RESUMO
OBJECTIVE: To analyze whether the expression and modulation of T cell receptor (TCR) signaling is dependent on Casitas B lineage lymphoma b (Cbl-b) in T cells from patients with systemic lupus erythematosus (SLE) upon stimulation with a tolerogenic substance. METHODS: Peripheral blood mononuclear cells were obtained from 20 patients with SLE (active disease or in remission) and 20 healthy controls. Levels of Cbl-b expression were measured using reverse transcription-polymerase chain reaction and Western blotting in peripheral CD4+ T cells from SLE patients and healthy controls upon anergy induction. Cell proliferation was measured using the carboxyfluorescein diacetate succinimidyl ester dilution method. Cytokine production was analyzed by luminometry, and surface expression of activation markers was assessed by flow cytometry. Transfection assays were performed to induce overexpression of Cbl-b, and phosphorylation of TCR-associated kinases was evaluated. RESULTS: CD4+ T cells from SLE patients displayed resistance to anergy (as evidenced by increased cell proliferation, interleukin-2 production, and expression of activation and costimulatory markers), and this was associated with altered Cbl-b expression. Upon ionomycin treatment, primary T cells showed enhanced MAPK activity and decreased Akt phosphorylation, which was representative of the anergic state. In T cells from lupus patients, Cbl-b overexpression led to increased expression of phosphorylated MAPK, thus indicating the reversibility of anergy resistance. CONCLUSION: These findings suggest that abnormal peripheral tolerance in SLE is caused by a deficiency in Cbl-b, and that this ubiquitin ligase plays a key role in regulating TCR signaling during the induction of peripheral tolerance.
Assuntos
Proteínas Adaptadoras de Transdução de Sinal/imunologia , Linfócitos T CD4-Positivos/imunologia , Lúpus Eritematoso Sistêmico/imunologia , Tolerância Periférica/imunologia , Proteínas Proto-Oncogênicas c-cbl/imunologia , Receptores de Antígenos de Linfócitos T/imunologia , Proteínas Adaptadoras de Transdução de Sinal/genética , Proteínas Adaptadoras de Transdução de Sinal/metabolismo , Adulto , Linfócitos T CD4-Positivos/citologia , Linfócitos T CD4-Positivos/metabolismo , Estudos de Casos e Controles , Proliferação de Células , Anergia Clonal , Citocinas/imunologia , Citocinas/metabolismo , Feminino , Perfilação da Expressão Gênica , Humanos , Masculino , Proteínas Proto-Oncogênicas c-cbl/genética , Proteínas Proto-Oncogênicas c-cbl/metabolismo , RNA Mensageiro/análise , Receptores de Antígenos de Linfócitos T/metabolismo , Reação em Cadeia da Polimerase Via Transcriptase Reversa , Transdução de Sinais/imunologiaRESUMO
Early Growth Response (EGR) zinc finger transcription factors are induced under diverse mitogenic signals on different cell types such as lymphocytes. Their genetic expression does not require de novo protein synthesis, which suggests its role as immediate response mediators between cell surface receptor signaling and gene expression regulation. EGR factors are involved in modulating the immune response, by means of the induction of differentiation of lymphocyte precursors, activation of T and B cells, as well as their involvement in central and peripheral tolerance. The maturation state, particularly for B cells, and signaling through the T or B cell receptors seems to be quite relevant for the induction of the expression of these transcription factors. EGR-1 functions as a positive regulatory factor for B and T cells mediated by transcriptional regulation of key cytokines and costimulatory molecules, and its interaction with NFAT. On the opposite, EGR-2 and 3 act as negative regulators involved in anergy induction and apoptosis. EGR-2 and 3 deficiency has been related to the development of lupus like disease in murine models. The deficiency of these transcription factors has been associated to deficient Cbl-b expression, a resistant to anergy phenotype, and expansion of effector and activated T cells.