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Cell Surface Interaction with Expression of Fas Ligand Mediates Prolactin-Induced Apoptosis In Rat Luteal Cell Culture (Rat 황체세포 배양에 있어서 Prolactin에 의한 황체퇴행 및 Fas Ligand의 발현)

  • 장규태;박미령;선동수;윤창현
    • Journal of Embryo Transfer
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    • v.13 no.2
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    • pp.179-190
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    • 1998
  • Prolactin (PRL) surge in cycling rats at proestrous afternoon has previously been reported as an inducer of apoptotic cell death of luteal cells. This death-inducing action of PRL seeins unusual, because PRL can he categorized as a cell-survival factor, if other known physiological functions of PRL are taken into account. In this study, the apoptotic action of PRL was assessed in cultured cells prepared from rat luteal tissue and underlying molecular /cellular mechanism of PRL-induced luteolysis was analyzed. The latest crop of corpora lutea (CLs) were enucleated from rat ovaries at 18:00 h on the proestrous day before the next ovulation. Donor rats were pretreated with CB154, a dopamine agonist, in order to he exempted from the endogenous PRL surge. The harvested GLs were dispersed and cultured with or without PRL (2$\mu$g /ml) for 24 or 48 h. An addition of PRL to the culture medium changed the parameters indicative of cell death via apoptosis: a decrease in cell viability (MTT) and an increase in chromatin condensation. Most of the DNA breakdown in nuclei induced by PRL occurred in steroidogenic cells which were identified by 3$\beta$-HSD activity staining, and the number of 3$\beta$-HSD-positivecells were significantly decreased. Interestingly, most of the cells with an apoptotic nucleus adhered to one or more intact and seemingly non-steroidogenic cells. Because the expression of Fas has heen shown to be abundant in murine ovary, and Fas is known to have an exact physiological role in occurrence of apoptotic cell death, the membrane form-Fas ligand (rnFasL) was quantified in the cell lysate. An addition of PRL increased expression of mFasL. Moreover, an addition of concanavalin A (ConA), a T-cell specific activator, in place of PRL, enhanced the apoptotic parameters. Cumulatively, the apoptotic PRL action was addressed to cells unknown than steroidogenic lute~ cells. The most prohable candidate for the direct target cells is Tcells in the luteal tissue that can express mFasL in response to PRL.

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Studies on the NO Production and Expression Induction Effect of NOS Gene by Salviae Radix (단삼에 의한 NO 생성 및 NOS 유전자의 발현 유도 효과에 관한 연구)

  • 조현주;원진희;문구;문석재;유기원;유봉하
    • The Journal of Korean Medicine
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    • v.21 no.3
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    • pp.20-30
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    • 2000
  • Objective : This experiment was performed in order to study the effect of an aqueous extract of Salviae radix root(SRRAE) on NO production and NOS gene induction from macrophages Methods : To investigate dose-dependent effects of SRRAE for NO release on the $rIFN-{\gamma}-treated$ macrophages, the cells were incubated for 6 hrs in a medium containing $rIFN-{\gamma}$ (5 U/ml), stimulated with SRRAE and incubated in a CO2 incubator. The cells were treated with 5 U/ml $rIFN-{\gamma}$ plus 100 g/ml of SRRAE, Then, the cells were incubated with various concentrations of NGMMA at $37^{\circ}C$ for 48 hrs, Results : SRRAE had no effect on NO production by itself, whereas recombinant $interferon-{\gamma}(rIFN-{\gamma})$ alone showed modest activity, When SRRAE was used in combination with $rIFN-{\gamma}$, there was a marked cooperative induction of NO production in a dose-dependent manner. The optimal effect of SRRAE on NO production was shown at 6hrs after treatment with $rIFN-{\gamma}$. The SRRAE-induced production of NO was inhibited by NG-monomethyl- L-arginine(NGMMA) and arginase. $rIFN-{\gamma}$ in combination with SRRAE showed a marked increase of the expression of the inducible NOS(iNOS) gene. In addition, the effect of SRRAE was mainly dependent on the SRRAE-induced tumor necrosis $factor-{\alpha}(TNF-{\alpha})$ secretion. Conclusions : SRRAE induces NO production from macrophages as a result of SRRAE-induced $TNF-{\alpha}$ secretion. SRRAE may provide a second signal for synergistic induction of NO production in macrophages already induced to express iNOS gene by $rIFN-{\gamma}$.

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Recombinant Glargine Insulin Production Process Using Escherichia coli

  • Hwang, Hae-Gwang;Kim, Kwang-Jin;Lee, Se-Hoon;Kim, Chang-Kyu;Min, Cheol-Ki;Yun, Jung-Mi;Lee, Su Ui;Son, Young-Jin
    • Journal of Microbiology and Biotechnology
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    • v.26 no.10
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    • pp.1781-1789
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    • 2016
  • Glargine insulin is a long-acting insulin analog that helps blood glucose maintenance in patients with diabetes. We constructed the pPT-GI vector to express prepeptide glargine insulin when transformed into Escherichia coli JM109. The transformed E. coli cells were cultured by fed-batch fermentation. The final dry cell mass was 18 g/l. The prepeptide glargine insulin was 38.52% of the total protein. It was expressed as an inclusion body and then refolded to recover the biological activity. To convert the prepeptide into glargine insulin, citraconylation and trypsin cleavage were performed. Using citraconylation, the yield of enzymatic conversion for glargine insulin increased by 3.2-fold compared with that without citraconylation. After the enzyme reaction, active glargine insulin was purified by two types of chromatography (ion-exchange chromatography and reverse-phase chromatography). We obtained recombinant human glargine insulin at 98.11% purity and verified that it is equal to the standard of human glargine insulin, based on High-performance liquid chromatography analysis and Matrix-assisted laser desorption/ionization Time-of-Flight Mass Spectrometry. We thus established a production process for high-purity recombinant human glargine insulin and a method to block Arg (B31)-insulin formation. This established process for recombinant human glargine insulin may be a model process for the production of other human insulin analogs.

Enhanced Coupling of $M_1$ Muscarinic Receptors to Activation of Phospholipase C upon Mutation of a Transposed Amino Acid Triplet Repeat

  • Lee, Seok-Yong;Sung, Ki-Wug;Kim, Ok-Nyu;Lee, Sang-Bok
    • The Korean Journal of Physiology and Pharmacology
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    • v.1 no.1
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    • pp.19-25
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    • 1997
  • The C-terminus ends of the second putative transmembrane domains of both $M_1$ and $M_2$ muscarinic receptors contain a triplet of amino acid residues consisting of leucine (L), tyrosine (Y) and threonine (T). This triplet is repeated as LYT-TYL in $M_1$ receptors at the interface between the second transmembrane domain and the first extracellular loop. Interestingly, however, it is repeated in a transposedfashion (LYT-LYT) in the sequence of $M_2$ receptors. In our previous work, we investigated the possible significance of this unique sequence diversity for determining the distinct differential receptor function at the two receptor subtypes. However, we found mutation of the LYTTYL sequence of $M_1$ receptors to the corresponding $M_2$ receptor LYTLYT sequence demonstrated markedly enhanced the stimulation of phosphoinositide (PI) hydrolysis by carbachol without a change in its coupling to increased cyclic AMP formation. In this work, thus, the enhanced stimulation of PI hydrolysis in the LYTLYT $M_1$ receptor mutant was further investigated. The stimulation of PI hydrolysis by carbachol was enhanced in the mutant $M_1$ receptor, and this change was not due to alterations in the rate of receptor desensitization or sequestration. The observed larger response to carbachol at mutant $M_1$ receptors was also not due to an artifact resulting from selection of CHO cells which express higher levels of G-proteins or phospholipase C. Our data suggest that although the LYTTYL sequence in $M_1$ muscarinic receptors is not involved in determining receptor pharmacology, mutation of the sequence enhanced the coupling of $M_1$ receptors to the stimulation of phospholipase C.

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Studies on the Ecological Behaviour of Forage Plants in Korea (우리나라 자생사초(自生飼草)의 생태적(生態的) 반응(反應)에 관한 연구(硏究))

  • Park, G.J.;Yoon, S.H.;Lee, J.K.
    • Journal of The Korean Society of Grassland and Forage Science
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    • v.19 no.2
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    • pp.95-104
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    • 1999
  • For the using as the guideline to evaluate the grassland based on the botanical composition, the ecological behaviour which was expressed of nine(1~9:reaction and nitrogen figure) or twelve(1~12:moisture figure) division scale was given to the each plant species in grassland. At first, the ecological behaviours(moisture, reaction and nitrogen figure) were given to the 141 main forage plants of grassland in Korea. In the improved grassland, moisture, reaction and nitrogen figure of the ecological behaviour were much more increased 0.73, 0.94 and 0.95 than those of the waste type grassland, respectively. Therefore, the ecological behaviour of grassland was not only possible to express the vegetational conditions of various plant communities in some simple figures, but also compared with different types of grassland at the same time.

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Nitric Oxide Donor, NOR-3, Increased Expression of Cyclooxygenase-2, but not of Cyclooxygenase-1 in Cultured VSMC

  • Lee, Dong-Hyup;Park, Ji-Eun;Kang, Young-Jin;Lee, Kwang-Youn;Choi, Hyoung-Chul
    • The Korean Journal of Physiology and Pharmacology
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    • v.10 no.3
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    • pp.161-165
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    • 2006
  • NO and cyclooxygenase-2 (COX-2) are contributes to vascular inflammation induced by various stimulation. The mechanism, which explains a linkage between NO and COX-2, could be of importance in promoting pathophysiological conditions of vessel. We investigated the effects of NO donors on the COX-l and COX-2 mRNA/protein expression, as well as the nitrite production in culture medium of vascular smooth muscle cell (VSMC). VSMC was primarily cultured from thoracic aorta of rat. In this experiments, COX-l and COX-2 mRNA/protein expressions were analysed and nitrite productions were investigated using Griess reagent. VSMC did not express COX-2 protein in basal condition (Nonlipopolysaccharide (LPS) stimulated). In LPS-stimulated experiments, after 3 hours of NO donor pretreatment, LPS $10{\mu}g/ml$ was treated for 24 hours. COX-l protein expressions were unchanged by SNP and NOR-3. NOR-3 significantly increased COX-2 mRNA/protein expression under LPS stimulation. In contrast, SNP did not increase COX-2 mRNA/protein expression under LPS stimulation. Nitrite production was higher in NOR-3 treatment than SNP treatment under LPS stimulation. These results suggest that the expression of COX-2 in VSMC is regulated by NOR-3, COX-2 expressions were depending on the types of NO donor and LPS stimulation in VSMC.

A Kinetic Study on the Ammonia Nitrogen Adsorption by Physical Characteristics of Activated Carbon (활성탄 물성에 따른 암모니아성 질소 흡착의 동력학적 연구)

  • Seo, Jeong-beom;Kang, Joon-won;Lee, Ik-soo
    • Journal of Korean Society on Water Environment
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    • v.24 no.3
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    • pp.311-316
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    • 2008
  • This study aimed to obtain equilibrium concentration on adsorption removal of ammonia nitrogen by activated carbon, to express the adsorption characteristics following Freundlich isotherm and also, based on the value obtained, to investigate the relationship between physical characteristics of activated carbon and dynamics of ammonia nitrogen removal by obtaining rate constant and effective pore diffusivity. The results summarized from this study are as follows. It was noted that powdered activated carbon showed better adsorption ability than granular activated carbon. The value of constant (f) of Freundlich isotherm of powered activated carbon was $4.6{\times}10^{-8}$ which is bigger than that of granular activated carbon. The adsorption rate constant on ammonia nitrogen of powered activated carbon with high porosity and low effective diameter was highest as 0.416 hr-1 and the effective pore diffusivity ($D_e$) was lowest as $1.17{\times}10^{-6}cm^2/hr$, and the value of ammonia nitrogen adsorption rate constant of granular activated carbon was $0.149{\sim}0.195hr^{-1}$. It was revealed that, with the same amount of dosage, the adsorptive power of activated carbon with lower effective diameter and bigger porosity was better and its rate constant was also high. With a little adsorbent dosage of 2 g, there was no difference removal ability of ammonia nitrogen as change of adsorption properties.

Ultrasonic-assisted Micellar Extraction and Cloud-point Pre-concentration of Major Saikosaponins in Radix Bupleuri using High Performance Liquid Chromatography with Evaporative Light Scattering Detection

  • Suh, Joon-Hyuk;Yang, Dong-Hyug;Han, Sang-Beom
    • Bulletin of the Korean Chemical Society
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    • v.32 no.8
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    • pp.2637-2642
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    • 2011
  • A new ultrasonic-assisted micellar extraction and cloud-point pre-concentration method was developed for the determination of major saikosaponins, namely saikosaponins -A, -C and -D, in Radix Bupleuri by high performance liquid chromatography with evaporative light scattering detection (HPLC-ELSD). The non-ionic surfactant Genapol X-080 (oligoethylene glycol monoalkyl ether) was chosen as the extraction additive and parameters affecting the extraction efficiency were optimized. The highest yield was obtained with 10% (w/v) Genapol X-080, a liquid/solid ratio of 200:1 (mL/g) and ultrasonic-assisted extraction for 40 min. In addition, the optimum cloud-point pre-concentration was reached with 10% sodium sulfate and equilibration at $60^{\circ}C$ for 30 min. Separation was achieved on an Ascentis Express C18 column (100 ${\times}$ 4.6 mm i.d., 2.7 ${\mu}M$) using a binary mobile phase composed of 0.1% acetic acid and acetonitrile. Saikosaponins were detected by ELSD, which was operated at a $50^{\circ}C$ drift tube temperature and 3.0 bar nebulizer gas ($N_2$) pressure. The water-based solvent modified with Genapol X-080 showed better extraction efficiency compared to that of the conventional solvent methanol. Recovery of saikosaponins ranged from 93.1 to 101.9%. An environmentally-friendly extraction method was successfully applied to extract and enrich major saikosaponins in Radix Bupleuri.

A Study on the Making of the Noise Map for Traffic Noise Level (도로교통 소음지도 작성에 관한 연구)

  • Park, Sang-Ill;Choi, Hyung-Il;Cheong, Kyung-Hoon;Yeom, Dong-Ick;Jin, Chang-Beom
    • Journal of Environmental Science International
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    • v.16 no.12
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    • pp.1393-1399
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    • 2007
  • This research helps you understand the road traffic noise levels by using a noise map. We have observed the change of the road traffic noise levels around $07:00{\sim}08:30\;and\;22:00{\sim}23:00$ using the noise map in the city. The road traffic noise level is very high both at noon and at night around a beltway and an interchange that is linked with a highway. It seems that the main route of so many vehicles, which are at neighboring cities such as N city and D and H districts and which avoid traffic jams in the city, is the beltway and interchange. The road traffic noise level of a nearby express bus terminal, railroad station, and airport is more than 75 dB at noon and 65 dB at night. The road traffic noise level of G city at night is observed to be more than 55 dB. The noise levels of a residence area and a university are higher than a road with high noise levels when the commuters drive to work. The end of the day exceeds 11 o'clock because of a culture level of development that arouses spare time, eating out, adults' drinking culture, nightlife of the youth, etc. Therefore, the road traffic noise level is high during late night hours, and it exceeds regulatory guidelines(55 dB(A)). It also damages the residence area that is located near the road.

Total Dynamic Analysis of Deep-Seabed Integrated Mining System (심해저 광물자원 채광시스템의 통합거동 해석)

  • Kim, Hyung-Woo;Hong, Sup;Choi, Jong-Su;Yeu, Tae-Kyeong
    • Proceedings of the Korea Committee for Ocean Resources and Engineering Conference
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    • 2006.11a
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    • pp.311-314
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    • 2006
  • This paper concerns about total dynamic analysis of integrated mining system. This system consists of vertical steel pipe, intermediate buffer station, flexible pipe and self-propelled miner. The self-propelled miner and buffer are assumed as rigid-body of 6-dof. Discrete models of vertical steel pipe and flexible pipe are adopted, which are obtained by means of lumped-parameter method. The motion of mining vessel is not considered. Instead, the motion of mining vessel is taken into account in form of various boundary conditions (e.g. forced excitation in slow motion and/or fast oscillation and so on). A terramechanics model of extremely soft cohesive soil is applied to the self-propelled miner. The hydrodynamic forces and moments are included in the dynamic models of vehicle and lifting pipe system. Hinged and fixed constraints are used to define the connections between sub-systems (vertical steel pipe, buffer, flexible pipe, miner). Equations of motion of the coupled model are derived with respect to the each local coordinates system. Four Euler parameters are used to express the orientations of the sub-systems. To solve the equations of motion of the total dynamic model, an incremental-iterative formulation is employed. Newmark-b method is used for time-domain integration. The total dynamic responses of integrated mining system are investigated.

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