• Title/Summary/Keyword: E2-like enzyme

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Hepaprotective Effect of Standardized Ecklonia stolonifera Formulation on CCl4-Induced Liver Injury in Sprague-Dawley Rats

  • Byun, Jae-Hyuk;Kim, Jun;Choung, Se-Young
    • Biomolecules & Therapeutics
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    • v.26 no.2
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    • pp.218-223
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    • 2018
  • The liver is an essential organ for the detoxification of exogenous xenobiotics, drugs and toxic substances. The incidence rate of non-alcoholic liver injury increases due to dietary habit change and drug use increase. Our previous study demonstrated that Ecklonia stolonifera (ES) formulation has hepatoprotective effect against alcohol-induced liver injury in rat and tacrine-induced hepatotoxicity in HepG2 cells. This present study was designated to elucidate hepatoprotective effects of ES formulation against carbon tetrachloride ($CCl_4$)-induced liver injury in Sprague Dawley rat. Sixty rats were randomly divided into six groups. The rats were treated orally with ES formulation and silymarin (served as positive control, only 100 mg/kg/day) at a dose of 50, 100, or 200 mg/kg/day for 21 days. Seven days after treatment, liver injury was induced by intraperitoneal injection of $CCl_4$ (1.5 ml/kg, twice a week for 14 days). The administration of $CCl_4$ exhibited significant elevation of hepatic enzymes (like AST and ALT), and decrease of antioxidant related enzymes (superoxide dismutase, glutathione peroxidase and catalase) and glutathione. Then, it leaded to DNA damages (8-oxo-2'-deoxyguanosine) and lipid peroxidation (malondialdehyde). Administration of ES formulation inhibited imbalance of above factors compared to $CCl_4$ induced rat in a dose dependent manner. Real time PCR analysis indicates that CYP2E1 was upregulated in $CCl_4$ induced rat. However, increased gene expression was compromised by ES formulation treatment. These findings suggests that ES formulation could protect hepatotoxicity caused by $CCl_4$ via two pathways: elevation of antioxidant enzymes and normalization of CYP2E1 enzyme.

Adenine attenuates lipopolysaccharide-induced inflammatory reactions

  • Silwal, Prashanta;Lim, Kyu;Heo, Jun-Young;Park, Jong IL;Namgung, Uk;Park, Seung-Kiel
    • The Korean Journal of Physiology and Pharmacology
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    • v.22 no.4
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    • pp.379-389
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    • 2018
  • A nucleobase adenine is a fundamental component of nucleic acids and adenine nucleotides. Various biological roles of adenine have been discovered. It is not produced from degradation of adenine nucleotides in mammals but produced mainly during polyamine synthesis by dividing cells. Anti-inflammatory roles of adenine have been supported in IgE-mediated allergic reactions, immunological functions of lymphocytes and dextran sodium sulfate-induced colitis. However adenine effects on Toll-like receptor 4 (TLR4)-mediated inflammation by lipopolysaccharide (LPS), a cell wall component of Gram negative bacteria, is not examined. Here we investigated anti-inflammatory roles of adenine in LPS-stimulated immune cells, including a macrophage cell line RAW264.7 and bone marrow derived mast cells (BMMCs) and peritoneal cells in mice. In RAW264.7 cells stimulated with LPS, adenine inhibited production of pro-inflammatory cytokines $TNF-{\alpha}$ and IL-6 and inflammatory lipid mediators, prostaglandin $E_2$ and leukotriene $B_4$. Adenine impeded signaling pathways eliciting production of these inflammatory mediators. It suppressed $I{\kappa}B$ phosphorylation, nuclear translocation of nuclear factor ${\kappa}B$ ($NF-{\kappa}B$), phosphorylation of Akt and mitogen activated protein kinases (MAPKs) JNK and ERK. Although adenine raised cellular AMP which could activate AMP-dependent protein kinase (AMPK), the enzyme activity was not enhanced. In BMMCs, adenine inhibited the LPS-induced production of $TNF-{\alpha}$, IL-6 and IL-13 and also hindered phosphorylation of $NF-{\kappa}B$ and Akt. In peritoneal cavity, adenine suppressed the LPS-induced production of $TNF-{\alpha}$ and IL-6 by peritoneal cells in mice. These results show that adenine attenuates the LPS-induced inflammatory reactions.

Genome-wide survey and expression analysis of F-box genes in wheat

  • Kim, Dae Yeon;Hong, Min Jeong;Seo, Yong Weon
    • Proceedings of the Korean Society of Crop Science Conference
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    • 2017.06a
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    • pp.141-141
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    • 2017
  • The ubiquitin-proteasome pathway is the major regulatory mechanism in a number of cellular processes for selective degradation of proteins and involves three steps: (1) ATP dependent activation of ubiquitin by E1 enzyme, (2) transfer of activated ubiquitin to E2 and (3) transfer of ubiquitin to the protein to be degraded by E3 complex. F-box proteins are subunit of SCF complex and involved in specificity for a target substrate to be degraded. F-box proteins regulate many important biological processes such as embryogenesis, floral development, plant growth and development, biotic and abiotic stress, hormonal responses and senescence. However, little is known about the F-box genes in wheat. The draft genome sequence of wheat (IWGSC Reference Sequence v1.0 assembly) used to analysis a genome-wide survey of the F-box gene family in wheat. The Hidden Markov Model (HMM) profiles of F-box (PF00646), F-box-like (PF12937), F-box-like 2 (PF13013), FBA (PF04300), FBA_1 (PF07734), FBA_2 (PF07735), FBA_3 (PF08268) and FBD (PF08387) domains were downloaded from Pfam database were searched against IWGSC Reference Sequence v1.0 assembly. RNA-seq paired-end libraries from different stages of wheat, such as stages of seedling, tillering, booting, day after flowering (DAF) 1, DAF 10, DAF 20, and DAF 30 were conducted and sequenced by Illumina HiSeq2000 for expression analysis of F-box protein genes. Basic analysis including Hisat, HTseq, DEseq, gene ontology analysis and KEGG mapping were conducted for differentially expressed gene analysis and their annotation mappings of DEGs from various stages. About 950 F-box domain proteins identified by Pfam were mapped to wheat reference genome sequence by blastX (e-value < 0.05). Among them, more than 140 putative F-box protein genes were selected by fold changes cut-offs of > 2, significance p-value < 0.01, and FDR<0.01. Expression profiling of selected F-box protein genes were shown by heatmap analysis, and average linkage and squared Euclidean distance of putative 144 F-box protein genes by expression patterns were calculated for clustering analysis. This work may provide valuable and basic information for further investigation of protein degradation mechanism by ubiquitin proteasome system using F-box proteins during wheat development stages.

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Antioxidant Ability and Hair Growth Effect of Samhwang-Sasimtang in C57BL/6 Model (삼황사심탕의 항산화능 및 C57BL/6 마우스 모델에서의 발모 촉진효과)

  • Kim, Min Ji;Park, Kyo Hyun;Lee, In Chul;Kim, Bae Hwan
    • Journal of Physiology & Pathology in Korean Medicine
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    • v.28 no.2
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    • pp.154-161
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    • 2014
  • The purpose of this study is to evaluate the antioxidant activity and hair growth effects of a natural herbal ethanol extract, Samhwang-Sasimtang(SS). In case of antioxidant ability of SS, the content of phenolic compounds was 28.44mg/g. The extract showed strong electron donating ability and free radical scavenging activity in a concentration-dependent manner. SOD-like activity also rose through increasing the concentrations of SS. In order to estimate the hair growth effects, the extract was applied to the back of seven-week-old C57BL/6 male mice ($150{\mu}{\ell}$ a day, five days a week, for four weeks) in four groups (C, control, saline; PC, positive control, 3% minoxidil; E1, experimental 1, 1% SS; E2, experimental 2, 2% SS). Ten mice were assigned to each group and five mice in each group were sacrificed at weeks 2 and 4, respectively. There was no a significant difference in body weight change among experimental groups. In macroscopic observation of hair growth at week 4, the scores of hair growth on the backs of mice were 80, 60, 40 and 20% in the PC, E2, E1 and C groups. In terms of histological observation, the ratio and thickness of hair follicles, the enzyme activities of ALP and ${\gamma}$-GT, immunohistological examination of IGF-1 and VEGF which are the hair growth factors, in each group were significantly high in this order: PC, E2, E1 and C group, at week 4. Meanwhile, hair growth inhibition factors, TGF-${\beta}1$ and Caspase-3, were reduced in PC, E1 and E2 groups compared with C group. These results indicate that SS extract may be effective in promoting hair growth, and suggest that it can be used practically as a superior natural agent for hair growth promotion.

The Relation of the Elevated Plasma Lipid Levels to Plasma Vitamin E Status and Activities of Erythrocyte Glutathione Perosicase in Smokers (흡연인들에서 증가된 혈장지질 농도가 비타민 E 영양상태와 글루타티온 과산화효 활성에 미치는 영향)

  • 윤군애
    • Journal of Nutrition and Health
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    • v.31 no.8
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    • pp.1254-1262
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    • 1998
  • This study has done to investigate the relationship between the icreased lipid oncentration caused by smoking and plama levels of vitamin A and vitamin E, antiodative enzyme activity, and lipid peroxidation , in 52 male smokers and 32 non-smokers, Dietary vitamin A and vitamin E intake was imilar in both smokers and non-smokers. Absolute plasma concentrations of vitamin A and vitamin E were not significantly different between two groups, whereas vitamin E/cholesterol ration in plasma was low or in smokers than in that of non-smokers(p<0.05). It was considered that this lowered effect was due to the elevated plasma lipid concentration rather than oxidant stress derived from smoking, in view of the fact that smokers had higher cholesterol (15.2%) adn LDL-C(26.6%) levels than non-smokers. In non-smokers, plasma thiobarbiturin acid reactive substances(TBARS) conrrelated positively with total cholesterol(r=0.63466, p<0.001), LDL-C level(r=0.57166, p<0.01) , and LDL-C/HDL-C ratio(r=0.45926, p<0.05) . Activities of glutathione perosidase(GSH-Px) , superoside dismutase(SOD), and catalse made no difference in both groups. However, it was observed in non-smokers that GSH-Px activity had negative correlations with total cholesterol(r=-0.67293, p<0.001), LDL-C level(r=-0.62878, p<0.001), and LDL-C/HDL-C ratio (r=-0.58824, p<0.01), indicating that there was a dependent relationship between lipid perosidation and plasma lipid level. The smokers also showed negative correlations for GSH-Px activity with total cholesterol (r=-0.29946, p<0.05) and LDL-c level (r=0.45914, p<0.001), and LDL-C/HDL-c ratio(r=-0.35438, p<0.05). It seemed that the lipid that the lipid level elevated by sustaines smoking resulted in reducing vitamin E/cholesterol ratio and proportion of antioxidant to oxidant load, and then GSH-Px activity, with insufficient removal of free radicals(TBARS 2.43$\pm$0.51 and 1.81$\pm$0.15nmol/ml in smokers and non-smokers, respectively). These findings suggest that higher plasma lipid levels may play a more important role in perturbing the antioxidant defense system including vitamin E status and GSH-Px activity, at least in circumstances that increase lipid concentration . In addition, in exposure to free radicals like those in cigarette smoke. In those cases the ratio of vitamin E/lipid in plasma can be a more indicator of vitamin E status than plasma levels of vitamin E alone.

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Selection and Characterization of Forest Soil Metagenome Genes Encoding Lipolytic Enzymes

  • Hong, Kyung-Sik;Lim, He-Kyoung;Chung, Eu-Jin;Park, Eun-Jin;Lee, Myung-Hwan;Kim, Jin-Cheol;Cho, Gyung-Ja;Cho, Kwang-Yun;Lee, Seon-Woo
    • Journal of Microbiology and Biotechnology
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    • v.17 no.10
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    • pp.1655-1660
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    • 2007
  • A metagenome is a unique resource to search for novel microbial enzymes from the unculturable microorganisms in soil. A forest soil metagenomic library using a fosmid and soil microbial DNA from Gwangneung forest, Korea, was constructed in Escherichia coli and screened to select lipolytic genes. A total of seven unique lipolytic clones were selected by screening of the 31,000-member forest soil metagenome library based on tributyrin hydrolysis. The ORFs for lipolytic activity were subcloned in a high copy number plasmid by screening the secondary shortgun libraries from the seven clones. Since the lipolytic enzymes were well secreted in E. coli into the culture broth, the lipolytic activity of the subclones was confirmed by the hydrolysis of p-nitrophenyl butyrate using culture supernatant. Deduced amino acid sequence analysis of the identified ORFs for lipolytic activity revealed that 4 genes encode hormone-sensitive lipase (HSL) in lipase family IV. Phylogenetic analysis indicated that 4 proteins were clustered with HSL in the database and other metagenomic HSLs. The other 2 genes and 1 gene encode non-heme peroxidase-like enzymes of lipase family V and a GDSL family esterase/lipase in family II, respectively. The gene for the GDSL enzyme is the first description of the enzyme from metagenomic screening.

The Effect of Estrogen Deficiency on Ethanol Metabolism in Ovariectomized Rats (난소절제 랫트에서 에스트로겐 저하가 알코올 대사에 미치는 영향)

  • Chang, Bo-Yoon;Kim, Sung-Yeon
    • YAKHAK HOEJI
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    • v.54 no.6
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    • pp.522-528
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    • 2010
  • This study was designed to examine the effect of estrogen deficiency on the metabolism of ethanol in ovariectomized rats. Female rats were assigned to an ovariectomy (OVX) and a sham (SHAM) surgery group. Gain body weight was greater in incresed in OVX group and especially uterus weight significantly decrease depending on the concentration of estrogen after 3 month of ovariectomy. Ethanol at the tolerative dose (6 g/kg) was injected to rats by oral administration to measure the concentration of ethanol in blood. The area under the blood concentration time curve (AUC) was significantly lower in OVX group than SHAM group. The significant decrease in AUC in OVX group indicates that the estrogen deficiency leads to changes of the factors related to ethanol metabolism. Activity of hepatic alcohol dehydrogenase was not significantly influenced by the ovariectomy and also the ethanol elimination rate in vivo was not different. Cytochrome P450 isozymes did not show any changes except CYP 1A1 and 2E1. Level of hepatic glutathione in OVX group was higher after treatment of ethanol. Therefore the reduction of AUC appears not to be directly associated with the difference of ethanol metabolizing enzyme, but to be related with the physical factors like body weight.

Effects of Xenoestrogens on Gene Expression of Cytochrome P450 Genes in in vitro Cultured Mice Spermatogenic Cells (체외배양 생쥐정소세포에서 합성에스트로겐이 P450 등위효소의 발현에 미치는 영향)

  • Lee, Ho-Joon;Kim, Myo-Kyung;Ko, Duck-Sung;Kim, Kil-Soo;Kang, Hee-Kyoo;Kim, Dong-Hoon
    • Clinical and Experimental Reproductive Medicine
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    • v.28 no.2
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    • pp.131-140
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    • 2001
  • Objective: To know the effects of xenoestrogen on spermatogenesis, we investigated the expression of cytochrome P450s enzymes (CYPscc, $CYP_{17{\alpha}}$, CYP19) and $3{\beta}$-HSD genes involved in steroidogenesis. Methods: Mouse testicular cells were prepared from 15-day-old ICR mice which had only pre-meiotic germ cells by enzyme digestion using collagenase and trypsin. Testicular cells were cultured in DMEM supplemented with FSH (0.1 IU/ml) and 10% FBS or medium with estrogen ($E_2$), bisphenol-A (BPA), octylphenol (OP; $10^{-9},\;10^{-7},\;10^{-6},\;10^{-5},\;10^{-4}M$, respectively) and aroclor 1254 (A1254) known as PCBs for 48 hours. The gene expression of cytochrome P450 enzymes were examined by semi-quantitive RT-PCR. The production of estrogen and testosterone was examined by RIA. Results: As results, expression of CYPscc mRNA was not significantly decreased, but $3{\beta}$-HSD and $CYP_{17{\alpha}}$. mRNA were significantly dose-dependent decreased. And production of testosterone and estrogen were not different except BPA and OP group ($10^{-5}M$). Conclusion: BPA, OP and A1254 might inhibit steroidogenesis by decreasing CYPscc, $3{\beta}$-HSD and $CYP_{17{\alpha}}$. mRNA expression in the mouse testis. These results suggest that BPA, OP and PCBs like as an endocrine disruptors inhibit the productions of steroidogenic enzymes and decrease the production of T and E by negative feedback mechanism. Therefore, these might disrupt steroidogenesis in Leydig cells of testis and would disturb testicular function and subsequently impair spermatogenesis.

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Resistance Function of Woody Landscape Plants to Air Pollutants(II) - POD Activity - (조경수목(造景樹木)의 대기오염물질(大氣汚染物質)에 대한 방어기능(防禦機能)(II) - POD 활성(活性)을 중심으로 -)

  • Kim, Myung Hee;Lee, Soo Wook
    • Journal of Korean Society of Forest Science
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    • v.81 no.3
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    • pp.234-246
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    • 1992
  • This study was conducted to determine the toxic effects of air pollutants on landscaping tress, Pinus densiflora, Pinus koraiensis, Ginkgo biloba, Liriodendron tulipifera, Platanus occidentalis and their resistance to the pollutant toxicity in urban and industrial regions of Seoul and Taejon, Korea. Total sulfur contents and enzyme activities such as superoxide dismutase and peroxidase were analyzed in tree foliage of Pinus densiflora, Pinus koraiensis, Ginkgo biloba, Liriodendron tulipifera, Platanus occidentalis. In addition, POD activity was analyzed in the foliage on tree seedlings, i.e. Pinus densiflora, Pinus koraiensis, Ginkgo biloba, Lirioderdron tulipifera, with the fumigation of $SO_2$ in gas chamber 4 hours a day for six days. In Ginkgo biloba total sulfur content and POD activity had a negative correlation while other species had a positive relationship in total sulfur content and enzyme activity. Air pollutants accumulated in tree tissues were supposed to enhance the enzyme activity like POD providing the resistance mechanisms. Especially Pinus koraiensis and Platanus occidentalis had higher POD activity than other species. The increase of temporary POD activity against environmental stress appeared in sensitive trees and prolonged increase of POD activity played an important role in resistance mechanism. SOD and POD activities in all species had a positive correlation except Ginkgo biloba. Changes of SOD and POD activities were different between species and in most species SOD as well as POD seemed to participate in resistance mechanism.

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Molecular Cloning and Function Analysis of an Anthocyanidin Synthase Gene from Ginkgo biloba, and Its Expression in Abiotic Stress Responses

  • Xu, Feng;Cheng, Hua;Cai, Rong;Li, Lin Ling;Chang, Jie;Zhu, Jun;Zhang, Feng Xia;Chen, Liu Ji;Wang, Yan;Cheng, Shu Han;Cheng, Shui Yuan
    • Molecules and Cells
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    • v.26 no.6
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    • pp.536-547
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    • 2008
  • Anthocyanidin synthase (ANS, leucoanthocyanidin oxygenase), a 2-oxoglutarate iron-dependent oxygenase, catalyzed the penultimate step in the biosynthesis of the anthocyanin class of flavonoids, from the colorless leucoanthocyanidins to the colored anthocyanidins. The full-length cDNA and genomic DNA sequences of ANS gene (designated as GbANS) were isolated from Ginkgo biloba for the first time. The full-length cDNA of GbANS contained a 1062-bp open reading frame (ORF) encoding a 354-amino-acid protein. The genomic DNA analysis showed that GbANS gene had three exons and two introns. The deduced GbANS protein showed high identities to other plant ANSs. The conserved amino acids (H-X-D) ligating ferrous iron and residues (R-X-S) participating in 2-oxoglutarate binding were found in GbANS at the similar positions like other ANSs. Southern blot analysis indicated that GbANS belonged to a multi-gene family. The expression analysis by real-time PCR showed that GbANS expressed in a tissue-specific manner in G. biloba. GbANS was also found to be up-regulated by all of the six tested abiotic stresses, UV-B, abscisic acid, sucrose, salicylic acid, cold and ethylene, consistent with the promoter region analysis of GbANS. The recombinant protein was successfully expressed in E. coli strain with pET-28a vector. The in vitro enzyme activity assay by HPLC indicated that recombinant GbANS protein could catalyze the formation the cyanidin from leucocyanidin and conversion of dihydroquercetin to quercetin, suggesting GbANS is a bifunctional enzyme within the anthocyanidin and flavonol biosynthetic pathway.