• Title/Summary/Keyword: RAF

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Effect of Yong-dam-sa-gan-tang on apoptosis in human hepatoma HepG2 (용담사간탕(龍膽瀉肝湯)에 의해 유도된 MAP kinases 활성화를 통한 간암 세포주 HepG2의 세포사멸)

  • Yun, Hyun-Jeong;Kim, Han-Seong;Heo, Sook-Kyoung;Hwang, Seong-Goo;Park, Won-Hwan;Park, Sun-Dong
    • Herbal Formula Science
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    • v.15 no.2
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    • pp.127-137
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    • 2007
  • The purpose of this study was to investigate the effect of Yong-dam-sa-gan-tang (YST) on apoptosis in HepG2 cells, First of all. to study the cytotoxic effect of methanol extract of YST on HepG2 cells, the cells were treated with various concentrations of YST and then cell viability was determined by XTT reduction method and trypan blue exclusion assay. YST reduced proliferation of HepG2 cells in a dose-dependent manner. To confirm the induction of apoptosis, HepG2 cells were treated with various concentrations of YST. The cleavage of poly AD P-ribose polymerase (P ARP), a substrate for caspase-3 and a typical sign of apoptosis, and the activation of caspase-3, procaspase-8 and procaspase-8 were examined by western blot analysis. YST decreased procaspase-3, procaspase-8 and procaspase-9 levels in a dose-dependent manner and induced the clevage of PARP. YST triggered the mitochondrial apoptotic signaling by increasing the release of cytochrome c from mitochondria to cytosol. Furthermore, YST also downregulated the anti-apoptotic Bcl-2 and upregulated the pro-apoptotic-Bax. Therefore, this result suggest that YST induced HepG2 cell death through the mitochondrial pathway. Sustained activation of the Ras/Raf/MEK/ERK cascade in cells results in a cell cycle arrest and has been implicated in the differentiation of certain cell types, in many cases acting to promote differentiation. YST decreased the activation of Ras/Raf/MEK/ERK cascade in a dose-dependent manner. These results suggest that YST is potentially useful as a chemo-therapeutic agent in HepG2.

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The Phospholipase-Protein Kinase C-MEK-ERK Pathway is Essential in Mycobacteria-induced CCL3 and CCL4 Expression in Human Monocytes (사람 단핵구에서 결핵균에 의해 유도되는 CCL3 및 CCL4 발현에 대한 Phospholipase-Protein Kinase C-MEK-ERK 경로의 역할 분석)

  • Yang, Chul-Su;Song, Chang-Hwa;Jung, Saet-Byel;Lee, Kil-Soo;Kim, Su-Young;Lee, Ji-Sook;Shin, A-Rum;Oh, Jae-Hee;Kwon, Yu-Mi;Kim, Hwa-Jung;Park, Jeong-Kyu;Paik, Tae-Hyun;Jo, Eun-Kyeong
    • IMMUNE NETWORK
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    • v.5 no.4
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    • pp.237-246
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    • 2005
  • Background: Little information is available on the identification and characterization of the upstream regulators of the signal transduction cascades for Mycobacterium tuberculosis (M. tbc)-induced ERK 1/2 activation and chemokine expression. We investigated the signaling mechanisms involved in expression of CCL3 /MIP-1 and CCL4/MIP-1 in human primary monocytes infected with M. tbc. Methods: MAP kinase phosphorylation was determined using western blot analysis with specific primary antibodies (ERK 1/2, and phospho-ERK1/2), and the upstream signaling pathways were further investigated using specific inhibitors. Results: An avirulent strain, M. tbc H37Ra, induced greater and more sustained ERK 1/2 phosphorylation, and higher CCL3 and CCL4 production, than did M. tbc H37Rv. Specific inhibitors for mitogen-activated protein kinase (MAPK) kinase (MEK; U0126 and PD98059) significantly inhibited the expression of CCL3 and CCL4 in human monocytes. Mycobactetia-mediated expression of CCL3 and CCL4 was not inhibited by the Ras inhibitor manumycin A or the Raf-1 inhibitor GW 5074. On the other hand, phospholipase C (PLC) inhibitor (U73122) and protein kinase C (PKC)specific inhibitors ($G\ddot{o}6976$ and Ro31-8220) significantly reduced M. tbc-induced activation of ERK 1/2 and chemokine synthesis. Conclusion: These results are the first to demonstrate that the PLC-PKC-MEK-ERK, not the Ras-Raf-MEK-ERK, pathway is the major signaling pathway inducing M. tbc-mediated CCL3 and CCL4 expression in human primary monocytes.

Effect of an Aqueous Extract of Poncirus trifoliate (L.) Raf. in Stroke Patient with Constipation (뇌졸중환자의 변비에서 지실 열수 추출물의 효과)

  • Moon, Hyo Jeong;Lee, Su Kyung;Noh, Se Eung;Joo, Min Cheol
    • Journal of Korean Medicine Rehabilitation
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    • v.26 no.2
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    • pp.97-103
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    • 2016
  • Objectives To evaluate the effects and safety of the aqueous extract of the dried, immature fruit of Poncirus trifoliate (L.) Raf. (Rutaceae) (PF) in stroke patients with constipation. Methods A total of 22 patients were recruited. Patients were interviewed about the clinical informations, constipation score and Bristol stool form scale at twice, before intake PF and after intake PF 2 weeks. The total and segmental colon transit time (CTT) were measured by using radio-opaque markers (Kolomark$^{(R)}$). The degree of stool retention was evaluated by the plain abdominal radiography and was scored by Leech score. Results Before intake PF, constipation scores ranged from 3 to 12, average $6.54{\pm}2.87$ and Bristol stool form scale ranged from 1 to 6, average $3.86{\pm}1.21$. CTTs were $9.05{\pm}6.89hours$, $14.29{\pm}10.68hours$, $12.11{\pm}7.19hours$ and $35.40{\pm}19.5hours$ in the right, left, rectosigmoid and total colon, respectively. Stool retention score was $2.45{\pm}0.61$, $2.3{\pm}0.86$, $1.9{\pm}0.85$, $6.65{\pm}1.56$ in the right, left, rectosigmoid and total colon, respectively. After 2 weeks, constipation scores ranged from 2 to 8, average $4.28{\pm}2.05$ and Bristol stool form scale ranged from 1 to 6, average $4.17{\pm}1.04$. CTTs were $7.41{\pm}8.86hours$, $11.12{\pm}9.12 hours$, $8.83{\pm}8.75hours$ and $27.3{\pm}20.2$ hours in the right, left, rectosigmoid and total colon, respectively. Stool retention score was $1.9{\pm}0.64$, $2.2{\pm}0.69$, $1.4{\pm}0.88$, $5.5{\pm}1.39$ in the right, left, rectosigmoid and total colon, respectively. There were statistically significant difference in the total and rectosigmoid colon CTT and constipation score, Stool retention score in right and rectosigmoid colon (p<0.05) after PF therapy. Conclusions These results suggest potential for PF therapy in stroke patient with constipation.

Taxifolin Inhibited the Nitric Oxide Production and Expression of Pro-inflammatory Cytokine mRNA in Lipopolysaccharide-stimulated RAW264.7 Cells

  • Rhee, Man-Hee;Endale, Mehari;Kamruzzaman, SM;Lee, Whi-Min;Park, Hwa-Jin;Yoo, Myung-Jo;Cho, Jae-Youl
    • Biomedical Science Letters
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    • v.14 no.3
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    • pp.147-155
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    • 2008
  • In previous works, we found that solvent extract of Opuntia humifusa Raf., a member of the lactaceae family, displayed potent anti-oxidative and anti-inflammatory activities. Thus, all solvent fractions, except for the water layer, showed potent scavenging effects. According to activity-guided fractionation, one of active radical scavenging principles in the ethyl acetate fraction was found to be taxifolin. In this study, we investigated whether taxifolin showed anti-oxidative activity. In addition, taxifolin modulated nitric oxide (NO) release and the expression of pro-inflammatory cytokine mRNA such as interleukin-$1{\beta}$ (IL-$1{\beta}$), IL-6, granulocyte-macrophage colony-stimulating factor (GM-CSF), and TNF-${\alpha}$. Taxifolin showed potent anti-oxidant activity with the $IC_{50}\;of\;8.5{\pm}1.4\;and\;9.3{\pm}1.0{\mu}M$ using xanthine/xanthine oxidase (XO) assay and 2,2-Diphenyl-lpicrylhydrazyl radical (DPPH) assay, respectively. We next determined the role of taxifolin on the immunomodulating activity using murine macrophage cell line RAW264.7 cells. Taxifolin dose-dependently inhibited NO production in lipopolysaccharide (LPS)-activated RAW264.7. It also significantly blocked the expression of inducible NO synthase (iNOS) mRNA in the LPS-stimulated RAW264.7 cells. In addition, taxifolin potently suppressed the expression of IL-$1{\beta}$, IL-6 and GM-CSF mRNA in LPS-activated RAW264.7 cells, but not that of TNF-${\alpha}$ Moreover, taxifolin significantly inhibited the transcriptional activity of nuclear factor-${\kappa}B$ (NF-${\kappa}B$) and activator protein -1 (AP-1). These results suggest that taxifolin may downregulate inflammatory iNOS, IL-$1{\beta}$, IL-6 and GM-CSF gene expressions through inhibition of NF-K and AP-1 activation in LPS-stimulated RAW264.7 cells.

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4-(N-Methyl-N-nitrosamino)-1(3-pyridyl)-1-butanone(NNK) Restored the Cap-dependent Protein Translation Blocked by Rapamycin

  • Kim Jun-Sung;Park Jin Hong;Park Sung-Jin;Kim Hyun Woo;Hua Jin;Cho Hyun Sun;Hwang Soon Kyung;Chang Seung Hee;Tehrani Arash Minai;Cho Myung Haing
    • Toxicological Research
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    • v.21 no.4
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    • pp.347-353
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    • 2005
  • Eukaryotic initiation factor 4E (elF4E) is a key element for cap-dependent protein translation controlled by affinity between elF4E and 4E-binding protein 1 (4E-BP1). Rapamycin can also affect protein translation by regulating 4E-BP1 phosphorylation. Tobacco-specific nitrosamine, 4(N-methyl-N-nitrosamino )-1-(3-pyridyl)-1-butanone (NNK) is a strong lung carcinogen, but its precise lung cancer induction mechanism remains unknown. Relative roles of cap-dependent and -independent protein translation in terms of NNK-induced lung carcinogenesis were elucidated using normal human bronchial epithelial cells. NNK concentrations applied in this study did not decrease cell viability. Addition of NNK restored rapamycin-induced decrease of protein synthesis and rapamycin-induced phosphorylation of 4E-BP1, and increased expression levels of mTOR, ERK1/2, p70S6K, and Raf-1 in a concentration-dependent manner. NNK also caused perturbation of normal cell cycle progression. Taken together, NNK might cause toxicity through the combination of restoration of 4E-BP1 phosphorylation and increase of elF4E as well as mTOR protein expression, interruption of Raf1/ERK as well as the cyclin G-associated p53 network. Our data could be applied towards elucidation of the molecular basis for lung cancer treatment.

Connection the Rhizomicrobiome and Plant MAPK Gene Expression Response to Pathogenic Fusarium oxysporum in Wild and Cultivated Soybean

  • Chang, Chunling;Xu, Shangqi;Tian, Lei;Shi, Shaohua;Nasir, Fahad;Chen, Deguo;Li, Xiujun;Tian, Chunjie
    • The Plant Pathology Journal
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    • v.35 no.6
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    • pp.623-634
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    • 2019
  • Little known the connections between soybeans mitogen-activated protein kinase (MAPK) gene expression and the rhizomicrobiome upon invasion of the root pathogen Fusarium oxysporum. To address this lack of knowledge, we assessed the rhizomicrobiome and root transcriptome sequencing of wild and cultivated soybean during the invasion of F. oxysporum. Results indicated F. oxysporum infection enriched Bradyrhizobium spp. and Glomus spp. and induced the expression of more MAPKs in the wild soybean than cultivated soybean. MAPK gene expression was positively correlated with Pseudomonadaceae but negatively correlated with Sphingomonadaceae and Glomeraceae in both cultivated and wild soybean. Specifically, correlation profiles revealed that Pseudomonadaceae was especially correlated with the induced expression of GmMAKKK13-2 (Glyma.14G195300) and GmMAPK3-2 (Glyma.12G073000) in wild and cultivated soybean during F. oxysporum invasion. Main fungal group Glomeraceae was positively correlated with GmMAPKKK14-1 (Glyma.18G060900) and negatively correlated with GmRaf6-4 (Glyma.02G215300) in the wild soybean response to pathogen infection; while there were positive correlations between Hypocreaceae and GmMAPK3-2 (Glyma.12G073000) and between Glomeraceae and GmRaf49-3 (Glyma.06G055300) in the wild soybean response, these correlations were strongly negative in the response of cultivated soybean to F. oxysporum. Taken together, MAPKs correlated with different rhizomicrobiomes indicating the host plant modulated by the host self-immune systems in response to F. oxysporum.

Antimicrobial and Antifungal Activities of Lisianthus (Eustoma grandiflorum) Essential Oil (리시안셔스 유래 에센셜 오일의 항세균 및 항진균 효과)

  • Ji, Keunho;Kim, Dong Kwang;Kim, Young Tae
    • Journal of Life Science
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    • v.27 no.4
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    • pp.430-434
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    • 2017
  • Essential oils are fragrant oils extracted from the leaves, stems, peels, petals and roots of aromatic plants cultivated by natural means or using organic agricultural techniques. Essential oils have commonly been used as antibacterial and antifungal agents. In the present study, essential oil was extracted from lisianthus (Eustoma grandiflorum [Raf.] Shinn.) and tested for antifungal activities against three eumycetes (Penicillium pinophilum, Chaetomium glogosum and Aspergillus niger). Lisianthus essential oil showed high antifungal activities against three eumycetes, especially against Aspergillus niger, for which the resulting minimum inhibitory concentration (MIC) was 0.005 mg/ml. In addition, the extracted essential oil was shown to have antimicrobial activity against ten intestinal pathogenic bacteria (Escherichia coli, Salmonella typhimurium, Klebsiella pneumonia, Staphylococcus aureus, Bacillus cereus, Listeria monocytogenes, Pseudomonas aeruginosa, Bacillus subtilis, Enterococcus faecalis and Vibrio parahaemolyticus) according to the disc diffusion method and was also shown to exhibit strong antibacterial activity against an additional three pathogenic bacteria (Bacillus subtilis, Listeria monocytogenes and Vibrio parahaemolyticus). These results indicate that lisianthus essential oil could be used as an antibiotic against harmful bacteria that produce intestinal illnesses. From the present study, we suggest that lisianthus extracts can be utilized as potential antifungal and antibacterial agents and for the development of pharmaceutical and cosmetic products.

Germination Effect of Soybean on Its Contents of Isoflavones and Oligosaccharides

  • Kim, Woo-Jung;Lee, Hye-Yeon;Won, Moo-Ho;Yoo, Sang-Ho
    • Food Science and Biotechnology
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    • v.14 no.4
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    • pp.498-502
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    • 2005
  • Three Korean soybean varieties - Shinpaldal-2, Seomoktae and Seoritae - were investigated for changes in their physical properties and the amount of functional components (i.e. isoflavones and oligosaccharides), during germination. Soybeans were germinated at $20^{\circ}C$ for 96 hr in complete darkness. The dry weights of cotyledone, hypocotyl, seed coat, and hilum of Seoritae were heavier than those of other varieties. The dry weights of the three bean varieties decreased steadily in spite of root growth. The largest amount of isoflavone content was observed from Shinpaldal-2 (1.824 mg/g), followed by Seoritae (1.216 mg/g) and Seomoktae (1.125 mg/g). Total isoflavone content increased by 13% during initial germination, and then decreased thereafter. Aglycone types such as daidzein and genistein dominated the increase in isoflavone contents. The increase in genistein content of Shinpaldal-2 was 17.5 fold compared with ungerminated soybean, while the amount of daidzein was 6.7 times as much as ungerminated Shinpaldal-2 over an 18-hr germination period. Oligosaccharide contents such as raffinose (Raf) and stachyose (Sta) rapidly decreased during germination, while the sucrose (Sue) content remained constant until 36-48 hr of germination. From these results, it was clearly shown that the germination process significantly changed the contents of functional nutrients in soybeans. Therefore, the optimization of germination process should be considered to improve the biological functionality of soybeans in food processing.

Induction of the Nuclear Proto-Oncogene c-fos by the Phorbol Ester TPA and c-H-Ras

  • Kazi, Julhash U.;Soh, Jae-Won
    • Molecules and Cells
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    • v.26 no.5
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    • pp.462-467
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    • 2008
  • TPA is known to cooperate with an activated Ras oncogene in the transformation of rodent fibroblasts, but the biochemical mechanisms responsible for this effect have not been established. In the present study we used c-fos promoter-luciferase constructs as reporters, in transient transfection assays, in NIH3T3 cells to assess the mechanism of this cooperation. We found a marked synergistic interaction between TPA and a transfected v-Ha-ras oncogene in the activation of c-fos promoter and SRE. SRE has binding sites for TCF and SRF. A dominant-negative Ras (ras-N17) inhibited the TPA-Ras synergy by blocking the PKC-MAPK-TCF pathway. Dominant-negative RhoA and Rac1 (but not Cdc42Hs) inhibited the TPA-Ras synergy by blocking the Ras-Rho-SRF signaling pathway. Constitutively active $PKC{\alpha}$ and $PKC{\varepsilon}$ showed synergy with v-Ras. These results suggest that the activation of two distinct pathways such as Ras-Raf-ERK-TCF pathway and Rho-SRF pathway are responsible for the induction of c-fos by TPA and Ras in mitogenic signaling pathways.

Overexpression of GAP Causes the Delay of NGF-induced Neuronal Differentiation and the Inhibition of Tyrosine Phosphorylation of SNT in PC12 Cells

  • Yang, Sung-Il;Kaplan, David
    • BMB Reports
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    • v.28 no.4
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    • pp.316-322
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    • 1995
  • The GTPase activating protein (GAP) can function both as a negative regulator and an effector of $p21^{ras}$. Overexpression of GAP in NIH-3T3 cells has been shown to inhibit transformation by ms or src. To investigate the function of GAP in a differentiative system, we overexpressed this protein in the nerve growth factor (NGF)-responsive PC12 cell line. Two-fold overexpression of GAP caused a delay of several days in the onset of NGF- but not FGF-induced neuronal differentiation of PC12 cells. However, the NGF-induced activation or tyrosine phosphorylation of upstream (Trk, PLC-${\gamma}1$, SHC) and downstream (B-Raf and $p44^{mapk/erk1}$) components of $p21^{ras}$, signalling cascade was not altered by GAP overexpression. Therefore, the change of phenotype induced by GAP was probably not due to GAP functioning as a negative regulator of $p21^{ras}$. Rather, we found that NGF-induced tyrosine phosphorylation of SNT, a specific target of neurotrophin-induced tyrosine kinase activity, was inhibited by GAP overexpression. SNT is thought to function upstream or independent of $p21^{ras}$. Thus in PC12 cells, overexpressed GAP may control the rate of neuronal differentiation through a pathway involving SNT rather than the $p21^{ras}$ signalling pathway.

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