• The Korean Journal of Physiology and Pharmacology
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• v.3 no.4
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• pp.375-382
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• 1999

Growing evidence indicates that enhanced generation or actions of nitric oxide (NO) are implicated in the pathogenesis of hypertension in spontaneously hypertensive rats and diabetic nephropathy in streptozotocin (STZ)-induced diabetic rats. We investigated whether inducible nitric oxide synthase (iNOS) expression in STZ-induced diabetic rats is responsible for the alterations of vascular reactivity. Diabetic state was confirmed 28 days after injection of STZ (i.p) in rats by measuring blood glucose. In order to evaluate whether short term (4 weeks) diabetic state is related with altered vascular reactivity caused by iNOS expression, isometric tension experiments were performed. In addition, plasma nitrite/nitrate (NOx) levels and expression of iNOS in the lung and aorta of control and STZ-treated rats were compared by using Griess reagent and Western analysis, respectively. Results indicated that STZ-treated rats increased the maximal contractile response of the aorta to phenylephrine (PE), and high $K^+,$ while the sensitivity remained unaltered. Endothelium-dependent relaxation, but not SNP-mediated relaxation, was reduced in STZ-treated rats. Plasma nitrite/nitrates are significantly increased in STZ-treated rats compared to controls. The malondialdehyde (MDA) contents of liver, serum, and aorta of diabetic rats were also significantly increased. Furthermore, nitrotyrosine, a specific foot print of peroxynitrite, was significantly increased in endothelial cells and smooth muscle layers in STZ-induced diabetic aorta. Taken together, the present findings indicate that enhanced release of NO by iNOS along with increased lipid peroxidation in diabetic conditions may be responsible, at least in part, for the augmented contractility, possibly through the modification of endothelial integrity or ecNOS activity of endothelium in STZ-diabetic rat aorta.

• The Korean Journal of Physiology and Pharmacology
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• v.10 no.3
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• pp.161-165
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• 2006

NO and cyclooxygenase-2 (COX-2) are contributes to vascular inflammation induced by various stimulation. The mechanism, which explains a linkage between NO and COX-2, could be of importance in promoting pathophysiological conditions of vessel. We investigated the effects of NO donors on the COX-l and COX-2 mRNA/protein expression, as well as the nitrite production in culture medium of vascular smooth muscle cell (VSMC). VSMC was primarily cultured from thoracic aorta of rat. In this experiments, COX-l and COX-2 mRNA/protein expressions were analysed and nitrite productions were investigated using Griess reagent. VSMC did not express COX-2 protein in basal condition (Nonlipopolysaccharide (LPS) stimulated). In LPS-stimulated experiments, after 3 hours of NO donor pretreatment, LPS $10{\mu}g/ml$ was treated for 24 hours. COX-l protein expressions were unchanged by SNP and NOR-3. NOR-3 significantly increased COX-2 mRNA/protein expression under LPS stimulation. In contrast, SNP did not increase COX-2 mRNA/protein expression under LPS stimulation. Nitrite production was higher in NOR-3 treatment than SNP treatment under LPS stimulation. These results suggest that the expression of COX-2 in VSMC is regulated by NOR-3, COX-2 expressions were depending on the types of NO donor and LPS stimulation in VSMC.

• Nutrition Research and Practice
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• v.8 no.3
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• pp.267-271
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• 2014

BACKGROUND/OBJECTIVES: Overproduction of nitric oxide (NO) by the inducible nitric oxide synthase (iNOS) enzyme can cause inflammation. Cyclooxygenase-2 (COX-2) is also involved in the inflammatory response through regulation of nuclear factor-kappa B $NF-{\kappa}B$(). Areca catechu is one of the known fruit plants of the Palmaceae family. It has been used for a long time as a source of herbal medicine in Indonesia. In this study, we explored the effect of Indonesian Areca catechu leaf ethanol extract (ACE) in lipopolysaccharide (LPS)-induced inflammation and carrageenan-induced paw edema models. Recently, this natural extract has been in the spotlight because of its efficacy and limited or no toxic side effects. However, the mechanism underlying its anti-inflammatory effect remains to be elucidated. MATERIALS/METHODS: We measured NO production by using the Griess reagent, and determined the expression levels of inflammation-related proteins, such as iNOS, COX2, and $NF-{\kappa}B$, by western blot. To confirm the effect of ACE in vivo, we used the carrageenan-induced paw edema model. RESULTS: Compared to untreated cells, LPS-stimulated RAW 264.7 cells treated with ACE showed reduced NO generation and reduced iNOS and COX-2 expression. We found that the acute inflammatory response was significantly reduced by ACE in the carrageenan-induced paw edema model. CONCLUSION: Taken together, these results suggest that ACE can inhibit inflammation and modulate NO generation via downregulation of iNOS levels and $NF-{\kappa}B$ signaling in vitro and in vivo. ACE may have a potential medical benefit as an anti-inflammation agent.

• The Journal of Korean Medicine Ophthalmology and Otolaryngology and Dermatology
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• v.24 no.1
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• pp.64-77
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• 2011

Objectives : 판람근(板藍根)은 십자화과에 속하는 대청(大靑) 또는 숭남의 근(根)을 건조한 것이다. 본 연구는 판람근(板藍根)이 청열해독(淸熱解毒)함에 근거하여, LPS로 활성화된 Raw264.7 cell에서 판람근(板藍根)과 그 성분중의 하나인 tryptanthrin이 염증매개물질에 미치는 효과를 살펴보고자 하였다. Methods : 세포생존율은 MTT, nitric oxide (NO)는 Griess reagent를 사용하여 측정하였으며, 각 단백질의 발현량은 Western blot 방법을 사용하였으며, cytokine 및 cyclooxygenase-2 (COX-2)는 ELISA방법을 사용하여 측정하였다. Results : LPS는 NO 및 prostaglandin E2 (PGE2)를 유의하게 상승시켰으며, 판람근(板藍根)추출물 (IRE) 및 tryptanthrin 은 이들을 유의하게 억제하였다. 그러나 판람근(板藍根)의 또 다른 성분인 indigo는 유의한 결과를 나타내지 못하였다. IRE와 tryptanthrin은 inhibitory kappa B alpha의 인산화를 억제하여, nuclear factor-${\kappa}$B (NF-${\kappa}$B)의 핵으로의 전위(轉位)를 억제하여, iNOS 및 cytokine을 억제하였다. IRE와 tryptanthrin의 PGE2 억제는, COX-2의 발현억제에서가 아니라, COX-2의 활성을 억제함에서 기인하였다. Conclusion : 이러한 결과는 판람근(板藍根)이 NF-${\kappa}$B pathway를 경유하여 iNOS의 발현 및 COX-2의 활성을 억제함을 나타내며, 이러한 판람근(板藍根)의 항염증효능은 일부 tryptanthrin의 작용에서 기인함을 시사한다.

• Korean Journal of Medicinal Crop Science
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• v.20 no.5
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• pp.381-386
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• 2012

Eucommia ulmoides OLIVER (EU) is a traditional Korean herbal used for the treatment of rheumatoid arthritis (RA). In the present study, the molecular pharmacology basis of its anti-inflammatory effect is revealed in this work, EU was studied in lipopolysaccharide (LPS)-activated macrophage cells (RAW 264.7) as an established inflammation model. After activation, nitric oxide (NO) production and iNOS mRNA were measured by using a colorimetric assay (Griess reagent), and reverse transcription polymerase chain reaction (RT-PCR), respectively. The change in the content of $PGE_2$, $TNF{\alpha}$, and IL-6 was concurrently monitored by ELISA. In results, we found that in the concentration range without showing cytotoxicity, EU produced a remarkable anti-inflammatory effect and showed a dose-dependent inhibition of LPSinduced NO production. Compared with indomethacin, EU has more potency and a specific action of NO inhibition, $PGE_2$, IL-6, and TNF-${\alpha}$ inhibition. These results suggest that EU may be a suitable herbal medicine to yield the greatest anti-inflammatory activity for food additives and medicine.

• The Korea Journal of Herbology
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• v.21 no.2
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• pp.135-142
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• 2006

Objectives : On-Bi-Tang(OB) has been prescribed Chinese traditional medicine for the treatment of inflammatory disease such as chronic renal failure. In this study, we investigated the anti-inflammatory effect of OB extract in the BV2 murine microglial cells. Methods : After the water extract of OB was treated in BV2 cells, murine microglial line, the cell viability was measured by MTT assay. The production of nitric oxide (NO) and $TNF-{\alpha}$ was determined based on Griess reagent and enzyme linked immunosorbant assay (ELISA). mRNA expression of inducible nitric oxide synthase (iNOS) and $TNF-{\alpha}$ was analyzed by RT-PCR. Results : OB extract significantly inhibited the LPS-induced production of NO and TNF-a in BV2 cells. OB extract also suppressed the mRNA expression of iNOS and $TNF-{\alpha}$ in BV2 cells activated with LPS. Conclusion : These data suggests that OB extract may have the anti-inflammatory effect through the modulation of NO production and inflammatory cytokine such as $TNF-{\alpha}$.

• The Korea Journal of Herbology
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• v.30 no.4
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• pp.71-79
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• 2015

Objectives : These present study was designed to investigate the relaxation effects of Alpiniae Oxyphyllae Fructus(AOF) on isolated corpus cavernosum smooth muscle.Methods : Rabbit corpus cavernous tissues were prepared in strip. Then relaxation responses of AOF at 0.01-3 ㎎/㎖ in contracted strips induced by phenylephrine(PE) were measured. To evaluate mechanisms, indomethacin(IM) tetraethylammonium chloride(TEA), Nω-nitro-L-arginine(_L-NNA), methylene blue(MB) were treated before AOF extract(0.1-3 ㎎/㎖) infused into precontracted strips induced by PE. And 1 mM Ca²⁺was infused into precontracted strips after pretreatment of AOF extract(3 ㎎/㎖) in Ca²⁺-free krebs-ringer solution. NO concentration was measured by Griess reagent system. Ratio of smooth muscles to collagen fibers and eNOS positive reaction were measured by histocheminal and immunohistochemical process.Results : The cavernous strips were significantly relaxed by AOF extract 0.1, 0.3, 1, 3 ㎎/㎖ and the pretreatment with IM 10 μM,_L-NNA 100 μM, MB 10 μM inhibited relaxation of AOF compared to non-pretreatment, but the pretreatment with TEA 100 μM didn't affect relaxation of AOF. In a Ca²⁺-free solution, pretreatment with AOF reduced increase on contraction of strips by Ca²⁺supply than non-pretreatment. On HUVEC, NO concentration was increased. On corpus cavernosum of penis in Spontaneous Hypertensive Rat, ratio of smooth muscles to collagen fibers and eNOS positive reaction in AOF group were increased compared to PE groupConclusions : Taken this results, we can suggest that AOF extract exerts a relaxation effects on rabbit corpus cavernosum smooth muscle in part by suppressing influx of extracellular Ca²⁺throughout prostacyclin, the NO-cGMP system.

• The Korea Journal of Herbology
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• v.31 no.1
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• pp.69-75
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• 2016

Objectives : The aim of this study is to investigate the various effects of individual or combined extract of GamiSaengmaeksan (GSS) on cell viability, anti-inflammatory and antioxidant activityMethods : In order to evaluate cytotoxicity, MTT assay was performed. We investigated the levels of proinflammatory cytokines such as tumor necrosis factor (TNF)-α and interleukin (IL)-6 and interleukin (IL)-1β, and nitric oxide(NO) in LPS-induced RAW 264.7 cells to check the effects on anti-inflammatory activity. The level of NO production in RAW 264.7 cells was measured by using Griess reagent. The levels of cytokines and ROS were measured by Luminex and Flow cytometry, respectively.Results : At concentration of 200 ㎍/㎖ GSS, cytotoxicity was observed in RAW 264.7 cells. However, at concentration less than 100 ㎍/㎖ of both combine and individual GSS, cytotoxicity was not observed in Raw 264.7 cells. However, the level of ROS in RAW 264.7 cells were decreased at both extract of 100 ㎍/㎖ GSS. Also, the level of NO in RAW 264.7 cells were decreased from extraction of concentration of 100 ug/ml in GSS and individual-extraction of Liriopis Tuber, White Ginseng and Glycyrrhizae Radix. In addition, productions of pro-inflammatory cytokines (TNF-α) in LPS-induced RAW 264.7 cells were decreased from extraction of concentration of 10 and 100 (㎍/㎖) in GSS and individual-extraction of Liriopis Tuber.Conclusions : It is concluded that combined extract of GSS appears to be more effective in anti-oxidation and anti-inflammatory effect than those in individual-extraction of GSS. These results may be developed as a raw material for new therapeutics to ease the symptoms related with inflammatory and oxidative stress.

• 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.

• The Journal of Korean Medicine Ophthalmology and Otolaryngology and Dermatology
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• v.29 no.3
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• pp.95-105
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• 2016

Objectives : Allergic diseases have a various symptoms of hyperresponsiveness and recently hyperresponsive reaction in the chronic phase is reported as the important mechanisms. Cheonggisan(CGS) is used in oriental clinics for curing various skin diseases due to effect of controlling of pruritus. There have been studies on the anti-allergic effect and anti-inflammatory effect of CGS, but there had no study of anti-allergic effects in allergic late inflammation of CGS, so we aimed to find out the effects of CGS in allergic late inflammation in our study.Methods : To investigate the anti-allergy effect and anti-inflammatory effect of CGS, RAW 264.7 macrophage cells and CSG water-extracts were used. Cytotoxic effect of CSG was examined by MTT assay, an oxidative product of NO was measured in the culture medium by the Griess reagent assay. The level of prostaglandin E2(PGE2) was measured by competitive enzyme-linked immunoassay. Cytokine(PGE2, IL-1β, IL-6, TNF-α) was measured by Bio-Plex suspension assay system and quantitative multiplexed cytokine/chemokine assay.Results : We investigated that there was no cytotoxic effect of CGS water-extract at any levels of concentration on RAW 264.7 macrophage cells by MTT assay. CGS water-extracts significantly suppressed the levels of the inflammatory mediators such as NO and PGE2, cytokine of IL-1β, TNF-α at the level of 400 ㎍/㎖ CGS concentration. But there was no significant effect on IL-6 production suppression.Conclusions : These results suggest that CSG water-extract has and anti-inflammatory effects in allergic reaction. These properties may contribute to the allergic diseases and inflammatory related disease care.