• The Korea Journal of Herbology
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• v.31 no.4
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• pp.1-10
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• 2016

Objectives: In Korean medicine, Curculiginis Rhizoma was treated for arthritis in remedy. But efficacy of Curculiginis Rhizoma on collagen induced arthritis was not revealed.Methods: Anti inflammatory effect of Curculiginis Rhizoma was researched in vitro with RAW264.7 cell and cell toxicity, levels of proinflammatory cytokines (TNF-α, IL-1β, IL-6 and IL-12) and PGE2 were analyzed by ELISA assay. Inflammatory protein were analyzed by western blotting assay (JNK, ERK, COX-2, TNF-α and IL-1β). In vivo, collagen induced arthritis mice model was used to evaluate anti-inflammation effect through arthritis index, immune cell number and cytokine levels (TNF-α, IL-6 and IL-1β) in serum.Results: ECR(Extract of Curculiginis Rhizoma) has not shown cell toxicity in 200 ㎍/㎖ on RAW264.7 cell. ECR suppressed releases of NO, TNF-α, IL-1β, IL-6, IL-12 and PGE2 on RAW264.7 cell treated with lipopolysacharide (1 ㎍/㎖). And ECR inhibited regulation of TNF-α, IL-1β and IL-6 mRNA, reduced protein release of JNK, ERK, iNOS, COX-2, IL-1β and TNF-α. AI of group treated with ECR 200 ㎎/㎏ and 100 ㎎/㎏ were significantly decreased compared to vihicle arthritis mice, the number of immune cell in foot joint was increased on control mice but those of group treated with ECR 200 ㎎/㎏ and 100 ㎎/㎏ were significantly reduced. This results correspond with contens of cytokines (TNF-α, IL-1β and IL-6) in serum.Conclusions: Curculiginis Rhizoma has anti-inflammation effect on RAW264.7 cell in vitro and collagen induced arthritis in vivo. So it is necessary to research more mechanism for cascade imfact.

• The Journal of Korean Obstetrics and Gynecology
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• v.27 no.3
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• pp.12-27
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• 2014

Objectives: This study was performed to evaluate the effect of Guibi-tang water extract (GB) on osteoporosis. Methods: We examined the effect of GB on osteoclast differentiation using murine pre-osteoclastic RAW 264.7 cells treated with receptor activator of nuclear factor kappa-B ligand (RANKL). The effect of GB on osteoclast was measured by counting TRAP (+) multinucleated cells and measuring TRAP activity. The mRNA expressions of osteoclastogenesis-related genes (Cathepsin K, MMP-9, TRAP, NFATc1, MITF, TNF-${\alpha}$, IL-6, COX-2) were measured by real-time PCR. We examined the effect of GB on osteoblast proliferation, ALP activity, bone matrix protein synthesis and collagen synthesis using murine calvarial cell. Results: GB decreased the number of TRAP (+) multinucleated cells and inhibited TRAP activity in RANKL-stimulated RAW 264.7 cell. GB decreased the expression of genes related osteoclastogenesis such as Cathepsin K, MMP-9, TRAP, NFATc1, MITF, COX-2 in RANKL-stimulated RAW 264.7 cell. But GB did not decrease the expression of iNOS and increased the expression of TNF-${\alpha}$, IL-6 in RANKL-stimulated RAW 264.7 cell. These genes (iNOS, TNF-${\alpha}$, IL-6) are thought to be related with the inflammatory bone destruction. GB increased cell proliferation of rat calvarial cell and also increased ALP activity in rat calvarial cell. GB did not increase bone matrix protein synthesis but increased collagen synthesis in rat calvarial cell. Conclusions: This study suggests that GB may be effective in treating osteoporosis by inhibiting osteoclast differentiation and its related gene expression and by increasing osteoblast proliferation.

• Journal of Applied Biological Chemistry
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• v.66
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• pp.23-28
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• 2023

Since the global shock caused by COVID-19, interest in immune-enhancing materials is rapidly increasing, therefore, the development of novel materials is necessary from the industrial and health perspectives. In this study, we selected Nelumbo nucifera Gaertner Seed Extract (NSE) and evaluated immune enhancement effect by using RAW 264.7 murine macrophage cells. NSE significantly up-regulated production of nitric oxide and reactive oxygen species without affecting cell viability in RAW 264.7 cells. Additionally, NSE exhibited an increase of inducible nitric oxide synthase and cyclooxygenase-2 expression in RAW 264.7 cells. The enzyme-linked immunosorbent assay results showed that NSE-treatment significantly enhanced production of interleukin 6 and tumor necrosis factor-α in RAW 264.7 cells. Furthermore, we observed that NSE significantly up-regulated phosphorylation of p65, I kappa B kinase α/β, and I kappa B (IκB) α as well as down-regulation of IκB α expression in RAW 264.7 cells. Our findings indicate that NSE could be the potential health-functional food material with capacity of improving immunity via Nuclear factor-kappa B signaling pathway.

• Microbiology and Biotechnology Letters
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• v.45 no.1
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• pp.19-26
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• 2017

Effects of resveratrol glucosylation on the immunomodulation properties of resveratrol and on the viability of macrophage cells have been studied by using murine macrophage RAW 264.7 cells. Nitric oxide (NO) and interleukin 6 (IL-6) expression in macrophages in vitro were studied after treatment with different concentrations of (E)-resveratrol, (E)-resveratrol 3-O-${\beta}$-${\small{D}}$-glucoside (R-3-G), or (E)-resveratrol 4'-O-${\beta}$-${\small{D}}$-glucoside (R-4'-G). In vitro viability of RAW 264.7 cells after treatment with the aforementioned three compounds was also studied. As demonstrated by macrophage cell viability assays, two different resveratrol monoglucosides, R-3-G and R-4'-G, exhibited 50-80% reduced cytotoxicity in comparison to (E)-resveratrol in A549 and HepG2 cells. Compared to the resveratrol aglycon, both glucosylated resveratrol derivatives positively modulated NO and IL-6 production in macrophages positively via transcriptionally up-regulating IL-6 and iNOS expression. Conjugation of a glucose moiety on resveratrol was found to enhance the immunomodulating activity of resveratrol and the viability of RAW 264.7 cells.

• Applied Chemistry for Engineering
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• v.30 no.4
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• pp.479-486
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• 2019

This study investigated the anti-inflammatory effects of mixed extracts of Achyranthes japonica Nakai (Aj) and Aralia continentalis Kitagawa (Ac) (ratios of 1 : 2, 1 : 3, 1 : 5, 2 : 1, 3 : 1 and 5 : 1) on RAW264.7 macrophages. Cell toxicity was determined using a cell counting kit (CCK) assay. We evaluated anti-inflammatory effects of the mixed extracts of Aj and Ac by measuring interleukin $(IL)-1{\beta}$, IL-6, and tumor necrosis factor $(TNF){\alpha}$ using an enzyme-linked immunosorbent assay (ELISA) kit assay. The mixed extracts of Aj and Ac inhibited lipopolysaccharide (LPS)-induced $IL-1{\beta}$ and $TNF{\alpha}$ in LPS-stimulated macrophages. Comparing different ratios of the mixed extracts, the 2 : 1 ratio of Aj and Ac has much more potency and inhibited the production of $TNF{\alpha}$ in LPS-induced RAW264.7 cells. The results of the present study showed that the mixed extracts of Aj and Ac have potential anti-inflammatory effects on RAW264.7 macrophages. Therefore, these extracts may be used as a good source of functional foods for the protection against inflammatory diseases.

• Journal of Life Science
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• v.30 no.4
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• pp.359-369
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• 2020

Although many studies on immune modulatory materials have used RAW 264.7 cells, few have used T cell-derived TK-1 cell lines. Moreover, although some studies have investigated the efficacy of plant-derived β-sitosterol, few have examined the immunomodulatory activity of its analogue, daucosterol. In this study, β-sitosterol and daucosterol were isolated from D. batatas and identified by nuclear magnetic resonance spectroscopy. To evaluate the immune-enhancing or inhibitory effects of the isolated phytosterols, the expression levels of the inflammatory response genes COX-2, TNF-α, IL-6, and iNOS were analyzed by RT-PCR. The relative expression levels of TNF-α and iNOS in RAW 264.7 cells were increased more than threefold with β-sitosterol treatment comparing to those of untreated control. In the case of TK-1 cells, the expression level of TNF-α was decreased and the expression level of iNOS was increased in a β-sitosterol concentration-dependent manner. The expression levels of COX-2, TNF-α, and IL-6 increased by approximately 0.7-1.2 times in RAW 264.7 cells treated with daucosterol compared to those of untreated control, but iNOS expression decreased by 0.8-0.18 times. In the case of daucosterol-treated TK-1 cells, the expression levels of TNF-α, IL-6, and iNOS were markedly reduced from those of TK-1 cells treated only with lipopolysaccaride. As a conclusion, β-sitosterol treatment increased TNF-α and iNOS expression levels in RAW 264.7 cells, thus exerting an immune- boosting effect. However, in TK-1 cells, iNOS expression increased while TNF-α expression decreased, indicating an immunosuppressive activity of β-sitosterol. Daucosterol appears to exert an immunosuppressive effect in both macrophages and T cell lines by inhibiting iNOS expression in RAW 264.7 cells and greatly inhibiting the expression of TNF-α, IL-6, and iNOS in TK-1 cells.

• Toxicological Research
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• v.26 no.4
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• pp.267-273
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• 2010

As the use of carbon fullerene increases in the chemical industry, the concern over its biological and toxicological effects is also increasing. In this study, the suspension of carbon fullerene (C60) in phosphate buffered saline was prepared and toxicity was investigated using cultured RAW 264.7 and in intraperitoneally injected mice, respectively. The average size of carbon fullerene in the suspension was $53.7{\pm}26.5nm$ when determined by particle size analyzer. Cell viability was significantly decreased by the exposure of carbon fullerene ($0.25\sim2.00\;{\mu}g/ml$) for 96 hrs in the cultured RAW 264.7 cells. Intracellular reduced glutathione (GSH) level was also decreased compared to the level of the non-treated control group during the exposure period, while the level of nitric oxide was increased. When mice were intraperitoneally injected with carbon fullerene, serum cytokine levels of IL-1 and IL-6 were increased with the increased expression of inflammatory genes in peritoneal macrophage and T cell distribution in blood lymphocytes. The results suggested inflammatory responses were induced by carbon fullerene.

• Journal of Periodontal and Implant Science
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• v.23 no.1
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• pp.48-55
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• 1993

Macrophage inflammatory $protein-1{\alpha}(MIP-1{\alpha})$ from activated T cell or macrophage, which is small inducible cytokine of unkown biological function, has been shown to display inflammation chemokinetic activities, as well as myelosuppressive effect on more immature progenitor cells. In this paper we show the $MIP-1{\alpha}$ mRNA expression and the presence of $MIP-1{\alpha}$ binding sites from murine macrophage cell line RAW 264.7, and primary cells of mouse bone marrow and spleen. $MIP-1{\alpha}$ mRNA was induced from LPS-stimulated RAW 264.7, but not inhibited by cyclosporin A treatment, and also was expressed from mouse splenocyted and bone marrow cell which were not increased by ferritin or lactoferrin treatment. The results of receptor binding assay showed that radiolabeled RAW 264.7 cell with kd value of 0.91 nM, and binding sites per cell of 378. bone marrow cell and splenocyte also appeared to have $MIP-1{\alpha}$ binding sites 33 and 11 per cell, respectiviely.

• Journal of Haehwa Medicine
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• v.23 no.2
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• pp.5-13
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• 2015

Objectives : The anti-inflammatory and antioxidant effects of water extracts of Sasangja-tang(SSJ) and Gami-sasangja-tang(GSJ) were investigated. The effects of SSJ and GSJ were compared. Methods : We performed cell viability assay in HaCaT cells and RAW 264.7 cells using 3-(4,5-dimethylthiazol-2-yl)-2, 5 diphenyltetrazolium bromide(MTT) assay. We measured chemokines(regulated on activation normal T-cell expression and secreted ; RANTES/CCL5, interferon-inducible protein; IP-10/CXCL10, macrophage-derived chemokine; MDC/CCL22) in HaCat cells, also we measured cytokines (tumor necrosis factor-${\alpha}$; TNF-${\alpha}$, interleukin-6; IL-6) and nitric oxide(NO) production in RAW 264.7 cells using enzyme-linked immunosorbent assay(ELISA) and NO assay. Western blot assay was used to evaluate the expression for inducible nitric oxide synthase(iNOS) in RAW 264.7 cells. Results : SSJ and GSJ did not affect the cell viability at the concentrations treated ($0-800{\mu}g/ml$). As a result of SSJ and GSJ treatment in HaCat cells stimulated by TNF-${\alpha}$(10 ng/ml) and interferon(IFN)-${\gamma}$(10 ng/ml), the production of RANTES and IP-10 was inhibited significantly. However there was no significant difference in the secretion of MDC. And in RAW 264. 7 cells stimulated by lipopolysaccharide(LPS, $1{\mu}g/ml$), SSJ and GSJ treatment significantly inhibited the secretion of TNF-${\alpha}$ and IL-6 and the production of NO. The expression of iNOS was also decresed by SSJ and GSJ treatment in RAW 264. 7 cells. Compared with SSJ, GSJ was superior to SSJ in inhibition of RANTES, IP-10, TNF-${\alpha}$, IL-6 and NO production at the concentration of $200{\mu}g/ml$. Conclusion : Both SSJ and GSJ have anti-inflammatory and antioxidant effects. And GSJ has better effects than SSJ.

• The Korea Journal of Herbology
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• v.30 no.4
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• pp.89-94
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• 2015

Objectives : It had been reported that herbal medicines containing polyphenol and flavonoid have been shown to be associated with decreased the cause of aging and variety of disease such as reactive nitrogen species and reactive oxygen species in several recent studies. Salviae miltiorrhizae Radix, origined fromSalvia miltiorrhizaBGE., is one of popular traditional herbal medicines that is commonly used by traditional medicine practitioners. To this date, Salviae miltiorrhizae Radix has more than 2000-year history of mature application. This study was conducted to investigate whether the Salviae miltiorrhizae Radix methanol extract has an inhibitory effect association with oxidation or inflammation.Methods : Cytotoxic activity of Salviae miltiorrhizae Radix methanol extract on RAW264.7 cells was evaluated by using 3-[4, 5-dimethylthiazol-2-yl]-2, 5-diphenyltetrazolium bromide solution. Nitric oxide production was measured using griess reagent system. Western blot analysis and measurement for changes of protein expression, nitric oxide synthase and cyclooxygenase-2, also performed.Results : The medicinal plant, Salviae miltiorrhizae Radix, does not impair the cell viability in tested concentration (25-100 μg/ml). SMR showed anti-oxiative effectsin vitroby decreasing electron donating ability, and also showed anti-inflammatory effects suppressing NO and COX-2 expressin in LPS induced RAW264.7 activation. SMR inhibited the generation of intracellular ROS production as dose dependant manner.Conclusions : These results indicate that Salviae miltiorrhizae Radix methanol extract has an anti-inflammatory activities via anti-oxidative effects, and the anti-inflammatory effect was presentedd as dose dependant manner.