• Journal of Physiology & Pathology in Korean Medicine
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• v.23 no.5
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• pp.1106-1115
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• 2009

Mori Ramulus has multiple applications in Korean traditional medicine prescription because it has antioxidant and anti-inflammatory effects by reducing macrophage activities. Yet, no studies on the anti-arthritic activity of EMR (extract of Mori Ramulus) have been reported in vitro and in vivo. Rheumatoid arthritis (RA) is a systemic autoimmune disease with chronic inflammation characterized by hyperplasia of synovial cells in affected joints, which ultimately leads to the destruction of cartilage and bone. Because collagen-induced arthritis (CIA) is similar to RA in pathological symptoms and immune reactions, there have been several reports concerning RA using CIA mouse model. Here, we investigated the effects of Mori Ramulus on RA using CIA mice. The importance of CD4+ Th1 cells in RA progress was previously indicated and studies further showed that Th17 cells play a prime role in severity of disease. Accordingly, the present study was focused on CIA associated with CD4+ Th1 cells and Th1 7 cells. DBA/1OlaHsd mice were immunized with bovine type II collagen (CII). After a second collagen immunization, mice were treated with EMR once a day for 4 weeks. The severity of arthritis within the paw joints was evaluated by histological assessment of cartilage destruction and pannus formation. Immune cells in peripheral blood mononuclear cells (PBMC), draining lymph node (DLN) and paw joints, cytokine production and gene expression were assessed from CIA mouse using ELISA, FACS and real-time PCR analysis. Administration of EMR significantly suppressed the progression of CIA and inhibited the production of TNF-$\alpha$, IL-6 and IL-17 in the serum. The erosion of cartilage was dramatically reduced in mouse knees after treatment with EMR. In conclusion, our results demonstrate that EMR significantly suppressed the progression of CIA and that this action was mediated by the decreased production of TNF-$\alpha$, IL-6, IL-17 and collagen II-specific antibody in the serum. EMR suppressed Th17 cells and reduced level of IL-6 via B cell suppression, and thus, the levels of autoantibodies produced from B cells were decreased. Furthermore, EMR suppressed NKT cells which directly stimulate B cells and develop imbalance of Th1/Th2 cell. Oral administration of EMR (100 mg/kg or 200 mg/kg) significantly suppressed the progression of CIA, which is comparable to that of methotrexate (MTX, 0.3 mg/kg) used as a positive control. We are currently studying the mechanism underlying the therapeutic role for EMR in CIA mice.

• Journal of Life Science
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• v.18 no.11
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• pp.1471-1478
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• 2008

In this study, nitric oxide (NO) production in a macrophage-lymphocyte co-culture system was used to assess the cytokine producing capability of cells during endotoxin tolerance in mice. Incubation of peritoneal macrophages with interferon-$\tau$ (IFN-$\tau$) in the presence of lipopolysaccharide (LPS) augmented NO synthesis. Exogenous tumor necrosis factor-$\alpha$(TNF-$\alpha$) could also replace LPS for the stimulation of NO production. Macrophages co-cultured with splenic lymphocytes showed augmented NO synthesis by LPS alone. However, pretreatment of mice with 2.5 mg/kg LPS completely prevented the lethality and the increase of blood TNF-$\alpha$ and IFN-$\tau$ after the second challenge with a lethal dose of LPS. In addition, when macrophages prepared from LPS-tolerant mice were co-cultured with normal splenocytes, LPS also could not induce the production of NO, even in the presence of exogenous TNF-$\alpha$. Moreover, when normal macrophages were co-cultured with splenocytes obtained from LPS-tolerant mice, stimulation with LPS could not evoke the NO production enhancement. However, this down-regulation was able to reverse by exogenous IFN-$\tau$ or concanavalin A (ConA), a stimulator of IFN-$\tau$ production. Our results indicate that not only macrophages but also lymphocytes contribute to LPS tolerance. As INF-$\tau$ can enhance the expression of TNF-$\alpha$, the decrease of INF-$\tau$synthesis from lymphocytes may orchestrate with the decrease of TNF-$\alpha$ synthesis from LPS-tolerant macrophages for the production of tolerant state and the prevention of excessive inflammation. Therefore, LPS tolerance may be exploited for prophylaxis of severe sepsis in patients at risk.

• Journal of Periodontal and Implant Science
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• v.27 no.2
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• pp.425-441
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• 1997

The neuropeptide substance P(SP) has been implicated in the mediation of inflammation and immune-mediated disease such as arthritis. Recently, it was reported that SP was markedly increased around the blood vessels in inflamed gingiva as well as in close association with the inflammatory cell infiltrate. These results support that SP may contribute to the pathophysiology of neuronal inflammation in human periodontal tissues. SP may regulate inflammatory/immune responses by stimulating the proliferation of human T cells, differentiation and antibody-secreting potential of B cells, macrophage respiratory burst, connective tissue proliferation, and the secretion of cytokines from monocytes and T cells. Here, I studied potential role of SP as a costimulatory chemical signal in inflammatory/immune responses, by determining the released proinflammatory cytokines such as $MIP-1{\alpha}$, $IL-1{\beta}$, and IL-6 from culture supernatants of homogeneous immune cell lines. Serum free cell supernatants were concentrated with TCA precipitation, fractionated with SDS-PAGE, and subjected into western blot analysis. Among 15 cell lines tested, macrophage/monocyte cell line RAW264.7 and WRl9m.1 showed the highest level of induction of $MIP-1{\alpha}$ when stimulated with LPS. Discrete IL-6 bands with multiple forms of molecular mass were detected from supernatants of B cell lines A20(32kDa), Daudi(32, 35kDa), and SKW6.4(29kDa), which were expressed constitutively. $IL-1{\beta}$ could not be detected by the method of western blot analysis from supernatants of all cell lines tested except RAW264.7, WRl9m.1, and erythroid cell line K562 which showed the least amount of $IL-{\beta}$ secretion. SP $10^{-9}M$ with suboptimal dose of LPS treatment showed synergistic induction of $MIP-1{\alpha}$ release from RAW264.7 or WR19m.1, and also IL-6 release from A20, but this synergism is not the case in costimulation of RAW264.7 or WRl9m.1 with SP $10^{-9}M$ and TPA. Although treatment of T cell line CTLL-R8 with SP $10^{-7}M$ or PHA+TPA induced modest level of $MIP-1{\alpha}$ secretion, synergism was not observed when they are applied together. These findings all together suggest the possibility of a regulatory role of SP in inflammatory/immune reaction through differential modulation of bioactivities of other chemical cosignals.

• The Journal of Pediatrics of Korean Medicine
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• v.25 no.1
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• pp.1-27
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• 2011

Objectives: The purpose of this study is to investigate the effects of Yijungtang(YJT) on atopic dermatitis in an in-vitro and in-vivo experiment using a RBL-2H3 mast cells and a NC/Nga atopic dermatitis mouse. Methods: In-vitro experiment, IL-4, IL-13 mRNA expression were evaluated by a real-time PCR, IL-4, IL-13 production by ELISA and transcription factor as GATA-1, GATA-2, NF-AT1, NF-AT2, AP-1 and NF-kB by western blotting. In-vivo experiment, clinical skin score we evaluated by, hematology and Serum total IgE and IgG1 of NC/Nga atopic dermatitis mouse, cytokine level, total number of cell, Immunohistochemical staining and Histological features of auxiliary lymph node(ALN), draining lymph node(DLN), peripheral blood mononuclear cells(PBMCs) and dorsal skin tissue in NC/Nga mouse. Results: YJT decreased IL-4, IL-13 mRNA expression, IL-4, IL-13 production and prominently decreased the expression of mast cell specific transcription factors including GATA-2, NF-AT2, c-Fos and NF-kB. YJT oral administration reduced the levels of skin severity scores. It also decreased the level of inflammatory cytokines such as IL-5, IL-13, histamine and IgE in the serum. It elevated IFN-gamma level in the spleenocyte culture supernatant but decreased. $CD3e^+$, $CD19^+$, $CD4^+$, $CD8^+$, $CD3e^+CD69^+$, $CD11b^+Gr-1^+$, $CCR3^+$ in the PBMCs, $CD4^+$, $CD8^+$, $CD3e^+CD69^+$, $B220^+CD23^+$ in the ALN, $CD4^+$, $CD3e^+CD69^+$ in the ALN and $CD4^+$, $CD11b^+Gr-1^+$ in the dorsal skin. Histological examination showed that infiltration levels of immune cells in the skin of AD-induced NC/Nga mice were much improved by YJT oral administration. Conclusions: The anti-allergic activities of YJT may be mediated by down-regulation of Th2 cytokines, such as IL-4 and IL-13, through the regulation GATA-2, NF-AT2 and NF-kB transcription factors in mast cells. YJT would be regulate molecular mediators and immune cells which are functionally associated with atopic dermatitis induced in NC/Nga mice, and may play an important role in recovering AD symptoms.

• Korean Journal of Acupuncture
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• v.26 no.3
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• pp.55-68
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• 2009

Objectives : To investigate the anti-inflammatory effects of Scutellaria baicalensis Georgi pharmacopuncture in lipopolysaccharide (LPS)-induced inflammatory rat model. Methods : Sprague-Dawley rats were divided into 4 groups; LPS control (n=6), LPS+Scutellaria baicalensis Georgi pharmacopuncture at BL23 (n=6, BL23), LPS+Scutellaria baicalensis Georgi pharmacopuncture at CV12 (n=6, CV12), and LPS+Scutellaria baicalensis Georgi pharmacopuncture at GV4 (n=6, GV4). Pharmacopuncture was given every two days for 4 weeks followed by inflammation induction by intraperitoneal LPS injection (5mg/kg). Blood, liver tissue, and peritoneal lavage fluid were taken and proinflammatory cytokines and other related factors were analysed. Results : For proinflammatory cytokines, CV12 pharmacopuncture group was significantly different compared with the control group in plasma IL-$1{\beta}$, IL-6, TNF-$\alpha$, and IL-10 5 h after LPS injection (P<0.05). For plasma IL-$1{\beta}$ and IL-6, CV12 pharmacopuncture group also showed significant difference at 2 h compared with the control (P<0.05). GV4 pharmacopuncture group was significantly different compared with the control at 5 h in plasma IL-$1{\beta}$, IL-6, and TNF-$\alpha$ and at 2 h in IL-10 (P<0.05). Liver cytokines were analyzed at 5 h after LPS injection; only CV12 pharmacopuncture group showed significant difference in IL-$1{\beta}$ (P<0.05) and others including IL-6, TNF-$\alpha$, and IL-10 had no difference compared with the control group. CD4/CD8 ratio and the phagocytic activities of polymorphonuclear neutrophils were not different from those of control group in all pharmacopuncture groups (P>0.05). Plasma NO3-/NO2- and intracellular adhesion molecule-1 of CV12 pharmacopuncture group were significantly lower than that of the control group (P<0.05). In the plasma concentration of prostaglandin E2, all 3 pharmacopuncture groups had significantly lower values than that of the control group (P<0.05), but there was no difference among pharmacopucnture groups. Monocyte chemoattractant protein-1 and cytokine-induced neutorphil chemoattractant-1 in peritoneal lavage fluid was significantly decreased in CV12 pharmacopuncture group compared with the control group (P<0.05). Conclusions : These results indicate that Scutellaria baicalensis Georgi pharmacopuncture at CV12 may have a potent anti-inflammatory effect in an LPS-induced inflammatory rat model.

• The Korean Journal of Physiology and Pharmacology
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• v.17 no.3
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• pp.209-216
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• 2013

Soybean polyunsaturated phosphatidylcholine (PC) is thought to exert anti-inflammatory activities and has potent effects in attenuating acute renal failure and liver dysfunction. The aim of this study was to investigate the effects of PC in protecting multiple organ injury (MOI) from lipopolysaccharide (LPS). Six groups of rats (N=8) were used in this study. Three groups acted as controls and received only saline, hydrocortisone (HC, 6 mg/kg, i.v.) or PC (600 mg/kg, i.p.) without LPS (15 mg/kg, i.p.) injections. Other 3 groups, as the test groups, were administered saline, HC or PC in the presence of LPS. Six hours after the LPS injection, blood and organs (lung, liver and kidney) were collected from each group to measure inflammatory cytokines and perform histopathology and myeloperoxidase (MPO) assessment. Serum cytokines (TNF-${\alpha}$, IL-6 and IL-10) and MPO activities were significantly increased, and significant histopathological changes in the organs were observed by LPS challenge. These findings were significantly attenuated by PC or HC. The treatment with PC or HC resulted in a significant attenuation on the increase in serum levels of TNF-${\alpha}$ and IL-6, pro-inflammatory cytokines, while neither PC nor HC significantly attenuated serum levels of IL-10, anti-inflammatory cytokine. In the organs, the enhanced infiltration of neutrophils and expression of ED2 positive macrophage were attenuated by PC or HC. Inductions of MPO activity were also significantly attenuated by PC or HC. From the findings, we suggest that PC may be a functional material for its use as an anti-inflammatory agent.

• BMB Reports
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• v.33 no.6
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• pp.454-462
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• 2000

Interleukin-4(IL-4) is known to be a major cytokine regulating immunoglobulin E(IgE) response by the induction of IgE production and type II IgE receptor(IgER II: CD23) expression. Recently, however, the role of neuroendocrine factors has been implicated in modulating the IgE response. Among various neuroendocrine growth factors, we investigated the effects of the insulin-like growth factor-1(IGF-1) since IL-4 and IGF-1 share common intracellular signaling molecules, such as the insulin receptor substrate-1/2(IRS-1/2) to induce a specific cellular response. In the human peripheral blood mononuclear cell (PBMC) cultures, IGF-1 was capable of inducing a substantial level of IgE production in a dose-dependent manner. It also noticeably upregulated the IL-4-induced or IL-4 plus anti-CD40-induced IgE production. Similarly, the IGF-1-induced IgE production was enhanced by IL-4 or anti-CD40 in an additive manner, which became saturated at high concentrations of IGF-1. Although IGF-1 alone did not induce IgER II (CD23) expression, it augmented the IL-4-induced surface CD23 expression in a manner similar to the action of anti-CD40. These results imply that IGF-1 is likely to utilize common signaling pathways with IL-4 and anti-CD40 to induce IgE and IgER II expression. In support of this notion, we observed that IGF-1 enhanced the IL-4-induced signal transducers and activators of transcription 6(STAT6) activation and independently induced $NF-{\kappa}B$ activation. Both of these bind to the IgE(C) or IgER II (CD23) promoters. Together, our data suggest that IL-4 and IGF-1 work cooperatively to activate STAT6 and $NF-{\kappa}B$. This leads to the subsequent binding of these transcription factors to the $C{\varepsilon}$ and CD23 promoters to enhance the expression of IgE and IgER II. The observed differential ability of IGF-1 on the induction of IgE vs. IgER II is discussed based on the different structure of the two promoters.

• Journal of Life Science
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• v.21 no.12
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• pp.1726-1731
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• 2011

Human Stem Cell Factor (hSCF) is a cytokine that binds to the c-Kit receptor and plays an important role in hematopoiesis, spermatogenesis, and melanogenesis. To produce the human Stem Cell Factor (hSCF) recombinant protein, we constructed a germline transgenic silkworm using the piggyback vector. The expression of the hSCF gene was driven by the Drosophila heat shock protein 70 (dHsp70) promoter. 3XP3 promotor-driven EGFP was used as a marker which allowed us to rapidly distinguish the transgenic silkworm. A mixture of the donor and helper vector was micro-injected into 1,020 eggs of bivoltin silkworms, Keomokjam. We obtained approximately 22 G1 broods that were EGFP-positive. The expression of the hSCF gene in the transgenic silkworm was analyzed by SDS-PAGE and immunoblotting. Also, analysis of insertion sites into the silkworm genome using inverse PCR showed that exogenous DNA was inserted into the transgenic silkworm genome. These results show that successfully constructed transgenic silkworm expresses the hSCF recombinant protein.

• Korean Journal of Environmental Biology
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• v.27 no.1
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• pp.88-94
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• 2009

The process of atherosclerosis begins through secretion of inflammatory cytokine or adhesion of leukocyte from damage in blood vessels and transmigration. This study was conducted to investigate the effects of delphinidin chloride (DC) in the initial process of atherosclerosis on the expression of ICAM-1 (Intracellular Adhesion Molecule-1) and VCAM-1 (Vascular Adhesion Molecule-1) related to adhesion of leukocyte at the HUVEC (human umbilical vein endothelial cell line. As a result, cell growth inhibition rate at 50 ${\mu}M$ was respectively 4, 3 and 5% without cell toxicity. As a result of morphological observation monocyte-endothelial cell adhesion assay and optical microscope carried out to measure attachment of mononuclear cells to endothelial cells induced by Tumor necrosis factor-alpha (TNF-$\alpha$) at concentrations without cell toxicity, DC concentration-dependently suppressed attachment. When effects on the expression of VCAM-1 and ICAM-1, cell adhesion molecules induced from endothelial cells by TNF-$\alpha$, were comparatively analyzed using western blot analysis and RT-PCR methods, protein of VCAM-1 and ICAM-1 and expression at the level of mRNA were concentration-dependently reduced. Taken together, the results of this studies provide evidence that DC possess an anti-metastatic activity.

• Journal of Ginseng Research
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• v.39 no.1
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• pp.29-37
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• 2015

Background: Panax ginseng (i.e., ginseng) root is extensively used in traditional oriental medicine. It is a modern pharmaceutical reagent for preventing various human diseases such as cancer. Ginsenosidesd-the major active components of ginsengd-exhibit immunomodulatory effects. However, the mechanism and function underlying such effects are not fully elucidated, especially in human monocytes and dendritic cells (DCs). Methods: We investigated the immunomodulatory effect of ginsenosides from Panax ginseng root on $CD14^+$ monocytes purified from human adult peripheral blood mononuclear cells (PBMCs) and on their differentiation into DCs that affect $CD4^+$ T cell activity. Results: After treatment with ginsenoside fractions, monocyte levels of tumor necrosis factor (TNF)-${\alpha}$, interleukin (IL)-6, and IL-10 increased through phosphorylation of extracellular signal-regulated kinase (ERK)1/2 and c-Jun N-terminal kinase (JNK), but not p38 mitogen-activated protein kinase (MAPK). After treatment with ginsenoside fractions, TNF-${\alpha}$ production and phosphorylation of ERK1/2 and JNK decreased in lipopolysaccharide (LPS)-sensitized monocytes.We confirmed that DCs derived from $CD14^+$ monocytes in the presence of ginsenoside fractions (Gin-DCs) contained decreased levels of the costimulatory molecules CD80 and CD86. The expression of these costimulatory molecules decreased in LPS-treated DCs exposed to ginsenoside fractions, compared to their expression in LPS-treated DCs in the absence of ginsenoside fractions. Furthermore, LPS-treated Gin-DCs could not induce proliferation and interferon gamma (IFN-${\gamma}$) production by $CD4^+$ T cells with the coculture of Gin-DCs with $CD4^+$ T cells. Conclusion: These results suggest that ginsenoside fractions from the ginseng root suppress cytokine production and maturation of LPS-treated DCs and downregulate $CD4^+$ T cells.