• Journal of Microbiology and Biotechnology
• /
• v.20 no.11
• /
• pp.1563-1570
• /
• 2010

To construct a highly sensitive detection system for endocrine disruptors (EDs), we have compared the activity of promoters with the n-alkane-inducible cytochrome P450 gene (ALK1), isocitrate lyase gene (ICL1), ribosomal protein S7 gene (RPS7), and the translation elongation factor-1${\alpha}$ gene (TEF1) for the heterologous gene in Yarrowia lipolytica. The promoters were introduced into the upstream of the lacZ or hERa reporter genes, respectively, and the activity was evaluated by ${\beta}$-galactosidase assay for lacZ and Western blot analysis for hER${\alpha}$. The expression analysis revealed that the ALK1 and ICL1 promoters were induced by n-decane and by EtOH, respectively. The constitutive promoter of RPS7 and TEF1 showed mostly a high level of expression in the presence of glucose and glycerol, respectively. In particular, the TEF1 promoter showed the highest ${\beta}$-galactosidase activity and a significant signal by Western blotting with the anti-estrogen receptor, compared with the other promoters. Moreover, the detection system was constructed with promoters linked to the upstream of the expression vector for the hER${\alpha}$ gene transformed into the Y. lipolytica with a chromosome-integrated lacZ reporter gene under the control of estrogen response elements (EREs). It was indicated that a combination of pTEF1p-hER${\alpha}$ and CXAU1-2XERE was the most effective system for the $E_2$-dependent induction of the ${\beta}$-galactosidase activity. This system showed the highest ${\beta}$-galactosidase activity at $10^{-6}\;M\;E_2$, and the activity could be detected at even the concentration of $10^{-10}\;M\;E_2$. As a result, we have constructed a strongly sensitive detection system with Y. lipolitica to evaluate recognized/suspected ED chemicals, such as natural/synthetic hormones, pesticides, and commercial chemicals. The results demonstrate the utility, sensitivity, and reproducibility of the system for identifying and characterizing environmental estrogens.

• Journal of Animal Science and Technology
• /
• v.44 no.6
• /
• pp.677-684
• /
• 2002

We have investigated the expression patterns of candidates for growth stage specifically expressed genes. The expression patterns of the EPV20, aldolase A, Translationally Controlled Tumor Protein (TCTP) and Adipocyte Differentiation Related Protein (ADRP) were examined by semiquantitative RT-PCR and northern blot analysis in skeletal muscle tissues of Hanwoo, especially in the longissimus dorsi at various growth stages. The EPV20 mRNA was expressed in longissimus dorsi tissue of Hanwoo, but there was no difference of expression levels during growth stages. Though the aldolase A gene was reported to be muscle-specific and regulated at developmental stages, the expression levels of aldolase A mRNA in the longissimus dorsi tissues showed little differences at various growth stages. The expression levels of TCTP which was reported as growth-related protein regulated at translation step were gradually increased during growth of Hanwoo. The expression levels of ADRP mRNA were rapidly increased at 24-month-old longissimus dorsi tissue of Hanwoo, and decreased at 30-month-old. Our data suggest that the ADRP gene show as growth-stage dependent expression and is related to fat deposition within muscular tissue.

• Journal of Microbiology and Biotechnology
• /
• v.14 no.1
• /
• pp.81-89
• /
• 2004

Tumor necrosis factor-$\alpha$ (TNF-$\alpha$) and lymphotoxin-$\alpha$ (LT-$\alpha$, TNF-$\beta$) can initiate and perpetuate human diseases such as multiple sclerosis (MS), rheumatoid arthritis (RA), and insulin-dependent diabetes mellitus (IDDM). TNFs can be blocked by the use of soluble TNF receptors. However, since monomeric soluble receptors generally exhibit low affinity or function as agonists, the use of monomeric soluble receptors has been limited in the case of cytokines such as TNF-$\alpha$, TNF-$\alpha$, interleukin (IL)-1, IL-4, IL-6, and IL-13, which have adapted to a multi component receptor system. For these reasons, very high-affinity inhibitors were created for the purpose of a TNFs antagonist to bind the TNFR and trigger cellular signal by using the multistep polymerase chain reaction method. First, recombinant simple TNFR-Ig fusion proteins were constructed from the cDNA sequences encoding the extracellular domain of the human p55 TNFR (CD120a) and the human p75 TNFR (CD120b), which were linked to hinge and constant regions of human $IgG_1$ heavy chain, respectively using complementary primers (CP) encoding the complementary sequences. Then, concatameric TNFR-Ig fusion proteins were constructed using recombinant PCR and a complementary primer base of recombinant simple TNFR-Ig fusion proteins. For high level expression of recombinant fusion proteins, Chinese hamster ovary (CHO) cells were used with a retroviral expression system. The transfected cells produced the simple concatameric TNFR-Ig fusion proteins capable of binding TNF and inactivating it. These soluble versions of simple concantameric TNFR-Ig fusion proteins gave rise to multiple forms such as simple dimers and concatameric homodimers. Simple TNFR-1g fusion proteins were shown to have much more reduced TNF inhibitory activity than concatameric TNFR-Ig fusion proteins. Concatameric TNFR-Ig fusion proteins showed higher affinity than simple TNFR-Ig fusion proteins in a receptor inhibitor binding assay (RIBA). Additionally, concatameric TNFR-Ig fusion proteins were shown to have a progressive effect as a TNF inhibitor compared to the simple TNFR-Ig fusion proteins and conventional TNFR-Fc in cytotoxicity assays, and showed the same results for collagen induced arthritis (CIA) in mice in vivo.

• Journal of Korean Society of Transportation
• /
• v.25 no.4
• /
• pp.79-87
• /
• 2007

The interchanges of urban freeways have many problems with traffic operation due to high off-ramp flows and frequent congestion at adjacent intersections. The flow exiting from off-ramps is affected by the operational status and traffic volume conditions of the nearest signalized intersection. As a result, off-ramp flow cannot exit and the queue backs up the freeway mainline when queues from the signalized intersection form up to the junction of the off-ramp and street. The spacing between an off-ramp and an adjacent intersection is likely to determine the traffic conditions at the adjacent intersection. However, the current design guidelines do not consider such a factor. This study is to develop a model calculating the spacing between off-ramps and adjacent intersections considering the signal, traffic, and road conditions. The variables affecting the model in this study are effective green time (g/C), volume-capacity ratio (v/c), the number of lanes, and off-ramp volume. Various scenarios are designed to represent the effects of the variables and the road networks are constructed using VISSIM, which is a common traffic micro-simulation software package. The queue length is derived from VISSIM and this length is considered as the recommended spacing between the off-ramp and the adjacent intersection. Through the simulation analysis, regression models are developed to calculate the queue length reflecting the various conditions such as signals, traffic, and road configurations. The developed model can be used to create road design guidelines to determine the location of off-ramps in the planning stage.

• Biomolecules & Therapeutics
• /
• v.21 no.4
• /
• pp.299-306
• /
• 2013

In the present study, we investigated the effect of ethanolic extract of the seed of Zizyphus jujuba var. spinosa (EEZS) on cholinergic blockade-induced memory impairment in mice. Male ICR mice were treated with EEZS. The behavioral tests were conducted using the passive avoidance, the Y-maze, and the Morris water maze tasks. EEZS (100 or 200 mg/kg, p.o.) significantly ameliorated the scopolamine-induced cognitive impairment in our present behavioral tasks without changes of locomotor activity. The ameliorating effect of EEZS on scopolamine-induced memory impairment was significantly reversed by a sub-effective dose of MK-801 (0.0125 mg/kg, s.c.). In addition, single administration of EEZS in normal naive mouse enhanced latency time in the passive avoidance task. Western blot analysis was employed to confirm the mechanism of memory-ameliorating effect of EEZS. Administration of EEZS (200 mg/kg) increased the level of memory-related signaling molecules, including phosphorylation of extracellular signal-regulated kinase or cAMP response element-binding protein in the hippocampal region. Also, the time-dependent expression level of brain-derived neurotrophic factor by the administration of EEZS was markedly increased from 3 to 9 h. These results suggest that EEZS has memory-ameliorating effect on scopolamine-induced cognitive impairment, which is mediated by the enhancement of the cholinergic neurotransmitter system, in part, via NMDA receptor signaling, and that EEZS would be useful agent against cognitive dysfunction such as Alzheimer's disease.

• Nuclear Medicine and Molecular Imaging
• /
• v.41 no.5
• /
• pp.343-349
• /
• 2007

Radioiodide transport has been extensively and successfully used in the evaluation and management of thyroid disease. The molecular characterization of the sodium/iodide symporter (NIS) and cloning of the NIS gene has led to the recent expansion of the use of radioiodide to cancers of the breast and other nonthyroidal tissues exogenously transduced with the NIS gene. More recently, discoveries regarding the functional analysis and regulatory processes of the NIS molecule are opening up exciting opportunities for new research and applications for NIS and radio iodide. The success of NIS based cancer therapy is dependent on achievement of maximal radioiodide transport sufficient to allow delivery of effective radiation doses. This in turn relies on high transcription rates of the NIS gene. However, newer discoveries indicate that nontranscriptional processes that regulate NIS trafficking to cell membrane are also critical determinants of radioiodide uptake. In this review, molecular mechanisms that underlie regulation of NIS transcription and stimuli that augment membrane trafficking and functional activation of NIS molecules will be discussed. A better understanding of how the expression and cell surface targeting of NIS proteins is controlled will hopefully aid in optimizing NIS gene based cancer treatment as well as NIS based reporter-gene imaging strategies.

• Journal of The Korean Society of Inherited Metabolic disease
• /
• v.12 no.1
• /
• pp.49-53
• /
• 2012

X-linked adrenoleukodystrophy (ALD) is a rare inherited metabolic disease which results in impaired peroxisomal ${\beta}$-oxidation and the accumulation of very long chain fatty acids (VLCFA) in the adrenal cortex, the myelin of the central nervous system, and the testes. X-linked ALD is caused by mutations in the ABCD1 gene encoding an ATP-binding cassette transporter superfamily located in the peroxisomal membrane. This disease is characterized by a variety of phenotypes. The classic childhood cerebral ALD is a rapidly progressive demyelinating condition affecting the cerebral white matter before the age of 10 years in boys. We report the case of a 8-year-old with childhood cerebral X-linked ALD who developed inattention, hyperactivity, motor incoordination and hemiparesis. We diagnosed ALD with elevated plasma very long chain fatty acid level and diffuse high signal intensity lesions in both parieto-occipital white matter and cerebellar white matter in brain MRI. We identified a novel c.983delT (p.Met329CysfsX7) mutation of the ABCD1 gene. There is no correlation between X-ALD phenotype and mutations in the ABCD1 gene. Further studies for searching additional non-genetic factor which determine the phenotypic variation will be needed.

• Journal of Physiology & Pathology in Korean Medicine
• /
• v.24 no.6
• /
• pp.983-988
• /
• 2010

The present study aims to investigate a possible mechanism of electroacupuncture (EA) in the spinal dorsal horn that may underlie N-methyl-D-aspartate (NMDA) receptor-associated extracellular signal-regulated kinase (ERK) mitogen-activated protein kinase (MAPK) signaling pathways. The hot plate latency of the ipsilateral hindpaw of EA-treated rats was significantly decreased compared with complete Freund's adjuvant (CFA)-injected ones. The expressions of NR1 and NR2B subuint mRNA of NMDA receptor in the whole L4-5 segments are decreased by CFA treatment, but NR2B subunit was significantly recovered by EA treatment. When we detected the expression of ERK, there were no significant difference between normal and CFA-treated rats with EA or NMDA receptor antagonist MK801. But phosphorylated ERK expressions were markedly induced by CFA, but these inductions were significantly modulated by EA treatment. Although hosphorylation of ERK was also arrested by MK801, these inductions of CFA-injected rats was markedly inhibited only by co-treatment with EA and MK801. Phosphorylated cAMP response element-binding protein (CREB), ERK-related transcriptional factor, showed a significant increase in CFA-treated rats and this increase was slightly inhibited by EA and MK801 treatments. But immunoreaction for phosphorylated CREB were significantly increased by CFA treatment in the superficial laminae of the dorsal horn and these inductions were significantly arrested by co-treatment of EA and MK801. Consequently, the hyperalgesia induced by CFA are associated NMDA receptor and EA and MK801 may showed anti-hyperalgesia via same mechanism for inhibition of ERK and CREB phosphorylation in the dorsal horn.

• Journal of Plant Biotechnology
• /
• v.38 no.2
• /
• pp.162-168
• /
• 2011

High salinity is a common stress condition that adversely affects plant growth and crop production. In response to various environmental stresses, plants activate a number of defense genes that function to increase the tolerance. To isolate Arabidopsis genes that are involved in abiotic stress responses, we carried out genetic screening using various mutant lines. Among them, the blh8 ($\b{B}$EL1-$\b{L}$ike $\b{H}$omeodomain $\underline{8}$) mutant specifically shows chlorotic phenotypes to ionic (specifically, $Na^+$ and $K^+$) stresses, but no differences in root growth. In addition, BLH8 is related to plant development and abiotic stress as predicted by a Graphical Gaussian Model (GGM) network program. It implies that BLH8 functions as a putative transcription factor related to abiotic stress responses. Collectively, our results show that gene network analysis is a useful tool for isolating genes involved in stress adaptation in plants.

• Development and Reproduction
• /
• v.5 no.1
• /
• pp.1-8
• /
• 2001

Golden hamsters show the reproductive activity that is determined by the photoperiod (length of light per day). Photoperiod is an environmental factor that is predictable through an entire year. The hamsters are sexually active in summer during which day length exceeds night time. The critical length is at least 12.5 hours of light in a day where reproductive function is maintained. The information of photoperiod is mediated by the pineal gland because removal of pineal gland blocks the influence of photoperiod on reproductive activity. The hamsters without pineal gland maintain sexual activity and promote it in a situation that suppresses gonadal activity. The pineal gland secretes melatonin that reflects the photoperiod. The appropriate administrations of melatonin into both pineal intact and pinealectomized hamsters lead to a gonadal reression. The results suggest that melatonin constitutes a part of control mechanism whereby environmental information is transduced to neuroendocrine signal respensible for the functional integrity of the reproductive system. Despite of the intense studies, the action site of melatonin is on the whole unknown. It is mainly due to the lack of acute efffct of melatonin on the secretion of reproductive hormones. However, sexually regressed animals display the low levelsof gonadotropins and the augmentation of the hypothalamic gonadotropin-releasing hormone (GnRH) content, implying that the antigonadotropic effects either by photoperiod and/or by the treatment of melatonin are mediated by the GnRH neuronal system. The action mechanism by which melatonin exerts its effect on GnRH neuron needs to be investigated. Recent cloning of melatonin receptor will contribute to examine various and putative potencies of melatonin via its anatomical identification and the action mechanism of melatonin on target tissues at the molecular level.