• IMMUNE NETWORK
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• v.7 no.4
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• pp.173-178
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• 2007

Background: We studied the role for expression of CD40 and CD40L by CD4 and CD8 T cells in the generation of CD8 T cell response to minor histocompatibility antigen, H60. H60 is a cellular antigen to which CD8 responses require CD4 T cell help. Methods: CD40- or CD40L-deficient mice were adoptively transferred with normal CD4 or CD8 T cells or with memory CD4 or CD8 T cells, and were immunized with male H60 congenic splenocytes to induce CD8 T cell response to H60. Peripheral blood CD8 T cell from the immunized mice were stained with the H60 tetramer. Results: CD8 T cell response to H60 was not induced in both CD40- and CD40L-deficient mice. Adoptive transfer of $CD40^{+/+}$ CD8 T cells into CD40-deficient mice did not compensate the defect in inducing CD8 T cell response to H60, while the H60-specific CD8 T cells were activated in the CD40-deficient mice that were adoptively transferred with $CD40^{+/+}$ CD4 T cells. Adoptive transfer of $CD40L^{+/+}$ CD4 T cells into CD40L-deficient mice induced primary CD8 T cell response for H60 and the presence of $CD40L^{+/+}$ CD4 T cells was required even for memory CD8 T cells response to H60. Conclusion: Our results suggest that the CD40-CD40L interaction mediates the delivery of CD4 T cell help to naive and memory H60-specific CD8 T cells. While the expression of CD40L by CD4 T cells is essential, signaling through CD40 on CD8 T cells is not required for the induction of CD8 T cell response to H60.

• IMMUNE NETWORK
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• v.7 no.2
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• pp.80-86
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• 2007

Background: CD40-activated B (CD40-B) cells might be an attractive source of autologous antigen-presenting cells (APCs) for immunotherapy due to the convenience to obtain from peripheral blood and expand in vitro. Moreover, CD40-B cells were found to be comparable with DCs in their capacity to raise antigen-specific CD8+ T cells. Here, we have established K562 cells expressing CD40L to expand CD40-activated B cells used for APCs. Methods: After activation of B cell by K562/CD40L, CD40-B cells were examined by counting B cell numbers. Surface expression of CD54, CD80, CD86 and HLA class II was measured by flow cytometry. The CD40-B cells were tested for its function as APC by mixed lymphocyte reactions (MLR) and by induction of T cell responses specific for pp65 peptide in vitro. Results: The expansion of B cells by K562/CD40L increased about 6-folds compared with anti-CD40 or K562. Furthermore, the expression of CD54, CD80, CD86 and HLA class II was up-regulated by K562/CD40L. B cells by K562/CD40L showed comparable antigen presentation activity with mature DCs as shown in MLR, INF-${\gamma}$ ELISPOT assay. Conclusion: These results suggest that K562/CD40L could be used to generate activated B cells as potent APCs which could be useful for cellular vaccination and adoptive immunotherapy.

• IMMUNE NETWORK
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• v.4 no.3
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• pp.143-154
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• 2004

Background: CD27 is recently known as a memory B cell marker and is mainly expressed in activated T cells, some B cell population and NK cells. CD27 is a member of tumor necrosis factor receptor family. Like CD40 molecule, CD27 has (P/S/T/A) X(Q/E)E motif for interacting with TNF receptor-associated factors (TRAFs), and TRAF2 and TRAF5 bindings to CD27 in 293T cells were reported. Methods: To investigate the CD27 signaling effect in B cells, human CD40 extracellular domain containing mouse CD27 cytoplamic domain construct (hCD40-mCD27) was transfected into mouse B cell line CH12.LX and M12.4.1. Results: Through the stimulation of hCD40-mCD27 molecule via anti-human CD40 antibody or CD154 ligation, expression of CD11a, CD23, CD54, CD70 and CD80 were increased and secretion of IgM was induced, which were comparable to the effect of CD40 stimulation. TRAF2 and TRAF3 were recruited into lipid-enriched membrane raft and were bound to CD27 in M12.4.1 cells. CD27 stimulation, however, did not increase TRAF2 or TRAF3 degradation. Conclusion: In contrast to CD40 signaling pathway, TRAF2 and TRAF3 degradation was not observed after CD27 stimulation and it might contribute to prolonged B cell activation through CD27 signaling.

• BMB Reports
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• v.30 no.6
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• pp.431-437
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• 1997

Since the role of CD40 on the interleukin-4(IL-4) -induced B cell activation has been strongly implicated in the agumentation of IgE production and response, we have investigated the intracelluar signaling pathways utilized by IL-4 and CD40 for type II IgE receptor (CD23) expression. IL-4 and anti-CD40 antibody treatment of human B cells, independently caused a rapid induction of CD23 gene activation within 2 h. There was a noticeable synergism between the action of the two agents inducing CD23 expression: the addition of anti-CD40 to the IL-4-treated culture significantly agumented the IL-4-induced CD23 on both mRNA and surface protein levels, and the inclusion of IL-4 in the anti-CD40-treated cells caused a further increase of CD23 expression far above the maximal level induced by anti-CD40. Protein tyrosine kinase (PTK) inhibitors effectively suppressed the both IL-4- and anti -CD40-induced CD23 expression. whereas protein kinase C (PKC) inhibitors had no effects. Electrophoretic mobility shift assays (EMSA) have shown that IL-4 and anti-CD40 induce the activation of NF-IL-4 and $NF-_{K}B$, respectively, binding to the CD23 promoter, both in a PKC-independent and PTK-dependent manner. These data suggest that the synergistic activation of CD23 gene expression by IL-4 and anti-CD40 is mediated by co-operative action of distinct nuclear factors. each of which is rapidly activated via PKC-independent and PTK-dependent process.

• The Korean Journal of Physiology and Pharmacology
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• v.7 no.5
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• pp.255-259
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• 2003

Dendritic cells (DCs) play a critical role in various immune responses involving $CD4^+$ T cells and have been used to generate anti-tumor immunity. Chemotherapy induces severe side effects including immunosuppression in patients with cancer. Although immunosuppression has been studied, the effects of anticancer drugs on DCs are not fully determined. In this study, we demonstrated that CD40 activation strongly protected DCs from 5-fluorouracil (5-FU) or mitomycin C-induced apoptosis. DCspecific surface markers, including CD11c and major histocompatibility complex (MHC) class II, were used for identifying DCs. CD 40 activation with anti-CD40 mAb significantly enhanced the viability of DCs treated with 5-FU or mitomycin C, assayed by MTT (3-[4,5-dimethylthiazol-2-yl]-2,5-diphenyltetrazolium bromide). Fluorescence staining and analysis clearly confirmed the enhancing effect of anti-CD40 mAb on the viability of DCs, suggesting that CD40 activation may transduce critical signals for the viability of DCs. Annexin V staining assay showed that CD40 significantly protected DCs from 5-FU or mitomycin C-induced apoptosis. Taken together, this study shows that CD40 activation with anti-CD40 mAb has strong anti-apoptosis effect on DCs, suggesting that CD40 activation may overcome the immunosuppression, especially downregulation of number and function of DCs in chemotherapy-treated cancer patients.

• IMMUNE NETWORK
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• v.8 no.3
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• pp.75-81
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• 2008

Background: Platelets take part in repairing the lesions of endothelial damage. To understand the molecular mechanism of this process, we tested the hypothesis that CD154 expressed on activated platelets stimulates proliferation of human endothelial cells. Methods: The expression levels of CD154 and CD40 on platelets and endothelial cells, respectively, were measured by flow cytometry and confocal microscopy. Function-blocking monoclonal antibody against CD154 was developed after immunization with CD154-transfected L cells. Results: An anti-CD40 agonist antibody and soluble CD154 both induced significant proliferation of endothelial cells. In addition, a function-blocking anti-CD154 antibody inhibited the platelet-induced proliferation of endothelial cells, indicating that the CD154-CD40 pathway is involved in these cellular interactions. An anti-VEGF antibody failed to inhibit the proliferation. This, in addition to the fact that very small amounts of VEGF are released from platelets or endothelial cells, suggests that VEGF does not play an important role in the platelet-stimulated proliferation of endothelial cells. Conclusion: Our results indicate that platelets induce proliferation of endothelial cells by CD154-CD40 interactions independently of VEGF.

• BMB Reports
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• v.48 no.1
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• pp.54-59
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• 2015

IL-10-producing B (Breg) cells regulate various immune responses. However, their phenotype remains unclear. CD40 expression was significantly increased in B cells by LPS, and the Breg cells were also enriched in $CD40^{hi}CD5^+$ B cells. Furthermore, CD40 expression on Breg cells was increased by IL-10, CD40 ligand, and B cell-activating factor, suggesting that $CD40^{hi}$ is a common phenotype of Breg cells. LPS-induced CD40 expression was largely suppressed by an anti-IL-10 receptor antibody and in IL-$10^{-/-}CD5^+CD19^+$ B cells. The autocrine effect of IL-10 on the CD40 expression was largely suppressed by an inhibitor of JAK/STAT3. In vivo, the LPS treatment increased the population of $CD40^{hi}CD5^+$ Breg cells in mice. However, the population of $CD40^{hi}CD5^+$ B cells was minimal in IL-$10^{-/-}$ mice by LPS. Altogether, our findings show that Breg cells are largely enriched in $CD40^{hi}CD5^+$ B cells and the autocrine effect of IL-10 is critical to the formation of $CD40^{hi}CD5^+$ Breg cells.

• IMMUNE NETWORK
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• v.2 no.3
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• pp.142-149
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• 2002

Background: Our previous study showed that purified protein derivative (PPD)-stimulated pleural mononuclear cells (PMC) from tuberculous pleurisy (Tbp) produced significantly more $IFN-{\gamma}$ (10- to 70-fold) after in vitro PPD stimulation than freshly isolated pleural cells from malignant pleurisy. The present study was designed to determine whether blocking the CD40-CD40 ligand (CD40L) interaction decreases $IFN-{\gamma}$ production by altering IL-12 levels. Methods: IL-12 and $IFN-{\gamma}$ production after neutralizing anti-CD40L antibody treatment was compared to the efficacy of anti-CD80, anti-CD86, and a combination of anti-CD80 and CD86 (CD80+86) monoclonal antibodies (mAb). These activities were measured by enzyme-linked immunosorbent assays (ELISAs) and reverse transcription-polymerase chain reaction (RT-PCR), after in vitro stimulation with PPO antigen (Ag). Results: Neutralization of CD80, CD86 and CD80+86 did not decrease $IFN-{\gamma}$ and IL-12 production in Tbp-PMC, whereas neutralization of CD40L significantly depressed IL-12 p40 and $IFN-{\gamma}$. In addition, neutralization of CD40L completely inhibited IL-12 p40 and $IFN-{\gamma}$ mRNA expression. Conclusion: The CD40-CD40L interaction might play a major role in IL-12 and $IFN-{\gamma}$ production in Tbp-PMC, thus contributing to protective immunity in human tuberculosis.

• The Korean Journal of Malacology
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• v.30 no.1
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• pp.41-49
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• 2014

This study was conducted to investigate the effects of cadmium (Cd) exposure on biochemical factors in the hemolymph and hepatopancreas of the abalone, Haliotis discus hannai. The abalone were exposed to 0, 5, 10, 20 and 40 ${\mu}g/L$ Cd for 4 weeks. The phenoloxidase (PO) activity was decreased in hemolymph of abalone exposed to 40 Cd ${\mu}g/L$ for 4 weeks compared to the control (P < 0.05). The hemolymph enzymes, alkaline phosphatase (ALP), aspartate aminotransferase (AST) and alanine aminotransferase (ALT) activities were markedly elevated in 40 Cd ${\mu}g/L$ after 4 weeks. The hemolymph calcium concentrations were significantly decreased in 20 and 40 Cd ${\mu}g/L$ for 4 weeks. Hepatopancreas superoxide dismutase (SOD) and catalase (CAT) activities were significantly increased by Cd. SOD was increased in both 20 and 40 Cd ${\mu}g/L$ and CAT, in 40 Cd ${\mu}g/L$ after 2 weeks (P < 0.05). These results suggested that the abalone SOD and CAT including PO may serve as a protective mechanism against oxidative stress by Cd. We conclude that a Cd concentration, 40 ${\mu}g/L$ in water may curtail hemolymph homeostasis and anti-oxidative reactions in abalone hepatopancreas. From these results, these biochemical factors may represent a convenient method of monitoring heavy metal pollution in coastal areas.

• Journal of the Korean Society of Food Science and Nutrition
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• v.25 no.4
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• pp.575-580
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• 1996

The purpose of this study was to investigate the effect of dietary vitamin E levels on the enzymes such as superoxide dismutase(SOD), glutathione peroxidase(GSH-Px) and glutathione S-transferase (G57) involved in antioxidative defense system and lipid peroxidation in brain of cadmium administered rats. Sprague-Dawely male rats weighing $60\pm5g$ were divided into control and experimental groups. The experimental groups were divided into Cd-0E(vitamin E free diet), Cd-40E(40mg vitamin E/kg diet) and Cd-400E(400mg vitamin E/kg diet) according to the level of vitamin E supplementation. After each group was fed diet ad libitum for 2 or 4 weeks, 2.5mg cadmium per kg body weight was injected intraperitoneally once a day for 4 days. The rats were sacrificed for examination on the next day after the last injection of cadmium. The results are as follows: SOD activities of rat brain were lower in Cd-0E, but had a similar tendency to Cd-40E, Cd-400E groups compared with control group. GSH-Px acivities of rat brain were decreased in Cd-400E, Cd-40E and Cd-0E groups. GST activities of rat brain were decreased in Cd-0E, Cd-40E groups and not significantly different in Cd-400E compared with control group. Thiobarbituric acid reactive substances(TBARS) of rat brain was increased in Cd-0E, Cd-40E, Cd-400E in that order, TBARS was lower in Cd-40E, and Cd-400E by 28.8% and 44%, respectively, than Cd-0E group. The present result suggests that high level of dietary vitamin E protects against lipid peroxidative damage in rat induced by cadmium.