• Title/Summary/Keyword: colony formation

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Radiosensitization Effect of Overexpression of Adenovirus-mediated SIRT6 on A549 Non-small Cell Lung Cancer Cells

  • Cai, Yong;Sheng, Zhao-Ying;Liang, Shi-Xiong
    • Asian Pacific Journal of Cancer Prevention
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    • v.15 no.17
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    • pp.7297-7301
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    • 2014
  • Objective: To explore the radiosensitization effect of overexpression of silent information regulator 6 (SIRT6) on A549 non-small cell lung cancer (NSCLC) cells. Methods: Adenovirus vector Ad-SIRT6 causing overexpression of SIRT6 was established. Western blotting and MTT assay were adopted to detect the level of SIRT6 protein and the inhibitory rate of A549 cell proliferation after different concentrations of adenovirus transduction (0, 25, 100, 200, and 400 pfu/cell) for 24 h. Control group, Ad-null group and Ad-SIRT6 group were designed in this experiment and virus concentration of the latter two groups was 200 pfu/cell. Colony formation assays were employed to test survival fraction (SF) of the 3 groups after 0, 2, 4, 6, 8, 10 X-ray irradiation. Flow cytometry was used to detect the status of cell cycle of 3 groups after 48 h of 4Gy X-ray irradiation and Western blotting was used to determine the expression of apoptosis-related genes of 3 groups after 48 h of 4GyX-ray irradiation. Results: In the range of 25~400 pfu/cell, the inhibitory rate of A549 cell proliferation increased as adenovirus concentration raised. The inhibitory rates under the concentrations of 0, 25, 100, 200, and 400 pfu/cell were 0%, $4.23{\pm}0.34%$, $12.7{\pm}2.57%$, $22.6{\pm}3.38%$, $32.2{\pm}3.22%$, $38.7{\pm}4.09%$ and $47.8{\pm}5.58%$ and there were significantly differences among groups (P<0.05). SF in Ad-SIRT6 group was lower than Ad-null and control groups after 4~10Gy X-ray irradiation (P<0.05) and the sensitization enhancement ratio (SER) was 1.35 when compared with control group. Moreover, after 48 h of 4Gy X-ray irradiation, there appeared a significant increase in G1-phase cell proportion, upregulated expression of the level of apoptosis-promoting genes (Bax and Cleaved caspase-3), but a obvious decline in S-phase and G2-phase cell proportion and a significant decrease of the level of apoptosis-inhibiting gene (Bal-2) in the Ad-SIRT6 group (P<0.05). Conclusion: The over-expression of adenovirus-mediated SIRT6, which has radiosensitization effect on A549 cells of NSCLC, can inhibit the proliferation of A549 cells and cause G0/G1 phase retardation as well as induce apoptosis of cells.

Knockdown of HMGN5 Expression by RNA Interference Induces Cell Cycle Arrest in Human Lung Cancer Cells

  • Chen, Peng;Wang, Xiu-Li;Ma, Zhong-Sen;Xu, Zhong;Jia, Bo;Ren, Jin;Hu, Yu-Xin;Zhang, Qing-Hua;Ma, Tian-Gang;Yan, Bing-Di;Yan, Qing-Zhu;Li, Yan-Lei;Li, Zhen;Yu, Jin-Yan;Gao, Rong;Fan, Na;Li, Bo;Yang, Jun-Ling
    • Asian Pacific Journal of Cancer Prevention
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    • v.13 no.7
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    • pp.3223-3228
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    • 2012
  • HMGN5 is a typical member of the HMGN (high mobility group nucleosome-binding protein) family which may function as a nucleosomal binding and transcriptional activating protein. Overexpression of HMGN5 has been observed in several human tumors but its role in tumorigenesis has not been fully clarified. To investigate its significance for human lung cancer progression, we successfully constructed a shRNA expression lentiviral vector in which sense and antisense sequences targeting the human HMGN5 were linked with a 9-nucleotide loop. Inhibitory effects of siRNA on endogenous HMGN5 gene expression and protein synthesis were demonstrated via real-time RT-PCR and western blotting. We found HMGN5 silencing to significantly inhibit A549 and H1299 cell proliferation assessed by MTT, BrdU incorporation and colony formation assays. Furthermore, flow cytometry analysis showed that specific knockdown of HMGN5 slowed down the cell cycle at the G0/G1 phase and decreased the populations of A549 and H1299 cells at the S and G2/M phases. Taken together, these results suggest that HMGN5 is directly involved in regulation cell proliferation in A549 and H1299 cells by influencing signaling pathways involved in cell cycle progression. Thus, our finding suggests that targeting HMGN5 may be an effective strategy for human lung cancer treatment.

Interspecific Protoplast Fusion of Ganoderma applanatum and Ganoderma lucidum and Fruit Body Formation of the Fusants (잔나비 걸상버섯과 영지(靈芝)의 종간원형질체(種間原形質體) 융합(融合)과 자실체형성(子實體形成)에 관한 연구(硏究))

  • Park, Young-Do;Yoo, Young-Bok;Shin, Pyung-Gyun;You, Chang-Hyun;Cha, Dong-Yeul;Park, Yong-Hwan;Lee, Jae-Sung
    • The Korean Journal of Mycology
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    • v.16 no.2
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    • pp.79-86
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    • 1988
  • Interspecific fusion products were obtained from fusion of Ganoderma applanatum and Ganoderma luridum. Frequency of fusion was 0.77-1.38%. Fusion products were selected by the comparision of morphology and color of colony. Fusion Products had chracteristics of two parental strains and generally grew faster than the parents. Some fusants were segregated on GCM. Fusion products were confirmed by mycelial morphology and electrophoretics pattern of esterase isozyme from mycelium. Most of fusion products were lack in clamp connection but those fusion products, that were segregated produced mycelium with and without clamp connection. Some of the fusion products produced fruit body on sawdust medium.

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A Study on the Formation and Change in the Mordern Sajik Park (근대 사직공원의 형성과 변천)

  • Kim, Seo-Lin;Kim, Hai-Gyoung;Park, Mi-Hyun
    • Journal of the Korean Institute of Traditional Landscape Architecture
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    • v.32 no.4
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    • pp.120-131
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    • 2014
  • Sajikdan(a sort of national shrine in Korea) built at the time of foundation of Joseon was entrenched into Sajik Park going through Japanese colonial era and recently the efforts to restore it is in progress. The details of change in Sajikdan in terms of diachronic analysis are as follows: Firstly, the first period refers to one prior to Japanese colonial era from the first king (also named as "Taejo" in Korean) of the Joseon Dynasty, during which it secured and strengthened the presence as a place for performing important national rites in a nation. It was built on the foot of Inwangsan Mt. at the time of the first king in Joseon Dynasty at first, was destroyed fully by fire during a Japanese Invasion period to Korea(1592-98) and afterward its ancestral ritual facilities were completed under the regime of Youngjo. However, as Japanese intervention coming to the fore, its place was destroyed and then ancestral rites were also abolished in 1908. Secondly, next period falls on 1910 to 1944 when it was transformed and entrenched into a park by the Japanese Empire. While facilities related to a park and an heterogeneous building around the part of boundary were set up, the area of altar, a ritual house and d door of Sajikdan were also designated as historical remains and treasures. Thirdly, this period refers to one from Korea's liberation year from Japanese colony(1945) to the year of 1984 when it had a mixed placeness with the statues, monuments and buildings with heterogeneous nature built. Furthermore, a door of Sajikdan was removed and reconstructed over twice due to opening of Sajik Tunnel. Fourthly, a final period falls on 1985 to the present when efforts are in progress to restore the historicity and symbolism of Sajikdan. A plan for restoration is promoted but now is a difficult time suffering from troubles caused by residents' resistance. Scrutinized historical researches through excavation investigation and residents' understanding are required altogether for restoration of Sajikdan.

Efficient Establishment of Presumptive Embryonic Stem Cells from Bovine Blastocysts by Exposure to Calcium Ionophore (Calcium Ionophore를 이용한 소 배반포로 부터의 배아주 유사세포의 효과적인 분리)

  • 김선욱;류재웅;이철상;한용만;박정선;유대열;이경광
    • Development and Reproduction
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    • v.3 no.1
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    • pp.45-51
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    • 1999
  • It is crucial to remove trophectoderm (TE) cells of blastocysts for an efficient isolation of pluripotent embryonic stem (ES)-like cells from bovine blastocysts. We evaluated the effectiveness of chemosurgery using calcium ionophore A23l87 (CIPA) by investigating the viability and pluripotency of ES-like cell lines isolated from in vitro-produced bovine blastocysts after CIPA treatment. The blastocysts treated with 50 $\mu$M CIPA for 25 min colonized most efficiently (51% of blastocysts) and developed to ES-like cell lines through 10 passages (4.8% of blastocysts) among CIPA-treated groups with different concentration and duration. In comparison with CIPA-untreated blastocysts, the colonization rate and overall viability of the CIPA-treated blastocysts were five times higher, suggesting that CIPA treatment condition defined in this study was highly efficient for establishing ES-like cell lines without apparent toxicity of CIPA. We evaluated in vitro pluripotency of the established three ES-like cell lines by examining alkaline phosphatase (AP) activity, capability of embryoid body formation, and chromosomal euploidity of the cells. Our cells showed a heterogeneous AP activity similarly to other reports. The cells were able to form simple embryoid bodies during suspension culture and majority of them showed a normal chromosome number of 60, the euploid chromosomal complement of bovine Therefore, our data suggest that CIPA treatment can be safely used for an efficient isolation of ES-like cell lines from bovine blastocysts.

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Assessment on Ecological Characteristics of Vegetation in the Trail of Adjacent to Molundae (몰운대 탐방로 주변 식생의 생태적 특성 평가)

  • Nam, Jung-Chil;Seo, Jung-Bum;Oh, Ju-Sung
    • Journal of Environmental Impact Assessment
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    • v.20 no.2
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    • pp.187-198
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    • 2011
  • The objective of this study is to analyze the flora and forest vegetation of trails with high visitor density at Molundae. Nine quadrats of $20{\times}20m$ were selected for the survey. The survey was conducted from April to October 2010. The obtained results are summarized as follows. Plot1, plot2, plot3, plot4, plot6, and plot7 were located at slopes of $5{\sim}20^{\circ}$, 17~40m above sea level, and were formed with the colony of Japanese black pine(Pinus thunbergii Parl) and Japanese black pine(Pinus thunbergii Parl)-white oak(Quercus aliena Blume). Tree layer had the height of 8~20m, and the coverage of 50~70%, while subtree layer had the height of 3-8m, and the coverage 30~80%. On the other hand, shrub layer had the height of 2~4m, and the coverage of 10~30%, and herb had the height of 0.2~1m and coverage 5~20%. Plot5, plot8, and plot9 were located at the summit areas of 57~78m above sea level, and $0^{\circ}$ slope. Japanese black pine(Pinus thunbergii Parl) formed a community there. Tree layer was 8~20m high, and covered 60~70%, of the area, and subtree layer was 6~8m high, and coverage 30~40%. Shrub layer had the height of 2~6m, and the coverage of 30%, while herb layer had the height 0.2~2m, and the coverage 20-80%. Succession does not occur in the surveyed areas which have high visitor density. Artificially planted sawtooth oak(Quercus acutissima) trees were found to disturb succession and formation of multi-layer vegetation, resulting in the ecologically unstable forest. Therefore, the researcher suggested the strategy of managing the vegetation in the conclusion. This study has the limit in that the plots selected for the survey reflected only part of various trails in the Molundae area. It is necessary to suggest the vegetation management plans by selecting more diverse trail areas in consideration of the visitor density and behaviors, and analyzing the changes in vegetation quantitatively in order to manage the vegetation in urban areas damaged by visitors more effectively.

MicroRNA-766-3p Inhibits Tumour Progression by Targeting Wnt3a in Hepatocellular Carcinoma

  • You, Yu;Que, Keting;Zhou, Yun;Zhang, Zhen;Zhao, Xiaoping;Gong, Jianpin;Liu, Zuojin
    • Molecules and Cells
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    • v.41 no.9
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    • pp.830-841
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    • 2018
  • Recent studies have indicated that microRNAs (miRNAs) play an important role in hepatocellular carcinoma (HCC) progression. In this study, we showed that miR-766-3p was decreased in approximately 72% of HCC tissues and cell lines, and its low expression level was significantly correlated with tumour size, TNM stage, metastasis, and poor prognosis in HCC. Ectopic miR-766-3p expression inhibited HCC cell proliferation, colony formation, migration and invasion. In addition, we showed that miR-766-3p repressed Wnt3a expression. A luciferase reporter assay revealed that Wnt3a was a direct target of miR-766-3p, and an inverse correlation between miR-766-3p and Wnt3a expression was observed. Moreover, Wnt3a up-regulation reversed the effects of miR766-3p on HCC progression. In addition, our study showed that miR-766-3p up-regulation decreased the nuclear ${\beta}-catenin$ level and expression of Wnt targets (TCF1 and Survivin) and reduced the level of MAP protein regulator of cytokinesis 1 (PRC1). However, these effects of miR-766-3p were reversed by Wnt3a up-regulation. In addition, PRC1 upregulation increased the nuclear ${\beta}-catenin$ level and protein expression of TCF1 and Survivin. iCRT3, which disrupts the ${\beta}-catenin-TCF4$ interaction, repressed the TCF1, Survivin and PRC1 protein levels. Taken together, our results suggest that miR-766-3p down-regulation promotes HCC cell progression, probably by targeting the Wnt3a/PRC1 pathway, and miR-766-3p may serve as a potential therapeutic target in HCC.

Characterization of TNNC1 as a Novel Tumor Suppressor of Lung Adenocarcinoma

  • Kim, Suyeon;Kim, Jaewon;Jung, Yeonjoo;Jun, Yukyung;Jung, Yeonhwa;Lee, Hee-Young;Keum, Juhee;Park, Byung Jo;Lee, Jinseon;Kim, Jhingook;Lee, Sanghyuk;Kim, Jaesang
    • Molecules and Cells
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    • v.43 no.7
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    • pp.619-631
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    • 2020
  • In this study, we describe a novel function of TNNC1 (Troponin C1, Slow Skeletal and Cardiac Type), a component of actin-bound troponin, as a tumor suppressor of lung adenocarcinoma (LUAD). First, the expression of TNNC1 was strongly down-regulated in cancer tissues compared to matched normal lung tissues, and down-regulation of TNNC1 was shown to be strongly correlated with increased mortality among LUAD patients. Interestingly, TNNC1 expression was enhanced by suppression of KRAS, and ectopic expression of TNNC1 in turn inhibited KRASG12D-mediated anchorage independent growth of NIH3T3 cells. Consistently, activation of KRAS pathway in LUAD patients was shown to be strongly correlated with down-regulation of TNNC1. In addition, ectopic expression of TNNC1 inhibited colony formation of multiple LUAD cell lines and induced DNA damage, cell cycle arrest and ultimately apoptosis. We further examined potential correlations between expression levels of TNNC1 and various clinical parameters and found that low-level expression is significantly associated with invasiveness of the tumor. Indeed, RNA interference-mediated down-regulation of TNNC1 led to significant enhancement of invasiveness in vitro. Collectively, our data indicate that TNNC1 has a novel function as a tumor suppressor and is targeted for down-regulation by KRAS pathway during the carcinogenesis of LUAD.

Effect of retinoic acid on the radiosensitivity of normal human oral keratinocyte (Retinoic acid가 사람 정상 구강각화세포의 방사선감수성에 미치는 영향에 관한 연구)

  • Lee Jean;Heo Min-Suk;Lee Sam-Sun;Oh Sung-Ook;Lee Sul-Mi;Choi Hang-Moon;Choi Soon-Chul;Park Tae-Won
    • Imaging Science in Dentistry
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    • v.33 no.2
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    • pp.97-105
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    • 2003
  • Purpose : To evaluate the effect of all-trans-retinoic acid (ATRA) on the radiosensitivity of normal human oral keratinocyte (NHOK). Materials and methods: Relative cell survival fraction including SF2 (survival fraction at 2 Gy) was calculated on the basis of colony formation assay. Data were fitted to the linear-quadratic model to establish the survival curve and calculate α and β values. Using flow cytometry at 1, 2, 3, 4, and 5 days after exposure to 2 and 10 Gy irradiation, cell cycle arrest and apoptosis were analysed. To understand the molecular mechanism of the radiosensitization of ATRA on NHOK, proteins related with apoptosis and cell cycle arrest were investigated by Western blot analysis. Results: Treatment with ATRA resulted in a significant decrease of SF2 value for NHOK from 0.63 to 0.27, and increased α and β value, indicating that ATRA increased radiosensitivity of NHOK. ATRA increased LDH significantly, but increasing irradiation dose decreased LDH, suggesting that the radiosensitizing effect of ATRA is not directly related with increasing cell necrosis by ATRA. ATRA did not induce appotosis but increased G2 arrest after 10 Gy irradiation, implying that the increased radiosensitivity of NHOK may be due to a decrease in mitosis casued by increasing G2 arrest. ATRA inhibited the reduction of p53 at 3 days after l0Gy irradiation and increased p21 at 1 day after 10 Gy irradiation. Further study is required to determine the precise relationship between this effect and the radiosensitizing effect of A TRA. Conclusion: These results suggested that ATRA increase radiosensitivity by inhibiting mitosis caused by increasing G2 arrest.

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Influence of Pesticides and Environmentally Friendly Agricultural Materials used in Tomato Cultivation on the Pathogenicity of the Entomopathogenic Fungus, Beauveria bassiana (토마토재배에 사용하는 농약과 친환경농자재가 곤충 병원성 곰팡이 Beauveria bassiana의 병원성에 미치는 영향)

  • Park, Jong-Ho;Hong, Sung-Jun;Han, Eun-Jung;Shim, Chang-Ki;Lee, Minho;Kim, Min-Jeong;Kim, JeongJun;Kim, Yong-Ki
    • Korean journal of applied entomology
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    • v.51 no.4
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    • pp.357-364
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    • 2012
  • This study was conducted to observe the influence of chemical pesticides and environmentally friendly agricultural materials (EFAMs) used in tomato cultivation on the pathogenicity of the entomopathogenic fungus, Beauveria bassiana. B. bassiana mycelium didn't grow on PDA media containing 13 fungicides including chlorothalonil and colonies were not formed on PDA media containing 12 fungicides. B. bassiana mycelium grew and colonies were formed on all PDA media containing insecticides and EFAMs, but mycelial growth and colony formation on most PDA media were significantly inhibited compared to the control. The insecticidal activity of B. bassiana against Trialeurodes vaporariorum was decreased when fungicides (polyoxin B, mandipropamid) and EFAMs containing sulfur were added, but insecticides (pyridaben, dinotefuran) and EFAMs originated from plant extracts did not have any influence on the insecticidal activity of B. bassiana. The pathogenicity of a mixture of B. bassiana and polyoxin B against T. vaporariorum was lower than that of B. bassiana alone under greenhouse conditions.