• Asian Pacific Journal of Cancer Prevention
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• 제16권17호
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• pp.7575-7582
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• 2015

Cyclin E, a key coordinator of the G1 to S transition in the cell cycle, may be deregulated in several malignancies, including breast cancer. The most significant aberration in cyclin E is its elastase mediated proteolytic cleavage into tumor specific low molecular weight isoforms (LMW-Es). LMW-Es are biochemically hyperactive and biologically drive tumorigenesis in transgenic mouse models. Additionally, expression of LMW-Es has been correlated with poor survival in breast cancer cases. Here we determine whether expression of LMW-Es in a breast cancer cell line that is naturally devoid of these deregulated forms would alter their progression through each phase of the cell cycle. The results revealed that LMW-Es expression resulted in an increased doubling time, concomitant with a predominant increase in the population in the S phase of the cell cycle. Moreover, downregulation of p53 in LMW-Es cells resulted in additional shortening of the doubling time and enrichment of cells in the S and G2/M phases of the cell cycle. Furthermore, expression of LMW-Es sensitized cells to ${\beta}$-estradiol (E2) mediated growth and changed expression patterns of estrogen receptor and Bcl-2. Intriguingly, expression of LMW-Es could surpass anti-apoptotic effects raised by p53 upregulation. Taken together these studies suggest that overexpression of LMW-Es in collaboration with p53 loss results in altered g rowth properties of MCF-7 cells, enhancing the oncogenic activity of these ER positive breast cancer cells.

• 한국발생생물학회지:발생과생식
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• 제23권4호
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• pp.333-344
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• 2019

In vertebrate reproductive system, estrogen receptor (ER) plays a pivotal role in mediation of estrogenic signaling pathways. In the present study, we report the cDNA cloning, expression analysis, and transcriptional activity of ERβ1 subtype from medaka Oryzias dancena. The deduced O. dancena ERβ1 (odERβ1; 519 amino acids) contained six characteristic A/B to E/F domains with very short activation function 2 region (called AF2). A phylogenetic analysis indicated that odERβ1 was highly conserved among teleost ERβ1 subgroup. A conventional RT-PCR revealed that the odERβ1 transcripts were widely distributed in the multiple tissues, the ovary, brain, gill, intestine, kidney, and muscle. Further, the relatively higher odERβ1 expressions in the ovary and brain were clearly reproduced in RT-qPCR assay. When HA-fused odERβ1 expression vector was transfected into HEK293 cells, an immunoreactivity for odERβ1 was mainly detected in the nucleus part. Finally, an estrogen responsive element driven luciferase reporter assays demonstrated that the transcriptional activity of odERβ1 significantly increased by estradiol-17β (E2) in a dose dependent manner (p<0.05). However, fold-activation of odERβ1 in the presence of E2 was markedly weak, when it compared with those of O. latipes ERβ1. Taken together, these data suggest that odERβ1 represents a functional variant of teleost ERβ subtype and provides a basic tool allowing future studies examining the function of F domain of ERβ1 subtype and expanding our knowledge of ERβ evolution.

• Molecules and Cells
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• 제39권2호
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• pp.119-128
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• 2016

Angelica polymorpha Maxim root extract (APRE) is a popular herbal medicine used for treating stomachache, abdominal pain, stomach ulcers, and rheumatism; however the effect of APRE on cancer cells has not yet been explored. Here, we examined APRE cytotoxicity seen on target neuroblastoma cells (NB) using cell viability assays, DAPI visualization of fragmented DNA, and Western blotting analysis of candidate signaling pathways involved in proliferation and apoptosis. We demonstrated that APRE reduced cell viability in NB to a greater extent than in fibroblast cells. In addition, we found that APRE could inhibit the three classes of MAPK proteins and could also down-regulate the PI3K/AKT/GSK-$3{\beta}$ activity all being relevant for proliferation and survival. APRE could also up-regulate Bax expression and down-regulate Bcl-2 and Mcl-1. With APRE treatment, depolarization of mitochondria membrane potential and activation of caspase-3 was demonstrated in the SH-SY5Y cells. We could not found increased activity of death receptor and caspase-8 as markers of the extrinsic apoptosis pathway for the APRE treated cells. In presence of a caspase-3 siRNA and a pan-caspase inhibitor, APRE could not reduce the viability of NB cells to a significant degree. So we predicted that with APRE, the intrinsic pathway was solely responsible for inducing apoptosis as we also showed that the non-caspase autophagy pathway or ER stress-ROS mediated pathways were not involved. These findings demonstrate that an intrinsic mitochondria-mediated apoptosis pathway mediates the apoptotic effects of APRE on SH-SY5Y cells, and that APRE shows promise as a novel agent for neuroblastoma therapy.

• 의료법학
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• 제12권1호
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• pp.11-39
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• 2011

[ ${\S}27$ ]Abs. 3 das Gesuntheitsdienstgesetz (the Medical Service Act) in Korea lautet: Niemand in der Absicht, sich oder einem Dritten einen $Verm{\ddot{o}}gensvorteil$ zu verschaffen, der Medizininstitut bzw. dem Mediziner (die Medizinerin) den Patienten vorstellen, ${\ddot{u}}bweweisen$, verleiten oder einen anderen zu dieser Handlung anstiften darf, wie z.B. die Selbstbeteiligung des Patienten nach dem Krankenkassengesetz (the National Health Insurance Act) oder dem Gesetz ${\ddot{u}}ber$ Beistand der ${\ddot{a}}rztlicher$ Betreuung (the Medical Care Assistance Act) skontieren oder befreien, Geld offerieren oder dem Allgemeinheit das Verkehrswesen anbieten usw. Nach dem Wortlaut ist jedoch unklar, ob unter diese Vorschriften der Fall subsumiert werden kann, wenn eine Medizininstitut bzw. ein(e) Mediziner(in) in der Absicht, sich einen $Verm{\ddot{o}}gensvorteil$ zu verschaffen, sich den Patienten verleitet. Nach dem Korean Supreme Court ist eine Medizininstitut bzw. ein(e) Mediziner(in) nur dann das Subjekt der Verleitungshandlung, wenn sie bzw. er ein Mittel gegen fairen oder $ordungsm{\ddot{a}}{\beta}ien$ Medizinmarkt verwendet oder dem Patienten eine ${\ddot{a}}rztlich$ rechtswidrige Behandlung (z.B. einen rechtswidrigen Schwangerschaftsabbruch) verspricht. In diesem Beitrag wird dagegen die Auffassung mittels der teleologischen Reduktion vertritt und argumentiert, dass ein ${\ddot{a}}rztlich$ rechtswidriges Behandlung nach dem Rechtsgut und dem Normzweck unter ${\S}27$ Abs. 3 das Gesuntheitsdienstgesetz nicht subsumiert werden, sondern allein nach eigenem Unrecht bestraft werden kann.

• Reproductive and Developmental Biology
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• 제33권4호
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• pp.243-248
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• 2009

To examine the differential expression of proteins during the cycling (70~80% confluences) and G0/G1 (full confluences) phases in porcine fetal fibroblast cells, we used a global proteomics approach by 2-D gel electrophoresis (2-DE) and MALDI-TOF-MS. Cycling cell were harvested at approximately 70% to 80% confluent state while cells in G0/G1 phase were recovered after maintenance of a confluent state for 48 hr. Cellular proteins with isoelectric points ranging between 3.0~10.0, were analyzed by 2-DE with 2 replicates of each sample. A total of approximately 700 spots were detected by 2.D gels stained with Coomassie brilliant blue. On comparing the cell samples obtained from the cycling and G0/G1 phases, a total of 13 spots were identified as differentially expressed proteins, of which 8 spots were up-regulated in the cycling cell and 5 were up-regulated in the G0/G1 phase. Differentially expressed proteins included K3 keratin, similar to serine protease 23 precursor, protein disulfide-isomerase A3, microsomal protease ER-60, alpha-actinin-2, and heat-shock protein 90 beta. The identified proteins were grouped on the basis of their basic functions such as molecular binding, catabolic, cell growth, and transcription regulatory proteins. Our results show expression profiles of key proteins in porcine fetal fibroblast cells during different cell cycle status.

• Animal cells and systems
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• 제7권3호
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• pp.227-235
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• 2003

Bisphenol A (BPA), nonylphenol (NP), and 4-tert-octylphenol (OP) are known endocrine disrupting chemicals (EDCs) with estrogenic activity in fish. This study compared the effects of BPA, NP and OP on in vitro vitellogenin (VTG) synthesis in primary cultures of hepatocytes of the Chinese minnow Phoxinus oxycephalus. The VTG secreted into the culture medium was measured using enzyme-linked immunosorbent assay (ELISA), which we developed in this study using an antibody prepared from homogenates of Chinese minnow egg. VTG synthesis was induced by estradiol-17$\beta$ ($E_2$) and phenols (BPA, NP and OP) treatment. $E_2$ at concentrations of 10$^{-6}$ M or higher increased VTG levels significantly (P < 0.05). Exposure to 10^5\;M\;BPA\;or\;10^-4$M NP and OPinduced in vitro VTG synthesis (P < 0.01). However, $10^-3$ M BPA, NP or OP did not induce VTG synthesis. These results suggest that SPA has the highest estrogenic potential in Chinese minnow hepatocytes. Tamoxifen, an anti-estrogen, drastically blocked the production of VTG by phenols (BPA, NP and OP) suggesting that phenols (BPA, NP and OP) may act via binding to estrogen receptor (ER) in Chinese minnow hepatocytes.

• Clinical and Experimental Reproductive Medicine
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• 제37권2호
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• pp.89-98
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• 2010

성호르몬은 lacrimal system, corneal anatomy and disease, aqueous humor dynamics and glaucoma, crystalline lens and cataract, retinal disease를 포함한 안과영역의 다양한 부위에 영향을 끼칠 수 있다. Androgen은 전신에 걸친 sebaceous glands의 발달, 분화, 지질생성 등에 관여하므로 androgen deficiency는 meibomian gland의 기능장애와 건성안에서 증발을 더욱더 심화시킬 수 있는 것으로 알려져 있다. 반면에 estrogens은 sebaceous glands의 크기, activity, 지질생성을 감소시켜 meibomian gland에서는 androgen과 반대되는 작용을 할 가능성이 있다. 성호르몬은 또한 각막의 구조와 질병경과에도 영향을 줄 수 있는데, 각막이 월경주기 제2일째 및 배란주위기에 두꺼워지며, 폐경 여성에서 여성호르몬 치료 시 각막두께가 증가한다는 사실은 estrogen levels과 각막 두께와의 연관을 시사하는 소견이라고 하겠다. Fuch's dystrophy 역시 폐경 후 여성에서 더 흔히 관찰되어, 나이에 따른 호르몬 변화가 그 원인 중 하나로 지목되고 있으며, vernal keratoconjunctivitis 환자의 각막조직에서는 estrogen과 progesterone 수용체의 overexpresson이 발견되어 역시 이들 질환에서의 성호르몬의 역할을 시사한다고 할 수 있겠다. 성호르몬은 안압에도 영향을 미칠 수 있는데, intraocular pressure와 혈중 luteinizing hormone, follicular stimulating hormone (FSH), estrone, estradiol, progesterone, testosterone과는 연관이 없는 것으로 보고한 결과도 있으나, 혈중 progesterone의 경우 특히 임신중인 여성에서 안압과의 연관관계가 보고되었으며, 또한 폐경 여성에서 혈중 남성호르몬 농도가 높을수록 안압이 상승하는 경향이 있음이 보고되었다. 한편 'Female sex'는 모든 종류의 백내장의 위험요소인 것으로 알려져 있다. 백내장으로 진단받은 여성의 경우 호르몬 측정 시 연령등을 보정하고도 폐경기에 가까운 호르몬 수치를 보였으며, 이들 백내장 환자군내에서는 estradiol과 FSH 농도 사이에 유의한 음의 상관관계가 보고되었다. 또한 DHEAS의 경우 백내장 위험도를 감소시키는 것으로 보고되었고, 그 기전으로 전구호르몬인 DHEA의 역할을 제시되고 있는데. 즉 백내장의 주요 기전 중 하나인 oxidative stress를 감소시키는 antioxidant activity를 DHEA가 보이기 때문일 것으로 추정되고 있다. 성호르몬은 망막 내 혈관의 흐름에도 영향을 줄 수 있는 것으로 알려져 있는데, 안구 내 혈류에서 여성호르몬의 긍정적인 효과와 남성호르몬의 부정적인 효과가 보고되었고, age-related macular degeneration (AMD)의 경우, estradiol이 ER-$\beta$ 수용체를 통해 보호효과가 있음이 보고되어, ER-$\beta$ 수용체가 향후 AMD 치료에서 치료적 target이 될 수도 있을 것으로 알려지고 있다. 또한 DHEAS의 경우 AMD 위험도를 증가시킬 가능성이 있으며, 당뇨로 진단받은 환자에서는 혈중 testosterone 농도는 당뇨병성 망막병증의 진행과 연관이 있는 것으로 보고되었다. 성호르몬은 안과영역의 다양한 질환과의 연관관계는 아직 추가연구가 필요한 실정이나 다양한 성호르몬이 안질환의 발생 및 경과에 영향을 줄 가능성이 있으며, 이와 같은 연관관계가 규명된다면 추후 호르몬을 통해 질환의 경과 및 예방의 가능성이 제시될 수도 있을 것으로 사료된다. 특히 폐경이후 다양한 안질환이 증가한다는 기존의 연구결과들을 고려 시, 안과영역에 있어 폐경기 여성호르몬 치료의 긍정적인 역할에 관해서는 지속적인 관심이 요구된다고 하겠다.

• 한국미생물생명공학회:학술대회논문집
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• 한국미생물생명공학회 2002년도 학술발표대회
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• pp.18-25
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• 2002

The pikromycin biosynthetic system in Streptomyces venezuleae is unique for its ability to produce two groups of antibiotics that include the 12-membered ring macrolides methymycin and neomethymycin, and the 14-membered ring macrolides narbomycin and pikromycin. The metabolic pathway also contains two post polyketide-modification enzymes, a glycosyltransferase and P450 hydroxylase that have unusually broad substrate specificities. In order to explore further the substrate flexibility of these enzymes a series of hybrid polyketide synthases were constructed and their metabolic products characterized. The plasmid-based replacement of the multifunctional protein subunits of the pikromycin PKS in S. venezuelae by the corresponding subunits from heterologous modular PKSs resulted in recombinant strains that produce both 12- and 14-membered ring macrolactones with predicted structural alterations. In all cases, novel macrolactones were produced and further modified by the DesVII glycosyltransferase and PikC hydroxylase leading to biologically active macrolide structures. These results demonstrate that hybrid PKSs in S. venezuelae can produce a multiplicity of new macrolactones that are modified further by the highly flexible DesVII glycosyltransferase and PikC hydroxylase tailoring enzymes. This work demonstrates the unique capacity of the S. venezuelae pikromycin pathway to expand the toolbox of combinatorial biosynthesis and to accelerate the creation of novel biologically active natural products. The polyketide backbone of rifamycin B is assembled through successive condensation and ${\beta}$-carbonyl processing of the extender units by the modular rifamycin PKS. The eighth module, in the RifD protein, contains nonfunctional DH domain and functional KR domain, which specify the reduction of the ${\beta}$-carbonyl group resulting in the C-21 bydroxyl of rifamycin B. A four amino acid substitution and one amino acid deletion were introduced in the putative NADPH binding motif in the proposed KR domain encoded by rifD. This strategy of mutation was based on the amino acid sequences of the corresponding motif of the KR domain of module 3 in the RifA protein, which is believed dysfunctional, so as to introduce a minimum alteration and retain the reading frame intact, yet ensure loss of function. The resulting strain produces linear polyketides, from tetraketide to octaketide, which are also produced by a rifD disrupted mutant as a consequence of premature termination of polyketide assembly. Much of the structural diversity within the polyketide superfamily of natural products is due to the ability of PKSs to vary the reduction level of every other alternate carbon atom in the backbone. Thus, the ability to introduce heterologous reductive segments such as ketoreductase (KR), dehydratase (DH), and enoylreductase (ER) into modules that naturally lack these activities would increase the power of the combinatorial biosynthetic toolbox. The dehydratase domain of module 7 of the rifamycin PKS, which is predicted to be nonfunctional in view of the sequence of the apparent active site, was replaced with its functional homolog from module 7 of rapamycin-producing polyketide synthase. The resulting mutant strain behaved like a rifC disrupted mutant, i.e., it accumulated the heptaketide intermediate and its precursors. This result points out a major difficulty we have encountered with all the Amycolatopsis mediterranei strain containing hybrid polyketide synthases: all the engineered strains prepared so far accumulate a plethora of products derived from the polyketide chain assembly intermediates as major products instead of just analogs of rifamycin B or its ansamycin precursors.

• Journal of Plant Biotechnology
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• 제1권1호
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• pp.20-30
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• 1999

In order to modify lipid production of Perilla qualitatively as well as quantitatively by genetic engineering, genes involved in carbon metabolism were isolated and characterized. These include acyl-ACP thioesterases from Perilla frutescens and Iris sp., four different $\beta$-ketoacyl- ACP synthases from Perilla frutescens, and two $\Delta$15 a-cyl-ACP desaturases(Pffad7, pffad3). Δ15 acyl-ACP desa turase (Δ15-DES) is responsible for the conversion of linoleic acid (18:2) to $\alpha$-linolenic acid (ALA, 18:3). pffad 3 encodes Δ15 acyl-desaturase which is localized in ER membrane. On the other hand, Pffad7 encodes a 50 kD plastid protein (438 residues), which showed highest sequence similarity to Sesamum indicum fad7 protein. Northern blot analysis revealed that the Pffad7 is highly expressed in leaves but not in roots and seeds. And Pffad3 is expressed throughout the seed developmental stage except very early and fully mature stage. We constructed Pffad7 gene under 355 promoter and Pffad3 gene under seed specific vicillin promoter. Using Pffad7 construct, Perilla, an oil seed crop in Korea, was transformed by Agrobacterium leaf disc method. $\alpha$-linolenic acid contents increased in leaves but decreased in seeds of transgenic Perilla. Currently, we are transforming Perilla with Pffad3 construct to change Perilla seed oil composition. We isolated three ADP-glucose pyrophosphorylase (AGP) genes from Perilla immature seed specific cDNA library. Nucleotide sequence analysis showed that two of three AGP (Psagpl, Psagp2) genes encode AGP small subunit polypeptides and the remaining (Plagp) encodes an AGP large subunit. PSAGPs, AGP small subunit peptide, form active heterotetramers with potato AGP large subunit in E. coli expressing plant AGP genes.

• 논리연구
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• 제7권2호
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• pp.15-46
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• 2004

표면은 서로 접촉하고 있는 두 개의 사물의 공통의 경계여야 한다; 그러므로 물의 표면은 물의 부분도 아니고 공기의 부분도 아니며, 또 이들 사이에 끼워진 그 어떤 물체도 아니다. 그렇다면 공기와 물을 갈라놓은 것은 무엇인가? 공기도 물도 아니며 실체가 없는 공동의 경계가 있어야만 한다는 것은 필연적이다. 왜냐하면 양자 사이에 끼워진 물체는 이들의 접촉을 막을 것이고, 물과 공기의 경우 이런 일은 일어나지 알고 있으며 그 어떤 매개체의 삽입 없이 접촉하고 있기 때문이다.