• Journal of Microbiology and Biotechnology
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• v.18 no.4
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• pp.686-694
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• 2008

The inhibitory effects of 5,6,3',5'-tetramethoxy 7,4'-hydroxyflavone (labeled as p7F) were elucidated on the productions of proinflammatory cytokines as well as inflammatory mediators in human synovial fibroblasts and macrophage cells. p7F inhibited IL-1${\beta}$ or TNF-${\alpha}$ induced expressions of inflammatory mediators (ICAM-1, COX-2, and iNOS). p7F also inhibited LPS-induced productions of nitric oxide and prostaglandin $E_2$ in RAW 264.7 cells. In order to investigate whether p7F would inhibit IL-1 signaling, p7F was added to the D10S Th2 cell line (which is responsive to only IL-1${\beta}$ and thus proliferates), revealing that p7F inhibited IL-1${\beta}$-induced proliferation of D10S Th2 cells in a dose-response manner. A flow cytometric analysis revealed that p7F reduced the intracellular level of free radical oxygen species in RAW 264.7 cells treated with hydrogen peroxide. p7F inhibited IkB degradation and NF-${\kappa}$B activation in macrophage cells treated with LPS, supporting that p7F could inhibit signaling mediated via toll-like receptor. Taken together, p7F has inhibitory effects on LPS-induced productions of inflammatory mediators on human synovial fibroblasts and macrophage cells and thus has the potential to be an anti-inflammatory agent for inhibiting inflammatory responses.

• Journal of Yeungnam Medical Science
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• v.40 no.1
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• pp.23-29
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• 2023

The pathological hallmark of rheumatoid arthritis (RA) is a synovial pannus that comprises proliferating and invasive fibroblast-like synoviocytes, infiltrating inflammatory cells, and an associated neoangiogenic response. Animal models have been established to study these pathological features of human RA. Spontaneous and induced animal models of RA primarily reflect inflammatory aspects of the disease. Among various induced animal models, collagen-induced arthritis (CIA) and collagen antibody-induced arthritis (CAIA) models are widely used to study the pathogenesis of RA. Improved transplantation techniques for severe combined immunodeficiency (SCID) mouse models of RA can be used to evaluate the effectiveness of potential therapeutics in human tissues and cells. This review provides basic information on various animal models of RA, including CIA and CAIA. In addition, we describe a SCID mouse coimplantation model that can measure the long-distance migration of human RA synoviocytes and cartilage destruction induced by these cells.

• Microbiology and Biotechnology Letters
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• v.23 no.5
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• pp.526-530
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• 1995

Phospholipase A$_{2}$ (PLA$_{2}$) is lipolytic enzyme that has known to be involved in inflammation. In the course of our screening for antiinflammatory compounds from natural products, a compound having PLA$_{2}$ inhibitory activities was isolated from the methanol extract of Umbilicaria esculenta. The compound was identified as lecanoric acid based on various NMR studies including DEPT, HETERO-COSY and HMBC experiments. Lecanoric acid inhibited human rheumatoid synovial PLA$_{2}$ activity with IC$_{50}$ of 0.17 mM and also exhibited antitumor activity (ED$_{50}$=2.7 $\mu $g/ml) against skin tumor cell line (LOX-IMVI).

• The Journal of Korean Medicine
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• v.31 no.4
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• pp.63-78
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• 2010

Objectives: This study was carried out to know the effects of Gwan-Jul-Bang-7 (hereafter referred to GJB-7) on the inhibition of arthritis induced by collagen on the mouse. Methods: To assess the effects of GJB-7 on mouse with arthritis induced by collagen II, we conducted several experiments such as analysis of cytotoxicity, hepatotoxicity, arthritis index, total cell number of draining lymph nodes and paw joints, value of immunocyte in PBMC (peripheral blood mononuclear cell), DLN (draining lymph node) and paw joint, measurement of cytokine and anti-collagen II, observation of the histological changes of joint. Results: 1. Cytotoxicity against HFC (human fibroblast cells) was not observed in any concentration and hepatotoxicity was not observed in the GJB-7 treated group. 2. The incidence of arthritis significantly decreased. 3. Total cell number of draining lymph nodes significantly increased and total cell number of paw joints significantly decreased. 4. The percentage of $CD8^+$ cells in PBMC (peripheral blood mononuclear cell) significantly increased. The percentage of $CD3^+/CD69^+$, and $CD3^+/CD49b^+$ cells in PBMC significantly decreased. 5. The percentage of $CD19^+,\;CD3^+$, and $CD4^+/CD25^+$ cells in DLN (draining lymph nodes) significantly increased. The percentage of $B220^+/CD23^+$ cells in DLN significantly decreased. 6. The percentage of $CD3^+,\;CD4^+$, and $CD11b^+/Gr-1^+$ cells in paw joints significantly decreased. 7. The production of TNF-$\alpha$, IL-6, and MCP-1 significantly decreased. 8. Anti-collagen II in serum significantly decreased. 9. With the hematoxylin and eosin stain, inflammation of joint decreased. Under Masson's trichrome stain, the cartilage destruction and synovial cell proliferation and the expression of collagen fibers decreased. Conclusions: Comparison of the results for this study showed that GJB-7 had immunomodulatory effects. So we expect that GJB-7 could be used as an effective drug for not only rheumatoid arthritis but also another auto-immune diseases.

• Biomolecules & Therapeutics
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• v.28 no.5
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• pp.414-422
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• 2020

Fangchinoline (FAN) is a bisbenzylisoquinoline alkaloid that is widely known for its anti-tumor properties. The goal of this study is to examine the effects of FAN on arthritis and the possible pathways it acts on. Human fibroblast-like synovial cells (FLS), carrageenan/kaolin arthritis rat model (C/K), and collagen-induced arthritis (CIA) mice model were used to establish the efficiency of FAN in arthritis. Human FLS cells were treated with FAN (1, 2.5, 5, 10 µM) 1 h before IL-1β (10 ng/mL) stimulation. Cell viability, reactive oxygen species measurement, and western blot analysis of inflammatory mediators and the MAPK and NF-κB pathways were performed. In the animal models, after induction of arthritis, the rodents were given 10 and 30 mg/kg of FAN orally 1 h before conducting behavioral experiments such as weight distribution ratio, knee thickness measurement, squeaking score, body weight measurement, paw volume measurement, and arthritis index measurement. Rodent knee joints were also analyzed histologically through H&E staining and safranin staining. FAN decreased the production of inflammatory cytokines and ROS in human FLS cells as well as the phosphorylation of the MAPK pathway and NF-κB pathway in human FLS cells. The behavioral parameters in the C/K rat model and CIA mouse model and inflammatory signs in the histological analysis were found to be ameliorated in FAN-treated groups. Cartilage degradation in CIA mice knee joints were shown to have been suppressed by FAN. These findings suggest that fangchinoline has the potential to be a therapeutic source for the treatment of rheumatoid arthritis.

• The Korean Journal of Physiology and Pharmacology
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• v.15 no.2
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• pp.83-88
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• 2011

A large body of evidence has indicated that induction of endogenous antioxidative proteins seems to be a reasonable strategy for delaying the progression of cell injury. In our previous study, cilostazol was found to increase the expression of the antioxidant enzyme heme oxygenase-1 (HO-1) in synovial cells. Thus, the present study was undertaken to examine whether cilostazol is able to counteract tumor necrosis factor-${\alpha}$ (TNF-${\alpha}$)-induced cell death in endothelial cells via the induction of HO-1 expression. We exposed human umbilical vein endothelial cells (HUVECs) to TNF-${\alpha}$ (50 ng/ml), with or without cilostazol ($10{\mu}M$). Pretreatment with cilostazol markedly reduced TNF-${\alpha}$-induced viability loss in the HUVECs, which was reversed by zinc protoporphyrine IX (ZnPP), an inhibitor of HO-1. Moreover, cilostazol increased HO-1 protein and mRNA expression. Cilostazol-induced HO-1 induction was markedly attenuated not only by ZnPP but also by copper-protoporphyrin IX (CuPP). In an assay measuring peroxisome proliferator-activated receptor-${\gamma}$ (PPAR-${\gamma}$) transcription activity, cilostazol directly increased PPAR-${\gamma}$ transcriptional activity which was completely abolished by HO-1 inhibitor. Furthermore, increased PPAR-${\gamma}$ activity by cilostazol and rosiglitazone was completely abolished in cells transfected with HO-1 siRNA. Taken together, these results indicate that cilostazol up-regulates HO-1 and protects cells against TNF-${\alpha}$-induced endothelial cytotoxicity via a PPAR-${\gamma}$-dependent pathway.

• BMB Reports
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• v.51 no.10
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• pp.508-513
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• 2018

Fibronectin fragments found in the synovial fluid of patients with osteoarthritis (OA) induce the catabolic responses in cartilage. Nuclear high-mobility group protein Box 1 (HMGB1), a damage-associated molecular pattern, is responsible for the regulation of signaling pathways related to cell death and survival in response to various stimuli. In this study, we investigated whether changes induced by 29-kDa amino-terminal fibronectin fragment (29-kDa FN-f) in HMGB1 expression influences the pathogenesis of OA via an HMGB1-modulated autophagy signaling pathway. Human articular chondrocytes were enzymatically isolated from articular cartilage. The level of mRNA was measured by quantitative real-time PCR. The expression of proteins was examined by western blot analysis, immnunofluorescence assay, and enzyme-linked immunosorbent assay. Interaction of proteins was evaluated by immunoprecipitation. The HMGB1 level was significantly lower in human OA cartilage than in normal cartilage. Although 29-kDa FN-f significantly reduced the HMGB1 expression at the mRNA and protein levels 6 h after treatment, the cytoplasmic level of HMGB1 was increased in chondrocytes treated with 29-kDa FN-f, which significantly inhibited the interaction of HMGB1 with Beclin-1, increased the interaction of Bcl-2 with Beclin-1, and decreased the levels of Beclin-1 and phosphorylated Bcl-2. In addition, the level of microtubule-associated protein 1 light chain 3-II, an autophagy marker, was down-regulated in chondrocytes treated with 29-kDa FN-f, whereas the effect was antagonized by mTOR knockdown. Furthermore, prolonged treatment with 29-kDa FN-f significantly increased the release of HMGB1 into the culture medium. These results demonstrated that 29-kDa FN-f inhibits chondrocyte autophagy by modulating the HMGB1 signaling pathway.

• IMMUNE NETWORK
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• v.1 no.1
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• pp.61-69
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• 2001

Background: To determine the molecular structure of type II collagen-specific T-cell receptors associated with rheumatoid arthritis (RA). Methods: We generated CII-specific T-cell lines of 8 RA patients by prolonged in vitro culture with bovine CII (bCII) and the immunogenic peptide (256-270) of human CII. The proliferation response towards CII stimulation was measured from the uptake of 3H-thymidine. Changes in the secretion of Th 1 and Th2 cytokines in the culture supernatent were measured by ELISA. The TCR clonotypes of these T-cells were examined by RT-PCR/SSCP analyses of all 22 $V_{\beta}$ chains. Results: T-cells from patients' tissue exhibited strong proliferation index upon CII stimulation, which was maintained up to 6 months in the culture. The secretion of INF-$\gamma$from these T-cells increased along with the duration of culture time, while the amount of IL-4 production did not show significant changes. The SSCP band patterns of patients' T-cells appear as discrete bands unlike the smeary streak produced from normal samples. Some SSCP bands, each representing selected expansion of a TCR containing certain subtype of $V_{\beta}$ peptides, appeared to be identical in more than one patients. Among these, the expansion of SSCP band representing the $V_{\beta}$ 14 CDR3 region persisted after switching the antigen to the immunogenic human peptide (256-270). Conclusion: CII-reactive T-cells expressing distinct CDR3 motifs are selectively expanded in the peripheral blood and synovial fluid of RA patients, and their persistent proliferation upon CII stimulation, as well as the production Th 1-type cytokines, may play pivotal roles in RA pathogenesis.

• Journal of Acupuncture Research
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• v.22 no.2
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• pp.71-81
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• 2005

Objective : Dear antler (Cervus korean TEMMINCK var. mantchuricus Swinhoe) used for traditional immunosuppressive and immuno-activating action. The effect of deer antler herbal-acupuncture(DAH) solution, prepared by water extract method, on cathepsin activities in bone tissues (cartilage and synovial) cells from mouse rheumatoid arthritis (RA) model was studied. The cysteine endoprotease cathepsin mediates degradation of the MHC class II invariant chain (Ii) in human and mouse antigen-presenting cells. The studies described here examine the functional significance of cathepsin inhibition on autoantigen presentation and organ-specific autoimmune diseases in a murine model for RA. Methods : An animal model for RA in BALB/c mice thymectomized 3 days after birth (3d-Tx) was constructed All 3d-Tx BALB/c mice developed autoimmune lesions in the bone tissue cells, starting at 3 weeks of age, and the disease mediated by CD4+ T cells was chronic and progressive. Significant inhibitory effects of DAH solution on cathepsin S and L were observed in each organ in a dose-dependent manner. Moreover, we confirmed that cathepsin S and L activity in each organ were clearly inhibited by DAB solution. When we examined the inhibitory effects of DAH solution against autoantigen-specific T cell responses in vitro, in regional lymph node cells, but not in spleens, from model mice, a significant inhibitory effect of DAB solution was observed in a dose-dependent manner. DAH solution do not block T cell proliferation to Con A, indicated that the dose of DAB solution 10 to $20\;{\mu}g/m{\ell}$ was sufficient to inactivate the autoantigen-specific T cell responses in vitro. In vivo therapeutic effects of DAB solution were examined in a murine model for RA, autoantigen-specific (C-II-specific) T cell response were significantly inhibited in LNCs from DAH solution-treated mice. Results : Iinhibition of cathepsin S and L in vivo alters autoantigen presentation and development of organ-specific autoimmunity in RA model. Conclusion : These data identify selective inhibition of cysteine protease cathepsin S and L as a potential therapeutic strategy for autoimmune disease process such RA. Thus, DAH solution will served as a potent anti-inflammatory and anti-arthritic agents for treatment of human RA.

• IMMUNE NETWORK
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• v.3 no.1
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• pp.1-7
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• 2003

Type II collagen (CII), major component of hyaline cartilage, has been considered as an auto-antigen in rheumatoid arthritis (RA). However, the clinical and biological significances with regard to the CII autoimmunity need to be clarified in human RA. The presence of antibodies to CII has been identified in sera, synovial fluid, and cartilage of patients with RA. In our study, the increased titer of IgG anti-CII in sera was well correlated with C-reactive protein, suggesting that this antibody may reflect the inflammatory status of RA. The titer of anti-CII antibodies (anti-CII Abs) tended to be higher in early stages of diseases. In our extending study, among 997 patients with RA, 269 (27.0%) were positive for circulatory IgG antibody to CII, those levels were fluctuated over time. It is hard to assess the significant amount of T cell responses to CII and CII (255~274) in RA. By using a sensitive method of antigen specific mixed lymphocyte culture, we can detect the presence of CII-reactive T cells in peripheral blood mononuclear cells of RA patients. Sixty seven (46.9%) of 143 patients showed positive CII reactive T cell responses to CII or CII (255~274). The frequencies of CII reactive T cells were more prominent in inflamed synovial fluid (SF) than in peripheral blood. These T cells could be clonally expanded after consecutive stimulation of CII with feeding of autologous irradiated antigen presenting cells (APC). Moreover, the production of Th1-related cytokine, such as IFN-${\gamma}$, was strongly up-regulated by CII reactive T cells. These data suggest that T cells responding to CII, which are probably presenting the IFN-${\gamma}$ producing cells, may play an important role in the perpetuation of inflammatory process in RA. To evaluate the effector function of CII reactive T cells, we investigated the effect of CII reactive T cells and fibroblasts-like synoviocytes (FLS) interaction on the production of pro-inflammatory cytokines. When the CII reactive T cells were co-cultured with FLS, the production of IL-15 and TNF-${\alpha}$ from FLS were significantly increased (2 to 3 fold increase) and this increase was clearly presented in accord to the expansion of CII reactive T cells. In addition, the production of IFN-${\gamma}$ and IL-17, T cell derived cytokines, were also increased by the co-incubation of CII reactive T cells with FLS. We also examined the impact of CII reactive T cells on chemokines production. When FLS were co-cultured with CII stimulated T cells, the production of IL-8, MCP-1, and MIP-1${\alpha}$ were significantly enhanced. The increased production of these chemokines was strongly correlated with increase the frequency of CII reactive T cells. Conclusively, immune response to CII was frequently found in RA. Activated T cells in response to CII contributed to increase the production of proinflammatory cytokines and chemokines, which were critical for inflammatory responses in RA. The interaction of CII-reactive T cells with FLS further augmented this phenomenon. Taken together, our recent studies have suggested that autoimmunity to CII could play a crucial role not only in the initiation but amplification/perpetuation of inflammatory process in human RA.