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1.
Turk J Haematol ; 37(1): 36-41, 2020 02 20.
Artigo em Inglês | MEDLINE | ID: mdl-31612695

RESUMO

Objective: CD4+CD8+ double-positive T-cells (DPTs) have been classified as a separate T-cell subpopulation, with two main phenotypes: CD4high CD8low and CD4low CD8high. In recent years, the relevance of DPTs in the pathogenesis of infections, tumors, and autoimmune diseases has been recognized. Reference values among healthy individuals remain unknown. Therefore, the aim of this study is to provide a reference value for DPTs in peripheral blood from healthy donors in a blood bank in Bogotá, Colombia, and to determine the activation status using a surface marker. Materials and Methods: One hundred healthy donors were enrolled in the study. Peripheral blood cells were stained for CD3, CD4, CD8, and CD154 (CD40L), and cellular viability was assessed with 7-aminoactinomycin D and analyzed by flow cytometry. Results: The median value for DPTs was 2.6% (interquartile range=1.70%-3.67%). Women had higher percentages of DPTs than men (3.3% vs. 2.1%). The subpopulation of CD4low CD8high showed higher expression of CD154 than the other T-cell subpopulations. Conclusion: DPT reference values were obtained from blood bank donors. A sex difference was found, and the CD4low CD8high subpopulation had the highest activation marker expression.


Assuntos
Bancos de Sangue , Doadores de Sangue , Antígenos CD4/metabolismo , Antígenos CD8/metabolismo , Contagem de Linfócitos , Subpopulações de Linfócitos T/metabolismo , Adolescente , Adulto , Idoso , Colômbia , Estudos Transversais , Feminino , Citometria de Fluxo , Humanos , Imunofenotipagem , Masculino , Pessoa de Meia-Idade , Adulto Jovem
2.
Cell Immunol ; 348: 103974, 2020 02.
Artigo em Inglês | MEDLINE | ID: mdl-31879029

RESUMO

Chronic infection by Trypanosoma cruzi decreases T cell proliferation and it is most likely accompanied by changes in signals required for activation. We assessed the effect of T. cruzi antigens on mitogen-induced proliferation of T cells from uninfected individuals and the association with the expression of molecules involved in antigen presentation, T cell costimulation and activation, and cytokine production. T. cruzi antigen exposure reduced mitogen-induced proliferation of CD4+ and CD8+ T cells in PBMC cultures, but only reduced mitogen-induced proliferation in the CD4+ T cells from sorted cell cultures cocultured with antigen-pulsed CD3- cells. CD40/CD80 and CD86 expression were reduced in antigen-pulsed DCs and monocytes, respectively. TNF-α, IL-10 and CCL17 levels were increased in cultures with antigen-pulsed CD3- cells, while CD3ζ chain expression was reduced in T cells from cultures with antigen. Our findings suggest that T. cruzi could alter T cell proliferation indirectly by downregulating costimulatory molecules and inducing the secretion of IL-10 and directly by decreasing TCR signaling.


Assuntos
Células Apresentadoras de Antígenos/imunologia , Antígenos de Protozoários/imunologia , Complexo CD3/imunologia , Doença de Chagas/imunologia , Linfócitos T/imunologia , Adulto , Proliferação de Células/fisiologia , Feminino , Humanos , Ativação Linfocitária/imunologia , Masculino , Transdução de Sinais/imunologia , Trypanosoma cruzi
3.
Front Immunol ; 10: 1671, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31379862

RESUMO

Monocytes are classified according to their CD14 and CD16 expression into classical (reparative), intermediate (inflammatory), and non-classical. This study assessed the frequency of monocyte and the relationship between monocyte subset percentages and the levels of blood cytokines in Colombian chagasic patients with different clinical forms. This study included chagasic patients in different clinical stages: indeterminate (IND) n = 14, chronic chagasic cardiomyopathy (CCC) n = 14, and heart transplant chagasic (HTCC) n = 9; controls with non-chagasic cardiopathy (NCC) n = 15, and healthy individuals (HI) n = 15. Peripheral blood mononuclear cells (PBMCs) were isolated, labeled for CD14, CD16, and HLA-DR, and analyzed by flow cytometry. Cytokines were measured with a bead-based immunoassay. Percentages of total CD14+ CD16+ and CD14+ HLA-DR+ monocytes were higher in patients with heart involvement (CCC, HTCC, and NCC) than controls. Percentages of intermediate monocytes increased in symptomatic chagasic patients (CCC and HTCC) compared to asymptomatic chagasic patients (IND) and controls (HI). Asymptomatic chagasic patients (IND) had higher percentages of classical monocytes, an increased production of CCL17 chemokine compared to chagasic symptomatic patients (CCC), and their levels of CCL17 was positively correlated with the percentage of classical monocyte subset. In CCC, the percentages of intermediate and classical monocytes were positively correlated with IL-6 levels, which were higher in this group compared to HI, and negatively with IL-12p40 concentration, respectively. Remarkably, there also was an important increased of classical monocytes frequency in three chronic chagasic patients who underwent cardiac transplant, of which one received anti-parasitic treatment. Our findings suggest that cardiac chagasic patients have an increased percentage of inflammatory monocytes and produce more IL-6, a biomarker of heart failure and left ventricular dysfunction, whereas asymptomatic chagasic individuals present a higher percentage of reparative monocytes and CCL17.


Assuntos
Doença de Chagas/imunologia , Citocinas/imunologia , Monócitos/imunologia , Adulto , Idoso , Doença de Chagas/sangue , Citocinas/sangue , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Índice de Gravidade de Doença
4.
Front Oncol ; 9: 245, 2019.
Artigo em Inglês | MEDLINE | ID: mdl-31024847

RESUMO

Acute leukemia is a heterogeneous set of diseases affecting children and adults. Current prognostic factors are not accurate predictors of the clinical outcome of adult patients and the stratification of risk groups remains insufficient. For that reason, this study proposes a multifactorial analysis which integrates clinical parameters, ex vivo tumor characterization and behavioral in vivo analysis in zebrafish. This model represents a new approach to understand leukemic primary cells behavior and features associated with aggressiveness and metastatic potential. Xenotransplantation of primary samples from patients newly diagnosed with acute leukemia in zebrafish embryos at 48 hpf was used to asses survival rate, dissemination pattern, and metastatic potential. Seven samples from young adults classified in adverse, favorable or intermediate risk group were characterized. Tumor heterogeneity defined by Leukemic stem cell (LSC) proportion, was performed by metabolic and cell membrane biomarkers characterization. Thus, our work combines all these parameters with a robust quantification strategy that provides important information about leukemia biology, their relationship with specific niches and the existent inter and intra-tumor heterogeneity in acute leukemia. In regard to prognostic factors, leukemic stem cell proportion and Patient-derived xenografts (PDX) migration into zebrafish were the variables with highest weights for the prediction analysis. Higher ALDH activity, less differentiated cells and a broader and random migration pattern are related with worse clinical outcome after induction chemotherapy. This model also recapitulates multiple aspects of human acute leukemia and therefore is a promising tool to be employed not only for preclinical studies but also supposes a new tool with a higher resolution compared to traditional methods for an accurate stratification of patients into worse or favorable clinical outcome.

5.
Immun Inflamm Dis ; 6(1): 47-57, 2018 03.
Artigo em Inglês | MEDLINE | ID: mdl-28967229

RESUMO

INTRODUCTION: Chagas disease is a parasitic infection whose pathogenesis is related to parasite persistence and a dysfunctional cellular immune response. Variability in cytokine secretion among chronic Trypanosoma cruzi-infected patients might preclude the identification of the pool of antigen specific T cells. The goal of this study was to determine the fraction of T cells responding to T. cruzi antigen measured by the expression of membrane TNF-α and CD154. METHODS: A total of 21 chagasic patients, 11 healthy and 5 non-chagasic cardiomyopathy controls were analyzed. PBMCs were short-term cultured in the presence of anti-CD28, anti-CD49d, anti-TNF-α, and TACE (TNF-α converting enzyme) inhibitor either under T. cruzi-lysate or polyclonal stimuli. Cells were stained with anti-CD3, anti-CD4, anti-CD8, and anti-CD154, and analyzed with flow cytometry. RESULTS: CD4+ and CD8+ T cells in chagasic patients displayed higher percentages of membrane-bound TNF-α+ and CD154+ compared with controls after T. cruzi-antigen stimulation. Both markers displayed a positive correlation in the T cell subpopulations analyzed. Symptomatic chagasic patients were differentiated from asymptomatic patients based on the expression of CD154 and membrane TNF-α in TCD4+ and TCD8+ compartments, respectively. CONCLUSIONS: These results show that both markers could be useful for assessing the pool of antigen-specific T cells in chronic chagasic patients.


Assuntos
Linfócitos T CD4-Positivos/imunologia , Ligante de CD40/imunologia , Linfócitos T CD8-Positivos/imunologia , Membrana Celular/imunologia , Doença de Chagas/imunologia , Trypanosoma cruzi/imunologia , Fator de Necrose Tumoral alfa/imunologia , Adulto , Linfócitos T CD4-Positivos/metabolismo , Linfócitos T CD4-Positivos/patologia , Ligante de CD40/sangue , Linfócitos T CD8-Positivos/metabolismo , Linfócitos T CD8-Positivos/patologia , Membrana Celular/metabolismo , Doença de Chagas/sangue , Feminino , Humanos , Masculino , Pessoa de Meia-Idade , Fator de Necrose Tumoral alfa/sangue
6.
Cytometry A ; 83(7): 627-35, 2013 Jul.
Artigo em Inglês | MEDLINE | ID: mdl-23671026

RESUMO

Astrocytes are involved in the pathogenesis of demyelinating diseases, where they actively regulate the secretion of proinflammatory factors, and trigger the recruitment of immune cells in the central nervous system (CNS). Antigen presentation of myelin-derived proteins has been shown to trigger astrocyte response, suggesting that astrocytes can directly sense demyelination. However, the direct response of astrocytes to lipid-debris generated during demyelination has not been investigated. The lipid composition of the myelin sheath is distinct, presenting significant amounts of cerebrosides, sulfocerebrosides (SCB), and ceramides. Studies have shown that microglia are activated in the presence of myelin-derived lipids, pointing to the possibility of lipid-induced astrocyte activation. In this study, a human astrocyte cell line was exposed to liposomes enriched in each myelin lipid component. Although liposome uptake was observed for all compositions, astrocytes had augmented uptake for liposomes containing sulfocerebroside (SCB). This enhanced uptake did not modify their expression of human leukocyte antigen (HLA) molecules or secretion of chemokines. This was in contrast to changes observed in astrocyte cells stimulated with IFNγ. Contrary to human monocytes, astrocytes did not internalize beads in the size-range of liposomes, indicating that liposome uptake is lipid specific. Epifluorescence microscopy corroborated that liposome uptake takes place through endocytosis. Soluble SCB were found to partially block uptake of liposomes containing this same lipid. Endocytosis was not decreased when cells were treated with cytochalasin D, but it was decreased by cold temperature incubation. The specific uptake of SCB in the absence of a proinflammatory response indicates that astrocytes may participate in the trafficking and regulation of sulfocerebroside metabolism and homeostasis in the CNS.


Assuntos
Astrócitos/metabolismo , Cerebrosídeos/metabolismo , Endocitose/efeitos dos fármacos , Lipossomos/metabolismo , Astrócitos/efeitos dos fármacos , Astrócitos/imunologia , Linhagem Celular , Cerebrosídeos/farmacologia , Quimiocinas/metabolismo , Colesterol/metabolismo , Antígenos HLA-DR/metabolismo , Antígenos de Histocompatibilidade Classe I/metabolismo , Humanos , Mediadores da Inflamação/metabolismo , Fosfatidilcolinas/metabolismo , Fosfatidiletanolaminas/metabolismo
7.
PLoS Negl Trop Dis ; 7(1): e2038, 2013.
Artigo em Inglês | MEDLINE | ID: mdl-23383358

RESUMO

BACKGROUND: Chronic persistent infections have been associated with T lymphocytes functional impairment. The aim of this study was to compare the activation status, the proliferative potential and the expression of CD28 and CD3ζ chain on T lymphocytes between chronic chagasic patients and uninfected controls. METHODOLOGY/PRINCIPAL FINDINGS: Forty-two chronic chagasic patients, 28 healthy individuals and 32 non-chagasic cardiomyopathy donors were included. Peripheral blood was marked for CD3, CD4, CD8, HLA-DR, CD28, CD38 and intracellular CD3ζ. Peripheral blood mononuclear cells were stained with carboxyfluorescein diacetate succinimidylester and incubated with T. cruzi lysate or phytohemagglutinin for five days. Cells from 3 healthy controls were incubated with T. cruzi trypomastigotes separated with transwells; and the expression of CD3ζ chain and proliferation index was determined. Heart-infiltrating cells from two chronic chagasic patients were tested for the aforementioned cellular markers. Chagasic patients displayed higher frequencies of CD4+/HLA-DR+/CD38+ (8.1% ± 6.1) and CD8+/HLA-DR+/CD38+ (19.8 ± 8.9) T cells in comparison with healthy (1.6 ± 1.0; 10.6 ± 8.0) and non-chagasic cardiomyopathy donors (2.9 ± 2.9; 5.8 ± 6.8). Furthermore, the percentage of CD4+ activated T cells was higher in chagasic patients with cardiac involvement. CD8+ T cells proliferation index in chagasic donors (1.7 ± 0.3) was lower when compared with healthy (2.3 ± 0.3) and non-chagasic cardiomyopathy individuals (3.1 ± 1.1). The frequencies of CD4+/CD28+ and CD8+/CD28+ T cells, as well as the CD3ζ(bright)/CD3ζ(dim)% ratios in CD4+ and CD8+ were lower in chagasic patients when compared with both control groups. The CD3ζ(bright)/CD3ζ(dim)% ratio and proliferative indexes for CD4+ and CD8+ T lymphocytes decreased gradually in those cells cultivated with parasites and displayed lower values than those incubated with medium alone. Finally, heart-infiltrating T cells from two T. cruzi infected patients also expressed activation markers and down-regulate CD28 and CD3ζ. CONCLUSIONS: CD8+ T lymphocytes from chagasic donors displayed reduced proliferative capacity, which might be associated with CD3ζ down-regulation and diminished CD28 expression on CD4 T cells.


Assuntos
Antígenos CD28/biossíntese , Complexo CD3/biossíntese , Linfócitos T CD8-Positivos/imunologia , Doença de Chagas/imunologia , Ativação Linfocitária , Adulto , Idoso , Idoso de 80 Anos ou mais , Proliferação de Células , Regulação para Baixo , Feminino , Humanos , Imunofenotipagem , Masculino , Pessoa de Meia-Idade
8.
PLoS Negl Trop Dis ; 5(8): e1294, 2011 Aug.
Artigo em Inglês | MEDLINE | ID: mdl-21886854

RESUMO

BACKGROUND: CD4+/CD8+ double positive (DP) T cells have been described in healthy individuals as well as in patients with autoimmune and chronic infectious diseases. In chronic viral infections, this cell subset has effector memory phenotype and displays antigen specificity. No previous studies of double positive T cells in parasite infections have been carried out. METHODOLOGY/PRINCIPAL FINDINGS: Seventeen chronic chagasic patients (7 asymptomatic and 10 symptomatic) and 24 non-infected donors, including 12 healthy and 12 with non-chagasic cardiomyopathy donors were analyzed. Peripheral blood was stained for CD3, CD4, CD8, HLA-DR and CD38, and lymphocytes for intracellular perforin. Antigen specificity was assessed using HLA*A2 tetramers loaded with T. cruzi K1 or influenza virus epitopes. Surface expression of CD107 and intracellular IFN-γ production were determined in K1-specific DP T cells from 11 chagasic donors. Heart tissue from a chronic chagasic patient was stained for both CD8 and CD4 by immunochemistry. Chagasic patients showed higher frequencies of DP T cells (2.1% ± 0.9) compared with healthy (1.1% ± 0.5) and non-chagasic cardiomyopathy (1.2% ± 0.4) donors. DP T cells from Chagasic patients also expressed more HLA-DR, CD38 and perforin and had higher frequencies of T. cruzi K1-specific cells. IFN-γ production in K1-specific cells was higher in asymptomatic patients after polyclonal stimulation, while these cells tended to degranulate more in symptomatic donors. Immunochemistry revealed that double positive T cells infiltrate the cardiac tissue of a chagasic donor. CONCLUSIONS: Chagasic patients have higher percentages of circulating double positive T cells expressing activation markers, potential effector molecules and greater class I antigenic specificity against T. cruzi. Although K1 tetramer positive DP T cell produced little IFN-γ, they displayed degranulation activity that was increased in symptomatic patients. Moreover, K1-specific DP T cells can migrate to the heart tissue.


Assuntos
Linfócitos T CD4-Positivos/imunologia , Linfócitos T CD8-Positivos/imunologia , Cardiomiopatia Chagásica/imunologia , Subpopulações de Linfócitos T/imunologia , Adulto , Idoso , Idoso de 80 Anos ou mais , Antígenos CD/análise , Linfócitos T CD4-Positivos/química , Linfócitos T CD8-Positivos/química , Cardiomiopatia Chagásica/patologia , Feminino , Antígenos HLA/análise , Humanos , Interferon gama/biossíntese , Masculino , Pessoa de Meia-Idade , Miocárdio/patologia , Perforina/análise , Subpopulações de Linfócitos T/química
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