• Title/Summary/Keyword: Real time PCR

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Anti-inflammatory Effect of Oxya chinensis sinuosa Ethanol Extract in LPS-induced RAW 264.7 Cells (LPS로 유도된 RAW 264.7세포에 대한 벼메뚜기(Oxya chinensis sinuosa) 에탄올 추출물의 항염증 효과)

  • Yoon, Young-Il;Chung, Mi Yeon;Hwang, Jae-Sam;Goo, Tae-Won;Ahn, Mi-Young;Lee, Young-Bo;Han, Myung-Sea;Yun, Eun-Young
    • Journal of Life Science
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    • v.24 no.4
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    • pp.370-376
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    • 2014
  • Although the grasshopper Oxya chinensis sinuosa has long been used as food in Korea, there is little data on its functional effects. In this study, we investigated the anti-inflammatory effect of O. c. sinuosa ethanol extract (OCE) in RAW 264.7 mouse macrophage cells treated with lipopolysaccharide (LPS) for induction of inflammation. First, we determined that there is no cytotoxicity at $2,000{\mu}g/ml$ or less of OCE in RAW 264.7 cells. To evaluate the anti-inflammatory effects of OCE, we investigated expression levels of pro-inflammatory cytokines such as tumor necrosis factor (TNF)-${\alpha}$ and interleukin (IL)-6, and pro-inflammatory enzymes such as inducible nitric oxide synthase (iNOS) and cyclo-oxygenase-2 (COX-2) in LPS-induced RAW 264.7 cells. In addition, we examined whether OCE could inhibit translocation of NF-${\kappa}B$ p65 into the nucleus in LPS induced RAW 264.7 cells. As a result, we found that the mRNA and protein levels of TNF-${\alpha}$ and IL-6 decreased in LPS-induced RAW 264.7 cells after treatment with OCE in a dose-dependent manner. In addition, we confirmed a $2,000{\mu}g/ml$ concentration of OCE inhibited translocation of NF-${\kappa}B$ p65 by immunnostaining and Western blot analysis, and a decrease in the protein expression levels of iNOS and COX-2. Accordingly, we suppose that OCE has an anti-inflammatory effect through down-regulation of TNF-${\alpha}$, IL-6, iNOS, and COX-2 related to ${\kappa}B$ p65 inflammatory signaling pathways.

Anti-wrinkle and Whitening Effects of Essential Oil from Abies koreana (Abies koreana 유래 정유의 항주름 및 미백 효과)

  • Song, Byeong-Wook;Song, Min-Jeong;Park, Mi-Jin;Choi, Don-Ha;Lee, Sung-Suk;Kim, Myungkil;Hwang, Ki-Chul;Kim, Il-Kwon
    • Journal of Life Science
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    • v.28 no.5
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    • pp.524-531
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    • 2018
  • The essential oil from Abies koreana E.H. Wilson had been developed, however, its efficacy has not yet been studied especially in terms of skin care research. The aim of this study is to investigate the effects of Abies koreana extracts (AKE) on melanogenesis and wrinkle formation in B16F10 melanoma cells (B16F10) and human dermal fibroblast cell line (HDF). The essential oil was extracted by hydrodistillation method and purified by anhydrous sodium sulfate. At a concentration of $10^{-5}$-fold, viability in these cells had been defined by cytotoxicity assays. Anti-melanogenic effects on B16F10 were evaluated using tyrosinase inhibition assay, and real-time PCR for verifying gene expression of tyrosinase, tyrosinase related protein-1 and -2 (TRP-1 and -2). AKEs reduced about 5-fold of tyrosinase inhibitory activity compared to ${\alpha}$-melanocyte-stimulating hormone (${\alpha}$-MSH)-induced group and about 30% reduction compared to Arbutin induced group. The mRNA levels of three melanin-related factors were increased, separately. To investigate the effects of anti-wrinkle, procollagen type I c peptide synthesis assay (PIP) and Western blot were performed. At AKE-treated group, PIP was up-regulated and the expression of collagen type 1 and matrix metalloproteinase (MMP)-1 were improved. Furthermore, AKE presented anti-wrinkle effects by increasing UVB-inhibited collagen type 1 expression, and reducing UVB-induced MMP-1 production at $60mJ/cm^2$ of UVB radiation. Therefore, Abies koreana extracts has potentials as a safe and an effective skin ingredient for whitening and anti-wrinkle.

The Comparison of the Effect of Cigarette and Stop Smoking-aiding Cigarette on Release of IL-6 from Bronchial Epithelial Cell (일반담배(Cigarette)와 금연 보조 담배(금연초, 허브담배, 쑥 담배)의 기관지 상피세포에서 IL-6유리 효과비교)

  • Kim, Myoung Chan;Jung, Jeil;Jung, Jong Hoon;Kim, Hak Ryul;Yang, Sei Hoon;Jeong, Eun Taik;Kim, Hui Jung
    • Tuberculosis and Respiratory Diseases
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    • v.59 no.5
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    • pp.530-535
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    • 2005
  • Background and Aims : Cigarette smoking induces an inflammatory response in the airways, which may play a key role in the pathogenesis of chronic obstructive pulmonary disease. Interleukin-6 (IL-6) is one of the cytokines that plays an important role in inducing bronchial inflammation. The aim of this study was to determine if the level of the pro-inflammatory cytokine, Interleukin-6, is increased when the bronchial epithelial cells are exposed to a cigarette smoke extract (CSE) and an extract from stop smoking-aiding cigarettes, and examined the safety of these commercially available stop smoking-aiding cigarettes. Method : Bronchial epithelial cells were exposed to CSE from cigarette and stop smoking-aiding cigarettes for 24 hours. ELISA was used to measure the IL-6 levels in the supernatant from each condition. The IL-6 mRNA levels were measured by Taqman Real time RT-PCR. N-acetyl-L-cysteine(NAC) was added to each condition to determine if NAC can inhibit the release of IL-6 from the bronchial epithelial cells when they are exposed to CSE from cigarette and stop smoking-aiding cigarettes. Result : When bronchial epithelial cells were exposed to a CSE from cigarettes and stop smoking-aiding cigarettes, each type of CSE stimulated IL-6 production from the bronchial epithelial cells. The IL-6 mRNA level in the Bronchial epithelial cells was also elevated and NAC was found to inhibit the release of IL-6 from bronchial epithelial cells when they were exposed to the CSE from cigarettes and stop smoking-aiding cigarettes. Conclusion : Commercially available stop smoking-aiding cigarette can induce bronchial inflammation and can be harmful to smokers. Therefore, the safety of these cigarettes for smoking cessation should be evaluated.

Effects of the mixture of fenugreek seeds and Lespedeza cuneata extracts on testosterone synthesis in TM3 cells oxidative stressed with H2O2 (호로파와 야관문 복합추출물이 과산화수소로 산화적 스트레스가 가해진 TM3 세포의 테스토스테론 합성에 미치는 영향)

  • Lee, Kyeong Soo;Lee, Eun Kyoung;Seo, Yoonhee;Choe, Soo Young
    • Journal of Applied Biological Chemistry
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    • v.59 no.4
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    • pp.305-311
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    • 2016
  • This study investigated the effects of a mixture of fenugreek seeds and Lespedeza cuneata extracts on testosterone synthesis in TM3 cells that were oxidatively stressed with $H_2O_2$. In order to oxidatively stress TM3 cells, the cells were treated with $50{\mu}M$ hydrogen peroxide for 4 hr in serum-free media. Yagwanmun-horopa mixture (YHM) showed neither cytotoxicity nor increment of cell proliferation in the oxidatively stressed TM3 cells in any concentration. When the cells were treated with hydrogen peroxide, testosterone levels decreased, but the testosterone level was returned to that of the control level in the presence of YHM. In order to find out the reasons for the increase of testosterone, the expression of the genes involved in the synthesis or disintegration of testosterone. On the other hand, the levels of $3{\beta}$-HSD4 and 17, 20-desmorase, which are involved in testosterone synthesis, were decreased through the use of hydrogen peroxide and were recovered through YHM treatment. Aromatase and $5{\alpha}$-reductase2, which convert testosterone to estradiol and dihydrotestosterone, respectively, were increased through the use of hydrogen peroxide, and were returned to control level through YHM treatment. These results suggest that YHM does not affect TM3 cell proliferation. However, YHM increases the expression of testosterone-synthesizing enzyme, which was decreased through oxidative stress, and decreases the expression of testosterone- converting enzyme, which was increased through oxidative stress. Therefore, it is reasonable that YHM has strong recovery activity on testosterone to normal level, even in the oxidatively stressed TM3 cells which mimics the andropause state.

Association of β-Catenin with Fat Accumulation in 3T3-L1 Adipocytes and Human Population (β-catenin 유전자의 3T3-L1 지방세포 및 인체에서의 지방축적 연관성 연구)

  • Bae, Sung-Min;Lee, Hae-Yong;Chae, Soo-Ahn;Oh, Dong-Jin;Park, Suk-Won;Yoon, Yoo-Sik
    • Journal of Life Science
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    • v.21 no.9
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    • pp.1301-1309
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    • 2011
  • The major function of adipocytes is to store fat in the form of triglycerides. One of the signaling pathways known to affect adipogenesis, i.e. fat formation, is the WNT/${\beta}$-catenin pathway which inhibits the expression and activity of key regulators of adipogenesis. The purpose of this research is to find genes among the WNT/${\beta}$-catenin pathway which regulate adipogenesis by using small interfering (si) RNA and to find the association of single nucleotide polymorphisms (SNPs) of the gene with serum triglyceride levels in the human population. To elucidate the effects of ${\beta}$-catenin siRNA on adipogenesis key factors, PPAR${\gamma}$ and C/EBP${\alpha}$, we performed real-time PCR and western blotting experiments for the analyses of mRNA and protein levels. It was found that the siRNA-mediated knockdown of ${\beta}$-catenin upregulates adipogenesis key factors. However, upstream regulators of the WNT/${\beta}$-catenin pathway, such as DVL2 and LRP6, had no significant effects compared to ${\beta}$-catenin. These results indicate that ${\beta}$-catenin is a candidate gene for human fat accumulation. In general, serum triglyceride level is a good indicator of fat accumulation in humans. According to statistical analyses of the association between serum triglyceride level and SNPs of ${\beta}$-catenin, -10,288 C>T SNP (rs7630377) in the promoter region was significantly associated with serum triglyceride levels (p<0.05) in 290 Korean subjects. On the other hand, serum cholesterol levels were not significantly associated with SNPs of the ${\beta}$-catenin gene. The results of this study showed that ${\beta}$-catenin is associated with fat accumulation both in vitro and in the human population.

A Study on the Gene Expression of Adipogenic Regulators by an Herbal Composition (생약복합물에 의한 지방세포형성 조절자의 유전자 발현 연구)

  • Lee, Hae-Yong;Kang, Ryun-Hwa;Bae, Sung-Min;Chae, Soo-Ahn;Lee, Jung-Ju;Oh, Dong-Jin;Park, Suk-Won;Cho, Soo-Hyun;Shim, Yae-Jie;Yoon, Yoo-Sik
    • Journal of Life Science
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    • v.20 no.5
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    • pp.729-735
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    • 2010
  • In our previous study, it was reported that an herbal mixture, SH21B, inhibits fat accumulation and adipogenesis both in vitro and in vivo models of obesity. SH21B is a mixture composed of seven herbs: Scutellaria baicalensis Georgi, Prunus armeniaca Maxim, Ephedra sinica Stapf, Acorus gramineus Soland, Typha orientalis Presl, Polygala tenuifolia Willd, and Nelumbo nucifera Gaertner (Ratio 3:3:3:3:3:2:2). The aim of this study was to investigate the detailed molecular mechanisms of the effects of SH21B on various regulators of the adipogenesis pathway. During the adipogenesis of 3T3-L1 cells, SH21B significantly decreased the expression levels of central transcription factors of adipogenesis, such as peroxisome proliferator-activated receptor (PPAR)$\gamma$ and CCAAT/enhancer binding protein (C/EBP)$\alpha$. To elucidate the detailed molecular mechanism of the anti-adipogenic effects of SH21B, we examined the expression levels of the various pro-adipogenic or anti-adipogenic regulators of adipogenesis upstream of $PPAR{\gamma}$ and C/$EBP{\alpha}$. The mRNA levels of Krox20 and Kruppel-like factor (KLF) 15, which are pro-adipogenic regulators, were significantly down-regulated by SH21B treatment, whereas the mRNA levels of C/$EBP{\gamma}$ and KLF5 were not changed. KLF2 and C/EBP homologous protein (CHOP), which are anti-adipogenic regulators, were significantly up-regulated by SH21B treatment. These results suggest that the molecular mechanism of the anti-adipogenic effect of SH21B involves both the down-regulations of pro-adipogenic regulators, such as Krox20 and KLF15, and the up-regulations of anti-adipogenic regulators, such as KLF2 and CHOP, which results in the suppression of central transcription factors of adipogenesis including $PPAR{\gamma}$ and C/$EBP{\alpha}$.

Anti-atopic Effects of Castanea crenata Inner Shell Extracts Fermented by Lactobacillus bifermentans (Lactobacillus bifermentans로 발효한 율피의 항아토피 효과)

  • Kim, Bae Jin;Son, Woo Rim;Choi, Mi Ok;Jo, Seung Kyeung;Jung, Hee Kyoung;Lee, Jin Tae;Kim, Hak Yoon;Kwoen, Dae Jun
    • Journal of the Korean Society of Food Science and Nutrition
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    • v.42 no.9
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    • pp.1378-1386
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    • 2013
  • Atopic dermatitis (AD) is a common skin disease characterized by chronic and relapsing inflammatory dermatitis with immunological disturbances. In spite of the continuous increase in the incidence of AD, it is regrettable that till date there is no effective treatment to treat the same. Therefore, the present study was designed to examine the possible anti-atopic effects of Castanea crenata inner shell extracts fermented by Lactobacillus bifermentans (FCS) in 2,4-dinitrochlorobenzene (DNCB) induced AD in NC/Nga mice. Based on the results of HPLC analysis, we found that FCS contains anti-inflammatory factors such as gallic acid (10.18 mg/g) and ellagic acid (2.14 mg/g). The groups that we have used in this study included 0.1%, 1%, 5% fermented Castanea crenata inner shell extracts (FCS 0.1, FCS 1, FCS 5), 1,3-butylene glycol treated control (AD), and normal mice. After topical FCS treatment, we observed that the clinical severity score for AD was lower in both the FCS 1 and FCS 5 groups than the AD group. We also proved beyond doubt that there was improvement of melanin, erythema and skin moisture indices in the FCS 5 group. Spleen index and gene expression levels of pro-inflammatory cytokines such as IL-$1{\beta}$ and TNF-${\alpha}$ were significantly decreased in the FCS 5 group compared to the AD group (P<0.05). Further, we also found that the level of serum immunoglobulin E (IgE) in the FCS-treated group was decreased in a concentration-dependent manner. The results of our study suggest that FCS can be effectively used as a cosmeceutical ingredient for both the prevention and improvement of AD.

Neuroprotective effects of Momordica charantia extract against hydrogen peroxide-induced cytotoxicity in human neuroblastoma SK-N-MC cells (산화적 스트레스에 대한 여주 (Momordica charantia) 추출물의 항산화 효과 및 세포사멸 억제 기전을 통한 신경세포보호효과)

  • Kim, Kkot Byeol;Lee, Seonah;Heo, Jae Hyeok;Kim, Jung hee
    • Journal of Nutrition and Health
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    • v.50 no.5
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    • pp.415-425
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    • 2017
  • Purpose: Many studies have suggested that neuronal cells protect against oxidative stress-induced apoptotic cell death by polyphenolic compounds. We investigated the neuroprotective effects and the mechanism of action of Momordica charantia ethanol extract (MCE) against $H_2O_2-induced$ cell death of human neuroblastoma SK-N-MC cells. Methods: The antioxidant activity of MCE was measured by the quantity of total phenolic acid compounds (TPC), quantity of total flavonoid compounds (TFC), and 2,2-Diphenyl-1-pycrylhydrazyl (DPPH) radical scavenging activity. Cytotoxicity and cell viability were determined by CCK-8 assay. The formation of reactive oxygen species (ROS) was measured using 2,7-dichlorofluorescein diacetate (DCF-DA) assay. Antioxidant enzyme (SOD-1,2 and GPx-1) expression was determined by real-time PCR. Mitogen-activated protein kinases (MAPK) pathway and apoptosis signal expression was measured by Western blotting. Results: The TPC and TFC quantities of MCE were 28.51 mg gallic acid equivalents/extract g and 3.95 mg catechin equivalents/extract g, respectively. The $IC_{50}$ value for DPPH radical scavenging activity was $506.95{\mu}g/ml$ for MCE. Pre-treatment with MCE showed protective effects against $H_2O_2-induced$ cell death and inhibited ROS generation by oxidative stress. SOD-1,2 and GPx-1 mRNA expression was recovered by pre-treatment with MCE compared with the presence of $H_2O_2$. Pre-treatment with MCE inhibited phosphorylation of p38 and the JNK pathway and down-regulated cleaved caspase-3 and cleaved PARP by $H_2O_2$. Conclusion: The neuroprotective effects of MCE in terms of recovery of antioxidant enzyme gene expression, down-regulation of MAPK pathways, and inhibition apoptosis is associated with reduced oxidative stress in SK-N-MC cells.

The Influence of Atopic Findings on Severity of Pneumonia in Children with 2009 Pandemic Influenza A (H1N1) Infection (2009 신종 인플루엔자 A (H1N1) 폐렴 환아에서 아토피 소견이 폐렴의 중증도에 미치는 영향)

  • Kim, Jong Hee;Kim, Hyun Jeong;Kang, Im Ju
    • Pediatric Infection and Vaccine
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    • v.18 no.2
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    • pp.182-192
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    • 2011
  • Purpose : Atopic findings may be associated with severity of pneumonia in 2009 pandemic influenza A (H1N1) infection, which could suggest a possible association between atopic findings and the severity of viral infections. Thus, we studied association between atopic findings and severity of disease in children with H1N1 influenza infection. Methods : A retrospective study was performed in 74 children admitted in a single tertiary institute and confirmed as H1N1 patients by reverse transcriptase (RT) - polymerase chain reaction (PCR). They were divided into 2 groups according to the severity of pneumonia. We evaluated whether the atopic finding is risk factor between the two groups. Results : Children with severe pneumonia had higher percentages of serum eosinophilia (88% vs 40%, P <0.001), asthma (65% vs 35%, P =0.011), allergic rhinitis (71% vs 40%, P =0.009), and IgE level (P =0.007). We found positive correlations between aeroallergen sensitizations and severity of pneumonia (82% vs 53%, P =0.007). Conclusion : Among patients with H1N1 pneumonia, asthma and atopic findings are risk factors for severity of pneumonia.

Anti-inflammation effect of blueberry (Vaccinium ashei) leaf extract on RAW 264.7 macrophages stimulated by lipopolysaccharide (Lipopolysaccharide에 의해 활성화된 RAW 264.7대식세포에서 블루베리 잎(Vaccinium ashei) 추출물의 항염증 효과)

  • Kim, Dong In;Kim, Hyun Jung;Yun, Jong Moon;Lee, Ji Hye;Han, So Jung;Kim, Ha Eun;Jang, Min Jung;An, Bong Jeun
    • Food Science and Preservation
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    • v.25 no.1
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    • pp.107-116
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    • 2018
  • The aim of this study is to investigate the antioxidant and intracellular anti-inflammatory efficacy of blueberry leaf extracted with hot water (BLW), 70% ethanol (BLE), and 70% acetone (BLA) in RAW 264.7 macrophages. In order to evaluate the anti-inflammatory effect of blueberry leaf extracts, RAW 264.7 macrophages were stimulated with lipopolysaccharide (LPS) to induce the production of inflammation-related factors, which were measure by Western blotting and real-time PCR methods. i-NOS, COX-2 protein, and mRNA expression showed concentration-dependent decrease. The decreases in the mRNA expression levels of interleukin-$1{\beta}$ (IL-$1{\beta}$), interleukin-6 (IL-6), tumor necrosis factor-${\alpha}$ (TNF-${\alpha}$), and prostaglandin $E_2$ ($PGE_2$) were concentration-dependent. Further, the antioxidant effects of blueberry leaf on total polyphenol contents, electron donating ability and $ABTS^+$ radical scavenging activity were evaluated. The total polyphenol contents of BLW, BLE, and BLA were $217.04{\pm}2.98$, $156.72{\pm}3.90$, and $182.88{\pm}3.02mg\;TAE/g$, respectively, while the electron donating abilities at $1,000{\mu}g/mL$ of BLW, BLE, and BLA were 81.7, 79.6, and 79.3%, respectively. The $ABTS^+$ radical scavenging activity was fond to be concentration dependent. The nitric oxide (NO) production inhibition activities at $50{\mu}g/mL$ of BLW, BLE, and BLA were 35.1, 42.4 and 42.7%, respectively. In conclusion, the antioxidant and anti-inflammatory test results indicate that blueberry leaf extracts (BLW, BLE, and BLA) can be used as potential anti-inflammatory agents.