• 대한본초학회지
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• 제29권4호
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• pp.13-20
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• 2014

Objectives : Atotang was composed of 10 kinds of traditional medicinal herb. This research was performed to examine biological effects of Atotang for the development of prescription on atopic dermatitis. Methods : Atotang was extracted with 80% EtOH. Free radical scavenging assay has tested for anti-oxidative activity as well as the contents of total polyphenol. We observed the production of ROS, nitric oxide(NO) and the inflammatory cytokines such as interleukin-1beta(IL-${\beta}$), IL-6, tumor necrosis factor-alpha(TNF-${\alpha}$), Prostaglandin E2($PGE_2$) in Raw 264.7 cells stimulated by LPS. We used Disc diffusion method to investigate antibacterial activity on Candida albicans, Staphylococcus aureus and Staphylococcus epidermis. Result : Content of total phenolic compound of Atotang was 36.3 mg/g ext. DPPH and ABTS scavenging activities were 77% and 46% at 200 ug/ml respectively, showing dose-dependent increase. The amounts of ROS and NO in RAW 264.7 cells were decreased by 30% and 19% at 200 ug/ml, respectively, showing dose-dependent decrease. The prodcution of IL-1beta, IL-6 and TNF-alpha in RAW 264.7 cells were decreased dose-dependently by 81%, 67%, and 20% at 200 ug/ml, respectively. Atotang was reduced LPS-stimulated production of $PGE_2$ by 33%. Atotang on C. albicans, S. aureus and S. epidermis was selected by a disc diffusion method and inhibition effect of the Atotang on the growth of S. epidermis was the greatest. Conclusion : The results indicated that Atotang showed biological activities showing anti-oxidant, anti-inflammatory and antibacterial effects. Based on these results, it is concluded that Atotang can be applied to the prescription on atopic dermatitis.

• 대한본초학회지
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• 제32권3호
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• pp.97-103
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• 2017

Objective : According to recent studies, Anemarrhenae Rhizoma has anti-inflammatory activities of DW extract, but it hasn't not yet conducted to evaluate inflammatory factors about 80% ethanol extract. Therefore, The aim of this study is to investigate the various effects of individual or combined 80% ethanol extract of Anemarrhenae Rhizoma on cell viability and various anti-inflammatory factors. Methods : Anemarrhenae Rhizoma extract was prepared with 80% ethanol. MTT assay, ELISA, and Luminex were performed in LPS-activated RAW 264.7 cell line to measure cytotoxicity, Nitric oxide (NO), cyclooxygenase-2 (COX-2), prostaglandin E2 ($PGE_2$), Leukotriene B4 ($LTB_4$), and cytokines ($IL-1{\beta}$, IL-6, and $TNF-{\alpha}$), respectively. Results : At concentration of $200{\mu}g/m{\ell}$ Anemarrhenae Rhizoma extract, cytotoxicity was observed in RAW 264.7 cells. However, at concentration less than $100{\mu}g/m{\ell}$ of Anemarrhenae Rhizoma, cytotoxicity was not observed in RAW 264.7 cells. All concentration of Anemarrhenae Rhizoma extract showed no difference of NO, and $IL-1{\beta}$level in RAW 264.7 cells compared with control group. In contrast, at concentration of $100{\mu}g/m{\ell}$ Anemarrhenae Rhizoma extract significantly inhibited LPS-induced production of COX-2, PGE2, and $LTB_4$ level in RAW 264.7 cells. In addition, the production of proinflammatory cytokines (IL-6, $TNF-{\alpha}$) in LPS-induced RAW 264.7 cells was significantly decreased at concentration of all or 10, and $100{\mu}g/m{\ell}$, respectively. Conclusion : These findings demonstrate that Anemarrhenae Rhizoma has inhibitory effect on inflammatory mediators in LPS-activated RAW 264.7 cells showing possible developed as a raw material for new therapeutics to ease the symptoms related with inflammatory.

• 대한본초학회지
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• 제32권4호
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• pp.61-68
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• 2017

Objectives : Jema-sunghyangjungkisan (JSGS, Jima Xingxiang Zhengqi san) and yeoldahanso-tang (YDHST, Reduo hanshao decoction) are traditional herbal formulas which commonly used to prevent and treat stroke in traditional korean medicine. However, JSGS and YDHST extracts have not been previously reported to have anti-inflammatory effects. Therefore, We measured the anti-inflammatory effects of JSGS and YDHST extracts on lipopolysaccharide (LPS)-stimulated murine macrophage cell line, RAW 264.7 cells. Methods : To investigate the anti-inflammatory activities of JSGS and YDHST extracts, tumor necrosis factor-alpha ($TNF-{\alpha}$), interleukin (IL)-6, nitric oxide (NO) and prostaglandin $E_2$ ($PGE_2$) were examined in RAW 264.7 cells with LPS of $1{\mu}g/m{\ell}$. Results : JSGS and YDHST extracts did not have any cytotoxicity in RAW 264.7 cells. $TNF-{\alpha}$ concentration decreased 49.67% at $500{\mu}g/m{\ell}$ by JSGS but, YDHST has no statistically significant effect at all concentration. IL-6 accumulation on JSGS and YDHST extracts in LPS-stimulated RAW 264.7 cells reduced 22.03% and 41.44% at $500{\mu}g/m{\ell}$ respectively. In addition, JSGS has no inhibitory effects on NO accumulation and YDHST reduced 10.08% at $500{\mu}g/m{\ell}$. Moreover, JSGS and YDHST treatment does-dependently reduced the $PGE_2$ production. In particular, YDHST ($500{\mu}g/m{\ell}$) extract was more effective compared with $10ng/m{\ell}$ of indomethacin which is the $PGE_2$ positive control. Conclusions : Our results suggest that treatment of JSGS and YDHST extracts decreased the LPS-stimulated inflammation. Therefore, in the present study, we demonstrated that JSGS and YDHS may be used as a potential anti-inflammatory therapeutic agent.

• 대한본초학회지
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• 제34권6호
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• pp.117-124
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• 2019

Objective : Broccoli is edible green plant that has a wide variety of health benefits including cancer prevention and cholesterol reduction. However, leaves of broccoli are not eaten and are mostly left as waste. This study was conducted to evaluate the effects of the broccoli leaf extract (BLE) on prostaglandin E₂ (PGE₂) production related to nuclear factor kappa B (NF-κB) signaling in lipopolysaccharide (LPS)-activated macrophages. Methods : BLE was prepared by extracting dried leaf with ethanol. Cell viability was determined by 3-(4,5-Dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assay. PGE₂ and inflammatory cytokines were detected by enzyme-linked immunosorbent assay (ELISA). Expression level of each protein was monitored by Western blot analysis. Results : In LPS-activated Raw264.7 cells, PGE₂ release into culture medium was dramatically enhanced compared to control cells. However, increased PGE₂ was attenuated dose-dependently by treatment with BLE. Inhibition of PGE₂ production by BLE was due to the suppression of cyclooxygenase-2 (COX-2) expression determined by Western blot analysis. BLE also inhibited the production of inflammatory cytokines such as interleukin-6 (IL-6) and tumor necrosis factor-alpha (TNF-α). Inhibition at PGE₂ and cytokine was mediated from inhibition of nuclear translocation of NF-κB due to the repression of inhibitory kappa B alpha (IκBα) phosphorylation and degradation. Conclusion : This study showed that BLE exerted inhibitory activities against PGE₂, which is critical for the initiation and resolution of inflammatory responses, and that inhibition of PGE₂ was mediated by suppression of NF-κB signaling. These results suggest that the waste broccoli leaves could be used for controlling inflammation.

• 대한본초학회지
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• 제37권6호
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• pp.37-44
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• 2022

Objectives : From this study, we sight to identify chondro-protective and anti-inflammatory effects of Sorbi Commixtae Fructus extract and its compound, chlorogenic acid. Methods : Sorbi Commixtae Fructus were extracted by 50% ethanol. And chlorogenic acid in Sorbi Commixtae Fructus 50% extract was quantified by high performance liquid chromatography (HPLC). To investigate chondro-protective effects, we treated Sorbi Commixtae Fructus 50% ethanol extract and chlorogenic acid in TNF𝛼-activated ATDC5 murine chondrogenic cells. After 24 hours, protein level of matrix metalloproteinase-3 (MMP3) and mRNA level of matrix metalloproteinase-13 (MMP13) were measured by using ELISA or reverse transcription PCR, respectively. To examine anti-inflammatory effects, we treated Sorbi Commixtae Fructus 50% ethanol extract and chlorogenic acid in LPS-induced RAW 264.7 murine macrophages. We measured the level of inflammatory mediators, such as Prostaglandin E2 (PGE2), Interleukin-6 (IL6) by ELISA and nitric oxide (NO) by Griess reagent assay. Results : A concentration of chlorogenic acid in Sorbi Commixtae Fructus 50% ethanol extract was 3.9 mg/g. Sorbi Commixtae Fructus 50% ethanol extract and chlorogenic acid attenuated protein level of MMP3 and mRNA level of MMP13 in TNF𝛼-activated ATDC5 cells. Sorbi commixtae Fructus 50% ethanol extract inhibited the level of PGE2, IL6 and NO in LPS-activated RAW 264.7 cells in dose dependent manner, but chlorogenic acid has no anti-inflammatory effects. Conclusions : These findings demonstrated that Sorbi Commixtae Fructus 50% ethanol extract has chondro-protective and anti-inflammatory effects showing possible therapeutics to ease the symptoms related with osteoarthritis.

• 대한본초학회지
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• 제37권6호
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• pp.1-8
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• 2022

Objectives : This research aimed to investigate chondro-protective and anti-inflammatory effects of Caraganae Sinicae Flos 50% ethanol extract and its compound, tilianin. Methods : Caraganae Sinicae Flos was extracted with 50% ethanol. Tilianin in Caraganae Sinicae Flos 50% ethanol extract was quantified by HPLC analysis method. To investigate chondro-protective effects of Caraganae Sinicae Flos 50% ethanol extract, ATDC5 chondrogenic cells were co-treated with Caraganae Sinicae Flos 50% ethanol extract (or tilianin) and tumor necrosis factor-𝛼 (TNF𝛼) for 24 hours. After treatement for 24 hours, media supernatant was used for quantifying protein level of matrix metalloproteinase-3 (MMP3) by ELISA and harvested cells were used for analyzing mRNA expression level of matrix metalloproteinase-13 (MMP13) by reverse transcription PCR. To identify anti-inflammatory effects of Caraganae Sinicae Flos 50% ethanol extract, RAW 264.7 macrophage cells were co-treated with Caraganae Sinicae Flos 50% ethanol extract (or tilianin) and lipopolysaccharide (LPS) for 24 hours. media was used for quantifying the level of prostaglandin E2 (PGE2), interleukin-6 (IL6) by ELISA and nitric oxide by Griess reagent asssay. Results : Caraganae Sinicae Flos 50% ethanol extract and tilianin attenuated protein level of MMP3 and mRNA expression level of MMP13 in TNF𝛼-activated ATDC5 cells. Caraganae Sinicae Flos 50% ethanol extract inhibited the level of PGE2, IL6 and NO in LPS-activated RAW 264.7 cells in dose dependent manner, though tilianin inhibited PGE2 only. Conclusions : These results presented that Caraganae Sinicae Flos 50% ethanol extract could be used as natural medicines for osteoarthritis.

• 대한한의학회지
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• 제42권4호
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• pp.10-24
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• 2021

Objectives: Yongdamsagan-tang (YST) and Paljung-san (PJS) in traditional medicine and finasteride in modern medicine are used to treat benign prostatic hyperplasia (BPH). In recent, the use of combination herbal remedies with conventional drugs has been increasing. Therefore, we investigated the anti-inflammatory effects of these drugs to treat BPH and the influence of herbal formulas on finasteride metabolism. Methods: The inhibitory effects of the herbal formulas and finasteride on the production of inflammatory mediators and cytokines were determined in lipopolysaccharide (LPS)-treated RAW 264.7 cells. Additionally, the influence of herbal formulas on activities of human drug metabolizing enzymes (DMEs) was assessed using human microsomal enzymes. Results: We observed that YST, PJS and finasteride inhibited the production of nitric oxide (NO), prostaglandin E₂ (PGE₂) and interleukin-6 (IL-6) in RAW 264.7 cells. The half maximal inhibitory concentration (IC₅₀) of YST on PGE₂ production was calculated to be below 25 ㎍/mL. YST inhibited the activity of uridine diphosphate-glucuronosyltransterase (UGT) 1A4 with an IC₅₀ value of 49.35 ㎍/mL. The activities of cytochrome P450 (CYP) 1A2, CYP2B6, CYP2C19, CYP3A4, and UGT1A1 were inhibited by PJS (IC₅₀ < 100 ㎍/mL, each). Although PJS and YST inhibited the activities of CYP3A4 and UGT1A4, respectively, these formulas may not influence the metabolism of finasteride because the IC₅₀ values of herbal formulas on DMEs are too high to affect metabolism. Conclusions: Our results suggest that the combination of finasteride and YST or PJS might not influence their drug metabolism and that the drugs may have synergistic effects against BPH.

• 대한본초학회지
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• 제37권6호
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• pp.19-28
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• 2022

Objectives : To develop natural ingredients that help prevent or treat anti-inflammatory-related diseases and use themas basic data, we investigated anti-inflammatory activity of Cynanchi Atrati Radix Et Rhizoma water extracts(CWE) in lipopolysaccharide(LPS)-induced murine macrophage cell line, RAW 264.7 cells. Methods : The cell viabilities were evaluated with RAW 264.7 cells. The production of nitric oxide(NO), prostaglandin E2(PGE2), pro-inflammatory cytokines such tumor necrotic factor(TNF)-α and interleukin(IL)-6 were assessed in LPS-induced RAW 264.7 cell treated with CWE. Furthermore, the protein expression of inducible nitric oxide synthase (iNOS), cyclooxygenase-2(COX-2), and mitogen-activated protein kinase(MAPK) were assessed by western blotting. Results : In RAW 264.7 cell, the cell viability by CWE treatment was more than 98.4% at a concentration of 100-400 ㎍/mL. At a concentration of 800 ug/ml of CWE, the cell viability was as low as 86%. At doses of 100, 200 and 400 ㎍/mL, CWE inhibited the production of NO, PGE2, TNF-𝛼 and IL-6 in a dose-dependent manner and also decreased the expression of iNOS and COX-2 from LPS-induced RAW 264.7 cells. In addition, CWE significantly inhibited the MAPK pathway including decreased the phosphorylation of the p38, c-Jun N-terminal kinase(JNK) and extracellular signal-regulated kinase(ERK1/2). Conclusions : Our study provides evidence that CWE inhibits the production of main pro-inflammatory molecules in LPS-induced RAW 264.7 cells via expression of p38, JNK, and ERK1/2 MAPK signaling pathways. Therefore, CWE is expected to be widely used as a natural ingredient for anti-inflammatory functional foods or pharmaceuticals in the future.

• 대한통합의학회지
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• 제11권3호
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• pp.159-169
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• 2023

Purpose : Though other Citrus spp. have reported their anti-inflammatory and antioxidative activities in previous studies, the biological activity of Kiyomi (Citrus unshiu × C. sinensis) has not been reported yet. Therefore, this study attempted to analyze the anti-inflammatory mechanisms of Kiyomi leaf ethanol extract (KLEE) in lipopolysaccharide (LPS) stimulated RAW 264.7 cells. Methods : The cytotoxic effect of KLEE in RAW 264.7 cells was determined by WST-1 assay. Bacterial endotoxin, the concentration of nitric oxide (NO) was analyzed by the Griess reaction. In addition, Western blot analysis was applied to measure the protein expression level of inducible NO synthase (iNOS). The phosphorylated status of the critical inflammatory transcription factor, nuclear factor (NF)-𝜅B, and its upstream signaling molecules, phosphoinositide 3-kinase (PI3K)/Akt as well as mitogen-activated protein kinases (MAPKs), were also measured by Western blot analysis. Results : KLEE was not cytotoxic up to a concentration of 200 ㎍/㎖, and protein expression levels of iNOS and cyclooxygenase (COX)-2, enzymes that counteract NO and prostaglandin (PG) E2 production, were inhibited by KLEE treatment. The phosphorylated status of PI3K/Akt as well as MAPKs including extracellular regulated kinase (ERK), c-jun NH2kinase (JNK), and p38, were significantly attenuated by KLEE treatment in LPS stimulated RAW 264.7 cells. Moreover, one of phase II enzymes, heme oxygenase (HO)-1 which has known for its anti-inflammatory capacity, was strongly induced by KLEE treatment. Conclusion : Consequently, KLEE treatment significantly attenuated the production of NO as well as the expression levels of iNOS and COX-2 in LPS-stimulated RAW 264.7 cells. The inflammatory transcription factor, NF-𝜅B, as well as its upstream signaling molecules, PI3K/Akt and MAPKs, were also diminished by KLEE treatment with statistical significance in LPS-stimulated RAW 264.7 cells. These results suggest that KLEE might be a promising candidate for the attenuation of inflammatory disorders.

• 대한본초학회지
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• 제24권4호
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• pp.181-188
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• 2009

Objectives : In this study, we investigated the effect of methyl gallate of Paeonia suffruticosa(Moutan Cortex Radicis) on inflammatory response in activated macrophages. Methods : RAW264.7 cells were incubated with different concentrations of methyl gallate of Paeonia suffruticosa for 30 min and then stimulated with or without LPS at indicated times. Cell toxicity was determined by MTT assay. The concentrations of nitric oxide (NO), prostaglandin $E_2$ ($PGE_2$) and inflammatory cytokines (TNF-$\alpha$, IL-6) were measured in culture medium by Griess assay, enzyme-immuno assay, and ELISA, respectively. The expressions of iNOS, COX-2 and cytokine mRNA and protein were determined by RT-PCR and Western blot, respectively. The $I{\kappa}-B{\alpha}$ degradation in cytosol and NF-${\kappa}B$ p65 translocation into nuclear of the cells were determined by Western blot. Results : Methyl gallate was significantly inhibited LPS-induced production of NO and PGE2 in RAW264.7 cells. Methyl gallate was also suppressed LPS-induced expression of iNOS and COX-2 mRNA and protein in the cells. Methyl gallate was inhibited LPS-induced production of TNF-$\alpha$ and IL-6 via suppression of their mRNA expressions. Methyl gallate blocked the NF-${\kappa}B$ pathway in LPS-stimulated RAW264.7 cells. Conclusions : This study suggests that methyl gallate of Paeonia suffruticosa may have an antiinflammatory property through suppressing inflammatory mediator production in activated macrophages.