• The Journal of Korean Medicine Ophthalmology and Otolaryngology and Dermatology
• /
• v.20 no.1 s.32
• /
• pp.1-15
• /
• 2007

Objectives : RVC has long been used for a useful natural agent ameliorating inflammation related symptoms in the folk medicine recipe. This study was performed to investigate effects of RVC on the inflammation and oxidation in RAW 264.7 cells. Methods : The RVC was extracted with 80% ethanol and sequentially partitioned with solvents in order to increase polarity. With the various fractions, we determined the activities on the inflammation and oxidation in RAW 264.7 cells. Results : 1. Among the various solvent extracts of RVC, the butanol fraction showed the most powerful inhibitory ability against nitric oxide (NO) production in lipopolysaccharide (LPS)-induced RAW 264.7 cells without affecting cell viability. 2. Butanol fraction showed a oxidation inhibition effect by decreasing the DPPH and OH radicals. 3. Butanol fraction exhibited the inhibitory avilities against iNOS and COX-2. 4. Reverse transcriptase polymerase chain reaction (RT-PCR) and Westem blotting analysis revealed that the BuOH fraction provided a primary inhibitor of the iNOS protein and mRNA expression in LPS-induced RAW 264.7 cells. Among the up-regulater molecules of iNOS and COX-2, the BuOh fraction of RVC was shown the inhibitory activity of phoshporylation of c-Jun N-terminal kinase (JNK) 1/2 and threonine protein kinase (AKT), the one of the MAPKs pathway. Conclusion : Thus, the present study suggests that the response of a component of the BuOH fraction to NO generation via iNOS expression provide a important clue to elucidate anti-inflammatory and anti-oxidation mechanism of RVC.

• The Korean Journal of Physiology and Pharmacology
• /
• v.25 no.4
• /
• pp.307-319
• /
• 2021

Dysregulation of the Wnt pathway causes various diseases including cancer, Parkinson's disease, Alzheimer's disease, schizophrenia, osteoporosis, obesity and chronic kidney diseases. The modulation of dysregulated Wnt pathway is absolutely necessary. In the present study, we evaluated the anti-inflammatory effect and the mechanism of action of Wnt-C59, a Wnt signaling inhibitor, in lipopolysaccharide (LPS)-stimulated epithelial cells and macrophage cells. Wnt-C59 showed a dose-dependent anti-inflammatory effect by suppressing the expression of proinflammatory cytokines including IL6, CCL2, IL1A, IL1B, and TNF in LPS-stimulated cells. The dysregulation of the Wnt/β-catenin pathway in LPS stimulated cells was suppressed by WntC59 treatment. The level of β-catenin, the executor protein of Wnt/β-catenin pathway, was elevated by LPS and suppressed by Wnt-C59. Overexpression of β-catenin rescued the suppressive effect of Wnt-C59 on proinflammatory cytokine expression and nuclear factor-kappa B (NF-κB) activity. We found that the interaction between β-catenin and NF-κB, measured by co-immunoprecipitation assay, was elevated by LPS and suppressed by Wnt-C59 treatment. Both NF-κB activity for its target DNA binding and the reporter activity of NF-κB-responsive promoter showed identical patterns with the interaction between β-catenin and NF-κB. Altogether, our findings suggest that the anti-inflammatory effect of Wnt-C59 is mediated by the reduction of the cellular level of β-catenin and the interaction between β-catenin and NF-κB, which results in the suppressions of the NF-κB activity and proinflammatory cytokine expression.

• The Korea Journal of Herbology
• /
• v.24 no.1
• /
• pp.169-178
• /
• 2009

Objectives : In this study, the effects of ethylacetate extract of Ostericum koreanum on inflammation in RAW264.7 cells were investigated. Methods : Dried roots of Ostericum koreanum was extracted with 80% methanol for 24 h, and then fractionated with n-butanol, n-hexan and ethylacetate. RAW264.7 cells, a mouse macrophage line were incubated with different concentrations of the extract for 30 min and then stimulated with LPS at indicated times. Cell toxicity was determined by MTT assay. The concentrations of nitric oxide (NO) and prostaglandin E2 ($PGE_2$) were measured by Griess assay and enzyme immunoassay (EIA), respectively. The expressions of inducible nitric oxide synthease (iNOS) and cyclooxyganase (COX) -2 mRNA and protein were determined by RT-PCR and Western blot. Results : The methanol extract of Ostericum koreanuman and its fractions were significantly inhibited the NO and PGE2 productions in LPS-stimulated RAW264.7 cells. Among the fractions of Ostericum koreanuman the ethylacetate fraction was more strongly inhibited NO and $PGE_2$ productions compared with other fractions. The ethylacetate fraction was also suppressed LPS-induced mRNA expressions of iNOS and its protein levels in RAW264.7 cells. Conclusions : This study suggests that the ethylacetate fraction of Ostericum koreanum may have an anti-inflammatory property through suppressing inflammatory mediator productions in activated macrophages, suggesting have a therapeutic potential for the treatment of various inflammatory diseases.

• Proceedings of the Korean Society of Toxicology Conference
• /
• 2002.11b
• /
• pp.184-184
• /
• 2002

In our previous study, we reported that PG, a polysaccharide isolated from Plyatycodon grandiflorum, activated macrophages and B cells, but not T cells. Here, we investigated in more detail the mechanism of action of PG in macrophage activation.(omitted)

• IMMUNE NETWORK
• /
• v.5 no.3
• /
• pp.144-149
• /
• 2005

Background: Fc receptor-mediated phagocytosis is a complex process involving the activation of kinases and phosphatases. FcgammaRIIB has been known to transduces inhibitory signals through an immunoreceptor tyrosine-based inhibitory motif (ITIM) in cytoplasmic domains. In this study, we examined the involvement of inositol-phosphatase in the Fc receptor-mediated phagocytosis. Methods: J774 cells were infected using vaccinia viral vector containing SH2 domain-containing inositol-phosphatase (SHIP) cDNA and stimulated with the sensitized sheep red blood cells. Results: Stimulation of J774 cells induced the tyrosine phosphorylation of SHIP which was maximal at 5 minutes. Phosphatidylinositol-3 (PI-3) kinase inhibitor (wortmannin) inhibits J774 cell phagocytosis of sensitized sheep red blood cells in a dose-dependent manner. Heterologious expression of SHIP in J774 cells inhibits phagocytosis of sensitized sheep red blood cells in a dose-dependency manner, but catalytically dead mutants of SHIP has no effect on phagocytosis. Conclusion: These results strongly suggest that the active signals mediated by PI-3 kinase are opposed by inhibitory signals through SHIP in the regulation of Fc receptor-mediated phagocytosis.

• Journal of Life Science
• /
• v.27 no.8
• /
• pp.873-878
• /
• 2017

We previously reported that NAD(P)H:quinone oxidoreductase 1 (NQO1)-knockout (KO) mice exhibited spontaneous inflammation in the gut. We also found that NQO1-KO mice showed highly increased inflammatory responses compared with NQO1-WT control mice when subjected to DSS-induced experimental colitis. In a Clostridium difficile toxin-induced mouse enteritis model, NQO1-KO mice were also sensitive compared with NQO1-WT mice. Moreover, numerous studies have shown that NQO1 is functionally associated with immune regulation. Here, we assessed whether NQO1 defects can alter macrophage activation. We found that peritoneal macrophages isolated from NQO1-KO mice produced more IL-6 and $TNF-{\alpha}$ than those isolated from NQO1-WT mice. Moreover, the dicumarol-induced inhibition of NQO1 significantly increased IL-6 and $TNF-{\alpha}$ production in peritoneal macrophages isolated from NQO1-WT mice, as well as in the cultured mouse macrophage cell line, RAW264.7. These results indicate that NQO1 may negatively regulate the activation of macrophages. Knockout or chemical inhibition of NQO1 markedly reduced the expression of $I{\kappa}B$ (inhibitor of $NF{\kappa}B$) in both mouse peritoneal macrophages and RAW264.7 cells. Finally, RAW264.7 cells treated with dicumarol exhibited morphological changes reflecting macrophage activation. Our results suggest that NQO1 may suppress the $NF{\kappa}B$ pathways in macrophages, thereby suppressing the activation of these cells. Thus, immunosuppressive activity may be among the many possible functions of NQO1.

• Journal of the Korean Society of Food Science and Nutrition
• /
• v.41 no.11
• /
• pp.1645-1648
• /
• 2012

Allium hookeri, a member of the onion family, has long been mainly cultivated for food and medicinal use in Southeast Asia countries owing to its various biological properties. However, no studies of the anti-inflammatory effects of A. hookeri extracts have been conducted to date. Therefore, this study was investigated the potential of the methanol extract of A. hookeri to suppress the inflammation in lipopolysaccharide (LPS)-induced mouse macrophage RAW264.7 cells. This study was performed on macrophage cells that were pretreated with $0{\sim}500{\mu}g/mL$ of methanol extract of A. hookeri root prior to LPS treatment. Treatment with methanol extract of A. hookeri root significantly inhibited LPS-induced nitric oxide formation in dose-dependent manner. Treatment of A. hookeri root also significantly decreased LPS-induced TNF-${\alpha}$ and IL-6 production. The results of this study provide new evidence of the anti-inflammatory properties of A. hookeri and indicate that it may have a potential therapeutic use for the prevention and treatment of macrophage derived chronic immune diseases.

• Tuberculosis and Respiratory Diseases
• /
• v.68 no.3
• /
• pp.168-174
• /
• 2010

Background: Sepsis still has a high mortality rate despite adequate supportive care. Newer therapeutic modalities have been developed but they have generally ended in failure. Recently, insulin was reported to have an anti-inflammatory effect by inhibiting the $I{\kappa}B/NF-{\kappa}B$ pathway, and may have therapeutic potential in sepsis. However, the precise mechanism of the anti-inflammatory effect of insulin is unclear. This study examined the role of insulin in activating $I{\kappa}B/NF-{\kappa}B$ in macrophage. Methods: Raw 264.7 cells, a murine macrophage cell line, were used in this experiment. Western blotting using $I{\kappa}B$ Ab and phosphor-specific $I{\kappa}B$ Ab was performed to evaluate the degradation and phosphorylation of $I{\kappa}B$ cells. For the $I{\kappa}B$ Kinase (IKK) activity, an immune complex kinase assay was performed. The level of interleukin-6 (IL-6) was measured by ELISA to determine the level of proinflammatory cytokine. Results: $I{\kappa}B{\alpha}$ degradation began 30 min after lipopolysaccharide (LPS) treatment. However, an insulin pretreatment suppressed the $I{\kappa}B{\alpha}$ degradation caused by the LPS treatment. The phosphorylation of $I{\kappa}B{\alpha}$ and IKK activity was also inhibited by the insulin pretreatment. Finally, the insulin pretreatment showed a tendency to suppress the induction of IL-6 by LPS. Conclusion: Insulin might have an anti-inflammatory effect though partial inhibition of the $I{\kappa}B/NF{\kappa}B$ pathway in macrophage cell lines.

• Journal of dental hygiene science
• /
• v.19 no.2
• /
• pp.133-140
• /
• 2019

Background: The light-emitting diode (LED) curing light used is presumed to be safe. However, the scientific basis for this is unclear, and the safety of LED curing light is still controversial. The purpose of this study was to investigate the effect of LED curing light irradiation according to the conditions applied for the polymerization of composite resins in dental clinic on the cell viability and inflammatory response in Raw264.7 macrophages and to confirm the stability of LED curing light. Methods: Cell viability and cell morphology of Raw264.7 macrophages treated with 100 ng/ml of lipopolysaccharide (LPS) or/and LED curing light with a wavelength of 440~490 nm for 20 seconds were confirmed by methylthiazolydiphenyl-tetrazolium bromide assay and microscopic observation. The production of nitric oxide (NO) and prostaglandin $E_2$ ($PGE_2$) was confirmed by NO assay and $PGE_2$ enzyme-linked immunosorbent assay kit. Expression of interleukin $(IL)-1{\beta}$ and tumor necrosis factor $(TNF)-{\alpha}$ in total RNA and protein was confirmed by reverse transcription polymerase chain reaction and Western blot analysis. Results: The LED curing light did not affect the viability and morphology of normal Raw264.7 cells but affected the cell viability and induced cytotoxicity in the inflammation-induced Raw264.7 cells by LPS. The irradiation of the LED curing light did not progress to the inflammatory state in the inflammation-induced Raw264.7 macrophage. However, LED curing light irradiation in normal Raw264.7 cells induced an increase in NO and $PGE_2$ production and mRNA and protein expression of $(IL)-1{\beta}$ and $(TNF)-{\alpha}$, indicating that it is possible to induce the inflammatory state. Conclusion: The irradiation of LED curing light in RAW264.7 macrophage may induce an excessive inflammatory reaction and damage oral tissues. Therefore, it is necessary to limit the long-term irradiation which is inappropriate when applying LED curing light in a dental clinic.

• Proceedings of the Korean Society of Embryo Transfer Conference
• /
• 2002.11a
• /
• pp.88-88
• /
• 2002

인공수정 및 수정란기술의 활성화에 따라 소에 있어서 인공수정은 90%이상이 실시되고 있으나, 수정란이식은 수정란의 생산이 안정적이지 않아 활성화에 많은 지장을 초래하고 있다. 이의 원인은 수정란이식에 의한 수태율의 저하가 가장 크며, 수태율 향상을 위하여 수란우에 progesterone, hCG 등의 주사가 실시되고 있다. 그러나 이는 수정란의 착상에 있어서 자궁의 환경을 개선한다고 하나, 착상의 정확한 기전의 구명은 미미한 상태이다. 한편 비장유래 macrophage가 황체를 자극하고 TGF-$\beta$의 생산을 유도하는 것으로 보고되고 있으며, IL-I $\alpha$와 $\beta$에 따라 TGF-$\beta$ 생산에 있어서 약간의 차이를 보이는 것으로 보고되고 있다. 따라서 본 연구는 비장유래 macrophage가 TGF-$\beta$의 생산시 임신관련 Cytokine인 IL-I$\alpha$와의 관계를 조사하기 위하여 실시하였다. 임신 및 비임신 도축 암소의 비장을 채취하여 얼음에 채워 실험실로 운반한 후 비장의 표면을 70%의 알콜로 세척하고, 표피를 벗겨 비장조직을 세절하여 10% FBS+DMEM에 넣어 조직을 눌러 짜면서 조직속의 세포를 분리하였다. 세척한 배양액은 4-5$m\ell$를 100mm 유리 petri dish에 넣고 39$^{\circ}C$, 5% $CO_2$, 95% 공기인 배양기에서 2시간이상 배양하였으며, 배양 후 냉장된 buffer A 용액으로 세척하여 유리 petri dish의 바닥에 부착된 macrophage만을 cell scraper로 분리하였다. 분리한 macrophage는 0.5-1 $\times$ $10^{6}$ cells/$m\ell$가 되게 조정하여, IL-I 을 0.001, 0.01, 0.1 또한 1 ng/$m\ell$를 첨가하여 농도에 따른 효과를 조사하였고, 각각 24, 48, 72, 96 또한 120시간을 배양하여 시간에 의한 효과도 실시하였다. 각 배치구에서 얻어진 배양액은 TGF-$\beta$를 조사하기 전까지 -2$0^{\circ}C$에 동결 보존하였다. TGF-$\beta$의 측정은 TGF-$\beta$ kit(promega, USA)를 이용하여 실시하였으며, 통계학적 분석은 Anova test를 Statview program을 이용하여 분석하였다. 시험의 결과 대조구에 비해 IL-I 첨가구는 2-3배의 TGF-$\beta$생산을 보였으며, 배양시간에 따른 생산은 시간이 지남에 따라 약간 상승하는 경향을 보였으나, 유의적인 차이를 보이지는 않았다. 또한 IL-I의 농도에 따른 생산의 변화는 IL-I의 농도에 따라 약간의 차이를 보였고 유의적인 차이는 인정되지 않았다. 임신 및 비임신의 경우 임신우의 비장 macrophage가 비임신보다는 약간 상승하는 거스로 나타났다. 이상의 결과로 볼 때 IL-I $\alpha$ 는$\beta$subunit 보다 TGF-$\beta$ 생산에 있어서 서로 다른 양상을 보일 것으로 추정되며, IL-I은 macrophage의 직접적인 영향을 주기보다는 황체세포를 매개로 한 자궁에 TGF-$\beta$의 생산을 유도하는 것으로 사료되며, 임신관련 cytokine에 대한 다양한 연구가 요구되고 있다.