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1.
Article in English | WPRIM | ID: wpr-186263

ABSTRACT

Previously, we reported that CD40-induced production of reactive oxygen species (ROS) by NADPH oxidase requires the TNF receptor-associated factor (TRAF) 3, as well as the activities of phosphatidylinositol 3-kinase (PI3K) and Rac1. Here we investigated the possible mechanisms of the production of ROS after CD40 ligation in B cells. We describe an alternative ROS production pathway that is triggered by CD40 ligation, involves 5-lipoxygenase (5-LO), and results in activation of p38 MAPK. Our studies in Raji human B lymphomas revealed that CD40-induced ROS production by 5-LO also requires the activities of PI3K and Rac1. In contrast to the NADPH oxidase pathway, however, TRAF molecules are not required for the CD40-induced ROS production by 5-LO. The association of CD40 with 5-LO is dependent on CD40 ligation in Raji B cells, and co-immunoprecipitation experiments using epitope-tagged proteins transiently expressed in human embryonic kidney 293T cells revealed the role of the regulatory subunit of PI3K, p85, in this association. Collectively, these data suggest a separate pathway for the CD40-induced ROS production in B cells and demonstrate that this pathway requires 5-LO via direct association of p85 with both CD40 and 5-LO.


Subject(s)
CD40 Antigens/metabolism , Arachidonate 5-Lipoxygenase/metabolism , B-Lymphocytes/enzymology , CD40 Ligand/metabolism , Cell Line, Tumor , Enzyme Activation , HEK293 Cells , Humans , Phosphatidylinositol 3-Kinases/metabolism , Protein Binding , Reactive Oxygen Species/metabolism , Signal Transduction , p38 Mitogen-Activated Protein Kinases/metabolism , rac GTP-Binding Proteins/metabolism
2.
Medical Principles and Practice. 2009; 18 (4): 266-271
in English | IMEMR | ID: emr-92166

ABSTRACT

To evaluate subclinical inflammation and fibrinolysis in low-risk type 2 diabetic subjects and to assess the efficacy of metformin and rosiglitazone in this group Sixty-one normotensive, normoalbuminuric type 2 diabetic subjects without diabetes-related complications were included in a 4-week standardization period with glimepiride. After the standardization period, 21 subjects were excluded and the remaining 40 were randomly divided into two groups matched for age, gender, body mass index and disease duration. The first group [n = 20] received metformin [1,700 mg/day], the second group [n = 20] rosiglitazone [4 mg/day] for 12 weeks. Patients with low-density lipoprotein-cholesterol higher than 130 mg/dl at the beginning of the randomization period were treated with simvastatin [maximum dose 20 mg/day]. Twenty-three healthy controls were also recruited. Cytokine measurements were performed with ELISA kits Baseline plasma plasminogen activator inhibitor-1 [PAI-1] level of type 2 diabetic subjects was significantly elevated [p = 0.038], but baseline levels of soluble CD40 ligand [sCD40L] and thrombin-activatable fibrinolysis inhibitor-1 [TAFI] antigen did not differ from healthy controls. Twelve weeks of metformin or rosiglitazone therapy did not cause significant changes in sCD40L, PAI-1 and TAFI antigen levels. In simvastatin-treated subjects [n = 9] significant reductions of PAI-1 were achieved [p = 0.028], while sCD40L and TAFI-Ag did not differ from baseline values. Our results showed that nonobese diabetic patients at low cardiovascular risk had similar levels of subclinical markers of inflammation and fibrinolysis as matched healthy controls. Neither metformin nor rosiglitazone caused marked changes in sCD40L, PAI-1 and TAFI antigen levels. A subset of patients who received simvastatin showed a modest decrease in PAI-1 level and could contribute to beneficial vasculoprotective effect of the drug in type 2 diabetics


Subject(s)
Humans , Male , Female , Plasminogen Activator Inhibitor 1/metabolism , Metformin , Thiazolidinediones , Simvastatin , CD40 Ligand/metabolism , Carboxypeptidase B2/metabolism , Fibrinolysis , Blood Pressure/drug effects
3.
Article in English | WPRIM | ID: wpr-43452

ABSTRACT

Several myeloid leukemia-derived cells have been reported to possess the ability to differentiate into dendritic cells (DC). MUTZ-3, a myeloid leukemia cell line, responds to GM-CSF, IL-4 and TNF-alpha, and acquires a phenotype similar to immature monocyte-derived DC (MoDC). In the present study, MUTZ-3-derived DC (MuDC) showed high level expression of HLA class II molecules, CD80 and CD86, and were able to function as potent antigen presenting cells as previously reported. Interestingly, MuDC maturation was induced by CD40-mediated stimulation, but not by LPS stimulation. We analyzed CCR1, CCR7 and Toll-like receptor (TLR) expressions in MuDC, and measured IL-10 and IL-12 production after maturation stimuli. Although MuDC expressed the mRNA for TLR4, a major component of the LPS receptor system, they did not show an enhanced level of CCR7 or cytokine production after LPS stimulation. In contrast, they responded to CD40 stimulation, which resulted in increased levels of CD83, CD86 and CCR7. Moreover, while LPSstimulated MoDC could potently stimulate NK cells in a DC-NK cell co-culture, LPS-stimulated MuDC failed to stimulate primary NK cells. Taken together, our findings suggest that, although MuDC express TLR4, unlike TNF-alpha and IL-1beta, LPS does not stimulate MuDC to acquire mature phenotypes, and they may have impaired activity to initiate innate immune response.


Subject(s)
CD40 Antigens/metabolism , B7-1 Antigen/metabolism , B7-2 Antigen/metabolism , Blotting, Western , CD40 Ligand/metabolism , Cell Differentiation , Cell Line, Tumor , Coculture Techniques , Dendritic Cells/drug effects , Enzyme-Linked Immunosorbent Assay , Fluorescein-5-isothiocyanate , Fluorescent Antibody Technique, Indirect , Fluorescent Dyes , Humans , Interleukin-10/analysis , Interleukin-12/analysis , Killer Cells, Natural/metabolism , Leukemia, Myeloid/pathology , Lipopolysaccharides/pharmacology , Mitogen-Activated Protein Kinase 3/metabolism , RNA, Messenger/metabolism , Reverse Transcriptase Polymerase Chain Reaction , Toll-Like Receptor 4/metabolism , Tumor Necrosis Factor-alpha/pharmacology , p38 Mitogen-Activated Protein Kinases/metabolism
4.
Mem. Inst. Oswaldo Cruz ; 96(suppl): 103-105, Sept. 2001.
Article in English | LILACS | ID: lil-295885

ABSTRACT

In this communication the authors analyzed the pattern of expression of IFN-gamma as a surrogate type 1 response in different clinical forms of schistosomiasis in response to stimulation involving T-cell dependent and T-cell independent pathways, to investigate which pathways were functional in human schistosomiasis, and to further characterize the nature of Th1 response impairment in this parasitic disease


Subject(s)
Humans , CD40 Antigens/physiology , CD40 Ligand/physiology , Interferon-gamma/biosynthesis , Schistosomiasis mansoni/metabolism , Staphylococcus aureus/physiology , CD40 Antigens/metabolism , CD40 Ligand/metabolism , Schistosomiasis mansoni/immunology , T-Lymphocytes, Helper-Inducer/metabolism
5.
Braz. j. med. biol. res ; 34(6): 779-84, Jun. 2001. tab, graf
Article in English | LILACS | ID: lil-285853

ABSTRACT

The purpose of the present study was to investigate the expression (mRNA) of CD40 ligand (CD40L), interferon-gamma (IFN-gamma) and Fas ligand (FasL) genes in human cardiac allografts in relation to the occurrence of acute cardiac allograft rejection as well as its possible value in predicting acute rejection. The mRNA levels were determined by a semiquantitative reverse transcriptase-polymerase chain reaction method in 39 samples of endomyocardial biopsies obtained from 10 adult cardiac transplant recipients within the first six months after transplantation. Biopsies with ongoing acute rejection showed significantly higher CD40L, IFN-gamma and FasL mRNA expression than biopsies without rejection. The median values of mRNA expression in biopsies with and without rejection were 0.116 and zero for CD40L (P<0.003), 0.080 and zero for IFN-gamma (P<0.0009), and 0.156 and zero for FasL (P<0.002), respectively. In addition, the levels of IFN-gamma mRNA were significantly increased 7 to 15 days before the appearance of histological evidence of rejection (median of 0.086 in pre-rejection biopsies), i.e., they presented a predictive value. This study provides further evidence of heightened expression of immune activation genes during rejection and shows that some of these markers may present predictive value for the occurrence of acute rejection.


Subject(s)
Humans , Adult , Endocardium/metabolism , Graft Rejection/immunology , Heart Transplantation/immunology , Myocardium/metabolism , Proteins/metabolism , RNA, Messenger/analysis , Biopsy , CD40 Ligand/genetics , CD40 Ligand/metabolism , Endocardium/pathology , Gene Expression , Interferon-gamma/genetics , Interferon-gamma/metabolism , Membrane Glycoproteins/genetics , Membrane Glycoproteins/metabolism , Myocardium/pathology , Predictive Value of Tests , Proteins/genetics , Reverse Transcriptase Polymerase Chain Reaction , Transplantation, Homologous
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