• 동의생리병리학회지
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• 제18권3호
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• pp.908-913
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• 2004

Farfrae Flos has been clinically used for the treatment of asthma in traditional oriental medicine. There is lack of studies regarding the effects of Farfrae Flos on the immunological activities. The present study was conducted to evaluate the effect of Farfrae Flos on the regulatory mechanism of cytokines and nitric oxide (NO) for the immunological activities in Raw 264.7 cells. In Raw 264.7 cells stimulated with lipopolysaccharide (LPS) to mimic inflammation, Farfrae Flos water extract inhibited nitric oxide production in a dose-dependent manner and abrogated inducible nitric oxide synthase (iNOS) and cyclooxygenase (COX-2). Farfrae Flos water extract did not affect on cell viability. To investigate the mechanism by which Farfrae Flos water extract inhibits iNOS and COX-2 gene expression, we examined the on the phospholylation of inhibitor κBα and production of TNF-α, IL-1β and IL-6. Results provided evidence that Farfrae Flos inhibited the production of interleukin-1β (IL-1β) and the activation of phospholylation of inhibitor κBα in Raw 264.7 cells activated with LPS. These findings suggest that Farfrae Flos can produce anti-inflammatory effect, which may play a role in adjunctive therapy in Gram-negative bacterial infections.

• Advances in Traditional Medicine
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• 제7권4호
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• pp.341-347
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• 2007

Red seaweed (Callophyllis japonica) has long formed part of the diet of Asians, but the pharmacological properties of this plant have not been evaluated. In this study, we examined the effect of an ethanol extract of C. japonica on the generation of nitric oxide (NO) in RAW 264.7 cells. The C. japonica extract increased the generation of NO and tumor necrosis $factor-{\alpha}$ ($TNF-{\alpha}$), which were detected by the Griess method and an enzyme-linked immunosorbent assay, respectively. The increased production of NO by C. japonica extract was inhibited by $N^G$-monomethyl-L-arginine ($100{\mu}M$), a specific inhibitor of NO production in the L-arginine-dependent pathway, and by the nuclear $factor-{\kappa}B$ ($NF-{\kappa}B$) inhibitor, pyrrolidine dithiocarbamate ($10-100{\mu}M$) in a dose-dependent manner. These findings demonstrate that C. japonica extract stimulates the production of NO and $TNF-{\alpha}$ in RAW 264.7 cells through the activation of $NF-{\kappa}B$ and that this extract might also inhibit the growth of the human leukemic cells.

• Journal of Acupuncture Research
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• 제40권4호
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• pp.368-376
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• 2023

Background: Although bee venom (BV) has clinical benefits in osteoarthritis and rheumatoid arthritis, it has not been tested as treatment for gouty arthritis. Moreover, in vitro, BV has been proven to exhibit anti-inflammatory and positive effects on osteoarthritis, but only limited evidence can confirm its beneficial effects on gout. Thus, this study aims to assess the anti-inflammatory effects of BV on monosodium urate (MSU)-induced THP-1 monocytes. Methods: THP-1 monocytes were differentiated into mature macrophages using phorbol 12-myristate 13-acetate (PMA) and pretreated for 6 hours with BV and a Caspase-1 inhibitor in a physiologically achievable range of concentrations (BV, 0.1-1 ㎍/mL; Caspase-1 inhibitor, 1-10 μM), followed by MSU crystal stimulation for 24 hours. The secretions of interleukin-1β (IL-1β), tumor necrosis factor-α (TNF-α), IL-6, IL-8, cyclooxygenase-2 (COX-2), prostaglandin E2 (PGE2), and nitric oxide (NO) were increased in the MSU crystal-stimulated THP-1 cells. Results: Caspase-1 inhibitors suppressed the production of all mediators in a dose-dependent manner. BV worked on equal terms with Caspase-1 inhibitors and showed more satisfactory effects on TNF-α, PGE2, COX-2, and inducible nitric oxide synthase (iNOS). Moreover, the western blot analysis revealed that BV regulated the transcriptional levels of these mediators via the suppression of extracellular signal-regulated kinase (ERK) pathway activation. Conclusion: The results of the present study clearly suggest that BV inhibits MSU-induced inflammation in vitro, suggesting a possible role for BV in gout treatment.

• 한방안이비인후피부과학회지
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• 제22권2호
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• pp.1-18
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• 2009

Objective : Moutan Cortex (the root bark of Paeonia suffruticosa Andr.) is widely used in oriental medicine as a remedy for inflammation. However, as yet there is no clear explanation of how MC(Moutan Cortex) affects the production of inflammatory cytokine. This study was to determine the effects of Essence extracted MC on the mast cell-mediated inflammatory responses. Method : We observed the effect of MC on compound 48/80-induced histamine release of rat peritoneal mast cells and the effect of administering MC on PCA in rat. We measured the amount of inflammatory cytokine production induced by the phorbol myristate acetate (PMA) plus calcium ionophore(A23187) in the human mast cell line (HMC-1) incubated with various concentrations of MC. The TNF-$\alpha$ protein levels were analysised by Western blot. The TNF-$\alpha$, IL-6 and IL-8 secreted protein levels were measured by the ELISA assay. The TNF-$\alpha$, IL-6 and IL-8 mRNA levels were measured by the RT-PCR analysis. NF-$\kappa$B, phospho-I$\kappa$B and MAPKs were exmined by Western blot analysis. The NF-$\kappa$B promoter activity was examined by luciferase assay. Result : 1. Enzyme immunoassay indicated that MC suppressed histamine secretion of rat peritoneal mast cells. 2. In PCA dependent on IgE, MC had anti-allergic effect of the internal surface of rat skin. 3. Western blot indicated that MC decreased TNF-$\alpha$ protein levels. 4. ELISA indicated that MC decreased TNF-$\alpha$, IL-6 but MC had no significant effect on IL-8 in HMC-1 cells. 5. RT-PCR indicated that MC decreased TNF-$\alpha$, IL-8 but MC had no significant effect on IL-6 in HMC-l cells. 6. Western blot indicated that MC suppressed the induction of MAPKs, NF-$\kappa$B & phospho-I$\kappa$B activity in HMC-1 cells. 7. Luciferase assay indicated that MC suppressed the PMA plus A23187-induced NF-$\kappa$B promoting activityin HMC-1 cells. Conclusion : In this study, we have found that MC is an inhibitor of NF-$\kappa$B, MAPKs & cytokines on the mast cell-mediated inflammatory responses.

• The Korean Journal of Physiology and Pharmacology
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• 제19권3호
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• pp.229-234
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• 2015

Nafamostat mesilate (NM) is a serine protease inhibitor with anticoagulant and anti-inflammatory effects. NM has been used in Asia for anticoagulation during extracorporeal circulation in patients undergoing continuous renal replacement therapy and extra corporeal membrane oxygenation. Oxidative stress is an independent risk factor for atherosclerotic vascular disease and is associated with vascular endothelial function. We investigated whether NM could inhibit endothelial dysfunction induced by tumor necrosis factor-${\alpha}$ (TNF-${\alpha}$ ). Human umbilical vein endothelial cells (HUVECs) were treated with TNF-${\alpha}$ for 24 h. The effects of NM on monocyte adhesion, vascular cell adhesion molecule-1 (VCAM-1) and intracellular adhesion molecule-1 (ICAM-1) protein expression, p38 mitogenactivated protein kinase (MAPK) activation, and intracellular superoxide production were then examined. NM ($0.01{\sim}100{\mu}g/mL$) did not affect HUVEC viability; however, it inhibited the increases in reactive oxygen species (ROS) production and p66shc expression elicited by TNF-${\alpha}$ (3 ng/mL), and it dose dependently prevented the TNF-${\alpha}$ -induced upregulation of endothelial VCAM-1 and ICAM-1. In addition, it mitigated TNF-${\alpha}$ -induced p38 MAPK phosphorylation and the adhesion of U937 monocytes. These data suggest that NM mitigates TNF-${\alpha}$ -induced monocyte adhesion and the expression of endothelial cell adhesion molecules, and that the anti-adhesive effect of NM is mediated through the inhibition of p66shc, ROS production, and p38 MAPK activation.

• 한국임상수의학회지
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• 제31권6호
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• pp.469-476
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• 2014

본 연구는 염증상태에서의 CLA의 효과와 작용기전을 알아보기 위해 LPS-자극 RAW 264.7 macrophages에 있어 ROS 생성과 TNF-${\alpha}$ 생산, NF-${\kappa}B$ 및 $PPAR{\gamma}$ 활성을 검토하였다. t10c12-CLA는 LPS로 자극하지 않은 비염증시의 RAW 세포에서는 ROS 생성을 증가시켜 TNF-${\alpha}$ 생산을 유도하였으며, 이 효과는 $PPAR{\gamma}$ 활성화에 의존해서 NF-${\kappa}B$ 활성 증가에 의해 매개되었다. 반면, LPS로 자극한 염증조건의 RAW 세포에서는 t10c12-CLA가 $PPAR{\gamma}$ 활성화에 의존하지 않는 경로로 ROS 생성 및 과도한 TNF-${\alpha}$ 생산을 억제하였다. 본 결과로부터 CLA는 ROS 생성을 통해 TNF-${\alpha}$ 생산 및 NF-${\kappa}B$ 활성을 염증 유무에 따라 조절하는 것으로 사료되었다.

• Biomolecules & Therapeutics
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• 제28권2호
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• pp.172-183
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• 2020

Phosphoinositide 3-kinase (PI3K) is considered as a promising therapeutic target for rheumatoid arthritis (RA) because of its involvement in inflammatory processes. However, limited studies have reported the involvement of PI3KC2γ in RA, and the underlying mechanism remains largely unknown. Therefore, we investigated the role of PI3KC2γ as a novel therapeutic target for RA and the effect of its selective inhibitor, PBT-6. In this study, we observed that PI3KC2γ was markedly increased in the synovial fluid and tissue as well as the PBMCs of patients with RA. PBT-6, a novel PI3KC2γ inhibitor, decreased the cell growth of TNF-mediated synovial fibroblasts and LPS-mediated macrophages. Furthermore, PBT-6 inhibited the PI3KC2γ expression and PI3K/AKT signaling pathway in both synovial fibroblasts and macrophages. In addition, PBT-6 suppressed macrophage migration via CCL2 and osteoclastogenesis. In CIA mice, it significantly inhibited the progression and development of RA by decreasing arthritis scores and paw swelling. Three-dimensional micro-computed tomography confirmed that PBT-6 enhanced the joint structures in CIA mice. Taken together, our findings suggest that PI3KC2γ is a therapeutic target for RA, and PBT-6 could be developed as a novel PI3KC2γ inhibitor to target inflammatory diseases including RA.

• Tuberculosis and Respiratory Diseases
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• 제56권3호
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• pp.289-296
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• 2004

연구배경 : 기도 질환에서 점액이 과량 분비되는 경우 환자에게 불편함을 줄뿐만 아니라 기도 질환 예후에도 나쁜 영향을 미친다. 그러나, 기도 질환에서 점액이 과량 분비되는 것을 효과적으로 막는 방법이 없다. 점액의 성분 중 mucin은 당화 단백질로서 점액이 점성을 띄게 만드는 주요 성분이다. 본 연구를 통해서 mucin 분비 기전에 proteinase가 관여하는지 확인하고 만일 proteinase가 mucin 분비기전에 관여 한다면 어느 proteinase가 그런 역할을 하는지 확인하고자 하였다. 방 법 : (1) mucin 분비 억제 실험 군 특이적 proteinase 억제제를 사용하여 어느 군에 속하는 proteinase가 mucin 분비를 억제하는지 mucin을 생산하는 폐 세포주인 Calu-3를 이용하여 알아보았다. 군 특이적 proteinase 억제제로 PMSF(phenylmethylsulfonyl fluoride, serine proteinase inhibitor), E-64(cysteine proteinase inhibitor), Pepstatin(aspartic proteinase inhibitor), 1,10-Phenanthroline(metalloproteinase inhibitor)를사용하였다. 군 특이적 억제제를 Calu-3에 24시간동안 처리하여 분비된 mucin양을 enzyme linked immunoabsorbant assay(MUC5AC)로 정량하였고 그 결과를 대조군과 비교하였다. (2) Mucin 분비 자극 실험 Metalloproteinase 중에서 기도 질환 발병과 관련 있다고 알려진 matrix metalloproteinase-9 (MMP-9), MMP-12 그리고 TNF-alpha converting enzyme(TACE)를 Calu-3에 24시간 처리하여 분비된 mucin양을 enzyme linked immunoabsorbant assay (MUC5AC)로 정량하였고 그 결과를 대조군과 비교하였다. 결 과 : (1) 군 특이적 proteinase 억제제인 PMSF($10^{-4}M$), E-64($10^{-4}M$), Pepstatin($10^{-6}M$), 1,10-Phenanthroline($10^{-4}M$)는 MUC5AC 분비를 각각 $1{\pm}4.9%$(평균${\pm}$표준오차; 대조군과 비교 시 P=1.0), $-6{\pm}3.9%$ (P=0.34), $-13{\pm}9.7%$(P=0.34), $41{\pm}8.2%$(P=0.03) 감소시켰다(실험 회수 4번). (2) MMP-9(250ng/ml), MMP-12(100ng/ml), TACE(200ng/ml)에 의한 MUC5AC 분비량은 대조군에 비하여 각각 $103{\pm}6%$(P=0.39), $102{\pm}8%$(P=1.0), $107{\pm}13%$(P=0.39)이었다(실험 회수 6번). 결 론 : mucin 분비 기전에 metalloproteinase가 관여함을 시사하지만 MMP-9, MMP-12, TACE는 in vitro 모델에서 mucin 분비에 영향을 미치지 않았다.

• Journal of Applied Biological Chemistry
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• 제61권1호
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• pp.9-15
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• 2018

우리는 황련에서 분리된 베르베린의 화장품소재로의 활성을 평가하기 위해 항산화와 항주름 효능을 분석하였다. 시료의 독성을 평가하기 위해 3-(4,5-dimethyl-2-thiazolyl)-2,5-diphenyl-2H-tetrazolium bromide (MTT) assay를 수행하였으며 항산화와 항주름 활성평가를 위해 ROS 생성능, hyaluronic acid 생성능, MMP-2, -9, TIMP-1,-2, $TNF-{\alpha}$의 발현을 각각 측정하였다. 베르베린에 대한 자외선의 세포 독성은 CCD-986sk 섬유아세포를 이용하여 MTT 분석으로 측정되었고, 세포 독성이 $25{\mu}g/mL$의 농도 미만에서 관찰되지 않았다. 또한 베르베린이 ROS 생산을 농도 의존적으로 감소시키고 히알루론산의 합성을 촉진한다는 것을 발견했다. 베르베린은 주름 형성과 관련된 MMP-2 및 MMP-9의 protein 단계 및 mRNA 발현을 감소시키고 TIMP-1 및 TIMP-2의 발현을 증가시켰다. 프로 pro-inflammatory cytokine 으로 알려진 $TNF-{\alpha}$에 대한 베르베린의 억제효능을 실험한 결과, 농도의존적으로 $TNF-{\alpha}$ 유전자를 억제하는 효과를 나타내었다. 이러한 결과를 통해 베르베린은 활성산소종의 억제와 TIMP-1과 -2의 발현 조절에 의한 MMP-2,-9의 감소로 항산화 및 항주름에 효과가 있는 화장품 소재로서의 가치를 가진다고 사료된다.

• 한국식품과학회지
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• 제53권6호
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• pp.722-730
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• 2021

본 연구는 인삼열매로부터 RG-II 형태의 다당(GBW-II)을 분리하고 대식세포 활성화에 대한 세포 내 신호전달의 세부 기작을 규명함으로써 새로운 건강기능성식품 소재 개발을 위한 기초자료를 제시하고자 진행되었다. GBW-II의 구성당을 확인한 결과, 전형적인 RG-II의 구성당인 2-methyl-xylose, apiose, aceric acid, KDO 및 DHA와 같은 특이 구성당을 함유함을 확인할 수 있었다. GBW-II는 대식세포 유래 세포주인 RAW 264.7 cell에 처리하였을 경우, 어떠한 세포 독성도 확인되지 않았으나 IL-6와 TNF-α와 같은 cytokine의 분비는 농도 의존적으로 증가시키는 것으로 나타났다. 또한 RAW 264.7 cell을 이용한 세포 내 신호전달에 관한 실험 결과들을 종합해 볼 때, GBW-II는 대식세포 표면에 발현된 TLR2, TLR4 및 SR에 결합하여 MAPKs (p38, ERK) 및 NF-κB를 경유하여 IL-6와 TNF-α와 같은 cytokine의 분비를 증가시키는 것으로 최종 확인되었다. 한편, RG-I, RG-II, β-glucan, arabinoxylan 및 xyloglucan과 같은 식물체 유래 고분자 다당체의 약리활성은 그들의 구조적 차이에서 기원하는 것으로 알려져 있기 때문에 건강기능성식품 소재로의 개발을 위해서는 활성물질의 미세구조에 대한 해명이 필수적이라 할 수 있다. 따라서 본 연구진은 추후 연구에서 효소적 및 화학적 가수분해, methylation, sequencing 등을 이용하여 인삼열매 유래 정제 다당 GBW-II의 미세구조를 규명하고자 한다.