inglês | MEDLINE | ID: mdl-31513797

RESUMO

BACKGROUND & AIMS: T-regulatory (Treg) cells suppress the immune response to maintain homeostasis. There are 2 main subsets of Treg cells: FOXP3 (forkhead box protein 3)-positive Treg cells, which do not produce high levels of effector cytokines, and type 1 Treg (Tr1) cells, which are FOXP3-negative and secrete interleukin (IL) 10. IL10 is an anti-inflammatory cytokine, so Tr1 cells might be used in the treatment of inflammatory bowel diseases. We aimed to develop methods to isolate and expand human Tr1 cells and define their functions. METHODS: We obtained blood and colon biopsy samples from patients with Crohn's disease or ulcerative colitis or healthy individuals (controls). CD4+ T cells were isolated from blood samples and stimulated with anti-CD3 and anti-CD28 beads, and Tr1 cells were purified by using an IL10 cytokine-capture assay and cell sorting. FOXP3-positive Treg cells were sorted as CD4+CD25highCD127low cells from unstimulated cells. Tr1 and FOXP3-positive Treg cells were expanded, and phenotypes and gene expression profiles were compared. T cells in peripheral blood mononuclear cells from healthy donors were stimulated with anti-CD3 and anti-CD28 beads, and the suppressive abilities of Tr1 and FOXP3-positive Treg cells were measured. Human colon organoid cultures were established, cultured with supernatants from Tr1 or FOXP3-positive cells, and analyzed by immunofluorescence and flow cytometry. T84 cells (human colon adenocarcinoma epithelial cells) were incubated with supernatants from Tr1 or FOXP3-positive cells, and transepithelial electrical resistance was measured to determine epithelial cell barrier function. RESULTS: Phenotypes of Tr1 cells isolated from control individuals vs patients with Crohn's disease or ulcerative colitis did not differ significantly after expansion. Tr1 cells and FOXP3-positive Treg cells suppressed proliferation of effector T cells, but only Tr1 cells suppressed secretion of IL1B and tumor necrosis factor from myeloid cells. Tr1 cells, but not FOXP3-positive Treg cells, isolated from healthy individuals and patients with Crohn's disease or ulcerative colitis secreted IL22, which promoted barrier function of human intestinal epithelial cells. Tr1 cell culture supernatants promoted differentiation of mucin-producing goblet cells in intestinal organoid cultures. CONCLUSIONS: Human Tr1 cells suppress proliferation of effector T cells (adaptive immune response) and production of IL1B and TNF by myeloid cells (inmate immune response). They also secrete IL22 to promote barrier function. They might be developed as a cell-based therapy for intestinal inflammatory disorders.


Assuntos
Colite Ulcerativa/imunologia , Doença de Crohn/imunologia , Interleucina-10/metabolismo , Mucosa Intestinal/patologia , Linfócitos T Reguladores/imunologia , Adulto , Idoso , Biópsia , Comunicação Celular/imunologia , Proliferação de Células , Células Cultivadas , Colite Ulcerativa/sangue , Colite Ulcerativa/terapia , Colo/citologia , Colo/imunologia , Colo/patologia , Doença de Crohn/sangue , Doença de Crohn/terapia , Feminino , Fatores de Transcrição Forkhead/imunologia , Fatores de Transcrição Forkhead/metabolismo , Voluntários Saudáveis , Humanos , Interleucina-10/imunologia , Interleucinas/imunologia , Interleucinas/metabolismo , Mucosa Intestinal/citologia , Mucosa Intestinal/imunologia , Masculino , Pessoa de Meia-Idade , Cultura Primária de Células , Linfócitos T Citotóxicos/imunologia , Linfócitos T Reguladores/metabolismo , Linfócitos T Reguladores/transplante
5.
Toxicol Lett ; 316: 27-34, 2019 Nov.
Artigo em Inglês | MEDLINE | ID: mdl-31513887

RESUMO

OBJECTIVE: Atherosclerosis is an autoimmune inflammatory disease that is closely associated with long-term exposure to fine particulate matter (PM2.5). CD4+CD25+Foxp3+ regulatory T cells (Tregs) play a critical role in the regulation of T cell-mediated immune responses, and the depletion of CD4+CD25+Foxp3+ Tregs has been thought to play a prominent role in atherosclerosis. Therefore, we investigated the association between the CD4+CD25+Foxp3+ Tregs population and atherosclerotic development in ApoE-/- mice exposed to PM2.5. METHODS: We employed a real-world system to subject 40 ApoE-/- mice to ambient inhalation of PM2.5 (PM2.5 group, n = 20) or filtered air (FA group, n = 20) for 12 weeks. PM2.5 source apportionment, atherosclerotic lesions within aorta, lipid deposition and plaque accumulation in whole artery, serum level of inflammatory factors and lipid profiles, CD4+CD25+Foxp3+ Tregs population in splenocytes, Foxp3 protein and mRNA expressions in descending aorta and spleen were quantified, respectively. RESULTS: The daily average concentration of PM2.5 was 57.4 ± 25.6 µg/m3. Atherosclerotic lesions within aorta, lipid deposition and plaque accumulation in whole artery, serum levels of IL-6, TNF-α, TC and LDL-C in the PM2.5 group increased significantly compared to the FA group. Whereas, serum levels of IL-10 and TGF-ß, CD4+CD25+Foxp3+ Tregs population in splenocytes, Foxp3 protein and mRNA expressions in descending aorta and spleen in the PM2.5 group decreased significantly compared to the FA group. CONCLUSION: These results suggest that PM2.5 could accelerate the development of atherosclerosis in ApoE-/- mice, which is related to CD4+CD25+Foxp3+ Tregs down-regulation, as well as lipid deposition and systemic inflammation.


Assuntos
Aorta/efeitos dos fármacos , Doenças da Aorta/induzido quimicamente , Aterosclerose/induzido quimicamente , Fatores de Transcrição Forkhead/metabolismo , Subunidade alfa de Receptor de Interleucina-2/metabolismo , Material Particulado/toxicidade , Linfócitos T Reguladores/efeitos dos fármacos , Animais , Aorta/imunologia , Aorta/metabolismo , Aorta/patologia , Doenças da Aorta/sangue , Doenças da Aorta/imunologia , Doenças da Aorta/patologia , Aterosclerose/sangue , Aterosclerose/imunologia , Aterosclerose/patologia , Biomarcadores/sangue , LDL-Colesterol/sangue , Citocinas/sangue , Modelos Animais de Doenças , Progressão da Doença , Fatores de Transcrição Forkhead/imunologia , Predisposição Genética para Doença , Mediadores da Inflamação/sangue , Subunidade alfa de Receptor de Interleucina-2/imunologia , Masculino , Camundongos Endogâmicos C57BL , Camundongos Knockout para ApoE , Tamanho da Partícula , Fenótipo , Placa Aterosclerótica , Linfócitos T Reguladores/imunologia , Linfócitos T Reguladores/metabolismo , Linfócitos T Reguladores/patologia , Fatores de Tempo
6.
Mol Immunol ; 114: 323-329, 2019 10.
Artigo em Inglês | MEDLINE | ID: mdl-31442916

RESUMO

Ulcerative colitis (UC) is a chronic relapsing inflammatory disease that occurs in the gastrointestinal tract, characterized by an upregulation in autoantibody production and antimicrobial antibody production. The interaction between follicular helper T cells (Tfh) and follicular regulatory T cells (Tfr) is critical to the induction and regulation of antibody production. In this study, we investigated the characteristics of Tfr cells in UC patients. We gated circulating Tfr cells as CD4+CXCR5+CD25+CD127- T cells, of which approximately 73% on average were Foxp3+. The circulating Tfh (CD4+CXCR5+CD25-) cells from control subjects and UC patients presented a comparable capacity to induce IgM production from naive B cells and to mediate class switching to IgG. Tfr cells significantly reduced Tfh-mediated B cell help in both healthy controls and UC patients in a concentration-dependent manner. However, the suppression capacity of Tfr cells was significantly lower in UC patients than in healthy controls. Subsequently, we found that the frequency of CTLA-4-expressing cells was only slightly lower in UC patients, but the MFI of CTLA-4, however, was markedly lower in UC patients. CTLA-4 blockade nearly abrogated Tfr-mediated suppression of IgM, and significantly reduced Tfr-mediated suppression of IgG. Moreover, CTLA-4 blockade removed the relative advantage of Tfr suppression capacity in healthy controls compared to UC patients. Overall, this study demonstrated that CTLA-4 was required for Tfr-mediated suppression of B cell help, but was expressed at lower levels in UC patients.


Assuntos
Antígeno CTLA-4/imunologia , Colite Ulcerativa/imunologia , Linfócitos T Reguladores/imunologia , Adulto , Formação de Anticorpos/imunologia , Linfócitos B/imunologia , Linfócitos T CD4-Positivos/imunologia , Feminino , Fatores de Transcrição Forkhead/imunologia , Humanos , Imunoglobulina M/imunologia , Subunidade alfa de Receptor de Interleucina-2/imunologia , Subunidade alfa de Receptor de Interleucina-7/imunologia , Masculino , Pessoa de Meia-Idade , Receptores CXCR5/imunologia , Linfócitos T Auxiliares-Indutores/imunologia
7.
Immunopharmacol Immunotoxicol ; 41(4): 463-468, 2019 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-31339393

RESUMO

Context: CD4+ T lymphocytes are able to differentiate into distinct subtypes according to several immunological scenarios, including T helper (Th)1, Th2, Th17 and regulatory T (Treg) cells. CD4+ T cells are phenotypically flexible and have specific ion channels, such as the nicotinic acetylcholine receptors (nAChR) that could be modulated by peptides produced by marine snails, known as conotoxins. Their effect on T lymphocytes has not been explored and emerging evidence suggests that these peptides may have immunomodulatory activities. Objective: This study investigated the effect of two Californiconus californicus-derived synthetic conotoxins on the proliferation and differentiation of T lymphocyte subpopulations Th1, Th2, Th17 and Treg. Methods: Cells from lymph nodes of BALB/c mice were cultured in the presence of conotoxins cal14.1b and cal14.2c (5.5 µM), during 96 h. Cell proliferation and intracellular cytokine production (IFN-γ, IL-4, IL-17 and IL-10) were analyzed by flow cytometry. Results and Discussion: cal14.1b and cal14.2c increased intracellular IL-10 production in Treg (CD3+CD4+Foxp3+) cells and decreased intracellular IL-17 production (CD3+CD4+) after 72 h of culture. Conotoxins did not show any effect on T cell proliferation nor Th1/Th2 balance. Conclusion: These results suggest that synthetic conotoxins exert immunomodulatory activity, especially by regulating specific functions on T lymphocytes.


Assuntos
Linfócitos T CD4-Positivos/imunologia , Conotoxinas/imunologia , Fatores de Transcrição Forkhead/imunologia , Fatores Imunológicos/imunologia , Interleucina-10/imunologia , Peptídeos/imunologia , Linfócitos T Reguladores/imunologia , Animais , Organismos Aquáticos/imunologia , Diferenciação Celular/imunologia , Proliferação de Células/efeitos dos fármacos , Células Cultivadas , Citocinas/imunologia , Ativação Linfocitária/imunologia , Masculino , Camundongos , Camundongos Endogâmicos BALB C
8.
Pediatr Hematol Oncol ; 36(4): 198-210, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-31287345

RESUMO

The pathogenesis of aplastic anemia (AA) in children is not clear. This study was conducted to investigate the changes in the proportion and function of regulatory T cells (Tregs) in pediatric AA. The proportion of Tregs, mRNA levels of transcription factors, and concentrations of cytokines were measured by flow cytometry, reverse transcription-PCR, and enzyme-linked immunosorbent assay, respectively. Tregs were co-cultured with effector T cells (Teff) to evaluate the function of Tregs. The proportion of Tregs after immunosuppressive therapy (IST) in pediatric AA was monitored dynamically. Compared to the control, the proportions of Tregs in peripheral blood and bone marrow lymphocytes of the untreated AA group were lower (1.31% ± 0.73% vs. 3.16% ± 0.92%, 1.49% ± 0.81% vs. 3.06% ± 0.82%, respectively, p < 0.001). The mRNA levels of FOXP3 and STAT3 in the AA group were lower (p = 0.014; p < 0.001). However, the mRNA levels of T-BET did not significantly differ between groups. The concentration of interferon-γ and interleukin-17 in the AA group were higher (p = 0.004; p = 0.003), whereas the concentration of TGF-ß decreased (p = 0.044). The immunosuppressive function of Tregs was impaired in the AA group. After IST, the proportion of Tregs was significantly lower than that in the control. The proportion of Tregs at the time of diagnosis in the nonresponsive group was lower than that in the responsive group, but the difference was not significant. Treg levels were significantly decreased and were functionally impaired at the time of diagnosis of pediatric AA. However, there was no significant change in Tregs at the resolution of AA.


Assuntos
Anemia Aplástica/imunologia , Células da Medula Óssea/imunologia , Linfócitos T Reguladores/imunologia , Anemia Aplástica/patologia , Células da Medula Óssea/patologia , Criança , Pré-Escolar , Feminino , Fatores de Transcrição Forkhead/imunologia , Humanos , Masculino , Proteínas de Neoplasias/imunologia , RNA Mensageiro/imunologia , Fator de Transcrição STAT3/imunologia , Proteínas com Domínio T/imunologia , Linfócitos T Reguladores/patologia
9.
Medicine (Baltimore) ; 98(30): e16345, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31348233

RESUMO

To evaluate the potential role of Pten and CD4FOXP3 T cells in prognosis from endometrial cancer.Tissue samples and clinical data were collected from 200 patients with endometrial cancer and 100 control patients with benign uterine diseases. The expressions of Pten and CD4FOXP3 T cells were quantified by immunohistochemistry and immunofluorescence. After surgery, all patients were followed up for an average of 56.3 months. Surgical effects were evaluated based on the patients' symptoms and signs. A two-sided P value < .05 was considered significant.Pten diminished and CD4FOXP3 T cells significantly accumulated with the progression of endometial cancer, in comparison to the controls. Moreover, Pten expression was negatively correlated with the count of CD4FOXP3 T cells. Pten and CD4FOXP3 T cells were correlated with clinical characteristics, including tumor stage, differentiation and associated with patients' disease-free survival.Limited data were available between the expressions of Pten and CD4FOXP3 T cells in patients with endometrial cancer. Our study findings suggested that the expressions of Pten and CD4FOXP3 T cells might become possible biomarkers for the diagnosis and prediction in endometrial cancer.


Assuntos
Neoplasias do Endométrio/patologia , Fatores de Transcrição Forkhead/imunologia , PTEN Fosfo-Hidrolase/biossíntese , Linfócitos T/imunologia , Adulto , Idoso , Biomarcadores Tumorais , Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD4-Positivos/metabolismo , Estudos de Casos e Controles , Neoplasias do Endométrio/genética , Feminino , Fatores de Transcrição Forkhead/biossíntese , Humanos , Imuno-Histoquímica , Pessoa de Meia-Idade , PTEN Fosfo-Hidrolase/genética , PTEN Fosfo-Hidrolase/imunologia , Prognóstico , Linfócitos T/metabolismo
10.
Nat Protoc ; 14(7): 1946-1969, 2019 07.
Artigo em Inglês | MEDLINE | ID: mdl-31160786

RESUMO

The interrogation of single cells is revolutionizing biology, especially our understanding of the immune system. Flow cytometry is still one of the most versatile and high-throughput approaches for single-cell analysis, and its capability has been recently extended to detect up to 28 colors, thus approaching the utility of cytometry by time of flight (CyTOF). However, flow cytometry suffers from autofluorescence and spreading error (SE) generated by errors in the measurement of photons mainly at red and far-red wavelengths, which limit barcoding and the detection of dim markers. Consequently, development of 28-color fluorescent antibody panels for flow cytometry is laborious and time consuming. Here, we describe the steps that are required to successfully achieve 28-color measurement capability. To do this, we provide a reference map of the fluorescence spreading errors in the 28-color space to simplify panel design and predict the success of fluorescent antibody combinations. Finally, we provide detailed instructions for the computational analysis of such complex data by existing, popular algorithms (PhenoGraph and FlowSOM). We exemplify our approach by designing a high-dimensional panel to characterize the immune system, but we anticipate that our approach can be used to design any high-dimensional flow cytometry panel of choice. The full protocol takes a few days to complete, depending on the time spent on panel design and data analysis.


Assuntos
Anticorpos Monoclonais , Citometria de Fluxo/métodos , Imunofluorescência/métodos , Análise de Célula Única/métodos , Algoritmos , Biomarcadores , Tampões (Química) , Corantes , Biologia Computacional/métodos , Fatores de Transcrição Forkhead/imunologia , Humanos , Família de Moléculas de Sinalização da Ativação Linfocitária/imunologia , Linfócitos T/citologia , Linfócitos T/imunologia
11.
Int Immunopharmacol ; 74: 105675, 2019 Sep.
Artigo em Inglês | MEDLINE | ID: mdl-31177017

RESUMO

Rheumatoid arthritis (RA) is an inflammatory autoimmune disorder. Autoreactive T cells play a very significant role in the pathogenesis of RA. However, the exact mechanisms of disease severity and pathogenesis are poorly understood. We attempted to correlate T-helper cell activities with sexes of newly diagnosed patients with RA. The patients were divided based on their sex and disease severity. Examination of the expression of various factors using quantitative real-time PCR and FACS analysis of peripheral blood mononuclear cells revealed that T-bet, ROR-γt, Foxp3, and the level of cytokines associated with Th1 cells were almost identical among male and female patients with RA. Interestingly, there was a high correlation between Th17 expression and disease severity in female patients with RA. In general, there was no significant correlation between Th1 cell population and the disease severity in newly diagnosed patients with RA. In contrast, the frequency of both Th17 and Treg cells was higher in patients with more severe disease. The results suggested that, in patients with RA, the T-helper cell balance within peripheral blood was skewed towards the Th17 and Treg phenotypes. Besides Th17- and Treg-associated cytokines, elevated expression of IL-27/IL-23 cytokines might also be responsible for increased disease severity in female patients with RA.


Assuntos
Artrite Reumatoide/imunologia , Adulto , Citocinas/imunologia , Feminino , Fatores de Transcrição Forkhead/imunologia , Humanos , Masculino , Pessoa de Meia-Idade , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/imunologia , Caracteres Sexuais , Proteínas com Domínio T/imunologia , Linfócitos T Reguladores/imunologia , Células Th1/imunologia , Células Th17/imunologia
12.
Medicine (Baltimore) ; 98(22): e15722, 2019 May.
Artigo em Inglês | MEDLINE | ID: mdl-31145286

RESUMO

BACKGROUND: Studies have shown that CD4CD25Foxp3Treg cells suppress NKG2D expression on NK cells via a cell contact-dependent mechanism and increased TGF-ß and IL-10 production in some cancer models. We herein aimed to explore whether CD4CD25Foxp3Tregs suppress NKG2D-mediated NK cell cytotoxicity in peripheral blood and elucidate the exact mechanism underlying this phenomenon. METHODS: To explore the function of NKG2D, NK cell cultures were treated with an NKG2D-blocking antibody to block these receptors. Additionally, TGF-ß- and IL-10-blocking antibodies were added to NK and CD4CD25Foxp3Treg cell cocultures to evaluate whether the latter cells suppress NKG2D expression of NK cells via increasing the production of TGF-ß and IL-10. The expression of NKG2D on NK cells was detected by 3-color flow cytometry, and NK cell activity was assessed by 3 assays: a nonradioactive cytotoxicity assay, an ELISA measuring IFN-γ production and a flow cytometry assay to evaluate CD107a expression. RESULTS: Blocking NKG2D decreased NK cell cytotoxicity, IFN-γ production and CD107a expression. Moreover, blocking TGF-ß and IL-10 substantially increased the NKG2D expression in NK and CD4CD25Foxp3Treg cell cocultures. Similarly, blocking TGF-ß and IL-10 enhanced NK cell cytotoxicity, IFN-γ production and CD107a expression; Transwell insert assays also revealed increased IFN-γ production and CD107a and NKG2D expression. CONCLUSION: CD4CD25Foxp3Tregs suppress NKG2D-mediated NK cell cytotoxicity in peripheral blood via a cell contact-dependent mechanism and increased TGF-ß and IL-10 production.


Assuntos
Fatores de Transcrição Forkhead/imunologia , Subunidade alfa de Receptor de Interleucina-2/imunologia , Células Matadoras Naturais/imunologia , Subfamília K de Receptores Semelhantes a Lectina de Células NK/fisiologia , Linfócitos T Reguladores/imunologia , Animais , Citotoxicidade Imunológica , Humanos , Ativação Linfocitária , Camundongos
13.
Immunopharmacol Immunotoxicol ; 41(2): 267-276, 2019 Apr.
Artigo em Inglês | MEDLINE | ID: mdl-31056985

RESUMO

Context: Mentha longifolia (L.) Huds., has shown anti-inflammatory effects. Objective: To evaluate the immunomodulatory effects of menthol, the major constituent of Mentha longifolia on T cells as the main cells affecting the inflammatory responses. Methods: Effect of menthol on: proliferation and viability of the peripheral blood human mononuclear cells (PBMCs) by BrdU and propidium iodide (PI) staining, respectively, interferone (IFN)γ and interleukin (IL)-4 cytokine production in lymphocytes stimulated with phytohemagglutinin (PHA) and phorbol myristate acetate/calcium ionophore (PMA/CI) by ELISA; intracellular staining of CD4+ cells for IFNγ expression by flow cytometry and gene expressions of T heper (Th) cell transcription factors was measured using real time-PCR. Results: Menthol dose-dependently inhibited lymphocytes proliferation from 88.7% at 50 µg/ml to 3.63% at 800 µg/ml (p < .05). According to the results of PI staining, this inhibitory effect was not due to cell death. Menthol dose-dependently decreased IFNγ but not IL-4 production in culture of PHA- and PMA/CI-stimulated lymphocytes to more than 80% at 800 µg/ml. In flow cytometry analysis, menthol reduced the number of IFN-γ-expressing CD4+T cells stimulated either with PHA or PMA/CI. Treatment of PBMCs with 800 µg/ml of menthol decreased levels of T-bet from 14.5 ± 2.26 fold in untreated control to 2.76 ± 1.74 fold (p < .001). Foxp-3 expression decreased to nearly half, but GATA3 did not significantly change. Ratios of T-bet to GATA3 and T-bet to Foxp3 gene expressions were dose-dependently declined. Conclusion: Decreased IFNγ expression plus T-bet down-regulation suggested the inhibitory effect of menthol on Th1 cells differentiation and hence imply its possible therapeutic usefulness in inflammatory diseases.


Assuntos
Regulação para Baixo/efeitos dos fármacos , Interferon gama/imunologia , Mentol/farmacologia , Proteínas com Domínio T/imunologia , Células Th1/imunologia , Adulto , Regulação para Baixo/imunologia , Fatores de Transcrição Forkhead/imunologia , Fator de Transcrição GATA3/imunologia , Humanos , Inflamação/tratamento farmacológico , Inflamação/imunologia , Inflamação/patologia , Masculino , Células Th1/patologia
14.
Mol Immunol ; 112: 51-58, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31078116

RESUMO

Particulate matter (PM)2.5 is a common air pollutant known to induce damages in the respiratory, cardiovascular, and nervous systems. Previous study has shown that acute and high-level PM insult could significantly aggravate the severity of LPS-induced acute lung injury (ALI). However, humans typically experience more chronic and low-level PM, of which the effect on ALI is yet unclear. Here, we varied the concentration of PM from low, medium, to high, which was given to mice via intratracheal instillation for a short period of time. Compared to the saline-treated mice, mice with medium or high PM treatment presented significantly higher mortality rate, weight reduction, and bronchoalveolar lavage (BAL) protein concentration during ALI, while mice with low PM treatment did not demonstrate significant differences from saline-treated mice. However, when the PM was given for an elongated period of time, PM, even at the low level, significantly aggravated ALI severity. Furthermore, the PM-mediated changes were sustained even after PM withdrawal. We also examined the CD4 T cells in saline- or PM-treated mice. We found that, although PM did not significantly change the number of lung-infiltrating CD4 T cells, it significantly altered the composition of lung-infiltrating CD4 T cells, characterized by having a higher T-bet/Foxp3 ratio in the PM-treated group compared to the saline-treated group. Additionally, the Treg-mediated suppression was reduced in PM-treated mice. The effect of PM on CD4 T cells depended on the concentration of PM and the duration of the treatment, and was independent of the PM withdrawal. Overall, these results demonstrated that chronic and low-level PM was sufficient at aggravating ALI and altering pulmonary CD4 T cells, and the effect could be sustained even after PM withdrawal.


Assuntos
Lesão Pulmonar Aguda/imunologia , Linfócitos T CD4-Positivos/imunologia , Pulmão/imunologia , Material Particulado/imunologia , Animais , Líquido da Lavagem Broncoalveolar/imunologia , Modelos Animais de Doenças , Fatores de Transcrição Forkhead/imunologia , Masculino , Camundongos , Camundongos Endogâmicos C57BL , Proteínas com Domínio T/imunologia , Linfócitos T Reguladores/imunologia
15.
Immunology ; 157(4): 304-311, 2019 08.
Artigo em Inglês | MEDLINE | ID: mdl-31141166

RESUMO

Pulmonary hypertension (PH) is a common but dangerous complication in chronic obstructive pulmonary disease (COPD). We hypothesized that dysregulation in the T helper type 17 (Th17) compartment could contribute to the development of COPD-associated PH (COPD-PH). To investigate this hypothesis, patients with COPD-PH and age- and sex-matched healthy controls were recruited, and their circulating CD4+ T cells were activated using anti-CD3/CD28 antibodies. The frequency of interleukin-17 (IL-17) -secreting cells was significantly higher in COPD-PH patients than in healthy controls. The secretion of IL-17 was significantly higher from COPD-PH CD4+ T cells than from control CD4+ T cells, whereas the secretion of interferon-γ and IL-4 was not significantly different. The expression of transforming growth factor-ß, on the other hand, was significantly higher in healthy controls than in COPD-PH patients. Activated CD4+ T cells from COPD-PH patients also presented significantly lower forkhead box P3 (FOXP3) and higher retinoic acid receptor-related orphan C2 (RORC2) expression than CD4+ T cells from healthy controls. In both controls and patients, a negative correlation between RORC2 and FOXP3 was found, ex vivo and after CD3/CD28 activation. The serum IL-6 level was slightly higher in COPD-PH patients than in controls, but the IL-6 transcription by monocytes was comparable in COPD-PH patients and controls. Interestingly, CD4+ T cells from COPD-PH patients presented significantly higher levels of Kirsten rat sarcoma viral oncogene homolog and neuroblastoma RAS viral oncogene homolog than CD4+ T cells from healthy controls. Inhibiting Ras-GTPases using farnesylthiosalicylic acid significantly reduced the ratio of RORC2/FOXP3 expression in CD4+ T cells. Overall, we demonstrated that an imbalance of Th17/regulatory T cells was a hallmark of COPD-PH.


Assuntos
Hipertensão Pulmonar/imunologia , Doença Pulmonar Obstrutiva Crônica/imunologia , Proteínas ras/imunologia , Idoso , Feminino , Fatores de Transcrição Forkhead/imunologia , Regulação da Expressão Gênica/imunologia , Humanos , Hipertensão Pulmonar/etiologia , Interferon gama/imunologia , Interleucina-4/imunologia , Masculino , Pessoa de Meia-Idade , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/imunologia , Doença Pulmonar Obstrutiva Crônica/complicações , Células Th17 , Fator de Crescimento Transformador beta
16.
Prostate ; 79(9): 969-979, 2019 06.
Artigo em Inglês | MEDLINE | ID: mdl-30999388

RESUMO

BACKGROUND: Accumulating evidence shows that tumor cell-specific genomic changes can influence the cross talk between cancer cells and the surrounding tumor microenvironment (TME). Loss of the PTEN tumor suppressor gene is observed in 20% to 30% of prostate cancers (PCa) when first detected and the rate increases with PCa progression and advanced disease. Recent findings implicate a role for PTEN in cellular type I interferon response and immunosuppression in PCa. However, the way that PTEN inactivation alters antitumor immune response in PCa is poorly understood. MATERIALS AND METHODS: To investigate the changes associated with PTEN loss and an immunosuppressive TME in PCa, we used CIBERSORT to estimate the relative abundance of 22 immune-cell types from 741 primary and 96 metastatic tumors. Our in silico findings were then validated by immunohistochemical analysis of immune cells and IDO1 and PDL1 checkpoint proteins in a cohort of 94 radical prostatectomy specimens. RESULTS: FoxP3+ T regulatory cells (Tregs) were significantly increased in PTEN-deficient PCa in all three public domain cohorts. Loss of PTEN in bone metastases was associated with lower CD8+ T-cell abundance, but in liver metastasis, FoxP3+ Tregs were present at higher levels. PTEN-deficient lymph node metastasis had a distinct profile, with high levels of CD8+ T cells. Moreover, we found that metastatic PCa presents higher abundance of FoxP3+ Treg when compared to primary lesions. Since PTEN-deficient tumors are likely to be immunosuppressed as a consequence of increased FoxP3+ Tregs, we then evaluated the localization and expression of IDO1, PDL1 immune checkpoints, and the corresponding density of FoxP3+ Treg and CD8+ T cells using our validation cohort (n = 94). We found that IDO1 protein expression and FoxP3+ Treg density were higher in neoplastic glands compared with benign adjacent tissue. Moreover, higher densities of FoxP3+ Treg cells in both stromal (P = 0.04) and tumor (P = 0.006) compartments were observed in PTEN-deficient tumors compared to tumors that retained PTEN activity. Similarly, IDO1 protein expression was significantly increased in the tumor glands of PTEN-deficient PCa (P < 0.0001). Spearman correlation analysis showed that IDO1 expression was significantly associated with FoxP3+ Treg and CD8+ T-cell density (P < 0.01). CONCLUSIONS: Our findings imply that PTEN deficiency is linked to an immunosuppressive state in PCa with distinct changes in the frequency of immune cell types in tumors from different metastatic sites. Our data suggest that determining PTEN status may also help guide the selection of patients for future immunotherapy trials in localized and metastatic PCa.


Assuntos
Fatores de Transcrição Forkhead/imunologia , Indolamina-Pirrol 2,3,-Dioxigenase/imunologia , Linfócitos do Interstício Tumoral/imunologia , PTEN Fosfo-Hidrolase/deficiência , Neoplasias de Próstata Resistentes à Castração/imunologia , Linfócitos T Reguladores/imunologia , Idoso , Antígeno B7-H1/imunologia , Estudos de Coortes , Fatores de Transcrição Forkhead/biossíntese , Humanos , Tolerância Imunológica , Indolamina-Pirrol 2,3,-Dioxigenase/biossíntese , Masculino , Pessoa de Meia-Idade , Metástase Neoplásica , PTEN Fosfo-Hidrolase/genética , PTEN Fosfo-Hidrolase/imunologia , Neoplasias de Próstata Resistentes à Castração/enzimologia , Neoplasias de Próstata Resistentes à Castração/genética , Análise Serial de Tecidos , Microambiente Tumoral/imunologia
17.
Cell Physiol Biochem ; 52(5): 1178-1192, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-30990587

RESUMO

BACKGROUND/AIMS: Rheumatoid arthritis (RA) is a progressive, chronic, even disabling systemic autoimmune disease. Imbalance between pathogenic immune cells and immunosuppressive cells is associated with the pathogenesis and development of RA and other autoimmune diseases. As Foxp3 is also expressed on activated CD4+ cells in the presence of inflammation, the identification of Treg cells in patients with RA remains a challenge. METHODS: Comprehensive analyses were carried out by Flow cytometry. Expression of Helios, CD226, T cell immunoreceptor with Ig and ITIM domains clinical samples and healthy controls. RESULTS: We have systemically examined three potential markers, Helios, CD226 and TIGIT, that are possibly related to Treg identification, and found that Helios expression on CD4+Foxp3+cells was decreased and negatively correlated with the disease activity of RA patients, while CD226 and TIGIT both showed elevated expression levels in CD4+Foxp3+cells in RA patients and they were not associated with disease activity of RA patients. CONCLUSION: Taken together, our findings indicate that CD4+CD25hiCD127low/-Foxp3+Helios+ may represent the real Treg cell population in patients with RA.


Assuntos
Antígenos de Diferenciação de Linfócitos T/imunologia , Antígenos de Diferenciação/imunologia , Artrite Reumatoide/imunologia , Fatores de Transcrição Forkhead/imunologia , Fator de Transcrição Ikaros/imunologia , Receptores Imunológicos/imunologia , Linfócitos T Reguladores/imunologia , Adulto , Artrite Reumatoide/patologia , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Linfócitos T Reguladores/patologia
18.
Int Immunopharmacol ; 72: 402-412, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31030096

RESUMO

BACKGROUND AND OBJECTIVE: Tangeretin demonstrates broad anti-inflammatory effects. The present study aimed to assess whether tangeretin functions in regulating T-regulatory cells (Tregs) and alleviating allergic rhinitis (AR). METHODS: An ovalbumin (OVA)-induced AR animal model was constructed to monitor the changes in the allergic symptom score, OVA-specific IgE titers, histopathological characteristics and T-helper cell (Th1, Th2, and Th17)-related cytokine levels under tangeretin or dexamethasone (DXM) administration. The expression levels of Notch1/Jagged1 and FOXP3, and the proportion of Tregs in the spleens of these animals, were also detected. Furthermore, purified naive CD4 + T cells were utilized to assess the effects of tangeretin on Notch1 expression and their differentiation in vitro. RESULTS: Both tangeretin and DXM administration alleviated airway inflammation, decreased the production of serum OVA-induced IgE, but only tangeretin administration restored the balance of cytokine profiles compared with those in the AR group. The abundance of splenic CD4 + CD25 + FOXP3 + Treg cells and the transcription factor FOXP3 were significantly increased under tangeretin treatment, either in AR mice or in naïve CD4 + T-cell differentiation, followed by a concomitant reduction in Notch1/Jagged1 expression. However, as a positive control, the treatment of allergic rhinitis with dexamethasone was not related to the expression of Notch1/Jagged1 or the differentiation of Treg cells. CONCLUSION: Tangeretin could promote regulatory T cell responses by inhibiting Notch1/Jagged1 expression, followed by promoting FOXP3/Treg cell differentiation and thus could serve as a novel curative therapeutic for AR.


Assuntos
Flavonas/farmacologia , Proteína Jagged-1/imunologia , Receptor Notch1/imunologia , Rinite Alérgica/imunologia , Linfócitos T Reguladores/efeitos dos fármacos , Animais , Diferenciação Celular/efeitos dos fármacos , Citocinas/imunologia , Feminino , Flavonas/uso terapêutico , Fatores de Transcrição Forkhead/imunologia , Camundongos Endogâmicos C57BL , Rinite Alérgica/tratamento farmacológico , Transdução de Sinais/efeitos dos fármacos , Baço/efeitos dos fármacos , Baço/imunologia , Linfócitos T Reguladores/imunologia
19.
Int Immunopharmacol ; 72: 422-428, 2019 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-31030098

RESUMO

Epigallocatechin gallate (EGCG) is a polyphenol that is found in green tea that has been shown to ameliorate airway inflammation in an ovalbumin-sensitized asthmatic mouse model. The purpose of this study was to investigate whether the immunomodulatory and anti-inflammatory effects of EGCG by regulating the regulatory T cell (Treg)/Th 17 cells balance in this model. Female BALB/c mice were sensitized and challenged with ovalbumin by intraperitoneal injection. EGCG was administered to asthmatic mice intraperitoneally 1 h before each OVA challenge. Airway hyperresponsiveness (AHR) was measured, and lung inflammatory infiltrations were assessed by hematoxylin and eosin (HE) staining. Serum OVA-specific IgE levels, Interleukin-10 (IL-10) levels and Interleukin-17A (IL-17A) levels in the bronchoalveolar lavage fluid (BALF), serum, and splenocyte culture supernatants were measured by ELISA. Flow cytometry was used to assess the effects of EGCG on the frequency of CD4+CD25+Foxp3+Treg cells in the splenocytes and real-time PCR method was used to measure the expression of Forkhead box P3 (Foxp3) mRNA and retinoid-related orphan receptor gammat (RORγt) mRNA in the lung tissue. The results showed that administration of EGCG significantly decreased AHR and OVA specific IgE in the serum, increased IL-10 levels in the BALF, serum, and splenocyte culture supernatant, and the frequency of CD4+CD25+Foxp3+Treg cells in the splenocytes in asthmatic mice. Administration of EGCG also ameliorated airway inflammation and eosinophil infiltrations in asthmatic mice. These results suggested that EGCG likely ameliorated OVA-induced airway inflammation by increasing the production of IL-10, the number of CD4+CD25+Foxp3+Treg cells and expression of Foxp3 mRNA in the lung tissue, and it could be an effective agent for treating asthma.


Assuntos
Anti-Inflamatórios/farmacologia , Anti-Inflamatórios/uso terapêutico , Asma/tratamento farmacológico , Catequina/análogos & derivados , Linfócitos T Reguladores/efeitos dos fármacos , Células Th17/efeitos dos fármacos , Animais , Asma/imunologia , Asma/patologia , Asma/fisiopatologia , Catequina/farmacologia , Catequina/uso terapêutico , Modelos Animais de Doenças , Feminino , Fatores de Transcrição Forkhead/imunologia , Imunoglobulina E/sangue , Interleucina-10/imunologia , Interleucina-17/imunologia , Pulmão/efeitos dos fármacos , Pulmão/imunologia , Pulmão/patologia , Camundongos Endogâmicos BALB C , Membro 3 do Grupo F da Subfamília 1 de Receptores Nucleares/imunologia , Linfócitos T Reguladores/imunologia , Células Th17/imunologia
20.
Nat Commun ; 10(1): 1823, 2019 04 23.
Artigo em Inglês | MEDLINE | ID: mdl-31015452

RESUMO

Granulomas are the pathological hallmark of tuberculosis (TB) and the niche where bacilli can grow and disseminate or the immunological microenvironment in which host cells interact to prevent bacterial dissemination. Here we show 34 immune transcripts align to the morphology of lung sections from Mycobacterium tuberculosis-infected mice at cellular resolution. Colocalizing transcript networks at <10 µm in C57BL/6 mouse granulomas increase complexity with time after infection. B-cell clusters develop late after infection. Transcripts from activated macrophages are enriched at subcellular distances from M. tuberculosis. Encapsulated C3HeB/FeJ granulomas show necrotic centers with transcripts associated with immunosuppression (Foxp3, Il10), whereas those in the granuloma rims associate with activated T cells and macrophages. We see highly diverse networks with common interactors in similar lesions. Different immune landscapes of M. tuberculosis granulomas depending on the time after infection, the histopathological features of the lesion, and the proximity to bacteria are here defined.


Assuntos
Linfócitos B/imunologia , Macrófagos/imunologia , Mycobacterium tuberculosis/imunologia , Tuberculoma/imunologia , Tuberculose Pulmonar/imunologia , Animais , Linfócitos B/metabolismo , Modelos Animais de Doenças , Fatores de Transcrição Forkhead/genética , Fatores de Transcrição Forkhead/imunologia , Fatores de Transcrição Forkhead/metabolismo , Humanos , Interleucina-10/genética , Interleucina-10/imunologia , Interleucina-10/metabolismo , Pulmão/imunologia , Pulmão/microbiologia , Pulmão/patologia , Macrófagos/metabolismo , Camundongos Endogâmicos C3H , Camundongos Endogâmicos C57BL , Mycobacterium tuberculosis/isolamento & purificação , RNA Mensageiro/isolamento & purificação , Fatores de Tempo , Tuberculoma/microbiologia , Tuberculoma/patologia , Tuberculose Pulmonar/microbiologia , Tuberculose Pulmonar/patologia
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