• The Korea Journal of Herbology
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• v.23 no.2
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• pp.179-192
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• 2008

Objectives : Draconis Resina (DR) has been used as a traditional Korean herbal medicine since ancient times, and today it is used as a medication for wounds, tumors, diarrhea, rheumatism, in the itching of insect bites and with other conditions in the folk medicine. The aim of this study was to determine whether fractionated extracts of DR inhibit free radical generation, intracellular oxidation, production of nitrite, an index of NO, PGE2, iNOS, COX-2 and proinflammatory cytokines in lipopolysaccharide (LPS)-treated RAW 264.7 macrophages, Methods : DR extract prepared with methanol, and then fractionated with hexane, dichloromethane, ethylacetate, n-butanol and water. Inhibitory effect of DR onto free radical generation was determined by measuring DPPH, superoxide anions and nitric oxide scavenging activities in vitro. Cytotoxic activity of extracts on RAW 264.7 cells was measured using 5-(3-caroboxymeth-oxyphenyl)-2H-tetra-zolium inner salt (MTS) assay. Intracelluar oxidation was analysed by DCF-DA assay. The nitric oxide (NO) production was measured by Griess reagent system. The levels of iNOS and COX-2 expression were confirmed by western blot. And proinflammatory cytokines were measured by ELISA kit. Results : Our results indicated that fractionated extracts, especially dichloromethane and ethyl acetate extracts, significantly inhibited free radical generation, the LPS-induced H202, NO, PGE2 production and iNOS, COX-2 expression accompanied by an attenuation of TNF-${\alpha}$, IL-$1{\beta}$ and IL-6 formation in macrophages. Conclusions : Our results indicate that dichloromethane and ethyl acetate extracts of DR have potential as an agent of chronic inflammatory diseases.

• The Korea Journal of Herbology
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• v.29 no.6
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• pp.21-26
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• 2014

Objectives : Taraxacum coreanum (TC) have been used as a traditional medicine to treat inflammatory diseases and anti-oxidant effect in Korea. However, the anti-inflammatory effect of TC water extract on lipopolysaccharide (LPS)-induced inflammation is not well-known. Therefore, this study was performed to identify the anti-inflammatory effect of TC on LPS induced inflammatory. Methods : RAW 264.7 cells were treated with 500 ng/mL of LPS. Water extracts of TC (0.1, 0.25, 0.5 mg/ml) was treated 1 h prior to LPS. Cell viability was measured by MTT assay. Levels of nitric oxide (NO) were measured with Griess reagent and pro-inflammatory cytokines were measured by enzyme-linked immunosorbent assay (ELISA) and real-time polymerase chain reaction (real-time PCR). We also examined molecular mechanisms such as mitogen-activated protein kinases (MAPKs) and nuclear factor-B ($NF-{\kappa}B$) activation by western blot. Results : Water Extract from TC itself did not have any cytotoxic effect in RAW 264.7 cells. TC treatment inhibited the production of NO production, and pro-inflamamtory cytokines such as interleukin (IL)-6 and $IL-1{\beta}$ on protein and mRNA levels. In addition, TC treatment inhibited the LPS-induced activation of MAPKs such as extracellular signal-regulated kinase1/2 (ERK1/2), p38 kinases (p38), c-Jun $NH_2$-terminal kinase (JNK) and $NF-{\kappa}B$. Conclusions : In summary, our result suggest that treatment of TC could reduce the LPS-induced inflammation. Thereby, TC could be used as a protective agent against inflammation. Also, this study could give a clinical basis that TC could be a drug or agent to prevent inflammation.

• Imaging Science in Dentistry
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• v.35 no.3
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• pp.127-131
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• 2005

Purpose : We performed the present study to investigate whether osteotropic hormomes play roles on the nitric oxide (NO) production in culture of ROS 17/12.8 osteoblastic cells. Materials and Methods : The osteoblastic cell line ROS17/2.8 cells were cultured In F12 medium supplemented with $5\%$ fetal bovine serum (FBS) at $37^{\circ}C$ in a humidified atmosphere of $5\%\;CO_2$ in air. ROS17/2.8 cells were plated in 96-well plates at a density of $2-3\times10^3cells/well$ and grown to confluence. Then the cells were pretreated with osteotropic hormones (parathyroid hormone (PTH) 20-500 ng/mL, 1, 25-dihydroxycholecalciferol $(1,\;25[OH]_2D_3)$ 1-100 nM; prostaglandin $E_2 (PGE_2)$ 20-500 ng/mL in the medium supplemented with $0.4\%$ FBS for 72 hours and the cells were treated with cytokines $(TNF{\alpha}\;and\;IFN{\gamma})$ in phenol red-free F12 medium for an additional 48 hours. NO synthesis was assessed by measuring the nitrite anion concentration, the reaction product of NO, in the cell culture medium using Griess reagent. Results : PTH and $1,\;25[OH]_2D_3$ pretreatment induced a significant increase in NO production in the presence of $TNF{\alpha}\;and\;IFN{\gamma}.\;PGE_2$ slightly induced NO production compared to the control group. But, $PGE_2$ pretreatment did not affect in NO production in the presence of $TNF{\alpha}\;and\;IFN{\gamma}$. Conclusions : These results suggest that the actions of osteotropic hormones In bone metabolism may be partially mediated by NO in the presence of cytokines.

• Journal of Physiology & Pathology in Korean Medicine
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• v.27 no.5
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• pp.602-610
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• 2013

This study was aimed to evaluate the cavernosal relaxation effect of Crataegii fructus(CF) in the contracted rabbit penile corpus cavernosum by agonists.In order to study the effect of CF on the vasoconstriction of rabbit penile corpus cavernosum, isolated rabbit penile corpus cavernosum tissues were used for the experiment using organ baths containing Krebs solution.To investigate the cavernosal relaxation of CF, CF extract at $0.01{\sim}3.0mg/m{\ell}$ was added after penile corpus cavernosum were contracted by norepinephrine(NE) $1{\mu}M$. To analyze the mechanism of CF's vasorelaxation, CF extract infused into contracted penile tissues by NE after each treatment of indomethacin(IM), $N{\omega}$-nitro-L-arginine(L-NNA), methylene blue(MB), tetraethylammonium chloride(TEA).To study the effect of CF on influx of extracellular calcium chloride($Ca^{2+}$) in penile tissues, in $Ca^{2+}$-free krebs solution, $Ca^{2+}$ 1 mM infused into contracted penile tissues by NE after pretreatment of CF. Cytotoxic activity of CF on human umbilical vein endothelial cell(HUVEC) was measured by MTT assay, and nitric oxide(NO) prodution was measured by Griess reagent. CF relaxed cavernosal strip with endothelium contracted by NE, but in the strips without endothelium, CF-induced relaxation was significantly inhibited. The pretreatment of L-NNA, MB, TEA decreased significantly on the cavernosal relaxation than not-treatment of them. But the pretreatment of IM had no significant effect on the cavernosal relaxation. In $Ca^{2+}$-free krebs solution, when $Ca^{2+}$ infused into contracted penile tissues by NE, pretreatment of CF inhibit contraction induced by adding $Ca^{2+}$.NO production wasn't increased by treatment of CF on HUVEC. This findings showed that CF is effective for the relaxation of rabbit penile corpus cavernosum, and we suggest that CF relax rabbit corpus cavernosal smooth muscle through multiple action mechanisms that include increasing the release of nitric oxide from corporal sinusoidal endothelium, inhibition of $Ca^{2+}$ mobilization into cytosol from the extracellular fluid, and maybe a hyperpolarizing action.

• The Korea Journal of Herbology
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• v.25 no.4
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• pp.11-16
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• 2010

Objective : Oriental medicines have been combined oriental medical theory which based on the seven modes of emotions. Notopterygium incisum (N. incisum) and Clematis manshurica (C. manshurica) have been used as an anti-rheumatic and analgesic medicine for the treatment of rheumatism, headache, cold, etc. In this study, we evaluate the synergistic anti-inflammatory effect of N. incisum and C. manshurica. Method : To evaluate the synergistic anti-inflammatory effect of a herbal mixture N. incisum and C. manshurica, we examined the changed ear thickness in 12-O-tetradecanoyl-phorbol-13-acetate (TPA)-induced mouse ear edema model after topical application of herbal mixture. In addition, the levels of markers for inflammation, such as tumor necrosis factor (TNF)-${\alpha}$, interleukin (IL)-$1{\beta}$, prostaglandin $G_2$ ($PGE_2$), and nitric oxide (NO) were measured by ELISA assay and Griess reagent in lipopolysaccharide-stimulated Raw 264.7 cells. Results : Our results showed that aqueous extracts of N. incisum and C. manshurica combination significantly inhibited the mouse ear edema induced by TPA. Moreover, the aqueous extracts of N. incisum and C. manshurica combination exhibited synergistic effects in down-regulating TNF-${\alpha}$, IL-$1{\beta}$, $PGE_2$ level, but not NO. Conclusions : This study suggested that combined treatment of N. incisum and C. manshurica, based on seven methods in prescription compatibility, has a synergistic effect in down-regulating inflammatory response both in vitro and in vivo models.

• Herbal Formula Science
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• v.28 no.4
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• pp.339-349
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• 2020

Objectives : Wiryeong-tang (WRT) is a traditional herbal medicine used to treat kidney-related diseases. However, the anti-inflammatory and anti-gastritis effect of Wiryeong-tang was not well known. Therefore, we experimented to confirmed the anti-inflammatory and anti-gastritis effects of Wiryeong-tang. Methods : The RAW 264.7 cells were pre treated with Wiryeong-tang mix soft extract (WRT-mse; 50, 100 and 200 ㎍/mL) for 1 hrs, and then incubated with lipopolysaccharide (LPS; 500 ng/mL). Cell viability was measured by the MTT method, and nitric oxide (NO) was measured with griess reagent. In addition, pro-inflammatory cytokines were measured by enzyme-linked immunosorbent assay (ELISA) and reverse transcription-polymerase chain reaction (RT-PCR). For anti-gastritis effect in vivo, acute gastritis was induced using 150 mM HCl/60% ethanol used ICR mice. WRT-mse (133 mg/kg) was pre treated for 3 days and then treated with 150 mM HCl/60% ethanol 1 hrs later. Then gastritis was observed and inflammatory cytokines in the gastric tissue was measured. Results : The 8 marker components of the WRT-mse were determined by simultaneous analysis using HPLC. WRT-mse was not toxic and inhibited pro-inflammatory cytokines such as IL-1β, IL-6 and TNF-α at NO production, protein and mRNA levels. Also, it was confirmed that WRT-mse improved bleeding and edema in gastritis, and suppresses inflammatory cytokines. Conclusion : In summary, our results suggest that the treatment of the WRT-mse reduced and improved the 150 mM HCl/60% ethanol induced acute gastritis and the inflammation caused by LPS stimulation in RAW 264.7 cells. Therefore, this study may provide useful drug or clinical evidence for WRT-mse to prevent inflammation.

• Journal of Physiology & Pathology in Korean Medicine
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• v.27 no.4
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• pp.400-408
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• 2013

This study aimed to investigate the relaxation effects and its underlying mechanisms of Rubus coreanus(RC) extract in contracted rabbit corpus cavernous tissues by phenylephrine(PE) $1{\mu}M$. In order to define the relaxation effects of RC, rabbit corpus cavernous tissues were prepared in $2{\times}2{\times}6mm$ sized strip. The dose-dependent relaxation responses of RC at 0.01-3.0 $mg/m{\ell}$ in contracted strips induced by PE were measured and also observed after endothelial denudation. To analyze the underlying mechanisms of RC-induced relaxation, indomethacin(IM), tetraethylammonium chloride(TEA), $N{\omega}$-nitro-L-arginine (L-NNA), methylene blue(MB) were treated before RC extract infused into precontracted strips induced by PE. To study the effect of RC extract on influx of extracellular $Ca^{2+}$ in corpus cavernous strips, calcium chloride(Ca) 1 mM infused into precontracted strips induced by PE after pretreatment of RC extract in $Ca^{2+}$-free krebs-ringer solution. To investigate cytotoxic activity and nitric oxide(NO) concentration of RC extract on human umbilical vein endothelial cell(HUVEC), cell viability on HUVEC was measured by MTT assay, and NO concentration was measured by Griess reagent system. The cavernous strips were significantly relaxed by RC extract at 1.0 $mg/m{\ell}$, 3.0 $mg/m{\ell}$ and the relaxation responses to RC were inhibited significantly by endothelial disruption. The pretreatment of IM, TEA didn't affect RC extract-induced endothelium-dependent relaxation, but the pretreatment of L-NNA, MB reduced RC extract-induced endothelium-dependent relaxation. When $Ca^{2+}$ was supplied the cavernous strips which were precontracted by PE in a $Ca^{2+}$-free krebs-ringer solution, contraction of strips was increased, but pretreatment of RC inhibited contractile response to $Ca^{2+}$. When RC extract was applicated on HUVEC, NO concentration was increased. Our findings show that RC extract exerts a relaxing effect on corpus cavernosum in part by suppressing influx of extracellular $Ca^{2+}$ through activating the NO-cGMP system.

• Journal of Pharmacopuncture
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• v.12 no.4
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• pp.89-96
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• 2009

Objectives : The purpose of this study is to investigate the effect of Water Extract from Lactobacillus pentosus-fermented ARTEMISIAE ARGI FOLIUM (AFL) on nitric oxide production of mouse macrophage Raw 264.7 cells impaired by various toxicants such as gallic acid, EtOH, nicotine, acetaminophen, and acetaldehyde. Methods : ARTEMISIAE ARGI FOLIUM was fermented with Lactobacillus pentosus and extracted by water. Nitric oxide production of mouse macrophage Raw 264.7 cells was measured by Griess reagent assay. Examined concentrations of AFL were 10, 50, 100, 200, 400 ug/mL. Results : The results of the experiment are as below. 1. AFL at the concentration of 400 ug/mL significantly recovered nitric oxide production which was reduced by gallic acid (100 uM) in Raw 264.7 cells. 2. AFL at the concentration of 200, 400 ug/mL significantly recovered nitric oxide production which was reduced by EtOH (100 uM) in Raw 264.7 cells. 3. AFL at the concentration of 400 ug/mL significantly recovered nitric oxide production which was reduced by nicotine (1mM) in Raw 264.7 cells. 4. AFL at the concentration of 200, 400 ug/mL significantly recovered nitric oxide production which was reduced by acetaminophen(2 mM) in Raw 264.7 cells. 5. AFL at the concentration of 200, 400 ug/mL significantly recovered nitric oxide production which was reduced by acetaldehyde (200 uM) in Raw 264.7 cells. Conclusions : AFL could be supposed to have the immune-enhancing activity related with nitric oxide production of macrophage impaired by various toxicants.

• Korean journal of applied entomology
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• v.45 no.1 s.142
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• pp.31-36
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• 2006

Inflammation in the brain has known to be associated with the development of a various neurologiacal diseases. The hallmark of neuro-inflammation is the activation of microglia, brain macrophage. Pro-inflammatory compounds including nitric oxide(NO) are the main cause of neuro-degenerative disease such as Alzheimer's disease. In the study, we examined whether Harmonia axyridis extracts inhibit the NO production by a direct method using Griess reagent, western blotting and by RT-PCR(Reverse Transcription-Polymerase Chain Reactionin) the gene expression of inducible nitric oxide synthase(iNOS). Distilled water$(H_2O)$ and methanol(MeOH) extracts of H. axyridis inhibited the protein expression of TNF-a(Tumor Necrosis Factor) and IL-6(Interleukin) in LPS (Lipopolysaccharide) stimulated BV-2 cells at the concentration of 100 ng/ml. Incubation of BV-2 cells with the extracts of $H_2O$ of MeOH inhibited the LPS induced NO and iNOS protein. And this inhibition of iNOS protein is concordant with the inhibition of iNOS mRNA expression. These data suggested that H. axyridis extracts may play a crucial role in inhibiting the NO production.

• Journal of Physiology & Pathology in Korean Medicine
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• v.28 no.2
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• pp.178-185
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• 2014

Leonuri Fructus, a semen of Leonuri Herba, has been used for the treatment of menstrual disorders such as amenorrhea, dysmenorrhea and leukorrhea and for the remedy of hyperemia. The present study was conducted to evaluate the anti-inflammatory effects of the Leonuri Fructus extract (Leonurus japonicus Houtt. EtOH extract; LJE) in vivo and in vitro. In vitro study, the MTT assay for cell viability was conducted to determine the non-cytotoxic concentration of LJE treatment in media. The levels of NO were measured with Griess reagent. Pro-inflammatory cytokines were detected by ELISA method. The inflammation-related proteins of this study were detected by immunoblot anlaysis. The increases of NO production and iNOS expression were detected in LPS-treated cells compared with control, but LJE attenuated the increases of NO and iNOS by LPS. LJE reduced the production of TNF-${\alpha}$ and IL-$1{\beta}$ induced by LPS stimulation. LJE suppresses the signaling pathways of NF-${\kappa}B$ and MAPKs in LPS-induced macrophage cells. In vivo study, carrageenan-induced hind paw acute edematous inflammation rat model was used for evaluation of anti-inflammatory activity of LJE. LJE significantly inhibited the increases of hind paw swelling, skin thicknesses and inflammatory cell infiltrations, and decreased the numbers of mast cell induced by carrageenan injection. These results suggest that LJE has an anti-inflammatory therapeutic potential, which is mediated through modulating NF-${\kappa}B$ activation and MAPK phosphorylation. Inhibition of the rat paw edema induced by carrageenan is considered as direct evidence that LJE may be a useful source to treat inflammation.