• Title/Summary/Keyword: in vitro reactivation

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Effect of Cholera Toxin, Dibutyryl cAMP and Adenosine on the In Vitro Reactivation of Latent Herpes Simplex Virus

  • Cheong, D.K.;Park, N.H.
    • Toxicological Research
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    • v.4 no.1
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    • pp.47-53
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    • 1988
  • Cholera toxin and dibutyryl cyclic adenosine 3', 5'-monophosphate(db-cAMP) increased the rate and number of infections units produced in the in vitro reactivation of latent herpes simplex virus, whereas adenosine diminished them. cAMP concentration in latently infected trigeminal ganglia of mice was greatly increased by cholera toxin but was not affected by adenosine.

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In vitro Screening of Oxime Reactivators on the Model of Paraoxon-inhibited Acetylcholinesterase-SAR Study

  • Holas, Ondrej;Musilek, Kamil;Pohanka, Miroslav;Kuca, Kamil;Opletalova, Veronika;Jung, Young-Sik
    • Bulletin of the Korean Chemical Society
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    • v.31 no.6
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    • pp.1609-1614
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    • 2010
  • Acetylcholinesterase reactivators are crucial antidotes for the treatment of organophosphate intoxication. Standard in vitro test was chosen using a rat brain homogenate as the source of AChE. Screening of reactivation potency was performed with two concentration of reactivator (1000 ${\mu}M$ and 10 ${\mu}M$). Results were compared to established reactivators pralidoxime, methoxime, HI-6, trimedoxime and obidoxime. More than 30 novel reactivators performed equal or better reactivation ability of POX-inhibited AChE compared to currently used reactivators. The structure-activity relationship for reactivators of paraoxon-inhibited AChE was developed.

Effect of n-Butyrate on the In Vitro Reactivation of Latent Herpes Simplex Virus (잠재성 Herpes Simplex Virus의 재활성화에 대한 n-Butyrate의 효과)

  • Chun, Yeon-Sook;Park, No-Hee
    • The Korean Journal of Pharmacology
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    • v.22 no.2
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    • pp.123-127
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    • 1986
  • n-Butyrate (n-BT A) increased the rate and number of infectious units produced in the in vitro reactivation of latent herpes simplex virus. While the mechanism of action of n-BT A is obscure, a continuous presence of n-BT A is necessary for its inductive effect.

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Latent Infection and Reactivation of Human Cytomegalovirus from Human Monocyte THP-1 Cells (인체단핵세포주 THP-1세포에서 Human Cytomegalovirus의 잠복감염과 재활성화)

  • 윤상임;문명숙;이찬희
    • Korean Journal of Microbiology
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    • v.37 no.2
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    • pp.145-150
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    • 2001
  • Reactivation of human cytomegalovirus (HCMV) from latency is often fatal to immunocompromised individuals. To understand the effect of HCMV on human monocytes where HCMV establishes latency, two human monocyte cell lines at different stages in differentiation, THP-1 and HL-60 were infected with HCMV. While the viability and morphology of HL-60 cells were not significantly affected by HCMV, the viability of THP-1 cells was dramatically decreased by HCMV infection. THP-1 cells infected with HCMV became aggregated and adhered to the surface of culture dishes, probably due to the increased expression of adherence molecules CD11b on the infected THP-1 cells. THP-1 cells established a latent HCMV infection were induced to differentiate by treatment with TPA and hydrocortisone. Recovery of infectious HCMV from the culture supernatant of differentiated THP-1 cells was dependent on the time of induction of differentiation after HCMV infection. Thus, in vitro model of reactivation of HCMV from latently infected monocytes was established.

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Reactivation of Silenced WT1 Transgene by Hypomethylating Agents - Implications for in vitro Modeling of Chemoimmunotherapy

  • Kwon, Yong-Rim;Son, Min-Jung;Kim, Hye-Jung;Kim, Yoo-Jin
    • IMMUNE NETWORK
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    • v.12 no.2
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    • pp.58-65
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    • 2012
  • Background: A cell line with transfected Wilms' tumor protein 1 (WT1) is has been used for the preclinical evaluation of novel treatment strategies of WT1 immunotherapy for leukemia due to the lack of appropriate murine leukemia cell line with endogenous WT1. However, silencing of the transgene occurs. Regarding the effects of hypomethylating agents (HMAs) on reactivation of silenced genes, HMAs are considered to be immune enhancers. Methods: We treated murine WT1- transfected C1498 (mWT1-C1498) with increasing doses of decitabine (DAC) and azacitidine (AZA) to analyze their effects on transgene reactivation. Results: DAC and AZA decreased the number of viable cells in a dose- or time-dependent manner. Quantification of WT1 mRNA level was analyzed by real-time polymerase chain reaction after mWT1-C1498 treated with increasing dose of HMA. DAC treatment for 48 h induced 1.4-, 14.6-, and 15.5-fold increment of WT1 mRNA level, compared to untreated sample, at 0.1, 1, and $10{\mu}M$, respectively. Further increment of WT1 expression in the presence of 1 and $10{\mu}M$ DAC was evident at 72 h. AZA treatment also induced up-regulation of mRNA, but not to the same degree as with DAC treatment. The correlation between the incremental increases in WT1 mRNA by DAC was confirmed by Western blot and concomitant down-regulation of WT1 promoter methylation was revealed. Conclusion: The in vitro data show that HMA can induce reactivation of WT1 transgene and that DAC is more effective, at least in mWT1-C1498 cells, which suggests that the combination of DAC and mWT1-C1498 can be used for the development of the experimental model of HMA-combined WT1 immunotherapy targeting leukemia.

Biphasic activation of extracellular signal-regulated kinase (ERK) 1/2 in epidermal growth factor (EGF)-stimulated SW480 colorectal cancer cells

  • Joo, Donghyun;Woo, Jong Soo;Cho, Kwang-Hyun;Han, Seung Hyun;Min, Tae Sun;Yang, Deok-Chun;Yun, Cheol-Heui
    • BMB Reports
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    • v.49 no.4
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    • pp.220-225
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    • 2016
  • Cancer cells have different characteristics due to the genetic differences where these unique features may strongly influence the effectiveness of therapeutic interventions. Here, we show that the spontaneous reactivation of extracellular signalregulated kinase (ERK), distinct from conventional ERK activation, represents a potent mechanism for cancer cell survival. We studied ERK1/2 activation in vitro in SW480 colorectal cancer cells. Although ERK signaling tends to be transiently activated, we observed the delayed reactivation of ERK1/2 in epidermal growth factor (EGF)-stimulated SW480 cells. This effect was observed even after EGF withdrawal. While phosphorylated ERK1/2 translocated into the nucleus following its primary activation, it remained in the cytoplasm during late-phase activation. The inhibition of primary ERK1/2 activation or protein trafficking, blocked reactivation and concurrently increased caspase 3 activity. Our results suggest that the biphasic activation of ERK1/2 plays a role in cancer cell survival; thus, regulation of ERK1/2 activation may improve the efficacy of cancer therapies that target ERK signaling.

Somatic Embryogenesis: Morphogenesis, Physiology, Biochemistry and Molecular Biology

  • Thorpe, Trevor A.
    • Korean Journal of Plant Tissue Culture
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    • v.27 no.4
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    • pp.245-258
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    • 2000
  • Somatic embryogenesis has become a major tool in the study of plant embryology, as it is possible in culture to manipulate cells of many plant species to produce somatic embryos in a process that is remarkably similar to zygotic embryogenesis. Traditionally, the process has been studied by an examination of the ex vitro factors which influence embryo formation. Later structural, physiological and biochemical approaches have been applied. Host recently, molecular tools are being used. Together, these various approaches are giving valuable information on the process. This article gives an overview of somatic embryogenesis by reviewing information on the morphogenesis, physiology, biochemistry and molecular biology of the process. Topics covered include a brief description of the factors involved in the production of embryogenic cells. Carrot cell suspension is most commonly used, and the development of a high frequency and synchronous system is outlined. At the physiological and biochemical lev-els various topics, including the reactivation of the cell cycle, changes in endogenous growth regulators, amino acid, polyamine, DNA, RNA and protein metabolism, and embryogenic factors in conditioned medium are all discussed. Lastly, recent information on genes and molecular markers of the embryogenic process are outlined. Somatic embryogenesis, the best example of totipotency in plant cells, is not only an important tool in studies in basic biology, but is potentially of equal significance in the micropropagation of economically important plants.

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In vitro Evaluation of New Acetylcholinesterase Reactivators as Casual Antidotes against Tabun and Cyclosarin

  • Kuca, Kamil;Jun, Daniel;Kim, Tae-Hyuk;Cabal, Jiri;Jung, Young-Sik
    • Bulletin of the Korean Chemical Society
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    • v.27 no.3
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    • pp.395-398
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    • 2006
  • Nerve agents (sarin, tabun, soman and VX) are class of military important substances able to cause many severe intoxications during few minutes. Currently, the threat of misuse of these agents is daily discussed. Unfortunately, there is no single antidote able to treat intoxication caused by all of these agents. Owing to this fact, new generation of antidotes, especially acetylcholinesterase (AChE; EC 3.1.1.7) reactivators, is still developed. In this study, we have tested four newly developed AChE reactivators: 1-(4-hydroxyiminomethylpyridinium)- 5-(4-carbamoylpyridinium)-3-oxa-pentane dibromide (1), 1-(3-hydroxyiminomethylpyridinium)-5-(4-carbamoylpyridinium)-3-oxa-pentane dibromide (2), 1,5-bis(2-hydroxyiminomethylpyridinium)-3-oxa-pentane dichloride (3) and 1,5-bis(4-hydroxyiminomethylpyridinium)-3-oxa-pentane dibromide (4) for their potency to reactivate in vitro tabun and cyclosarin-inhibited AChE. Their reactivation efficacy was compared with currently the most promising oxime HI-6 (1-(2-hydroxyiminomethylpyridinium)-3-(4-carbamoylpyridinium)-2-oxa-propane dichloride). According to obtained results, two AChE reactivators 1 and 4 were able to reactivate tabun-inhibited AChE. On the contrary, there was no better AChE reactivator than HI-6 able to reactivate cyclosarin-inhibited AChE.

Change of X Chromosome Status during Development and Reprogramming

  • Jung, Yong-Wook;Park, In-Hyun
    • Development and Reproduction
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    • v.15 no.3
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    • pp.187-195
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    • 2011
  • X chromosome inactivation (XCI) is a process that enables mammalian females to ensure the dosage compensation for X-linked genes. Investigating the mechanism of XCI might provide deeper understandings of chromosomal silencing, epigenetic regulation of gene expressions, and even the course of evolution. Studies on mammalian XCI conducted with mice have revealed many fundamental findings on XCI. However, difference of murine and human XCI necessitates the further investigation in human XCI. Recent success in reprogramming of differentiated cells into pluripotent stem cells showed the reversibility of XCI in vitro, X chromosome reactivation (XCR), which provides another tool to study the change in X chromosome status. This review summarizes the current knowledge of XCI during early embryonic development and describes recent achievements in studies of XCI in reprogramming process.

A Comparison between Low- and High-Passage Strains of Human CytomegalovirusS

  • Wang, Wen-Dan;Lee, Gyu-Cheol;Kim, Yu Young;Lee, Chan Hee
    • Journal of Microbiology and Biotechnology
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    • v.26 no.10
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    • pp.1800-1807
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    • 2016
  • To understand how human cytomegalovirus (HCMV) might change and evolve after reactivation, it is very important to understand how the nucleotide sequence of cultured HCMV changes after in vitro passaging in cell culture, and how these changes affect the genome of HCMV and the consequent variation in amino acid sequence. Strain JHC of HCMV was propagated in vitro for more than 40 passages and its biological and genetic changes were monitored. For each passage, real-time PCR was performed in order to determine the genome copy number, and a plaque assay was employed to get virus infection titers. The infectious virus titers gradually increased with passaging in cell culture, whereas the number of virus genome copies remained relatively unchanged. A linear correlation was observed between the passage number and the log10 infectious virus titer per virus genome copy number. To understand the genetic basis underlying the increase in HCMV infectivity with increasing passage, the whole-genome DNA sequence of the high-passage strain was determined and compared with the genome sequence of the low-passage strain. Out of 100 mutations found in the high-passage strain, only two were located in an open reading frame. A G-T substitution in the RL13 gene resulted in a nonsense mutation and caused an early stop. A G-A substitution in the UL122 gene generated an S-F nonsynonymous mutation. The mutations in the RL13 and UL122 genes might be related to the increase in virus infectivity, although the role of the mutations found in noncoding regions could not be excluded.