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Differentially Expressed Genes by Methylmercury in Neuroblastoma cell line using suppression subtractive hybridization (SSH) and cDNA Microarray

  • Kim, Youn-Jung;Chang, Suk-Tai;Yun, Hye-Jung;Ryu, Jae-Chun
    • Proceedings of the Korea Society of Environmental Toocicology Conference
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    • 2003.05a
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    • pp.187-187
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    • 2003
  • Methylmercury (MeHg), one of the heavy metal compounds, can cause severe damage to the central nervous system in humans. Many reports have shown that MeHg is poisonous to human body through contaminated foods and has released into the environment. Despite many studies on the pathogenesis of MeHg-induced central neuropathy, no useful mechanism of toxicity has been established so far. In this study, two methods, cDNA Microarray and SSH, were performed to assess the expression profile against MeHg and to identify differentially expressed genes by MeHg in neuroblastoma cell line. TwinChip Human-8K (Digital Genomics) was used with total RNA from SH-SY5Y (human neuroblastoma cell line) treated with solvent (DMSO) and 6.25 uM (IC50) MeHg. And we performed forward and reverse SSH method on mRNA derived from SH-SY5Y treated with DMSO and MeHg (6.25 uM). Differentially expressed cDNA clones were sequenced and were screened by dot blot and ribonuclease protection assay to confirm that individual clones indeed represent differentially expressed genes. These sequences were identified by BLAST homology search to known genes or expressed sequence tags (ESTs). Analysis of these sequences may provide an insight into the biological effects of MeHg in the pathogenesis of neurodegenerative disease and a possibility to develop more efficient and exact monitoring system of heavy metals as environmental pollutants.

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Identification of Genes that are Induced after Cadmium Exposure by Suppression Subtractive Hybridization

  • 이미옥
    • Proceedings of the Korea Society of Environmental Toocicology Conference
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    • 2003.05a
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    • pp.107-107
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    • 2003
  • The heavy metal cadmium is a xenobiotic toxicant of environmental and occupational concern and it has been classified as a human carcinogen. Inhalation of cadmium has been implicated in the development of emphysema and pulmonary fibrosis, but, the detailed mechanism by which cadmium induces adverse biological effects is not yet known. Therefore, we undertook the investigation of genes that are induced after cadmium exposure to illustrate the mechanism of cadmium toxicity For this purpose, we employed the polymerase chain reaction-based suppression subtractive hybridization technique. We identified 29 different cadmium-inducible genes in human peripheral mononuclear cells, such as macrophage migration inhibitory factor, lysophosphatidic acid acyltransferase-${\alpha}$, enolase-1${\alpha}$, VEGF, Bax, neuron-derived orphan receptor-1, and Nur77, which are known to be associated with inflammation, cell survival, and apoptosis. Induction of these genes by cadmium treatment was further confirmed by semi-quantitative reverse-transcription polymerase chain reaction. Further, we found that these genes were also induced after cadmium exposure in normal human lung fibroblast cell line, WI-38, suggesting potential use of this induction profile to monitor cadmium toxicity in the lung. Next, Nur77, one of cadmium-inducible genes, was further studied since the products of Nur77 are known to be involved in the apoptotic process of lung cells. Following cadmium treatment, Nur77 gene expression was increased at protein-level in A549 cells. Consistently, the reporter containing Nur77 binding sequence was activated by 2.5-fold after exposure to cadmium in reporter gene analysis by transient transfection experiments. When the plasmid encoding dominant negative Nur77 that represses the transcriptional function of wild-type Nur77 was transfected into A549 cells, the expression of Bax was significantly reduced, suggesting that induction of Nur77 was an important process in cadmium-induced apoptosis in the cells. Cadmium induced the expression of Nur77 in vivo, confirming the relevance of the data obtained in viro. Together our results suggest that Nur77 gene expression in exposure to cadmium leads apoptosis of lung cells which may cause pathological changes in lung.

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A study on the Normal Steady State Operation Characteristics of PV System Based on the Test Device (태양광전원용 시험장치를 이용한 정상상태 운용특성에 관한 연구)

  • Hasan, Md.Mubdiul;Munkbaht, Munkbaht;Kim, Byung-Ki;Rho, Dae-Seok
    • Proceedings of the KAIS Fall Conference
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    • 2012.05b
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    • pp.512-516
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    • 2012
  • Recently the Korean government's green energy growth policy has been taken at the national level due to the sufficient supply of renewable energy. Some specific technique should be taken in consideration for the operation of the grid voltage and power quality management. In this case, there may have some chance of operational problems. Typical problems arise when grid-connected solar power produced by Pacific sunshine. The power flow in the reverse direction can create overvoltage on the distribution line and gives value of malfunction on the system. Line voltage and overvoltage adjustment practice can stop these symptoms occurred. Under these circumstances, this paper presents an interconnection test devices for photovoltaic(PV) systems composed of distribution system simulator, PV system simulator and control and monitoring systems using the LabVIEW S/W, and simulates the customer voltage characteristics considering the 3 parameters on the introduction capacity for PV systems, system configuration and Power factor. This paper also proposes a new calculation algorithm for voltage profile to make comparison between calculation values and test device values. The results show that the simulation results for the normal operation characteristics of PV systems which are very practical and effective.

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Profiling Total Viable Bacteria in a Hemodialysis Water Treatment System

  • Chen, Lihua;Zhu, Xuan;Zhang, Menglu;Wang, Yuxin;Lv, Tianyu;Zhang, Shenghua;Yu, Xin
    • Journal of Microbiology and Biotechnology
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    • v.27 no.5
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    • pp.995-1004
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    • 2017
  • Culture-dependent methods, such as heterotrophic plate counting (HPC), are usually applied to evaluate the bacteriological quality of hemodialysis water. However, these methods cannot detect the uncultured or viable but non-culturable (VBNC) bacteria, both of which may be quantitatively predominant throughout the hemodialysis water treatment system. Therefore, propidium monoazide (PMA)-qPCR associated with HPC was used together to profile the distribution of the total viable bacteria in such a system. Moreover, high-throughput sequencing of 16S rRNA gene amplicons was utilized to analyze the microbial community structure and diversity. The HPC results indicated that the total bacterial counts conformed to the standards, yet the bacteria amounts were abruptly enhanced after carbon filter treatment. Nevertheless, the bacterial counts detected by PMA-qPCR, with the highest levels of $2.14{\times}10^7copies/100ml$ in softener water, were much higher than the corresponding HPC results, which demonstrated the occurrence of numerous uncultured or VBNC bacteria among the entire system before reverse osmosis (RO). In addition, the microbial community structure was very different and the diversity was enhanced after the carbon filter. Although the diversity was minimized after RO treatment, pathogens such as Escherichia could still be detected in the RO effluent. In general, both the amounts of bacteria and the complexity of microbial community in the hemodialysis water treatment system revealed by molecular approaches were much higher than by traditional method. These results suggested the higher health risk potential for hemodialysis patients from the up-to-standard water. The treatment process could also be optimized, based on the results of this study.

Upregulation and Clinicopathological Significance of Long Non-coding NEAT1 RNA in NSCLC Tissues

  • Pan, Lin-Jiang;Zhong, Teng-Fei;Tang, Rui-Xue;Li, Ping;Dang, Yi-Wu;Huang, Su-Ning;Chen, Gang
    • Asian Pacific Journal of Cancer Prevention
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    • v.16 no.7
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    • pp.2851-2855
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    • 2015
  • Background: Recent reports have shown that nuclear enriched abundant transcript 1 (NEAT1), a long noncoding RNA (lncRNA), contributes to the precise control of gene expression and is related to several human malignancies. However, limited data are available on the expression and function of NEAT1 in lung cancer. The major objective of the current study was to profile the expression and clinicopathological significance of NEAT1 in non-small cell lung cancers (NSCLCs). Materials and Methods: NEAT1 expression in 125 NSCLC cases and paired adjacent non-cancer tissues was assessed by real-time quantitative reverse transcription-PCR (qRT-PCR). Relationships between NEAT1 and clinicopathological factors were also investigated. Results: The relative level of NEAT1 was $6.98{\pm}3.74$ in NSCLC tissues, significantly elevated as compared to that of the adjacent non-cancer lung tissues ($4.83{\pm}2.98$, p<0.001). The area under curve (AUC) of high expression of NEAT1 to diagnose NSCLC was 0.684 (95% CI: 0.619~0.750, p<0.001). NEAT1 expression was positively correlated with patient age (r=-2.007, p=0.047), lymphatic metastasis (r=-2.731, p=0.007), vascular invasion (r=-3.617, p=0.001) and clinical TNM stage (r=-4.134, p<0.001). Conclusions: This study indicates that NEAT1 might be associated with oncogenesis and progression in NSCLC, and suggests application in molecular targeted therapy.

A Gas-Phase Investigation of Oxygen-Hydrogen Exchange Reaction of O(3P) + C2H5 → H(2S) + C2H4O

  • Jang, Su-Chan;Park, Min-Jin;Choi, Jong-Ho
    • Bulletin of the Korean Chemical Society
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    • v.35 no.3
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    • pp.839-844
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    • 2014
  • The gas-phase radical-radical reaction $O(^3P)$ + $C_2H_5$ (ethyl) ${\rightarrow}$ $H(^2S)$ + $CH_3CHO$(acetaldehyde) was investigated by applying a combination of vacuum-ultraviolet laser-induced fluorescence spectroscopy in a crossed beam configuration and ab initio calculations. The two radical reactants $O(^3P)$ and $C_2H_5$ were respectively produced by photolysis of $NO_2$ and supersonic flash pyrolysis of the synthesized precursor azoethane. Doppler profile analysis of the nascent H-atom products in the Lyman-${\alpha}$ region revealed that the average translational energy of the products and the average fraction of the total available energy released as translational energy were $5.01{\pm}0.72kcalmol^{-1}$ and 6.1%, respectively. The empirical data combined with CBS-QB3 level ab initio theory and statistical calculations demonstrated that the title exchange reaction is a major channel and proceeds via an addition-elimination mechanism through the formation of a short-lived, dynamical addition complex on the doublet potential energy surface. On the basis of systematic comparison with several exchange reactions of hydrocarbon radicals, the observed small kinetic energy release can be explained in terms of the loose transition state with a product-like geometry and a small reverse activation barrier along the reaction coordinate.

Comparative Analysis on Positive Bias Stress-Induced Instability under High VGS/Low VDS and Low VGS/High VDS in Amorphous InGaZnO Thin-Film Transistors

  • Kang, Hara;Jang, Jun Tae;Kim, Jonghwa;Choi, Sung-Jin;Kim, Dong Myong;Kim, Dae Hwan
    • JSTS:Journal of Semiconductor Technology and Science
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    • v.15 no.5
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    • pp.519-525
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    • 2015
  • Positive bias stress-induced instability in amorphous indium-gallium-zinc-oxide (a-IGZO) bottom-gate thin-film transistors (TFTs) was investigated under high $V_{GS}$/low $V_{DS}$ and low $V_{GS}$/high $V_{DS}$ stress conditions through incorporating a forward/reverse $V_{GS}$ sweep and a low/high $V_{DS}$ read-out conditions. Our results showed that the electron trapping into the gate insulator dominantly occurs when high $V_{GS}$/low $V_{DS}$ stress is applied. On the other hand, when low $V_{GS}$/high $V_{DS}$ stress is applied, it was found that holes are uniformly trapped into the etch stopper and electrons are locally trapped into the gate insulator simultaneously. During a recovery after the high $V_{GS}$/low $V_{DS}$ stress, the trapped electrons were detrapped from the gate insulator. In the case of recovery after the low $V_{GS}$/high $V_{DS}$ stress, it was observed that the electrons in the gate insulator diffuse to a direction toward the source electrode and the holes were detrapped to out of the etch stopper. Also, we found that the potential profile in the a-IGZO bottom-gate TFT becomes complicatedly modulated during the positive $V_{GS}/V_{DS}$ stress and the recovery causing various threshold voltages and subthreshold swings under various read-out conditions, and this modulation needs to be fully considered in the design of oxide TFT-based active matrix organic light emitting diode display backplane.

Forskolin-Induced Stimulation of RGS2 mRNA in C6 Astrocytoma Cells

  • Kim Sung-Dae;Cho Jae-Youl;Park Hwa-Jin;Kim Sang-Keun;Rhee Man-Hee
    • Biomedical Science Letters
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    • v.12 no.3
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    • pp.131-137
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    • 2006
  • RGS is a negative regulator of G-protein signaling and can be identified by the presence of a conserved $120{sim}125$ amino acid motif, which is referred to as the RGS box. A number of RGSs are induced in response to a wide variety of stimuli. Increased levels of RGSs lead to significant decreases in GPCR responsiveness. To obtain further evidence of a role of RGS proteins in rat C6 astrocytoma cells, we first determined the expression profile of RGS-specific mRNA in C6 cells using reverse transcription-polymerase chain reaction (RT-PCR) with a poly dT18 primer and transcript-specific primers. We found that RGS2, RGS3, RGS6, RGS9, RGS10, RGS12, and RGS16 were differentially expressed in C6 astrocytoma cells. The highest expression rate was found for RGS3, followed by RGS16, RGS10 and RGS9, whereas the expression level for RGS2 was barely detectable. We next assessed whether forskolin regulated the expression of RGSs expressed in C6 astrocytoma cells. The present study found that forskolin dose-dependently stimulated the expression of RGS2 transcripts. This up-regulation of RGS2 gene was abrogated by H-89, potent and broad-spectrum protein kinase A (PKA) inhibitors. Actinomycin D completely inhibited the up-regulation of RGS2 gene induced by forskolin $(10{\mu}M)$, indicating that the regulation of RGS2 gene is controlled at the transcriptional level. In addition, forskolin did significantly activate transcriptional cAMP response element (CRE) in either HEK 293 cells or C6 cells and did not modulate the $NF-{\kappa}B$ and AP-l activity as measured by luciferase reporter gene assay. Finally, forskolin induced the expression of RGS2 mRNA in C6 astrocytoma cells, which depend on the PKA pathway and CRE transcriptional pathways.

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Characterization of the MicroRNA Expression Profile of Cervical Squamous Cell Carcinoma Metastases

  • Ding, Hui;Wu, Yi-Lin;Wang, Ying-Xia;Zhu, Fu-Fan
    • Asian Pacific Journal of Cancer Prevention
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    • v.15 no.4
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    • pp.1675-1679
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    • 2014
  • Objectives: MicroRNAs (miRNAs) are important regulators of many physiological and pathological processes, including tumorigenesis and metastasis. In this study, we sought to determine the underlying molecular mechanisms of metastatic cervical carcinoma by performing miRNA profiling. Methods: Tissue samples were collected from ten cervical squamous cancer patients who underwent hysterectomy and pelvic lymph node (PLN) dissection in our hospital, including four PLN-positive (metastatic) cases and six PLN-negative (non-metastatic) cases. A miRNA microarray platform with 1223 probes was used to determine the miRNA expression profiles of these two tissue types and case groups. MiRNAs having at least 4-fold differential expression between PLN-positive and PLN-negative cervical cancer tissues were bioinformatically analyzed for target gene prediction. MiRNAs with tumor-associated target genes were validated by quantitative reverse transcription-polymerase chain reaction (RT-PCR). Results: Thirty-nine miRNAs were differentially expressed (>4-fold) between the PLN-positive and PLN-negative groups, of which, 22 were up-regulated and 17 were down-regulated. Sixty-nine percent of the miRNAs (27/39) had tumor-associated target genes, and the expression levels of six of those (miR-126, miR-96, miR-144, miR-657, miR-490-5p, and miR-323-3p) were confirmed by quantitative (q)RT-PCR. Conclusions: Six MiRNAs with predicted tumor-associated target genes encoding proteins that are known to be involved in cell adhesion, cytoskeletal remodeling, cell proliferation, cell migration, and apoptosis were identified. These findings suggest that a panel of miRNAs may regulate multiple and various steps of the metastasis cascade by targeting metastasis-associated genes. Since these six miRNAs are predicted to target tumor-associated genes, it is likely that they contribute to the metastatic potential of cervical cancer and may aid in prognosis or molecular therapy.

Sequence Characterization, Expression Profile, Chromosomal Localization and Polymorphism of the Porcine SMPX Gene

  • Guan, H.P.;Fan, B.;Li, K.;Zhu, M.J.;Yerle, M.;Liu, Bang
    • Asian-Australasian Journal of Animal Sciences
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    • v.19 no.7
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    • pp.931-937
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    • 2006
  • The full-length cDNA of the porcine SMPX gene was obtained by the rapid amplification of cDNA ends (RACE). The nucleotide sequences and the predicted protein sequences share high sequence identity with both human and mouse. The promoter of SMPX was sequenced and then analyzed to find the promoter binding sites. The reverse transcriptase-polymerase chain reaction (RT-PCR) revealed that SMPX has a high level of expression in heart and skeletal muscle, a very low expression in lung and spleen and no expression in liver, kidney, fat and brain. Moreover, SMPX has a differential expression level in skeletal muscle, the expression in 65-day embryos being higher than other stages. The porcine SMPX was mapped to SSCXp24 by using a somatic cell hybrid panel (SCHP) and was found closely linked to SW1903 using the radiation hybrid panel IMpRH. An A/G single nucleotide polymorphism (PCR-RFLP) in the 3'-untranslated region (3'-UTR) was detected in eight breeds. The analysis of allele frequency distribution showed that introduced pig breeds (Duroc and Large White) have a higher frequency of allele A while in the Chinese indigenous pig breeds (Qingping pig, Lantang pig, YushanBlack pig, Large Black-White pig, Small Meishan) have a higher frequencies of allele G. The association analysis using an experimental population (188 pigs), which included two cross-bred groups and three pure-blood groups, suggested that the SNP genotype was associated with intramuscular fat content.