• Journal of Acupuncture Research
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• 제24권6호
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• pp.15-27
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• 2007

Objectives : The purpose of this study is to investigate the effectiveness of Ulmus davidiana Planch herbal acupuncture(UA) to inhibit nuclear $factor(NF)-{\kappa}B$ activation on type II collagen-induced arthritis (CIA) in mice. Methods : Using an in vitro test, the synoviocytes picked out from the experimental CIA mice were subcultured. The synoviocyte cells were treated with phorbol-12-myristate-13-acetate(PMA) for 1 hour prior to the addition of indicated concentrations($0.4\;-\;1.0mg/m{\ell}$) of UA, and the cells were further incubated for 24 hours. The in vivo test, $NF-{\kappa}B$ p65, inducible nitric oxide synthase(iNOS), cyclooxygenase-2(COX-2), vascular cell adhesion molecule(VCAM)-1 production and apoptosis was observed by immunohistochemical staining. Results : The PMA-induced $I{\kappa}B$ kinase(IKK), iNOS and COX-2 mRNA expression were dose-dependently decreased in UA treated synoviocytes. Using the in vivo test, the number of eosinophils in mice treated with UA noticeably decreased in the the CIA group(P<0.05 using student t test). In mice treated with UA, there was less cartilage erosion. less bone destruction, mild synovial hyperplasia, mild fibrosis, and mild angiogenesis with less MIP-2 production. By immunohistochemical staining, suppression of $NF-{\kappa}B$ p65, iNOS production, inhibition of COX-2 production, inhibition of VCAM-1 production and inducing apoptosis were observed. Conclusions : These results suggest that UA might be applicable to the therapy of RA to suppress $NF-{\kappa}B$ activation.

• Asian Pacific Journal of Cancer Prevention
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• 제16권14호
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• pp.5835-5842
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• 2015

Background: Melatonin, which is mainly produced by the pineal gland, has a good inhibitory effect on cell growth of multiple cancer types. However, the underlying molecular mechanisms of anti-tumor activity for colon cancer have not been fully elucidated. In this study, we investigated the effects of melatonin on migration in human colon cancer RKO cells and the potential molecular mechanisms. Materials and Methods: The viability of RKO cells was investigated by MTT assay after treatment with melatonin, SB203580 (p38 inhibitor) and phorbol 12-myristate 13-acetate (PMA, MAPK activator) alone or in combination for 48h. The effects of melatonin, and ML-7, a selective inhibitor of myosin light chain kinase (MLCK), and SB203580, and PMA on the migration of RKO cells were analyzed by in vitro scratch-wound assay. The relative mRNA levels of MLCK was assessed by real-time quantitative RT-PCR. Western blotting analysis was performed to examine the expression of MLCK, phosphorylation of myosin light chain (pMLC) and p38 (pp38). Results: The proliferation and migration of human colon cancer RKO cells were inhibited significantly after treatment with melatonin. The expression levels of MLCK and phosphorylation of MLC of RKO cells were reduced, and real-time quantitative RT-PCR showed that melatonin had significant effects on suppressing the expression of MLCK. Furthermore, the phosphorylation level of p38, which showed the same trend, was also reduced when cells were treated by melatonin. In addition, ML-7 (25umol/l) could down-regulate the phosphorylation of p38. Conclusions: Melatonin could inhibit the proliferation and migration of RKO cells, and further experiments confirmed that p38 MAPK plays an important role in regulating melatonin-induced migration inhibition through down-regulating the expression and activity of MLCK.

• 대한약침학회지
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• 제20권1호
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• pp.52-56
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• 2017

Objectives: Neutrophils represent the front line of human defense against infections. Immediately after stimulation, neutrophilic enzymes are activated and produce toxic mediators such as pro-inflammatory cytokines, nitric oxide (NO) and myeloperoxidase (MPO). These mediators can be toxic not only to infectious agents but also to host tissues. Because flavonoids exhibit antioxidant and anti-inflammatory effects, they are subjects of interest for pharmacological modulation of inflammation. In the present study, the effects of rutin on stimulus-induced NO and tumor necrosis factor $(TNF)-{\alpha}$ productions and MPO activity in human neutrophils were investigated. Methods: Human peripheral blood neutrophils were isolated using Ficoll-Hypaque density gradient centrifugation coupled with dextran T500 sedimentation. The cell preparations containing > 98% granulocytes were determined by morphological examination through Giemsa staining. Neutrophils were cultured in complete Roswell Park Memorial Institute (RPMI) medium, pre-incubated with or without rutin ($25{\mu}M$) for 45 minutes, and stimulated with phorbol 12-myristate 13-acetate (PMA). Then, the $TNF-{\alpha}$, NO and MPO productions were analyzed using enzyme-linked immunosorbent assay (ELISA), Griess Reagent, and MPO assay kits, respectively. Also, the viability of human neutrophils was assessed using tetrazolium salt 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl tetrazolium bromide (MTT), and neutrophils were treated with various concentrations of rutin ($1-100{\mu}M$), after which MTT was appended and incubated at $37^{\circ}C$ for 4 hour. Results: Rutin at concentrations up to $100{\mu}M$ did not affect neutrophil viability during the 4-hour incubation period. Rutin significantly decreased the NO and $TNF-{\alpha}$ productions in human peripheral blood neutrophils compared to PMA-control cells (P < 0.001). Also, MPO activity was significantly reduced by rutin (P < 0.001). Conclusion: In this in vitro study, rutin had an anti-inflammatory effect due to its inhibiting NO and $TNF-{\alpha}$ productions, as well as MPO activity, in activated human neutrophils. Treatment with rutin may be considered as a therapeutic strategy for neutrophil-mediated inflammatory/autoimmune diseases.

• Journal of Ginseng Research
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• 제33권4호
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• pp.330-336
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• 2009

G-Rb1에 의한 단핵구 세포주인 U937 세포와 대식세포주인 RAW264.7 세포의 유착조절 능, 세포표면 단백질 발현율 및 세포모양 변화능 조절효과를 조사하여 다음과 같은 결론을 얻었다. 1. G-Rb1은 CD29항체 (MEM101A) 및 CD43 항체(161-46) 처리에 의해 유도된 세포-세포간 유착현상을 20%에서 35%까지 유의적으로 상승하였다. 2. G-Rb1은 fibronectin처리에 의해 유도된 U937 세포-fibronectin간 유착현상을 상승시켰다. 3. G-Rb1은 CD29 및 CD43의 세포표면 발현 수준을 유의적으로 증가시켰으나 CD82의 발현은 억제시켰다. 4. G-Rb1은 PMA에 의해 유도된 형태변화는 촉진경향을 반면에 LPS에 의한 변화는 억제경향을 나타내었다. 인삼의 주요사포닌 성분인 G-Rb1의 약리작용이 활발히 연구되고 있다. 특별히, 본 화합물이 갖는 면역증강 및 항암 효능은 인삼이 갖는 이들 효과를 대신하는 것으로 보고 되어져 있다. 그러나, 세포유착과정이 다양한 염증과정, 알러지 반응 및 암세포의 이동과 전이성 등에 관련이 있다는 관점에서 볼 때 본 연구결과는 이들 약물이 갖는 치료기전을 설명하기에는 충분하지 않는 것으로 판단된다. 오히려, 본 결과는 G-Rb1이 특정 농도 ($25-50\;{\mu}M$ 수준)에서 세포의 활성을 촉진할 수 있다는 가능성을 시사한다고 하겠다. 특별히, G-Rb1의 세포유착 촉진 기능에 관한 기전 이해를 위해, 이후, 세포유착 유도 신호전달 단백질의 활성을 포함한 다양한 분자적 수준에서의 추가적인 실험들을 진행하고 한다.

• 대한수의학회지
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• 제38권1호
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• pp.29-42
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• 1998

$Mg^{2+}$ is one of the most abundant divalent cations in mammalian body(0.2~1.0mM) and the important physiological roles are : first, the cofactor of many enzyme activities, second, the regulator of glycolysis and DNA synthesis, third, the important role of bioenergetics by regulating of phosphorylation, fourth, the influence of cardiac metabolism and function. In this work we have investigated the regulation of the $Mg^{2+}$ induced by ${\alpha}_1-adrenoceptor$ stimulation in perfused guinea pig hearts and isolated myocytes. The $Mg^{2+}$ content of the perfusate or the supernatant was measured by atomic absorbance spectrophotometry. The elimination of $Mg^{2+}$ in the medium increased the force of contraction of right ventricular papillary muscles, and the left ventricular pressure. Phenylephrine also enhanced the force of contraction in the presence of $Mg^{2+}-free$ medium. ${\alpha}_1-Agonists$ such as phenylephrine and methoxamine were found to induce $Mg^{2+}$ efflux in both perfused hearts and myocytes. These effects were blocked by prazosin, an ${\alpha}_1-adrenoceptor$ antagonist. The $Mg^{2+}$ influx could also be induced by phenylephrine and R59022, a diacylglycerol kinase inhibitor. In the presence of protein kinase C(PKC) inhibitors, phenylephrine produced an increase in $Mg^{2+}$ efflux from perfused hearts. Furthermore, $Mg^{2+}$ efflux by phenylephrine was amplified by phorbol 12-myristate 13-acetate(PMA). This enhancement of $Mg^{2+}$ efflux by PMA was blocked by prazosin in perfused hearts. By contrast, the $Mg^{2+}$ influx could be induced by verapamil, nifedipine, ryanodine in perfused hearts, but not in myocytes. $W^7$, a $Ca^{2+}$/calmodulin antagonist, completely blocked the phenylephrine-induced $Mg^{2+}$ efflux in perfused hearts. In conclusion, $Mg^{2+}$ is responsible for the cardiac activity associated with ${\alpha}_1-adrenoceptor$ stimulation. The mobilization of $Mg^{2+}$ is decreased or increased by ${\alpha}_1-adrenoceptor$ stimulation in guinea pig hearts. These responses may be related specifically to the respective pathways of signal transduction. A decrease in $Mg^{2+}$ efflux by ${\alpha}_1-adrenoceptor$ stimulation in hearts can be through PKC dependent and intracellular $Ca^{2+}$ levels.

• Tuberculosis and Respiratory Diseases
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• 제77권2호
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• pp.65-72
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• 2014

Background: It is valuable to find the potential activity of regulating the excessive mucin secretion by the compounds derived from various medicinal plants. We investigated whether aqueous extract of the root bark of Morus alba L. (AMA), kuwanon E, kuwanon G, mulberrofuran G, and morusin significantly affect the secretion and production of airway mucin using in vivo and in vitro experimental models. Methods: Effect of AMA was examined on hypersecretion of airway mucin in sulfur dioxide-induced acute bronchitis in rats. Confluent NCI-H292 cells were pretreated with ethanolic extract, kuwanon E, kuwanon G, mulberrofuran G, or morusin for 30 minutes and then stimulated with phorbol 12-myristate 13-acetate (PMA) for 24 hours. The MUC5AC mucin secretion and production were measured by enzyme-linked immunosorbent assay. Results: AMA stimulated the secretion of airway mucin in sulfur dioxide-induced bronchitis rat model; aqueous extract, ethanolic extract, kuwanon E, kuwanon G, mulberrofuran G and morusin inhibited the production of MUC5AC mucin induced by PMA from NCI-H292 cells, respectively. Conclusion: These results suggest that extract of the root bark and the natural products derived from Morus alba L. can regulate the secretion and production of airway mucin and, at least in part, explains the folk use of extract of Morus alba L. as mucoregulators in diverse inflammatory pulmonary diseases.

• 생명과학회지
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• 제29권3호
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• pp.287-294
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• 2019

갯메꽃은 해안 사구에 분포하며 식물로 높은 환경 적응력을 가지고 있으며 갯메꽃은 항산화, 해열, 살균, 이뇨작용 등에 효과가 있는 것으로 알려져 있다. 본 연구에서는 암전이 과정에서 중요한 역할을 한다고 알려진 matrix metalloproteinases (MMPs)의 일종인 MMP-2와 MMP-9의 활성에 대한 갯메꽃의 억제효과를 methylene chloride (MC) 및 methanol (MeOH) 추출물과 조추출물, 유기용매 분획물을 시료로 하여 인체 섬유육종 HT-1080 세포를 이용하여 ELISA (Enzyme-linked immunosorbent), gelatin zymography, Wound healing assay, RT-PCR, western blot 실험을 통해 조사하였다. 갯메꽃의 MC 및 MeOH 추출물, 조추출물과 4가지 유기용매 분획물이 HT-1080 세포에서 MMP-2와 MMP-9 생성 억제 활성을 나타냄을 확인하였다. 그 중에서도 n-hexane, 85% aq.MeOH 분획물이 전반적으로 MMP-2와 MMP-9에 대한 높은 억제활성을 보였다. 이들 결과로부터 85% aq.MeOH과 n-hexane 분획물에 MMP 억제 활성이 높은 물질이 존재할 것으로 추측되어지며, 갯메꽃 추출물 및 분획물이 암침윤 및 전이를 억제하는 효과적인 항암소재로서의 가능성이 있음을 제시한다. 이에 본 연구팀은 85% aq.MeOH과 n-hexane 분획물에 있는 활성물질을 분리하기 위한 연구를 추가적으로 수행할 계획이다.

• Journal of Ginseng Research
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• 제47권1호
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• pp.97-105
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• 2023

Background: Hyperactivated airway mucosa cells overproduce mucin and cause severe breathing complications. Here, we aimed to identify the effects of saponins derived from Panax ginseng on inflammation and mucin overproduction. Methods: NCI-H292 cells were pre-incubated with 16 saponins derived from P. ginseng, and mucin overproduction was induced by treatment with phorbol 12-myristate 13-acetate (PMA). Mucin protein MUC5AC was quantified by enzyme-linked immunosorbent assay, and mRNA levels were analyzed using quantitative polymerase chain reaction (qPCR). Moreover, we performed a transcriptome analysis of PMA-treated NCI-H292 cells in the absence or presence of Rg5, and differential gene expression was confirmed using qPCR. Phosphorylation levels of signaling molecules, and the abundance of lipid droplets, were measured by western blotting, flow cytometry, and confocal microscopy. Results: Ginsenoside Rg5 effectively reduced MUC5AC secretion and decreased MUC5AC mRNA levels. A systematic functional network analysis revealed that Rg5 upregulated cholesterol and glycerolipid metabolism, resulting in the production of lipid droplets to clear reactive oxygen species (ROS), and modulated the mitogen-activated protein kinase and nuclear factor (NF)-kB signaling pathways to regulate inflammatory responses. Rg5 induced the accumulation of lipid droplets and decreased cellular ROS levels, and N-acetyl-ⳑ-cysteine, a ROS inhibitor, reduced MUC5AC secretion via Rg5. Furthermore, Rg5 hampered the phosphorylation of extracellular signal-regulated kinase and p38 proteins, affecting the NF-kB signaling pathway and pro-inflammatory responses. Conclusion: Rg5 alleviated inflammatory responses by reducing mucin secretion and promoting lipid droplet-mediated ROS clearance. Therefore, Rg5 may have potential as a therapeutic agent to alleviate respiratory disorders caused by hyperactivation of mucosa cells.

• 대한한방부인과학회지
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• 제34권3호
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• pp.29-48
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• 2021

Objectives: Banchong-san (BCS) is a herbal formula composed of 13 korean medicinal herbs and is traditionally used to treat inflammatory diseases and pain. The object of this study was to research the antioxidant, anti-inflammatory, antipruritic and antimicrobial effects of the BCS in lipopolysaccharide (LPS)-stimulated RAW 264.7 macrophages. Methods: In this experiment, effects of BCS on the following four were measured as follows: (1) Anti-oxidative effects were evaluated by 1,1-diphenyl-2-picryl-hydrazyl (DPPH) Radical scavenging activity, 2,2-azino-bis-3-ethylbenzothiazoline-6-sulfonic acid (ABTS) Radical scavenging activity. (2) Anti-inflammatory effects were evaluated by the production amount of Reactive oxygen species (ROS), Nitric oxide (NO), Interleukin-1β (IL-1β), Interleukin 6 (IL-6), tumor necrosis factor-α (TNF-α), Prostaglandin E2 (PGE₂), inducible nitric oxide synthase (iNOS), cyclooxygenase-2 (COX-2)(the previous two are "mRNA"), extracellular signal-regulated kinase (ERK), c-Jun N-terminal kinase (JNK), p38 mitogen-activated protein kinases (p38), inhibitor of nuclear factor kappa B (IκBα), nuclear factor kappa B (NF-κB) (the previous five are "Protein") in LPS-Stimulated RAW 264.7 cells. (3)Antipruritic effects were evaluated by the production amount of histamine, Leukotriene B4 (LTB4), LeukotrieneC4 (LTC4) Levels in phorbol 12-myristate 13-acetate(PMA)/ionomycin-stimulated MC/9 mast cell. (4) Anti-microbial effects were evaluated by the growth suppression of Escherichia coli, Staphylococcus aureus, Pseudomonas aeruginosa and Aspergillus niger. Results: The following results were obtained through each measurement: (1) DPPH Radical Scavenging Activity, ABTS Radical Scavenging Activity evoked a significant concentration-dependent increase. (2) ROS, NO, IL-1β, IL-6, TNF-α, PGE₂ production amount, iNOS, COX-2 mRNA expression were significantly reduced in the BCS extraction group compared with the control group and significantly decreased the amount of ERK, JNK, p38, NF-κB Protein expression. The amount of IκB-α Protein Expression have increased significantly. (3) The amounts of histamine, LTB4, LTC4 were significantly decreased. (4) The antibacterial efficacy, BCS inhibited the growth of Escherichia coli, Pseudomonas aeruginosa at concentrations of 5 ㎍/ml, but did not suppress the growth of staphylococcus aureus and aspergillus niger. Conclusions: The experimental results show that BCS has anti-oxidant, anti-inflammatory, antipruritic and antimicrobial properties.

• 대한본초학회지
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• 제29권6호
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• pp.117-123
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• 2014

Objectives : Sinhyowoldo-san (SHWDS) is said to be a traditional medicine used for shigellosis, abdominal pain, diarrhea. But mechanism of SHWDS mediated-modulation of immune function is not sufficiently understood. To ascertain the molecular mechanisms of SHWDS 70% EtOH extract on pharmacological and biochemical actions in inflammation, we researched the effect of pro-inflammatory mediators in phorbol-12-myristate-13-acetate (PMA)+ A23187-activated human mast cell line (HMC-1). Methods : In the present research, cell viability was measured by MTS assay. pro-inflammatory cytokine production was measured by performing enzyme-linked immunosorbent assay (ELISA), reverse transcription polymerase chain reaction (RT-PCR), and western blot analysis to analyze the activation of mitogen-activated protein kinases (MAPKs), nuclear factor kappa-light-chain-enhancer of activated B cells ($NF-{\kappa}B$). The investigation focused on whether SHWDS inhibited the expressions of interleukin-6 (IL-6), interleukin-8 (IL-8), MAPKs and $NF-{\kappa}B$ in PMA+A23187-activated HMC-1 cells. Results : SHWDS has no cytotoxicity at measured concentration (50, 100, and $250{\mu}g/ml$). SHWDS ($250{\mu}g/ml$) inhibits pro-inflammatory cytokine expression in PMA+ A23187-activated HMC-1 cells. Moreover, SHWDS inhibited cyclooxygenase (COX)-2 expression. In activated HMC-1 cells, SHWDS suppressed phosphorylation of extracellular signal-regulated kinase (ERK 1/2) and c-jun N-terminal Kinase (JNK 1/2). Then, SHWDS suppressed activation of nuclear factor $NF-{\kappa}B$ in nuclear, degradation of IkB ${\alpha}$ in cytoplasm. Conclusions : We propose that SHWDS has an anti-inflammatory therapeutic potential, which may result from inhibition of ERK 1/2, JNK 1/2 phosphorylation and $NF-{\kappa}B$ activation, thereby decreasing the expression of pro-inflammatory genes.