• Biomolecules & Therapeutics
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• v.2 no.3
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• pp.236-243
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• 1994

A splenocyte culture system supplemented with liver microsomes was developed to detect immunotoxic chemicals which require metabolic activation using cyclophosphamide as a positive standard. When liver microsomes were added to splenocyte cultures isolated from female B6C3Fl mice, the proliferation of splenocytes by lipopolysaccharide (LPS) was increased and the proliferation by concanavalin A (Con A) was decreased. However, when compared with each corresponding control, cyclophophamide was successfully activated to metabolites capable of suppressing Iymphoproliferative responses. This suppression was clearly dependent upon the amounts of microsomes added and/or the concentration of cyclophosphamide exposed. In these cultures, the proliferation of splenocytes was suppressed when the cells were exposed to cyclophosphamide on the day of culture initiation. On the other hand, microsome was responsible for the increase in LPS mitogenicity and NADPH was responsible for the decrease in Con A mitogenicity. Finally, our present culture system was compared with the hepatocyte-splenocyte coculture system which we had developed earlier. We found that the hepatocyte-splenocyte coculture was better able to activate cyclophosphamide to metabolites capable of suppressing the antibody response to sheep erythrocytes. Although our present culture system was relatively poor to activate cyclophosphamide in cultures for antibody response, it will be useful as a simple screening method to detect suppression of certain in vitro immunotoxic parameters like LPS mitogenicity by chemicals which require metabolism.

• The Journal of Internal Korean Medicine
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• v.30 no.1
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• pp.9-23
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• 2009

Objectives : Asthma is a chronic inflammatory disorder of the airways by many cells such as mast cells, Th2 lymphocytes and eosinophile. The present study was aimed to evaluate the effects of Salviae miltiorrhizae (SM) on T cell cytokine production, mast cells. and eosinophils, Methods : We screened 13 herbs to find compounds with potential to control Th cytokine production. using concanavalin A (con A)-activated splenocyte cultures. Con A-activated $IFN-\gamma$ and IL-4 levels in supernatants of splenocyte cultures. Bone marrow derived mast cells (BMMC) were incubated with SM and then the expressions of membrane proteins of BMMC were analyzed by fluorescence activated cell sorter (FACS). BALB/c mice sensitized to ovalbumin (OVA) were challenged with aerosolized OVA for 6 weeks. During the last weeks some mice were treated with SM. Then eosinophils in bronchoalveolar lavage fluid (BALf) were counted and pathologic changes of lung tissue were observed with hematoxylin-eosin stain. Results : SM increased $IFN-\gamma$ level on splenocyte culture significantly. but had no significant effects on expressions of ICAM-1, CD62L, integrin $a_4$. c-kit, IL-3 receptors. CD11a, or IgE receptors of BMMC. SM treatment significantly inhibited eosinophil infiltrates in BALf and peribronchial lung inflammation. Conculusions : The present data suggested that SM may have an effect on Th cytokine secretion and eosinophils associated with asthma responses. Therefore SM might be of therapeutic value in treating asthma.

• Journal of Life Science
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• v.20 no.8
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• pp.1276-1280
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• 2010

Berberine has shown a number of beneficial effects, including anti-tumor, anti-inflammation, and vasodilatory effects. In this work we investigated the effects of berberine on the production of proinflammatory cytokines such as TNF-$\alpha$, IL-$1{\beta}$, and IL-6 in mice. The supernatants of cultured splenocytes exposed with berberine or berberine plus LPS were harvested to assay TNF-$\alpha$, IL-$1{\beta}$, and IL-6. The sera from the mice injected with berberine or berberine plus LPS were then isolated to assay these cytokines. The TNF-$\alpha$ production in mice splenocyte cultures exposed to berberine was inhibited compared to the PBS control. The sera from LPS plus berberine injected mice showed lower levels of TNF-$\alpha$ compared to those of LPS only injected mice. The IL-$1{\beta}$ production in mice splenocyte cultures exposed to berberine was inhibited at a high dose (3.0 ${\mu}g/ml$) compared to the PBS control. Also, the increase of IL-$1{\beta}$ by LPS exposure in splenocyte cultures was inhibited by a high dose of berberine. The IL-6 in splenocyte culture supernatants showed lower levels after berberine compared to the PBS control. Also, production of IL-6 after LPS exposure in splenocyte cultures was inhibited by a low dose of berberine (0.3 ${\mu}g/ml$). These findings suggest the probability that berberine down-regulates the production of proinflammatory cytokines such as TNF-$\alpha$, IL-$1{\beta}$, and IL-6.

• Biomolecules & Therapeutics
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• v.20 no.2
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• pp.196-200
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• 2012

Role of metabolism by intestinal bacteria in arbutin-induced immunotoxicity was investigated in splenocyte cultures. Following an incubation of arbutin with 5 different intestinal bacteria for 24 hr, its aglycone hydroquinone could be produced and detected in the bacterial culture media with different amounts. Toxic effects of activated arbutin by intestinal bacteria on lymphoproliferative response were tested in splenocyte cultures from normal mice. Lipopolysaccharide and concanavalin A were used as mitogens for B- and T-cells, respectively. When bacteria cultured medium with arbutin was treated into the splenocytes for 3 days, the medium cultured with bacteria producing large amounts of hydroquinone induced suppression of lymphoproliferative responses, indicating that metabolic activation by intestinal bacteria might be required in arbutin-induced toxicity. The results indicated that the present testing system might be applied for determining the possible role of metabolism by intestinal bacteria in certain chemical-induced immunotoxicity in animal cell cultures.

• Parasites, Hosts and Diseases
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• v.49 no.3
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• pp.303-308
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• 2011

This study investigated whether elevated host immune capacity can inhibit T. gondii infection. For this purpose, we used silk protein extracted from Bombyx mori cocoons as a natural supplement to augment immune capacity. After silk protein administration to BALB/c mice for 6 weeks, ratios of T lymphocytes ($CD4^+$ and $CD8^+$ T-cells) and splenocyte proliferative capacities in response to Con A or T. gondii lysate antigen (TLA) were increased. Of various cytokines, which regulate immune systems, Th1 cytokines, such as IFN-${\gamma}$, IL-2, and IL-12, were obviously increased in splenocyte primary cell cultures. Furthermore, the survival of T. gondii (RH strain)-infected mice increased from 2 days to 5 or more days. In a state of immunosuppression induced by methylprednisolone acetate, silk protein-administered mice were resistant to reduction in T-lymphocyte ($CD4^+$ and $CD8^+$ T-cells) numbers and the splenocyte proliferative capacity induced by Con A or TLA with a statistical significance. Taken together, our results suggest that silk protein augments immune capacity in mice and the increased cellular immunity by silk protein administration increases host protection against acute T. gondii infection.

• Journal of Life Science
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• v.25 no.10
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• pp.1176-1183
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• 2015

Ulmus macrocarpa has been used in Korean medicinal food material to physical disorder or tonic material. The purpose of the present study was to evaluate splenocyte life span expansion effects of Ulmus macrocarpa water extract (UMWE) in general cell culture condition. Splenocytes were handled in the presence of 100 μg/ml UMWE for several different time conditions. Live cells were detected with Hoechst 33,342 dye and cell survival molecules were identified through Western blot. Changes in level of cytokine synthesis were evaluated by ELISA. UMWE showed an effect on increased splenocyte survival. UMWE elevated slightly PI3K phosphorylation and ERK1/2 phosphorylation used at 48 hr and 96 hr. Moreover, Bcl-2 was elevated at 48 hr and 96 hr in UMWE-treated splenocytes. UMWE decreased caspase-3 level at 48 hr and 96 hr. ICAD protein increased at 48 hr culturing time. Hematopoietin IL-2 cytokine was down-regulated, however IL-4 hematopoietin cytokine was up-regulated in UMWE treated cell culture media. Increased IFN-γ levels were verified in the supernatant of UMWE-treated cells in all periods (48 hr and 96 hr). Increased patterns in the production of IL-12 cytokine occurred as compared with control after 48 and 96 hr in UMWE-treated-cell cultures. These results suggested that UMWE can prolong splenocyte life span by controlling various signal factors and cytokines.

• Parasites, Hosts and Diseases
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• v.25 no.2
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• pp.195-198
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• 1987

In order to test the function of Iymphocytes in Naegleria fowleri-nniected mice, the in nitro blastogenic response of splenocyte cultures to non-specific mitogens was studied. Concanavalin A and lipopolysaccharide stimulation were used as tests of T cell and B cell function. For the first 14 days following N. fowleri infection, Iymphoblastic transformation induced by T-cell mitogen was markedly reduced in comparison to the uninfected control mice. The blastogenic response to B-cell mitogen remained depressed in the infected mice up to 14 days after infection. The fluorescent antibody titers of sera of N. fowleri infected mice were between 1 : 4 and 1 : 32. The results suggest that there is a suppression of cell mediated immunity during the acute course of experimental Naegleria meningoencephalitis in mice.

• The Korea Journal of Herbology
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• v.25 no.2
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• pp.129-136
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• 2010

Objective : Sinapis alba L. (SA) is a korean traditional herbal medicine that is usually used to prevent or treat inflammatory diseases, such as respiratory infection and rheumatoid arthritis. However, the effects of SA supplementation in vitro on serum antibody levels, splenocyte and peritoneal macrophage immune responses have not yet been determined. In this study, we examined the effect of SA on the production of Th1/Th2 cytokines. Methods : Splenocytes were isolated from naive C57BL/6 mice. Cells were enriched for CD4+ cell populations by first staining the cells with anti-CD4 (BD PharMingen, Calif, USA). CD4+ T cells were selected on a (CS) column, and the flow-through was collected as CD4+ T cells. Isolated cells were activated by overnight incubation on 24-well plates coated with $1{\mu}g/mL$ anti-CD3, $1{\mu}g/mL$ anti-CD28 and with SA ($100{\mu}g/mL$). Primary macrophages were collected from the peritoneal cavities of mice (8-week-old female C57BL/6). The peritoneal macrophages were washed and plated with RPMI-1640 overnight for the experiments. After 48-hours cultures, samples were centrifuged at 2000 rpm for 10 minutes, and the supernatants were stored at $-80^{\circ}C$. Mouse IL-4, IFN-$\gamma$ and TNF-$\alpha$ were quantified using ELISA kits (BioSource International, Camarillo, Calif, USA) according to the manufacturer's protocols. Results : SA at 100ug/ml decreased the generation of Th1 cytokine (IFN-$\gamma$) by 0.5-fold. However, SA has no effect on Th2 (IL-4) production. Conclusions : These results suggest that SA may play an important role in the control of T-cell-mediated autoimmunity by down-regulation of Th1 cytokine (especially IFN-$\gamma$, TNF-$\alpha$). These data may contribute to the design of new immunomodulating treatments for a group of autoimmune diseases.