• Korean Journal of Microbiology
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• v.50 no.1
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• pp.27-32
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• 2014

Autophagy is one of the lysosomal degradation pathways to maintain cellular homeostasis. The damaged proteins or organelles are uptaken through extra- and intra-cellular stress, starvation and infected pathogens, subsequently, autophagosomes are fused with lysosomes to break down the molecules. Salmonella enterica serovar Typhimurium (S. Typhimurium), intracellular bacteria, cause acute gastroenteritis and food poisoning. Given that autophagy induced by S. Typhimurium plays an important role in the cells to control the infection, we identify whether the induction of autophagy with rapamycin, chemical inducer of autophagy, before infection regulates S. Typhimurium infection. After treatment of rapamycin or 3-methyladenine (3-MA), autophagy inhibitor, RAW264.7 cells were infected with S. Typhimurium. Pretretment of rapamycin decreased the growth rate of S. Typhimurium in the cells; otherwise, pretreatment of 3-MA increased the growth rate of S. Typhimurium. The expression of autophagy-related genes was significantly increased in the S. Typhimurium-infected cells pretreated with rapamycin. To examine whether induced autophagy by rapamycin control the infection with increase the production of reactive oxygen species (ROS) and nitric oxide (NO), antibacterial radical substrates were measured in infected cells followed by the treatment with either rapamycin or 3-MA. NO production increased in RAW264.7 cells; otherwise, ROS production remained unchanged during the infection. These findings suggest that inducing autophagy with rapamycin reveals antimicrobial activity as producing NO against S. Typhimurium infection in mouse macrophages.

• Journal of Life Science
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• v.29 no.9
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• pp.977-985
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• 2019

Aster yomena (Kitam.) Honda is an edible vegetable and perennial herb belonging to the Asteraceae family, and has been used for a long time for the prevention and treatment of various diseases. Although leaf extracts of A. yomena are known to have antioxidant and anti-inflammatory effects, accurate efficacy assessments are still inadequate. In this study, we investigated whether the antioxidant efficacy of ethanol extract of A. yomena leaf (EEAY) is correlated with the anti-inflammatory effect in RAW 264.7 macrophages. The results showed that EEAY significantly inhibited the hydrogen peroxide ($H_2O_2$)-induced growth inhibition in RAW 264.7 cells, which was associated with increased expression of nuclear factor erythroid 2-related factor-2 (Nrf2) and heme oxygenase-1 (HO-1). EEAY pretreatment also effectively prevented $H_2O_2$-induced reactive oxygen species generation and apoptosis through inhibition of caspase-3 activation and poly (ADP-ribose) polymerase degradation. Additionally, EEAY significantly increased the expression and production of interleukin-10, a representative anti-inflammatory cytokine, which was associated with increased expression of toll-like receptor 4 and myeloid differentiation factor 88 at transcriptional and translational levels. Furthermore, the increased production of nitric oxide (NO) by lipopolysaccharide was markedly abolished under the condition of EEAY pretreatment, and the inhibitory effect of NO production by EEAY was further increased by hemin, an HO-1 inducer. Overall, our results suggest that EEAY is able to activate the Nrf2/HO-1 signaling pathway to protect RAW 264.7 macrophages from oxidative and inflammatory stress.

• Korean Journal of Microbiology
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• v.43 no.4
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• pp.292-297
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• 2007

This study was performed to compare in vitro immune enhancing effects of polysaccharides extracted from cultivated mycelia of Agaricus blazei Murill. Carbohydrate contents of semi-purified polysaccharides were 85.6% and 95.3%, while ${\beta}$-glucan conents were 67.9% and 88.1% for intracellular and extracellular polysaccharide, respectively. Samples were adjusted to the same in their carbohydrate contents before efficacy tests. Both intracellular and extracellular polysaccharide increased nitric oxide (NO) synthesis of macrophage RAW 264.7 in dose dependent manner, and the maximum increase rate was 53.9 and 53.1% in intracellular and extraceltular polysaccharide, respectively. The polysaccharides also increased synthesis of cytokines such as interleukin (IL)-$1{\beta}$, IL-6 and tumor necrosis factor (TNF)-${\alpha}$ in RAW 264.7. For all the 3 cytokines, the increase rate of synthesis was much higher in extracellular polysaccharide compared to intracellular polysaccharide, especially at low concentration. Both polysaccarides increased the proliferation of splenocytes in vitro, intracellular polysaccharide showed increase in dose dependent manner while extraceltular polysaccharide showed increase untill medium concentration ($250\;{\mu}g/ml$). They did not show direct cytotoxicity against cancer cells such as B16F0 melanoma. As results, it was regarded that the both intracellular and extracellular polysaccharide from A. blazei showed immune enhancing effects in vitro, but the activity is higher in extracellular polysaccharide compared to intracellular polysaccharide.

• Microbiology and Biotechnology Letters
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• v.43 no.4
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• pp.322-329
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• 2015

The effects of water extracts of Gallus gallus var. domesticus (Yeonsan ogolgye, GD) on the activities of osteoblast differentiation and the restraint of osteoclast formation were investigated. The water extract of GD in the human osteoblast "MG-63" cell, was examined in relation to alkaline phosphatase (ALP) activity and alizarin red stains. In order to observe the effects of osteoclasts formation, we analyzed RAW 264.7 cell tartrate-resistant acid phosphatase (TRAP) activity and TRAP stains. The ALP activity of the water extract of hen and cock flesh (3 years) were 133.8% and 129.6%, respectively. The ALP activity of flesh extracts was also higher than that of the skin extracts. Concerning the effects of age, the 3 years old flesh extracts had a higher activity than that of the one year old extracts. However the activity of the 3 years old skin extracts was lower than that of the one year old extracts. For gender conditions, the ALP activity of the hen extract was higher than that of the cock. The degree bone mineralization in the three years old hen flesh exhibited the highest rate, at 124.3%, amongst all the groups. The TRAP activity of the flesh extracts of the three years old cock revealed the lowest rate, at 31.8%, compared to the control. Our results demonstrate that the water extract of GD increases bone mineralization and osteoblast differentiation activity in MG-63 cells and enhances the inhibitory activity of bone-resorption in RAW 264.7 cells. In conclusion, the water extracts of GD seem to be effective in the prevention and treatment of bone related disorders.

• The Journal of Korean Obstetrics and Gynecology
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• v.26 no.2
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• pp.1-16
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• 2013

Objectives: This study was performed to evaluate the effect of Dokhwalgisaengtang-gami water extract(DGG) on osteoporosis. Methods: The osteoclastogenesis and gene expression were determined in RANKL-stimulated RAW 264.7 cell. And osteoblastogenesis was also determined in rat calvarial cell. Results: The results were summarized as followes. 1. DGG decreased the number of TRAP positive cell in RANKL-stimulated RAW 264.7 cell. 2. DGG inhibited TRAP activity in RANKL-stimulated RAW 264.7 cell. 3. DGG decreased the expression of NAFTc1, MITF in RANKL-stimulated RAW 264.7 cell. 4. DGG increased the expression of iNOS, COX-2, IL-6 in RANKL-stimulated RAW 264.7 cell. 5. DGG decreased the expression of cathepsin K, MMP-9, TRAP in RANKL-stimulated RAW 264.7 cell. 6. DGG increased cell proliferation of rat calvarial cell. 7. DGG increased ALP activity in rat calvarial cell 8. DGG increased bone matrix protein, collagen synthesis and nodule formation in rat calvarial cell. Conclusions: It is concluded that DGG might decrease the bone resorption resulted from decrease of osteoclast differentiation and it's related gene expression. And DGG might increase the bone formation resulted from increase of osteoblast function.

• Journal of Applied Biological Chemistry
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• v.63 no.4
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• pp.421-428
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• 2020

This study investigated a mixture of Astragalus membranaceus (AM) and Lithospermum erythrorhizon (LE) extracts (ALM16), exerts anti-inflammatory effects in lipopolysaccharide (LPS)-induced RAW264.7 macrophage cells, and its underlying mechanism. ALM16 was prepared by mixing AM and LE extracts in a ratio of 7:3 (w/w). Cytotoxicity of ALM16 in RAW264.7 cells was not shown up to 200 ㎍/mL of ALM16. The results of this study showed that ALM16 does-dependently inhibits the production of nitric oxide, prostaglandin E2 and pro-inflammatory cytokines (interleukin-1β, interleukin-6, and tumor necrosis factor-α) in LPS-induced RAW264.7 cells. ALM16 not only markedly reduced the protein expression levels of inducible nitric oxide synthase and cyclooxygenase-2 (COX-2) in LPS-stimulated RAW264.7 cells, but also inhibited the nuclear translocation and DNA-binding activity of nuclear factor-kappa B (NF-κB). In addition, ALM16 specifically inhibited the phosphorylation of c-Jun N-terminal kinase and extracellular signal-regulated kinases in LPS-stimulated RAW264.7 cells. In conclusion, these results suggest that ALM16 may exert anti-inflammatory effect by modulating mitogen-activated protein kinase and NF-κB signaling pathways.

• Korean Journal of Food Science and Technology
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• v.53 no.2
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• pp.115-120
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• 2021

Scutellaria baicalensis has been used as a traditional medicine for diarrhea, dysentery, hypertension, hemorrhaging, insomnia, inflammation, and respiratory infections. This study examined the anti-inflammatory effect of Scutellaria baicalensis water extract (SWE) in lipopolysaccharide (LPS)-induced RAW 264.7 cells. To evaluate the anti-inflammatory effect of SWE, RAW 264.7 macrophages were stimulated with LPS to induce the production of inflammation-related factors, which were measured by western blotting. In RAW 264.7 cells, SWE inhibited the production of nitric oxide (NO) without causing cell toxicity. SWE also reduced the expression of inducible NO synthase and cyclooxygenase-2 protein, as well as the production of pro-inflammatory cytokines (such as tumor necrosis factor-α). The phosphorylation levels of the mitogen-activated protein kinase family members, such as JNK and p38, were also reduced by SWE. Thus, SWE could be used as a potential anti-inflammatory agent.

• Journal of Life Science
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• v.27 no.8
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• pp.873-878
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• 2017

We previously reported that NAD(P)H:quinone oxidoreductase 1 (NQO1)-knockout (KO) mice exhibited spontaneous inflammation in the gut. We also found that NQO1-KO mice showed highly increased inflammatory responses compared with NQO1-WT control mice when subjected to DSS-induced experimental colitis. In a Clostridium difficile toxin-induced mouse enteritis model, NQO1-KO mice were also sensitive compared with NQO1-WT mice. Moreover, numerous studies have shown that NQO1 is functionally associated with immune regulation. Here, we assessed whether NQO1 defects can alter macrophage activation. We found that peritoneal macrophages isolated from NQO1-KO mice produced more IL-6 and $TNF-{\alpha}$ than those isolated from NQO1-WT mice. Moreover, the dicumarol-induced inhibition of NQO1 significantly increased IL-6 and $TNF-{\alpha}$ production in peritoneal macrophages isolated from NQO1-WT mice, as well as in the cultured mouse macrophage cell line, RAW264.7. These results indicate that NQO1 may negatively regulate the activation of macrophages. Knockout or chemical inhibition of NQO1 markedly reduced the expression of $I{\kappa}B$ (inhibitor of $NF{\kappa}B$) in both mouse peritoneal macrophages and RAW264.7 cells. Finally, RAW264.7 cells treated with dicumarol exhibited morphological changes reflecting macrophage activation. Our results suggest that NQO1 may suppress the $NF{\kappa}B$ pathways in macrophages, thereby suppressing the activation of these cells. Thus, immunosuppressive activity may be among the many possible functions of NQO1.

• Journal of Life Science
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• v.25 no.7
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• pp.773-779
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• 2015

This study examined extracts from five different kinds of lees and nuruk and their organic solvent fractions in terms of several biological functions, such as anti-adipogenic, anti-inflammatory, and anti-proliferative activities. The anti-adipogenic activity was investigated by treating mouse pre-adipocyte 3T3-L1 cells with one extract (YE) and four organic solvent fractions (YAc, PAc, RAc, and WPAc) during adipogenesis. Among the treated samples, the ethyl acetate fraction of W-Ju lees (WPAc) showed the strongest anti-adipogenic effect, which was confirmed with oil red O staining and down-regulation of pro-adipogenic genes such as PPAR-gamma and SCD-1. Treatment with WPAc also reduced the expression of PPAR-gamma in a time-dependent manner. The effects of five different extracts were examined on nitric oxide (NO) production in mouse RAW 264.7 cells to determine anti-inflammatory activity. The ethyl acetate fraction of B-Ju lees (PAc) significantly decreased NO production in LPS-stimulated RAW 264.7 cells and it also inhibited NO production in a dose-dependent manner. The PAc fraction also dramatically decreased the viability of human colorectal cancer HCT116 cells in a dose-dependent manner. In addition, PAc increased the expression of NAG-1 and ATF3 genes in a dose dependent manner. Overall, these results indicate that lees and nuruk have several biological functions, including anti-adipogenic, anti-inflammatory, and anti-proliferative activities.

• Korean Journal of Environmental Biology
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• v.33 no.2
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• pp.256-261
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• 2015

This study was conducted to assess the influence of hazard pattern in the surface structure of pollen grains of Campsis grandiflora, and cytotoxicity of different part extracts and nectar on RAW264.7 macrophages. The pollen grains were medium sized ($21.8{\mu}m$) with tricolpate aperture type. In equatorial view, the pollens were prolate (P/E=1.8) and the exine pattern was smooth and reticulate. This result contradict with the rumor of having a hook-shaped protuberance that can damage the cornea because we couldn't observed any protuberance on the surface of the outer wall. Furthermore, we investigated the 70% MeOH extracts (flower, leaf, stem) and nectar of C. grandiflora for their cell viability in temporal basis via MTT analysis on RAW264.7 macrophage cells. There was no significant difference in the cytotoxicity among the MeOH extracts and nectar of C. grandiflora after 24 h. However, nectar showed the dosedependent cytotoxicity on RAW264.7 macrophage cells after 48 h.