• The Korean Journal of Physiology and Pharmacology
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• v.23 no.6
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• pp.493-499
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• 2019

Macrophage-associated inflammation is crucial for the pathogenesis of diverse diseases including metabolic disorders. Rhodanthpyrone (Rho) is an active component of Gentiana rhodantha, which has been used in traditional Chinese medicine to treat inflammation. Although synthesis procedures of RhoA and RhoB were reported, the biological effects of the specific compounds have never been explored. In this study, the anti-inflammatory activity and mechanisms of action of RhoA and RhoB were studied in lipopolysaccharide (LPS)-stimulated macrophages. Pretreatment with RhoA and RhoB decreased inducible nitric oxide synthase and cyclooxygenase-2 expressions in RAW 264.7 cells and in thioglycollate-elicited mouse peritoneal macrophages. In addition, it downregulated transcript levels of several inflammatory genes in LPS-stimulated RAW 264.7 cells, including inflammatory cytokines/chemokines (Tnfa, Il6, and Ccl2) and inflammatory mediators (Nos2 and Ptgs2). Macrophage chemotaxis was also inhibited by treatment with the compounds. Mechanistic studies revealed that RhoA and RhoB suppressed the nuclear factor $(NF)-{\kappa}B$ pathway, but not the canonical mitogen activated protein kinase pathway, in LPS-stimulated condition. Moreover, the inhibitory effect of RhoA and RhoB on inflammatory gene expressions was attenuated by treatment with an $NF-{\kappa}B$ inhibitor. Our findings suggest that RhoA and RhoB play an anti-inflammatory role at least in part by suppressing the $NF-{\kappa}B$ pathway during macrophage-mediated inflammation.

• The Journal of Korean Obstetrics and Gynecology
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• v.21 no.4
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• pp.36-48
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• 2008

Purpose: The purpose of this study is to investigate anti-inflammatory effect and immune responses of aqueous extract from Fructus Chaenomelis (FC). Methods: We studied anti-inflammatory effect by means of examining the production of NO(nitric oxide) and expressions of pro-inflammatory cytokine (TNF-$\alpha$(tumor necrosis factor-alpha), IL(Interleukin)-6, IL-12) in the LPS-induced peritoneal macrophages of mice. Also, The western blot analysis has been done to look into the mechanism of anti-inflammatory effect. Results: 1. The FC extract did not have any cytotoxicity in the peritoneal macrophages. 2. The FC extract inhibits the productions of NO, IL-6. IL-12 in the LPS-stimulated peritoneal macrophages of mice, but not of TNF-$\alpha$. 3. The FC extract inhibits the activation of NF-${\kappa}B$(nuclear factor-kappa B) by keeping $I{\kappa}B-\alpha$(inhibitory kappa B-alpha) from degradating, but not of MAPKs(mitogen-activated protein kinases) such as ERK(extracelluar signa 1-regulated kinase), JNK(c-Jun N-terminal kinase), p38. Conclusion: These results show that FC extract inhibits the production of pro-inflammatory cytokines such as IL-6. IL-12. NO by inhibiting NF-${\kappa}B$ activation in the peritoneal macrophages of mice. In conclusion, this experiment suggests that FC extract may be effective for the treatment of acute and chronic inflammation including genitourinary infection.

• Journal of Veterinary Science
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• v.22 no.2
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• pp.18.1-18.12
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• 2021

Background: We previously elucidated the protective mechanism of Korean red ginseng oil (RGO) against Brucella abortus infection, and our phytochemical analysis revealed that palmitic acid (PA) was an abundant component of RGO. Consequently, we investigated the contribution of PA against B. abortus. Objectives: We aimed to investigate the efficacy of PA against B. abortus infection using a murine cell line and a murine model. Methods: Cell viability, bactericidal, internalization, and intracellular replication, western blot, nitric oxide (NO), and superoxide (O₂^-) analyses and flow cytometry were performed to determine the effects of PA on the progression of B. abortus infection in macrophages. Flow cytometry for cytokine analysis of serum samples and bacterial counts from the spleens were performed to determine the effect of PA in a mouse model. Results: PA did not affect the growth of B. abortus. PA treatment in macrophages did not change B. abortus uptake but it did attenuate the intracellular survivability of B. abortus. Incubation of cells with PA resulted in a modest increase in sirtuin 1 (SIRT1) expression. Compared to control cells, reduced nitrite accumulation, augmented O₂^-, and enhanced pro-inflammatory cytokine production were observed in PA-treated B. abortus-infected cells. Mice orally treated with PA displayed a decreased serum interleukin-10 level and enhanced bacterial resistance. Conclusions: Our results suggest that PA participates in the control of B. abortus within murine macrophages, and the in vivo study results confirm its efficacy against the infection. However, further investigations are encouraged to completely characterize the mechanisms involved in the inhibition of B. abortus infection by fatty acids.

• The Korea Journal of Herbology
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• v.36 no.4
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• pp.23-30
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• 2021

Objectives : The purpose of this study was to investigate the antioxidant activities and nitric oxide (NO) production in RAW 264.7 macrophages in extract of Alisma orientale Juzep (EAOJ) and fermented extract (FAOJ) by Paenibacillus kribbensis AM49 (P. kribbensis AM49). Methods : The Alisma orientale Juzep was fermented with P. kribbensis AM49 at 37℃ for 72 hours. We measured total polyphenol and total flavonoid, DPPH radical scavenging activity, FRAP activity and reducing power by spectrometric assay in EAOJ and FAOJ at concentrations at 0.5, 1, 5, 10 mg/㎖. Positive control was used ascorbic acid. Furthermore, we examined effect of EAOJ and FAOJ on the cell viability and NO production in RAW 264.7 macrophages. Results : The total polyphenol and total flavonoids content of FAOJ were increased 9.16 mg/g, 2.59 mg/g to 12.58 mg/g, 3.45 mg/g. DPPH radical scavenging activity, FRAP activity and reducing power were dose dependently increased according to the treatment concentration (0.5, 1, 5, 10 mg/㎖) of EAOJ and FAOJ. In particular, DPPH radical scavenging activity, FRAP activity of FAOJ was significantly increased at 5, 10 mg/㎖. Reducing power of FAOJ at 10 mg/㎖ was similar to ascorbic acid at 0.1 mg/㎖. In addition, the cell viability and NO production in RAW 264.7 macrophages were significantly increased at the concentrations of 250, 500, 1000 ㎍/㎖. Conclusions : These results suggest that FAOJ by P. kribbensis AM49 has effects to antioxidant activity. In addition, the cell viability and NO production in RAW 264.7 macrophages were significantly increased.

• Journal of Ginseng Research
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• v.43 no.3
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• pp.394-401
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• 2019

Background: Ginsenoside Rb1, a triterpene saponin, is derived from the Panax ginseng root and has potent antiinflammatory activity. In this study, we determined if Rb1 can increase macrophage phagocytosis and elucidated the underlying mechanisms. Methods: To measure macrophage phagocytosis, mouse peritoneal macrophages or RAW 264.7 cells were cultured with fluorescein isothiocyanate-conjugated Escherichia coli, and the phagocytic index was determined by flow cytometry. Western blot analyses were performed. Results: Ginsenoside Rb1 increased macrophage phagocytosis and phosphorylation of p38 mitogenactivated protein kinase (MAPK), but inhibition of p38 MAPK activity with SB203580 decreased the phagocytic ability of macrophages. Rb1 also increased Akt phosphorylation, which was suppressed by LY294002, a phosphoinositide 3-kinase inhibitor. Rb1-induced Akt phosphorylation was inhibited by SB203580, (5Z)-7-oxozeaenol, and small-interfering RNA (siRNA)-mediated knockdown of $p38{\alpha}$ MAPK in macrophages. However, Rb1-induced p38 MAPK phosphorylation was not blocked by LY294002 or siRNA-mediated knockdown of Akt. The inhibition of Akt activation with siRNA or LY294002 also inhibited the Rb1-induced increase in phagocytosis. Rb1 increased macrophage phagocytosis of IgG-opsonized beads but not unopsonized beads. The phosphorylation of p21 activated kinase 1/2 and actin polymerization induced by IgG-opsonized beads and Rb1 were inhibited by SB203580 and LY294002. Intraperitoneal injection of Rb1 increased phosphorylation of p38 MAPK and Akt and the phagocytosis of bacteria in bronchoalveolar cells. Conclusion: These results suggest that ginsenoside Rb1 enhances the phagocytic capacity of macrophages for bacteria via activation of the p38/Akt pathway. Rb1 may be a useful pharmacological adjuvant for the treatment of bacterial infections in clinically relevant conditions.

• Food Science of Animal Resources
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• v.38 no.6
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• pp.1226-1236
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• 2018

Ovotransferrin (OTF) is a well-known protein of the transferrin family with strong iron chelating activity, resulting in its antimicrobial activity. Furthermore, OTF is known to have antioxidant, anticancer, and antihypertensive activities. However, there have been few studies about the immune-enhancing activity of OTF. In current study, we investigated the immune-enhancing activity of OTF using the murine macrophage cells in vitro. The effect of OTF on production of pro-inflammatory mediators and cytokines were determined using Griess assay and quantitative real-time PCR. Using Neutral Red uptake assay, we confirmed the effect of OTF on phagocytic activity of macrophages. Ovotransferrin significantly increased the production of nitric oxide (NO) and secretion of inducible nitric oxide synthase (iNOS) mRNA with no cytotoxic activity. Ovotransferrin (2 mg/mL) stimulated NO production up to $31.9{\pm}3.5{\mu}M$. Ovotransferrin significantly increased the mRNA expression levels of pro-inflammatory cytokines which are tumor necrosis factor-${\alpha}$ (TNF-${\alpha}$), Interleukin-$1{\beta}$ (IL-$1{\beta}$), and IL-6: OTF (2 mg/mL) treatment increased the secretion of mRNA for TNF-${\alpha}$, IL-$1{\beta}$, and IL-6 by 22.20-, 37.91-, and 6.17-fold of the negative control, respectively. The phagocytic activity of macrophages was also increased by OTF treatment significantly compared with negative control. Also, OTF treatment increased phosphorylation level of MAPK signaling pathways. These results indicated that OTF has immune-enhancing activity by activating RAW 264.7 macrophages via MAPK pathways.

• Fisheries and Aquatic Sciences
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• v.22 no.2
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• pp.6.1-6.10
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• 2019

Background: This study is aimed at identifying the anti-inflammatory properties of 70% ethanol extract produced from an edible brown seaweed Sargassum horneri (SJB-SHE) with industrial-scale production by Seojin Biotech Co. Ltd. S. horneri is a rich source of nutrient and abundantly growing along the shores of Jeju, South Korea. Methods: Here, we investigated the effect of SJB-SHE on LPS-activated RAW 264.7 macrophages. The cytotoxicity and NO production of SJB-SHE were evaluated using MTT and Griess assays, respectively. Additionally, protein expression and gene expression levels were quantified using ELISA, Western blots, and RT-qPCR. Results: Our results indicated that pre-treatment of RAW 264.7 macrophages with SJB-SHE significantly inhibited LPS-induced NO and $PGE_2$ production. SJB-SHE downregulated the proteins and genes expression of LPS-induced iNOS and COX2. Additionally, SJB-SHE downregulated LPS-induced production of pro-inflammatory cytokines (tumor necrosis factor-${\alpha}$, interleukin (IL)-6, and IL-$1{\beta}$). Furthermore, SJB-SHE inhibited nuclear factor kappa-B (NF-${\kappa}B$) activation and translocation to the nucleus. SJB-SHE also suppressed the phosphorylation of mitogen-activated protein kinases (ERK1/2 and JNK). Conclusions: Collectively, our results demonstrated that SJB-SHE has a potential anti-inflammatory property to use as a functional food ingredient in the future.

• The Korean Journal of Physiology and Pharmacology
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• v.25 no.5
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• pp.395-401
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• 2021

Extended inflammation and cytokine production pathogenically contribute to a number of inflammatory disorders. Formosanin C (FC) is the major diosgenin saponin found in herb Paris formosana Hayata (Liliaceae), which has been shown to exert anti-cancer and immunomodulatory functions. In this study, we aimed to investigate anti-inflammatory activity of FC and the underlying molecular mechanism. RAW264.7 macrophages were stimulated with lipopolysaccharide (LPS) or pretreated with FC prior to being stimulated with LPS. Thereafter, the macrophages were subjected to analysis of the expression levels of pro-inflammatory mediators, including nitric oxide (NO), prostaglandin E₂ (PGE), tumor necrosis factor-α (TNF-α), interleukin-1β (IL-1β), and IL-6, as well as two relevant enzymes, inducible nitric oxide synthase (iNOS), and cyclooxygenase-2 (COX-2). The analysis revealed that FC administration blunted LPS-induced production of NO and PGE in a dose-dependent manner, while the expression of iNOS and COX-2 at both mRNA and protein levels was inhibited in LPS-stimulated macrophages pre-treated with FC. Moreover, LPS stimulation upregulated mRNA expression and medium release of TNF-α, IL-1β, and IL-6, whereas this effect was blocked upon FC pre-administration. Mechanistic studies showed that inhibitory effects of FC on LPS-induced inflammation were associated with a downregulation of IκB kinase, IκB, and p65/NF-κB pathway. Taken together, these data suggest that FC possesses an inflammation-suppressing activity, thus being a potential agent for the treatment of inflammation-associated disorders.

• Journal of dental hygiene science
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• v.20 no.4
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• pp.213-220
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• 2020

Background: The leaves of Perilla frutescens, commonly called perilla and used for food in Korea, contain components with a variety of biological effects and potential therapeutic applications. The purpose of this study was to identify the components of 70% ethanol extracted Perilla frutescens (EEPF) and determine its inhibitory effects on oral microbial activity and production of nitric oxide (NO) and prostaglandin E2 (PGE2) in lipopolysaccharides (LPS)-stimulated Raw264.7 macrophages, consequently, to confirm the possibility of using EEPF as a functional component for improving the oral environment and preventing inflammation. Methods: One kg of P. frutescens leaves was extracted with 70% ethanol and dried at -70℃. EEPF was analyzed using high-performance liquid chromatography analysis, and antimicrobial activity against oral microorganisms was revealed using the disk diffusion test. Cell viability was elucidated using a methylthiazolydiphenyl-tetrazolium bromide assay, and the effect of EEPF on LPS-induced morphological variation was confirmed through microscopic observation. The effect of EEPF on LPS-induced production of pro-inflammatory mediators, NO and PGE2 was confirmed by the NO assay and PGE2 enzyme-linked immunosorbent assay. Results: The main component of EEPF was rosemarinic acid, and EEPF showed weak anti-bacterial and anti-fungal effects against microorganisms living in the oral cavity. EEPF did not show toxicity to Raw264.7 macrophages and had inhibitory effects on the morphological variations and production of pro-inflammatory mediators, NO and PGE2 in LPS-stimulated Raw264.7 macrophages. Conclusion: EEPF can be used as a functional material for improving the oral environment through the control of oral microorganisms and for modulating inflammation by inhibiting the production of inflammatory mediators.

• Journal of Physiology & Pathology in Korean Medicine
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• v.35 no.5
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• pp.169-176
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• 2021

This study was conducted to suggest the Cudrania tricuspidata leaf and Achyranthes japonica Nakai Complex (CAC) possibility of use as a functional natural material for improving bone disease. Cudrania tricuspidata leaf and Achyranthes japonica Nakai were mixed in the same amount, extracted with hot water, and then powdered and used in the study. After, the cytotoxicity of CAC for osteoblasts (MG63 cell), osteoclasts (differentiated RAW264.7 cell), and macrophages (RAW264.7 cell) were evaluated by MTT assay, and ALP assay and TRAP assay were performed to confirm the differentiation capacity of osteoblasts and osteoclasts, respectively. In addition, the anti-inflammatory effect in macrophages was evaluated by ELISA, qRT-PCR, and western blot assay. CAC did not proliferated osteoblasts and osteoclasts, but increased ALP activity against osteoblasts differentiation and decreased TRAP activity against osteoclasts differentiation. CAC did not proliferated macrophages but decreased nitric oxide production. Also, decreased NOS2, IL1B, IL6, PTGS2, and TNFA gene expression, and JNK and p38 protein phosphorylation in a concentration-dependent manner, but ERK protein phosphorylation was not changed. As a result, CAC increased the differentiation and activation of osteoblasts, inhibited the differentiation and activation of osteoclasts, and regulated the expression of inflammatory cytokines in macrophages. Therefore, it is thought that CAC can be used as a functional natural material that prevents bone disease and has an anti-inflammatory effect.