RESUMO
Trypanosoma cruzi acute infection leads to thymic atrophy, largely as a result of death of immature DP T cells. In a second vein, the glucocorticoid hormone imbalance promotes DP T cell apoptosis in infected mice. Herein, we assessed the involvement of caspase signaling in thymocyte death during T. cruzi acute infection. BALB/c mice were infected i.p. with 10(2) trypomastigote forms of T. cruzi and analyzed from 7 to 19 dpi. Thymocyte apoptosis was observed in early stages of infection, increasing along with time postinfection. Immature DN and DP as well as CD4(+) and CD8(+) thymocytes from infected mice showed increased activation of caspase-8, -9, and -3. In vitro treatment of thymocytes from infected mice with a general caspase inhibitor or the combination of caspase-8- and caspase-9-specific inhibitors increased the number of living thymocytes. Intrathymic injection of the general caspase inhibitor, but not caspase-8 or -9 inhibitors individually, prevented thymic atrophy and thymocyte depletion in infected mice. Moreover, blockade of glucocorticoid receptor activity with RU486 prevented DP thymocyte apoptosis, together with caspase-8 and -9 activation. These findings indicate that DP T cell apoptosis following experimental T. cruzi acute infection is dependent on glucocorticoid stimulation, promoting caspase-8 and -9 activation.
Assuntos
Caspase 8/metabolismo , Caspase 9/metabolismo , Doença de Chagas/enzimologia , Linfócitos T/patologia , Animais , Apoptose , Doença de Chagas/imunologia , Doença de Chagas/patologia , Citometria de Fluxo , Masculino , Camundongos , Camundongos Endogâmicos BALB C , Linfócitos T/enzimologia , Linfócitos T/parasitologia , Timo/enzimologia , Timo/imunologia , Timo/patologia , Trypanosoma cruzi/imunologiaRESUMO
BACKGROUND & AIMS: Defective apoptosis of lamina propria T cells (LPTs) is involved in the pathogenesis of Crohn's disease. Survivin, a member of the inhibitors of apoptosis family, prevents cell death and regulates cell division. Survivin has been studied extensively in cancer, but little is known about its role in Crohn's disease. METHODS: LPTs were isolated from mucosal samples of patients with Crohn's disease or ulcerative colitis and healthy individuals (controls). LPTs were activated with interleukin-2 or via CD3, CD2, and CD28 signaling, and cultured at 42°C to induce heat shock. Survivin expression was assessed by immunohistochemistry, confocal microscopy, and immunoblotting; survivin levels were reduced by RNA interference. Cell viability, apoptosis, and proliferation were measured by trypan blue exclusion, annexin-V/7-Aminoactinomycin D staining, and uptake of [3]thymidine, respectively. RESULTS: LPTs from patients with Crohn's disease had higher levels of survivin than LPTs from patients with ulcerative colitis or controls. RNA knockdown of survivin in LPTs inhibited their proliferation and promoted apoptosis. Levels of survivin were low in LPTs from patients with ulcerative colitis and controls as a result of ubiquitin-mediated proteasome degradation. In LPTs from patients with Crohn's disease, survivin bound to the heat shock protein (HSP)90, and therefore was resistant to proteasome degradation. Incubating LPTs with 17-N-allylamino-17-demethoxygeldanamycin, an inhibitor of HSP90, reduced levels of survivin and induced apoptosis. CONCLUSIONS: Levels of survivin are increased in LPTs from patients with Crohn's disease (compared with ulcerative colitis and controls) because survivin interacts with HSP90 and prevents proteasome degradation. This allows LPTs to avoid apoptosis. Strategies to restore apoptosis to these cells might be developed to treat patients with Crohn's disease.
Assuntos
Colite Ulcerativa/metabolismo , Doença de Crohn/metabolismo , Proteínas de Choque Térmico HSP90/metabolismo , Proteínas Inibidoras de Apoptose/metabolismo , Linfócitos T/metabolismo , Adolescente , Adulto , Idoso , Idoso de 80 Anos ou mais , Proteínas Reguladoras de Apoptose , Antígenos CD2/metabolismo , Antígenos CD28/metabolismo , Complexo CD3/metabolismo , Complexo CD3/farmacologia , Núcleo Celular/metabolismo , Proliferação de Células , Sobrevivência Celular , Células Cultivadas , Colite Ulcerativa/imunologia , Doença de Crohn/imunologia , Citoplasma/metabolismo , Endopeptidase K/farmacologia , Feminino , Humanos , Proteínas Inibidoras de Apoptose/genética , Interleucina-2/farmacologia , Mucosa Intestinal/imunologia , Mucosa Intestinal/metabolismo , Masculino , Pessoa de Meia-Idade , Proteínas Mitocondriais , Fosforilação/efeitos dos fármacos , Proteólise , Interferência de RNA , RNA Mensageiro/metabolismo , Transdução de Sinais , Survivina , Linfócitos T/efeitos dos fármacos , Linfócitos T/enzimologia , Ubiquitinação , Adulto JovemRESUMO
Thyroid hormones (THs) exert a broad range of actions on development, growth, and cell differentiation by both genomic and nongenomic mechanisms. THs regulate lymphocyte function, but the participation of nongenomic actions is still unknown. Here the contribution of both genomic and nongenomic effects on TH-induced division of T cells was studied by using free and noncell permeable THs coupled to agarose (TH-ag). THs-ag led to cell division, but to a lesser extent than free hormones. THs induced nongenomically the rapid translocation of protein kinase C (PKC) ζ isoform to cell membranes, extracellular-signal-regulated kinases (ERK1/2) phosphorylation and nuclear factor-κB (NF-κB) activation. The signaling cascade include sphingomyelinases acting up-stream the activation of PKCζ isoform, while ERK and NF-κB are activated downstream this PKC isoenzyme. Both free and THs-ag increased the protein and mRNA levels of TH nuclear receptor TRα1, while only free hormones incremented the inducible NOS gene and protein levels as well as a calcium independent NOS activity. Both effects were blunted by PKCζ inhibition. These results indicate that THs, by triggering a nongenomic signaling cascade that involves Smases-mediated activation of PKCζ, lead to ERK 1/2 and NF-κB activation and to the genomic increase of TRs and the inducible nitric oxide synthase protein and mRNA levels, improving T lymphocyte proliferation. These finding not only contribute to the understanding of the mechanisms involved in TH modulation of lymphocyte physiology, but would also point out for the first time the interplay between genomic and nongenomic TH actions in T cells.
Assuntos
Proliferação de Células , Óxido Nítrico Sintase Tipo II/metabolismo , Linfócitos T/enzimologia , Receptores alfa dos Hormônios Tireóideos/metabolismo , Tiroxina/metabolismo , Tri-Iodotironina/metabolismo , Animais , Linhagem Celular Tumoral , Proliferação de Células/efeitos dos fármacos , Regulação Enzimológica da Expressão Gênica , Camundongos , Proteína Quinase 1 Ativada por Mitógeno/metabolismo , Proteína Quinase 3 Ativada por Mitógeno/metabolismo , NF-kappa B/metabolismo , Óxido Nítrico Sintase Tipo II/genética , Fosforilação , Proteína Quinase C/antagonistas & inibidores , Proteína Quinase C/metabolismo , Inibidores de Proteínas Quinases/farmacologia , Transporte Proteico , RNA Mensageiro/metabolismo , Transdução de Sinais , Esfingomielina Fosfodiesterase/metabolismo , Linfócitos T/efeitos dos fármacos , Receptores alfa dos Hormônios Tireóideos/genética , Fatores de Tempo , Regulação para CimaRESUMO
The thymus plays a crucial role in the development of T lymphocytes by providing an inductive microenvironment in which committed progenitors undergo proliferation, T-cell receptor gene rearrangements and thymocyte differentiate into mature T cells. The thymus microenvironment forms a complex network of interaction that comprises non lymphoid cells (e.g., thymic epithelial cells, TEC), cytokines, chemokines, extracellular matrix elements (ECM), matrix metalloproteinases and other soluble proteins. The thymic epithelial meshwork is the major component of the thymic microenvironment, both morphologically and phenotypically limiting heterogeneous regions in thymic lobules and fulfilling an important role during specific stages of T-cell maturation. The process starts when bone marrow-derived lymphocyte precursors arrive at the outer cortical region of the thymic gland and begin to mature into functional T lymphocytes that will finally exit the thymus and populate the peripheral lymphoid organs. During their journey inside the thymus, thymocytes must interact with stromal cells (and their soluble products) and extracellular matrix proteins to receive appropriate signals for survival, proliferation and differentiation. The crucial components of the thymus microenvironment, and their complex interactions during the T-cell maturation process are summarized here with the objective of contributing to a better understanding of the function of the thymus, as well as assisting in the search for new therapeutic approaches to improve the immune response in various pathological conditions.
Assuntos
Diferenciação Celular , Linfócitos T/citologia , Timo/patologia , Animais , Movimento Celular , Quimiocinas/metabolismo , Matriz Extracelular/metabolismo , Galectinas/metabolismo , Humanos , Metaloproteases/metabolismo , Receptores de Quimiocinas/metabolismo , Linfócitos T/enzimologia , Timo/enzimologiaRESUMO
BACKGROUND: Pityriasis lichenoides (PL) is an inflammatory skin disease of unknown etiology. Nitric oxide (NO) has emerged as an important mediator of many physiological functions. The importance of NO-mediated signaling in skin diseases has been reported by several studies. METHODS: A review of clinical records and histopathological slides of 34 patients diagnosed with PL was performed. Three different groups of skin biopsies including PL chronica (24 patients), PL et varioliformis acuta (10 patients) and 15 normal skin samples were subjected to the immunohistochemistry technique for inducible nitric oxide synthase (iNOS) detection. RESULTS: Normal skin group exhibited a few number of iNOS-positive cells in the dermis and rare positive cells in the upper epidermis, unlike abundant epidermal and dermal iNOS expression observed in both PL groups. CONCLUSION: According to our results, we hypothesize that NO produced by iNOS could participate in PL pathogenesis. Abnormal and persistent responses to unknown antigens, probably a pathogen, associated with NO immunoregulatory functions could contribute to the relapsing course observed in PL. NO anti-apoptotic effect on T-cell lymphocytes could play a role on maintenance of reactive T cells, leading to a T-cell lymphoid dyscrasia.
Assuntos
Derme/enzimologia , Regulação Enzimológica da Expressão Gênica , Óxido Nítrico Sintase Tipo II/biossíntese , Óxido Nítrico/biossíntese , Pitiríase Liquenoide/enzimologia , Derme/patologia , Feminino , Humanos , Inflamação/enzimologia , Inflamação/patologia , Masculino , Pitiríase Liquenoide/patologia , Transdução de Sinais , Linfócitos T/enzimologia , Linfócitos T/patologiaRESUMO
This study investigated the effects of (-)-epicatechin (EC), its oligomers, dimer B2 (B2) and trimer C1 (C1), on calcium-dependent cell oxidation. Jurkat T cells were subjected to a Ca(2+) mobilization challenge by replacing Na(+) with K(+) in the incubation media. A 249+/-38% increase in intracellular Ca(2+) concentration ([Ca(2+)](i)) was observed and that effect was prevented by the presence of inhibitors of Ca(2+) mobilization and entrance. The pre-incubation of the cells in the presence of EC, B2 or C1 prevented K(+)-mediated increase in [Ca(2+)](i). IC(50) were 10, 24 and 196 nMfor EC, B2 and C1, respectively. Cell membrane depolarization was affected by K(+), but neither inhibitors of Ca(2+) mobilization nor EC, B2 or C1 modified the increase in membrane potential. An 84+/-7% increase in cell oxidants was observed after cell exposure to K(+). This increase was prevented by the inhibition of Ca(2+) mobilization, NADPH oxidase and protein kinase C, as well as by 10 nM EC, 10 nM B2 or 100 nM C1. In addition, EC and B2 (100 nM) significantly inhibited the activation of the [Ca(2+)](i)-regulated transcription factor NFAT. These results indicate that EC and related oligomers, assayed at physiologically achievable concentrations, can modulate [Ca(2+)](i) and then prevent cell oxidation and other Ca(2+)-regulated events.
Assuntos
Antioxidantes/farmacologia , Cálcio/metabolismo , Catequina/farmacologia , Estresse Oxidativo/efeitos dos fármacos , Proantocianidinas/farmacologia , Linfócitos T/efeitos dos fármacos , Humanos , Células Jurkat , Potenciais da Membrana , NADPH Oxidases/metabolismo , Fatores de Transcrição NFATC/metabolismo , Potássio/metabolismo , Proteína Quinase C/metabolismo , Linfócitos T/enzimologia , Linfócitos T/metabolismoRESUMO
The aim of the present study was to assess the effect of dehydroepiandrosterone (DHEA: 10 microM) and metformin (10 microM and 100 microM) in regulating proliferation of cultured T lymphocytes. T cells were isolated from lymph nodes of prepuberal BALB/c mice. We found that DHEA, metformin and DHEA + metformin added to the incubation media diminished proliferation of T cells. The inhibition by DHEA was higher than that produced by metformin, while the combined treatment showed a synergistic action that allowed us to speculate distinct regulatory pathways. This was supported later by other findings in which the addition of DHEA to the incubation media did not modify T lymphocyte viability, while treatment with metformin and DHEA + metformin diminished cellular viability and increased both early and late apoptosis. Moreover, DHEA diminished the content of the anti-oxidant molecule glutathione (GSH), whereas M and DHEA + metformin increased GSH levels and diminished lipid peroxidation. We conclude that DHEA and metformin diminish proliferation of T cells through different pathways and that not only the increase, but also the decrease of oxidative stress inhibited proliferation of T cells, i.e. a minimal status of oxidative stress, is necessary to trigger cellular response.
Assuntos
Antioxidantes/farmacologia , Desidroepiandrosterona/farmacologia , Metformina/farmacologia , Linfócitos T/citologia , Animais , Apoptose/efeitos dos fármacos , Proliferação de Células/efeitos dos fármacos , Sobrevivência Celular/efeitos dos fármacos , Células Cultivadas , Feminino , Glutationa/metabolismo , Peroxidação de Lipídeos/efeitos dos fármacos , Ativação Linfocitária , Camundongos , Camundongos Endogâmicos BALB C , Necrose , Óxido Nítrico Sintase/metabolismo , Estresse Oxidativo/efeitos dos fármacos , Síndrome do Ovário Policístico/imunologia , Linfócitos T/efeitos dos fármacos , Linfócitos T/enzimologiaRESUMO
OBJECTIVE: Data from the literature demonstrate that the local and systemic immune responses seem to play an important role in the progression of cervical intraepithelial neoplasia (CIN). Our aim was to investigate whether recurrences among CIN III patients might be related to the presence of local lymphocytes, macrophage and enzyme iNOS. METHODS: We analyzed 35 patients with CIN III who underwent conization and followed up for a minimum of 4 years. Using immunohistochemistry, the presence of T lymphocytes (CD3, CD8 and CD45RO), B lymphocytes (CD20), macrophages (CD68) and the expression of the enzyme iNOS were investigated. The quantity of marked cells is graded as: 0, absence of cells; 1, rare cells; 2, moderate number of cells; 3, many cells. For statistical purposes, we took the scores 0 and 1 to indicate weak marking and the scores 2 and 3 to indicate strong marking. RESULTS: We found strong positive expression of CD3-positive T lymphocytes among CIN III patients with recurrence following conization (100 vs. 50% without recurrence, p=0.02). We did not find any statistical differences in the expression of CD20, CD68, CD45RO, CD8 or iNOS. CONCLUSIONS: It is concluded that strong positive findings of CD3 T lymphocytes were related to recurrence following conization due to CIN III.
Assuntos
Complexo CD3/imunologia , Recidiva Local de Neoplasia/imunologia , Linfócitos T/imunologia , Displasia do Colo do Útero/imunologia , Neoplasias do Colo do Útero/imunologia , Adulto , Linfócitos B/enzimologia , Linfócitos B/imunologia , Linfócitos T CD8-Positivos/enzimologia , Linfócitos T CD8-Positivos/imunologia , Feminino , Seguimentos , Humanos , Imuno-Histoquímica , Macrófagos/enzimologia , Macrófagos/imunologia , Pessoa de Meia-Idade , Recidiva Local de Neoplasia/patologia , Óxido Nítrico Sintase Tipo II/biossíntese , Linfócitos T/enzimologia , Neoplasias do Colo do Útero/enzimologia , Neoplasias do Colo do Útero/patologia , Neoplasias do Colo do Útero/cirurgia , Adulto Jovem , Displasia do Colo do Útero/enzimologia , Displasia do Colo do Útero/patologia , Displasia do Colo do Útero/cirurgiaRESUMO
Purine nucleoside phosphorylase (PNP) catalyzes the reversible phosphorolysis of nucleosides and deoxynucleosides, generating ribose 1-phosphate and the purine base, which is an important step of purine catabolism pathway. The lack of such an activity in humans, owing to a genetic disorder, causes T-cell impairment, and thus drugs that inhibit human PNP activity have the potential of being utilized as modulators of the immunological system to treat leukemia, autoimmune diseases, and rejection in organ transplantation. Besides, the purine salvage pathway is the only possible way for apicomplexan parasites to obtain the building blocks for RNA and DNA synthesis, which makes PNP from these parasites an attractive target for drug development against diseases such as malaria. Hence, a number of research groups have made efforts to elucidate the mechanism of action of PNP based on structural and kinetic studies. It is conceivable that the mechanism may be different for PNPs from diverse sources, and influenced by the oligomeric state of the enzyme in solution. Furthermore, distinct transition state structures can make possible the rational design of specific inhibitors for human and apicomplexan enzymes. Here, we review the current status of these research efforts to elucidate the mechanism of PNP-catalyzed chemical reaction, focusing on the mammalian and Plamodium falciparum enzymes, targets for drug development against, respectively, T-Cell- and Apicomplexan parasites-mediated diseases.
Assuntos
Apicomplexa/enzimologia , Sistemas de Liberação de Medicamentos/métodos , Infecções por Protozoários/enzimologia , Purina-Núcleosídeo Fosforilase/metabolismo , Linfócitos T/enzimologia , Animais , Apicomplexa/patogenicidade , Humanos , Infecções por Protozoários/tratamento farmacológico , Infecções por Protozoários/parasitologia , Purina-Núcleosídeo Fosforilase/antagonistas & inibidores , Linfócitos T/parasitologiaRESUMO
Adhesive interactions play important roles in coordinating T cell migration and activation, which are mediated by binding of integrins to RGD motif found on extracellular matrix proteins. Disintegrins, isolated from snake venoms, contain the RGD sequence that confers selectivity to integrin interaction. We have investigated the ability of three RGD-disintegrins, ligands of alpha(5)beta(1) and alpha(v)beta(3), Flavoridin (Fl), Kistrin (Kr) and Echistatin (Ech), in modulating the activation of human T lymphocyte. The disintegrins induced T cell proliferation and CD69 expression. This activation parallels with actin cytoskeleton reorganization and tyrosine phosphorylation. Furthermore, the peptides induced focal adhesion kinase (FAK) and phosphoinositide 3-kinase (PI3K) activation. Finally, RGD-disintegrins were capable of driving NF-kappaB nuclear translocation and c-Fos expression, in a PI3K and ERK1/2 activities dependent manner. This report is the first to show that RGD-disintegrins interact with integrins on human T lymphocyte surface, modulating cell proliferation and activation of specific pathways coupled to integrin receptor.