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Effects of Hypobaric Conditions on Apoptosis Signalling Pathways in HeLa Cells

  • Arican, Gul Ozcan;Khalilia, Walid;Serbes, Ugur;Akman, Gizem;Cetin, Idil;Arican, Ercan
    • Asian Pacific Journal of Cancer Prevention
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    • v.15 no.12
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    • pp.5043-5047
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    • 2014
  • Nowadays increasing effectiveness in cancer therapy and investigation of formation of new strategies that enhance antiproliferative activity against target organs has become a subject of interest. Although the molecular mechanisms of apoptosis can not be fully explained, it is known that cell suicide program existing in their memory genetically is activated by pathophysiological conditions and events such as oxidative stress. Low pressure (hypobaric) conditions that create hypoxia promote apoptosis by inhibiting cell cycling. In this study, determination of the effects of fractional hypobaric applications at different times on HeLa cells at cellular and molecular levels were targeted. Experiments were carried out under hypobaric conditions (35.2 kPa) in a specially designed hypobaric cabin including 2% $O_2$ and 98% N. Application of fractional hypobaric conditions was repeated two times for 3 hours with an interval of 24 hours. At the end of the implementation period cells were allowed to incubate for 24 hours for activation of repair mechanisms. Cell kinetic parameters such as growth rate (MTT) and apoptotic index were used in determination of the effect of hypobaric conditions on HeLa cells. Also in our study expression levels of the Bcl-2 gene family that have regulatory roles in apoptosis were determined by the RT-PCR technique to evaluate molecular mechanisms. The results showed that antiproliferative effect of hypobaric conditions on HeLa cells started three hours from the time of application and increased depending on the period of exposure. While there was a significant decrease in growth rate values, there was a significant increase in apoptotic index values (p<0.01). Also molecular studies showed that hypobaric conditions caused a significant increase in expression level of proapoptotic gene Bax and significant decrease in antiapoptotic Bfl-1. Consequently fractional application of hypobaric conditions on HeLa cell cultures increased both antiproliferative and apoptotic effects and these effects were triggered by the Bax gene.

Isolation of Human and Mouse Orthologue HPRT Genes by Transformation-Associated Recombination (TAR) cloning (TAR cloning 법에 의한 인간 및 마우스의 상동성 HPRT 유전자의 분리)

  • Do, Eun-Ju;Kim, Jae-Woo;Chung, Chung-Nam;Park, In-Ho;Leem, Sun-Hee
    • Journal of Life Science
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    • v.16 no.6
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    • pp.1036-1043
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    • 2006
  • The transformation-associated recombination (TAR) cloning technique allows selective isolation of chromosome regions or genes from complex genome. The procedure requires knowledge of relatively small genomic sequences that reside adjacent to the chromosome region of interest. This method involves homologous recombination during spheroplast transformation between genomic DNA and a TAR vector that has 5' and 3' gene targeting sequences (hooks). To examine whether TAR cloning can be applied to the isolation of gene homologues, we chose the HPRT genes from human and mouse genome. As results, the yield of positive clones for HPRT gene from human and mouse genome when using a TAR vector containing mHPRT hook or hHPRT hook was almost same level. Analysis of the gap regions in mHPRT revealed that they contain abnormalities that could result in instability of the sequences. In conclusion, we were able to use the TAR cloning technology to isolate gene homologue (orthologue) from nonidentical genome. Moreover, the use of the TAR cloning system may accelerate work on closing the remaining gaps in mammalian genome to achieve the goal of annotation of all mammalian genes.

The effects of chitosan on the human periodontal ligament fibroblasts in vitro (키토산이 치주인대 섬유아세포에 미치는 영향)

  • Paik, Jeong-Won;Lee, Hyun-jung;Yoo, Yun-Jung;Cho, Kyoo-Sung;Kim, Chong-Kwan;Choi, Seong-Ho
    • Journal of Periodontal and Implant Science
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    • v.31 no.4
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    • pp.823-832
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    • 2001
  • Periodontal therapy has dealt primarily with attempts at arresting progression of disease, however, more recent techniques have focused on regenerating the periodontal ligament having the capacity to regenerate the periodontium. The effect of chitosan(poly-N-acetyl glucosaminoglycan), a carbohydrate biopolymer extracted from chitin, on periodontal ligament regeneration is of particular interest. The purpose of this study was to evaluate the effect of chitosan on the human periodontal ligament fibroblasts(hPDLFs) in vitro, with special focus on their proliferative properties by M'IT assay, the synthesis of type I collagen by reverse transcription-polymerase chain reaction(RT-PCR) and the activity of alkaline phosphatase(ALP). Fibroblast populations were obtained from individuals with a healthy periodontium and cultured with ${\alpha}MEM$ as the control group. The experimental groups were cultured with chitosan in concentration of 0.01,0.1, 1,2mg/ml. The results are as follows; 1. Chitosan-induced proliferative responses of hPDLFs reached a plateau at the concentration of O.lmg/ml(p<0.05). 2. When hPDLFs were stimulated with 0.lmg/ml chitosan, mRNA expression of type I collagen was up-regulated. 3. When hPDLFs were stimulated with 0.lmg/ml chitosan, ALP activity was significantly up-regulated(p<0.05). In summary, chitosan(0.lmg/ml) enhanced the type I collagen synthesis in the early stage, and afterwards, facilitated differentiation into osteogenic cells. The results of this in vitro experiment suggest that chitosan potentiates the differentiation of osteoprogenitor cells and may facilitate the formation of bone.

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Convolutional neural network of age-related trends digital radiographs of medial clavicle in a Thai population: a preliminary study

  • Phisamon Kengkard;Jirachaya Choovuthayakorn;Chollada Mahakkanukrauh;Nadee Chitapanarux;Pittayarat Intasuwan;Yanumart Malatong;Apichat Sinthubua;Patison Palee;Sakarat Na Lampang;Pasuk Mahakkanukrauh
    • Anatomy and Cell Biology
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    • v.56 no.1
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    • pp.86-93
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    • 2023
  • Age at death estimation has always been a crucial yet challenging part of identification process in forensic field. The use of human skeletons have long been explored using the principle of macro and micro-architecture change in correlation with increasing age. The clavicle is recommended as the best candidate for accurate age estimation because of its accessibility, time to maturation and minimal effect from weight. Our study applies pre-trained convolutional neural network in order to achieve the most accurate and cost effective age estimation model using clavicular bone. The total of 988 clavicles of Thai population with known age and sex were radiographed using Kodak 9000 Extra-oral Imaging System. The radiographs then went through preprocessing protocol which include region of interest selection and quality assessment. Additional samples were generated using generative adversarial network. The total clavicular images used in this study were 3,999 which were then separated into training and test set, and the test set were subsequently categorized into 7 age groups. GoogLeNet was modified at two layers and fine tuned the parameters. The highest validation accuracy was 89.02% but the test set achieved only 30% accuracy. Our results show that the use of medial clavicular radiographs has a potential in the field of age at death estimation, thus, further study is recommended.

Antinociceptive, anti-inflammatory, and cytotoxic properties of Origanum vulgare essential oil, rich with β-caryophyllene and β-caryophyllene oxide

  • Moghrovyan, Armenuhi;Parseghyan, Lilya;Sevoyan, Gohar;Darbinyan, Anna;Sahakyan, Naira;Gaboyan, Monica;Karabekian, Zaruhi;Voskanyan, Armen
    • The Korean Journal of Pain
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    • v.35 no.2
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    • pp.140-151
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    • 2022
  • Background: Essential oils are of great interest for their analgesic and anti-inflammatory properties. We aimed to study the content of the essential oil of the Origanum vulgare of the Armenian highlands (OVA) in different periods of vegetation and to investigate its antinociceptive and anti-inflammatory effects in mice (in vivo) and cytotoxic action in cultured cells (in vitro). OVA essential oil was extracted from fresh plant material by hydro-distillation. Methods: For OVA essential oil contents determination the gas chromatography-mass spectrometry method was used. Formalin and hot plate tests and analysis of cell viability using the methyl-thiazolyl-tetrazolium (MTT) assay were used. Results: The maximal content of β-caryophyllene and β-caryophyllene oxide in OVA essential oil was revealed in the period of blossoming (8.18% and 13.36%, correspondently). In the formalin test, 4% OVA essential oil solution (3.5 mg/mouse) exerts significant antinociceptive and anti-inflammatory effects (P = 0.003). MTT assay shows approximately 60% cytotoxicity in HeLa and Vero cells for 2.0 µL/mL OVA essential oil in media. Conclusions: The wild oregano herb of Armenian highlands, harvested in the blossoming period, may be considered as a valuable source for developing pain-relieving preparations.

Coloration Study of Red/Yellow β-FeOOH Nanorod using NH4OH Solution (NH4OH를 이용한 적황색 β-FeOOH 나노로드 길이에 따른 색상제어 연구)

  • Yu, Ri;Kim, IllJoo;Yun, JiYeon;Choi, Eun-Young;Pee, Jae-Hwan;Kim, YooJin
    • Journal of Powder Materials
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    • v.23 no.5
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    • pp.343-347
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    • 2016
  • Fe-based pigments have attracted much interest owing to their eco-friendliness. In particular, the color of nanosized pigments can be tuned by controlling their size and morphology. This study reports on the effect of length on the coloration of ${\beta}$-FeOOH pigments prepared using an $NH_4OH$ solution. First, rod-type ${\beta}$-FeOOH is prepared by the hydrolysis of $FeCl_3{\cdot}6H_2O$ and $NH_4OH$. When the amount of $NH_4OH$ is increased, the length of the rods decreases. Thus, the length of the nanorods can be adjusted from 10 nm to 300 nm. The color of ${\beta}$-FeOOH changes from orangered to yellow depending on the length of ${\beta}$-FeOOH. The color and phase structure of ${\beta}$-FeOOH is characterized by UV-vis spectroscopy, CIE Lab color parameter measurements, transmission electron microscopy (TEM), scanning electron microscopy (SEM), and powder X-ray diffraction (XRD).

In Vitro and In Vivo Anti-Tobacco Mosaic Virus Activities of Essential Oils and Individual Compounds

  • Lu, Min;Han, Zhiqiang;Xu, Yun;Yao, Lei
    • Journal of Microbiology and Biotechnology
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    • v.23 no.6
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    • pp.771-778
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    • 2013
  • Essential oils are increasingly of interest for use as novel drugs acting as antimicrobial and antiviral agents. In the present study, we report the in vitro antiviral activities of 29 essential oils, extracted from Chinese indigenous aromatic plants, against the tobacco mosaic virus (TMV). Of these essential oils, those oils from ginger, lemon, tea tree, tangerine peel, artemisia, and lemongrass effected a more than 50% inhibition of TMV at 100 ${\mu}g/ml$. In addition, the mode of antiviral action of the active essential oils was also determined. Essential oils isolated from artemisia and lemongrass possessed potent inactivation and curative effects in vivo and had a directly passivating effect on TMV infection in a dose-dependent manner. However, all other active essential oils exhibited a moderate protective effect in vivo. The chemical constitutions of the essential oils from ginger, lemon, tea tree, tangerine peel, artemisia, and lemongrass were identified by gas chromatography and gas chromatography-mass spectrometry. The major components of these essential oils were ${\alpha}$-zingiberene (35.21%), limonene (76.25%), terpinen-4-ol (41.20%), limonene (80.95%), 1,8-cineole (27.45%), and terpinolene (10.67%). The curative effects of 10 individual compounds from the active essential oils on TMV infection were also examined in vivo. The compounds from citronellal, limonene, 1,8-cineole, and ${\alpha}$-zingiberene effected a more than 40% inhibition rate for TMV infection, and the other compounds demonstrated moderate activities at 320 ${\mu}g/ml$ in vivo. There results indicate that the essential oils isolated from artemisia and lemongrass, and the individual compound citronellal, have the potential to be used as an effective alternative for the treatment of tobacco plants infected with TMV under greenhouse conditions.

Mature silkworm powder reduces blood alcohol concentration and liver injury in ethanol-treated rats

  • Lee, Da-Young;Hong, Kyung-Sook;Yun, Sun-Mi;Song, Moon-Young;Ji, Sang-Deok;Son, Jong-Gon;Kim, Eun-Hee
    • International Journal of Industrial Entomology and Biomaterials
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    • v.35 no.2
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    • pp.123-128
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    • 2017
  • Hangover due to alcohol consumption causes social and physical problems. There is a growing interest in edible insects worldwide. We have previously published a new technology to make hard mature silkworm, Bombyx mori, into edible form, steamed and freeze-dried mature silkworm larval powder (SMSP). In this study, AIN-76 or SMSP (0.1 and 1 g/kg rat body weight) containing diets in SD rats were pretreated for 2 weeks, and ethanol (3 g/kg rat body weight) was administered as an oral gavage and sacrificed after 3 hours. As a result, blood alcohol and aldehyde levels were significantly decreased in SMSP fed rats. In addition, liver injury markers, alanine aminotransferase (ALT) and alkaline phosphatase (ALP) were significantly decreased in SMSP group compared to ethanol group. $TNF-{\alpha}$, an inflammatory cytokine, and malondialdehyde (MDA), an oxidative stress marker, also showed a dose-dependent decrease in the group receiving SMSP. Conclusively, consumption of SMSP not only reduced hangover induced by ethanol, but also decreased liver damage, oxidative stress, and inflammatory response.

Gametophytic Abortion in Heterozygotes but Not in Homozygotes: Implied Chromosome Rearrangement during T-DNA Insertion at the ASF1 Locus in Arabidopsis

  • Min, Yunsook;Frost, Jennifer M.;Choi, Yeonhee
    • Molecules and Cells
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    • v.43 no.5
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    • pp.448-458
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    • 2020
  • T-DNA insertional mutations in Arabidopsis genes have conferred huge benefits to the research community, greatly facilitating gene function analyses. However, the insertion process can cause chromosomal rearrangements. Here, we show an example of a likely rearrangement following T-DNA insertion in the Anti-Silencing Function 1B (ASF1B) gene locus on Arabidopsis chromosome 5, so that the phenotype was not relevant to the gene of interest, ASF1B. ASF1 is a histone H3/H4 chaperone involved in chromatin remodeling in the sporophyte and during reproduction. Plants that were homozygous for mutant alleles asf1a or asf1b were developmentally normal. However, following self-fertilization of double heterozygotes (ASF1A/asf1a ASF1B/asf1b, hereafter AaBb), defects were visible in both male and female gametes. Half of the AaBb and aaBb ovules displayed arrested embryo sacs with functional megaspore identity. Similarly, half of the AaBb and aaBb pollen grains showed centromere defects, resulting in pollen abortion at the bi-cellular stage of the male gametophyte. However, inheritance of the mutant allele in a given gamete did not solely determine the abortion phenotype. Introducing functional ASF1B failed to rescue the AaBb- and aaBb-mediated abortion, suggesting that heterozygosity in the ASF1B gene causes gametophytic defects, rather than the loss of ASF1. The presence of reproductive defects in heterozygous mutants but not in homozygotes, and the characteristic all-or-nothing pollen viability within tetrads, were both indicative of commonly-observed T-DNA-mediated translocation activity for this allele. Our observations reinforce the importance of complementation tests in assigning gene function using reverse genetics.

Analysis of Polycyclic Aromatic Hydrocarbons in Agricultural Waterways in Chungbuk and Gyeongbuk Provinces in Korea

  • Kim, Leesun;Lee, Jong-Hwa;Kim, Jeong-Han;Lee, Sung-Eun
    • Korean Journal of Environmental Biology
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    • v.36 no.3
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    • pp.345-351
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    • 2018
  • An efficient, quick and low-cost extraction and clean up method for the determination of 14 polycyclic aromatic hydrocarbons (PAHs) in the agricultural water samples was optimized using gas chromatography-tandem mass spectrometry (GC-MS/MS). The extraction of the target compounds in water sample was carried out with acetonitrile, followed by partitioning promoted by the addition of salt. As a clean-up procedure, dispersive solid phase extraction was employed to purify the analytes of interest for GC-MS/MS analysis. This method was successfully applied for the quantification of PAHs in real water samples collected for the purpose of monitoring from the waterways located in Chungbuk (15 sites) and Gyeongbuk (6 sites), S. Korea. Phenanthrene (0.54 to $2.53{\mu}gL^{-1}$) was detected in all the water samples collected from both the sites. Fluoranthene was detected in the water samples collected from the two sites in Gyeongbuk province, but other PAHs were not determined in these water sampling sites. Based on these results, the determined PAHs were conducted using an environmental risk assessment. The risk characterization ratios (RCRs) for phenanthrene ranged from 0.37 to 3.21. These RCR values referred to as risk was not controlled because RCR values of some sites were greater than 1. In conclusion, it is proposed that the optimized method in combination with GC-MS/MS could be successfully employed for the determination of PAHs in any environmental samples including water samples.