• Title/Summary/Keyword: Realtime PCR

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Gene Expression Analysis of So Called Asian Dust Extracts in Human Acute Myeloid Leukemia Cells

  • Choi, You-Jin;Yin, Hu-Quan;Park, Eun-Jung;Park, Kwang-Sik;Kim, Dae-Seon;Lee, Byung-Hoon
    • Toxicological Research
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    • v.26 no.1
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    • pp.21-28
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    • 2010
  • As the frequency and the intensity of so called Asian dust (AD) events have increased, public concerns about the adverse health effects has spiked sharply over the last two decades. Despite the recent reports on the correlation between AD events and the risk for cardiovascular and respiratory disease, the nature of the toxicity and the degree of the risk are yet largely unknown. In the present study, we investigated the effects of the dichloromethane extract of AD (AD-X) and that of urban dust (NAD-X) collected during a non-AD period on gene expression in HL-60 cells using Illumina Sentrix HumanRef-8 Expression BeadChips. Global changes in gene expression were analyzed after 24 h of incubation with 50 or 100 ${\mu}g$/ml AD-X and NAD-X. By one-way analysis of variance (p < 0.05) and Benjamini-Hochberg multiple testing correction for false discovery rate of the results, 573 and 297 genes were identified as AD-X- and NAD-X-responsive, respectively. The genes were classified into three groups by Venn diagram analysis of their expression profile, i.e., 290 AD-X-specific, 14 NAD-X-specific, and 283 overlapping genes. Quantitative realtime PCR confirmed the changes in the expression levels of the selected genes. The expression patterns of five genes, namely SORL1, RABEPK, DDIT4, AZU1, and NUDT1 differed significantly between the two groups. Following rigorous validation process, these genes may provide information in developing biomarker for AD exposure.

Down-regulated MYH11 Expression Correlates with Poor Prognosis in Stage II and III Colorectal Cancer

  • Wang, Ren-Jie;Wu, Peng;Cai, Guo-Xiang;Wang, Zhi-Min;Xu, Ye;Peng, Jun-Jie;Sheng, Wei-Qi;Lu, Hong-Fen;Cai, San-Jun
    • Asian Pacific Journal of Cancer Prevention
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    • v.15 no.17
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    • pp.7223-7228
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    • 2014
  • The MYH11 gene may be related to cell migration and adhesion, intracellular transport, and signal transduction. However, its relationship with prognosis is still uncertain. The aim of this study was to investigate correlations between MYH11 gene expression and prognosis in 58 patients with stage II and III colorectal cancer. Quantitative real-time polymerase chain reaction was performed in fresh CRC tissues to examine mRNA expression, and immunohistochemistry was performed with paraffin-embedded specimens for protein expression. On univariate analysis, MYH11 expression at both mRNA and protein levels, perineural invasion and lymphovascular invasion were related to disease-free survival (p<0.05; log-rank test). Cancers with lower MYH11 expression were more likely to have a poor prognosis. Otherwise, MYH11 expression was unrelated to patient clinicopathological features. On multivariate analysis, low MYH11 expression proved to be an independent adverse prognosticator (p<0.05). These findings show that MYH11 can contribute to predicting prognosis in stage II and III colorectal cancers.

High Expression of Forkhead Box Protein C2 is Related to Poor Prognosis in Human Gliomas

  • Wang, Yao-Wu;Yin, Chun-Li;Zhang, Hong-Yi;Hao, Jin-Min;Yang, Yue-Ye;Liao, Heng;Jiao, Bao-Hua
    • Asian Pacific Journal of Cancer Prevention
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    • v.15 no.24
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    • pp.10621-10625
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    • 2015
  • Background: Increasing evidence has indicated that high Forkhead box protein C2 (FOXC2) level is closely associated with the development, progression, and poor prognosis of a variety of tumors. However, the relationship between FOXC2 and the progression of human gliomas remains to be clarified. The aim of present study was to assess FOXC2 expression and to explore its contribution in human gliomas. Materials and Methods: Realtime quantitative PCR was performed to examine FOXC2 expression in 85 pairs of fresh frozen glioma tissues and corresponding non-neoplastic brain tissues. Associations of FOXC2 expression with clinicopathological factors and prognosis of glioma patients were statistically analyzed. Results: The relative mRNA expression of FOXC2 was significantly higher in glioma tissues than the corresponding non-neoplastic brain tissues (p<0.001). In addition, high FOXC2 expression was significantly associated with advanced pathological grade (P=0.005) and the low Karnofsky performance score (KPS) (p=0.003), correlating with poor survival (p<0.001). Furthermore, multivariate Cox regression analysis showed that high FOXC2 expression was an independent predictor of overall survival (p=0.006). Conclusions: FOXC2 may act as an oncogenic gene and represent a potential regulator of aggressive development and a candidate prognostic marker in human gliomas.

Effects of Achyranthis Japonicae Radix-containing mixture on monosodium iodoacetate-induced osteoarthritis in rats (우슬(牛膝) 등 복합 추출물의 monosodium iodoacetate로 유발한 흰쥐 골관절염에 대한 효과)

  • Kim, Myung-Gyou;Seo, Il-Bok;Leem, Kang-Hyun;Jeong, Taejin;Kim, Jinseok
    • The Korea Journal of Herbology
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    • v.32 no.1
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    • pp.83-89
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    • 2017
  • Objectives : The present study was designed to determine the effects of mixture of Achyranthis Japonicae Radix, Scutellariae Radix, and Acanthopanacis Cortex on monosodium iodoacetate (MIA)-induced osteoarthritis in rats. The mixture was composed of Achyranthis Japonicae Radix, Scutellariae Radix, and Acanthopanacis Cortex extracts. Methods : Arthritis was induced by injection of MIA into knee joints of rats. At the end of experiment, gross examination on the articular structures of knee joints were performed. Proteoglycan (PG) contents in articular cartilages were analysed as well. Tumor necrosis $factor-{\alpha}$($TNF-{\alpha}$) and $interleukin-1{\beta}$ ($IL-1{\beta}$) contents in synovial fluids were measured by ELISA method and matrix metalloproteinase 2 (MMP2), MMP9, and tissue inhibitors of metalloproteinase 1 (TIMP1) mRNA were measured by a realtime PCR. Results : The surfaces of the articular cartilage were observed. The severity of osteoarthritis in the treated group were alleviated compared with control group. PG contents in articular cartilages of the treated group were increased compared with control group. $IL-1{\beta}$ contents in synovial fluids of the treated group were significantly decreased compared with control group. MMP2 and MMP9 mRNA contents in articular cartilages were significantly decreased compared with control group and TIMP1 mRNA contents were increased compared with control group. Conclusions : On the basis of these results, we concluded that Achyranthis Japonicae Radix-containing mixture treatment has anti-arthritic effects on the MIA-induced osteoarthritis in rats. And the effects were related with the reduction of $IL-1{\beta}$ in synovial membranes and the consequent reduction of MMP2 and MMP9 expressions.

Developmental Characteristics of SCNT Pig Embryos Knocked-out of Alpha-1,3-Galactosyltransferase Gene

  • Shim, Joo-Hyun;Park, Mi-Rung;Yang, Byoung-Chul;Ko, Yeoung-Gyu;Oh, Keon-Bong;Lee, Jeong-Woong;Woo, Jae-Seok;Park, Eung-Woo;Park, Soo-Bong;Hwang, Seong-Soo
    • Reproductive and Developmental Biology
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    • v.33 no.3
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    • pp.157-162
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    • 2009
  • This study was performed to comprehend the developmental characteristics of cloned embryos knocked out (KO) of $\alpha$-1,3-galactosyltransferase (GalT) gene. Immature oocytes were collected and cultured for 40 hrs (1-step) or 20hrs (with hormone) + 20hrs (without hormone) (2-step). The embryos transferred with miniature pig ear fibroblast cell were used as control. The reconstructed embryos were cultured in PZM-3 with 5% $CO_2$ in air at $38.5^{\circ}C$ for 6 days. To determine the quality of the blstocysts, TUNEL and quantitative realtime RT-PCR were performed. The embryos were transferred to a surrogate (Landrace) at an earlier stage of the estrus cycle. The maturation rate was significantly higher in 2-step method than that of 1-step (p<0.05). The blastocyst development of GalT KO embryos was significantly lower than that of normal cloned embryos (p<0.05). The total and apoptotic cell number of GalT KO blastocysts was not different statistically from control. The relative abundance of Bax-$\alpha$/Bcl-xl ratio was significantly higher in both cloned blastocysts than that of in vivo blastocysts (p<0.05). Taken together, it can be postulated that the lower developmental potential and higher expression of apoptosis related genes in GalT KO SCNT embryos might be a cause of a low efficiency of GalT KO cloned miniature pig production.

Mutation in clpxoo4158 Reduces Virulence and Resistance to Oxidative Stress in Xanthomonas oryzae pv. oryzae KACC10859

  • Cho, Jung-Hee;Jeong, Kyu-Sik;Han, Jong-Woo;Kim, Woo-Jae;Cha, Jae-Soon
    • The Plant Pathology Journal
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    • v.27 no.1
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    • pp.89-92
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    • 2011
  • Cyclic AMP receptor-like protein (Clp), is known to be a global transcriptional regulator for the expression of virulence factors in Xanthomonas campestris pv. campestris (Xcc). Sequence analysis showed that Xanthomonas oryzae pv. oryzae (Xoo) contains a gene that is strongly homologous to the Xcc clp. In order to determine the role of the Clp homolog in Xoo, a marker exchange mutant of $clp_{xoo4158}$ was generated. Virulence and virulence factors, such as the production of cellulase, xylanase, and extracellular polysaccharides (EPS) and swarming motility were significantly decreased in the $clp_{xoo4158}$ mutant. Moreover, the mutation caused the strain to be more sensitive to hydrogen peroxide and to over-produce siderophores. Complementation of the mutant restored the mutation-related phenotypes. Expression of $clp_{xoo4158}$, assessed by reverse-transcription realtime PCR and clp promoter activity, was significantly reduced in the rpfB, rpfF, rpfC, and rpfG mutants. These results suggest that the clp homolog, $clp_{xoo4158}$, is involved in the control of virulence and resistance against oxidative stress, and that expression of the gene is controlled by RpfC and RpfG through a diffusible signal factor (DSF) signal in Xanthomonas oryzae pv. oryzae KACC10859.

Effects of Temperature on Systemic Infection and Symptom Expression of Turnip mosaic virus in Chinese cabbage (Brassica campestris)

  • Chung, Bong Nam;Choi, Kyung San;Ahn, Jeong Joon;Joa, Jae Ho;Do, Ki Seck;Park, Kyo-Sun
    • The Plant Pathology Journal
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    • v.31 no.4
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    • pp.363-370
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    • 2015
  • Using the Chinese cabbage (Brassica campestris) cultivar 'Chun-goang' as a host and turnip mosaic virus (TuMV) as a pathogen, we studied the effects of ambient temperature ($13^{\circ}C$, $18^{\circ}C$, $23^{\circ}C$, $28^{\circ}C$ and $33^{\circ}C$) on disease intensity and the speed of systemic infection. The optimal temperature for symptom expression of TuMV was $18-28^{\circ}C$. However, symptoms of viral infection were initiated at $23-28^{\circ}C$ and 6 days post infection (dpi). Plants maintained at $33^{\circ}C$ were systemically infected as early as 6 dpi and remained symptomless until 12 or 22 dpi, depending on growth stage at the time of inoculation. It took 45 days for infection of plants grown at $13^{\circ}C$. Quantitative realtime polymerase chain reaction (q-PCR) results showed that the accumulation of virus coat protein was greater in plants grown at $23-28^{\circ}C$. The speed of systemic infection increased linearly with rising ambient temperature, up to $23^{\circ}C$. The zero-infection temperature was $10.1^{\circ}C$. To study the effects of abruptly elevated temperatures on systemic infection, plants inoculated with TuMV were maintained at $10^{\circ}C$ for 20 d; transferred to a growth chamber at temperatures of $13^{\circ}C$, $18^{\circ}C$, $23^{\circ}C$, $28^{\circ}C$, or $33^{\circ}C$ for 1, 2, or 3 d; and then moved back to $10^{\circ}C$. The numbers of plants infected increased as duration of exposure to higher temperatures and dpi increased.

Species composition, seasonal prevalence and flavivirus occurrence of mosquitoes in Daegu and Gunwi, South Korea during the period of 2015-2016

  • KIM, Da Yeong;CHOI, Moon Bo;LEE, Wook-Gyo;KWON, Ohseok
    • Entomological Research
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    • v.48 no.6
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    • pp.522-532
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    • 2018
  • As the vector-borne diseases rapidly increased due to climate change, we investigated the incidence of flavivirus occurrence among female mosquito species in urban and rural study sites in Daegu and Gunwi, South Korea using Realtime PCR. We collected more individuals and more species of mosquito in 2016 than in 2015. In 2015, we recorded a total of 22,033 mosquitoes (Trap Index, 224.8) representing 10 species from 5 genera, whereas in 2016, a total of 27,137 mosquitoes (Trap Index, 278.7) representing 13 species from 6 genera were collected. The number of mosquitoes in 2016 was higher in all study sites, except in the migratory bird habitats where the number decreased. Of the 14 species collected over the two years, Aedes vexans nipponii and Anopheles spp. were dominant in the cowsheds in rural farmland; Culex pipiens complex, in urban residential areas; and, Anopheles spp. and Cu. pipiens complex in migratory bird habitats caught in a black light trap. Cu. pipiens complex and Ae. albopictus in urban parks and Ae. albopictus and Armigeres subalbatus in migratory bird habitats were the dominant species caught in BG-sentinel traps. We found that the number of mosquitoes was highest during July-September. We analyzed 1,725 pools to detect flavivirus, where Chaoyang virus was detected among Ae. vexans niponii collected from cowsheds in rural farmland in 2016. The increased number of mosquitoes recorded in 2016 was mainly due to increased temperatures in the study areas, and reflected the change in study sites.

Stimulating effect of modified Goa-Gi-Um herbal remedy on melanogenesis in B16F10 melanoma cells (B16F10 멜라노마세포에서 과기음가미방의 멜라닌 생성 촉진 효과)

  • Moon, Na-Rang;Kim, Se Yoon;Lee, Jin Hyuk;Lee, Jung Bok;Park, Sunmin
    • The Korea Journal of Herbology
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    • v.28 no.3
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    • pp.69-74
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    • 2013
  • Objectives : Since hypopigmentation is known to increase the risk of skin cancer, melanogenesis in the skin needs to be regulated. Here, we evaluated the melanogenesis stimulatory effects of a modified Goagium herbal remedy (HR) and HR+ox bile (Bos taurus domesticus) extract (OBE) to address hypopigmentation disorders. Methods : B16F10 melanoma cells were treated with different dosages of HR and HR+OBE for 24 to 48 h after 1 h of 10 nM ${\alpha}$-melanocyte stimulating hormone (${\alpha}$-MSH). After the treatment, cell viability, tyrosinase activity, melanin synthesis and the expression of genes related to melanin synthesis were measured and the regulation of the ${\alpha}$-MSH signalling through cAMP responding element binding protein (CREB) was determined. Results : HR and HR+OBE with the ranges of $15{\sim}100{\mu}g/mL$ did not affect cell viability in melanoma cells. The 1 h treatment of HR+OBE (50 and $100{\mu}g/mL$) potentiated the phosphorylation of CREB by enhancing ${\alpha}$-MSH signaling and its 24 h treatment increased CREB expression. Consistent with CREB potentiation, their treatment for 24 h, the expression of microphthalmia-associated transcription factor (MIFT), tyrosinase, tyrosinase related protein (TRP)-1 and TRP-2 were increased in realtime PCR. Ultimately, the 48 h treatment of HR+OBE (50 and $100{\mu}g/mL$) increased tyrosniase activity and melanin contents in the melanoma cells in comparison to the control. Conclusions : HR+OBE (50 and $100{\mu}g/mL$) increases melanin synthesis in B16F10 melanoma cells via the stimulation of tyrosinase activity and expression of MIFT, tyrosinase, TRP-1 and TRP-2. HR+OBE can be used as the a possible treatment for hypopigmentation of the skin.

RNA polymerase I subunit D activated by Yin Yang 1 transcription promote cell proliferation and angiogenesis of colorectal cancer cells

  • Jianfeng Shan;Yuanxiao Liang;Zhili Yang;Wenshan Chen;Yun Chen;Ke Sun
    • The Korean Journal of Physiology and Pharmacology
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    • v.28 no.3
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    • pp.265-273
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    • 2024
  • This study aims to explore possible effect of RNA polymerase I subunit D (POLR1D) on proliferation and angiogenesis ability of colorectal cancer (CRC) cells and mechanism herein. The correlation of POLR1D and Yin Yang 1 (YY1) expressions with prognosis of CRC patients in TCGA database was analyzed. Quantitative realtime polymerase chain reaction (qRT-PCR) and Western blot were applied to detect expression levels of POLR1D and YY1 in CRC cell lines and CRC tissues. SW480 and HT-29 cells were transfected with si-POLR1D or pcDNA3.1-POLR1D to achieve POLR1D suppression or overexpression before cell migration, angiogenesis of human umbilical vein endothelial cells were assessed. Western blot was used to detect expressions of p38 MAPK signal pathway related proteins and interaction of YY1 with POLR1D was confirmed by dual luciferase reporter gene assay and chromatin immunoprecipitation (ChIP). TCGA data showed that both POLR1D and YY1 expressions were up-regulated in CRC patients. High expression of POLR1D was associated with poor prognosis of CRC patients. The results showed that POLR1D and YY1 were highly expressed in CRC cell lines. Inhibition or overexpression of POLR1D can respectively suppress or enhance proliferation and angiogenesis of CRC cells. YY1 inhibition can suppress CRC progression and deactivate p38 MAPK signal pathway, which can be counteracted by POLR1D overexpression. JASPAR predicted YY1 can bind with POLR1D promoter, which was confirmed by dual luciferase reporter gene assay and ChIP. YY1 transcription can up-regulate POLR1D expression to activate p38 MAPK signal pathway, thus promoting proliferation and angiogenesis ability of CRC cells.