• Title/Summary/Keyword: RNA amplification

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Expression of Fatty Acid Synthase Negatively Correlates with PTEN and Predicts Peritoneal Dissemination of Human Gastric Cancer

  • Xiang, Hong-Gang;Hao, Jun;Zhang, Wen-Jie;Lu, Wen-Jie;Dong, Ping;Liu, Ying-Bin;Chen, Lei
    • Asian Pacific Journal of Cancer Prevention
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    • v.16 no.16
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    • pp.6851-6855
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    • 2015
  • Background: This study aimed to examine the clinical significance of fatty acid synthase (FASN) expression in gastric cancer (GC), and investigate any prognostic role. Materials and Methods: FASN expression was assessed in gastric cancers by immunohistochemistry using 60 paraffin-embedded tissue specimens, and clinical data were collected by retrospective chart review. Moreover, FASN mRNA expression in 15 fresh resected specimens was evaluated by the reverse transcription-polymerase chain reaction (RT-PCR). Immunohistochemical staining of PTEN was performed to assess the correlation of PTEN with FASN in gastric cancer. Results: Increased expression of FASN was noted in gastric cancers. The frequency of FASN gene amplification was also significantly higher in gastric cancer than in adjacent normal tissue. FASN expression in human gastric cancer tissues was significantly correlated with patient TNM stage and peritoneal dissemination (p<0.05). Moreover, higher FASN expression significantly correlated with shorter overall survival (p<0.05). Here, upregulation of FASN negatively correlated with PTEN expression in gastric cancer. Conclusions: These findings indicate that FASN expression is upregulated in gastric cancer, and increased FASN may be critical to th peritoneal metastasis and survival. Our results suggest that FASN upregulation and PTEN downregualtion may be involved in peritoneal dissemination for gastric cancer progression.

MicroRNA-214 Regulates the Acquired Resistance to Gefitinib via the PTEN/AKT Pathway in EGFR-mutant Cell Lines

  • Wang, Yong-Sheng;Wang, Yin-Hua;Xia, Hong-Ping;Zhou, Song-Wen;Schmid-Bindert, Gerald;Zhou, Cai-Cun
    • Asian Pacific Journal of Cancer Prevention
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    • v.13 no.1
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    • pp.255-260
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    • 2012
  • Patients with non-small cell lung cancer (NSCLC) who have activating epidermal growth factor receptor (EGFR) mutations derive clinical benefit from treatment with EGFR-tyrosine kinase inhibitors ((EGFR-TKIs)-namely gefitinib and erlotinib. However, these patients eventually develop resistance to EGFR-TKIs. Despite the fact that this acquired resistance may be the result of a secondary mutation in the EGFR gene, such as T790M or amplification of the MET proto-oncogene, there are other mechanisms which need to be explored. MicroRNAs (miRs) are a class of small non-coding RNAs that play pivotal roles in tumorigenesis, tumor progression and chemo-resistance. In this study, we firstly successfully established a gefitinib resistant cell line-HCC827/GR, by exposing normal HCC827 cells (an NSCLC cell line with a 746E-750A in-frame deletion of EGFR gene) to increasing concentrations of gefitinib. Then, we found that miR-214 was significantly up-regulated in HCC827/GR. We also showed that miR-214 and PTEN were inversely expressed in HCC827/GR. Knockdown of miR-214 altered the expression of PTEN and p-AKT and re-sensitized HCC827/GR to gefitinib. Taken together, miR-214 may regulate the acquired resistance to gefitinib in HCC827 via PTEN/AKT signaling pathway. Suppression of miR-214 may thus reverse the acquired resistance to EGFR-TKIs therapy.

Molecular pathological interactions between Apple stem grooving virus (ASGV) and its fungi.

  • Hyekyung Shim;Lee, Hyunjeong;Seungbeom Hong;Park, Dae-Sup;DaeRobert A Samson;Hyeongjin Jee;Lee, Sukchan
    • Proceedings of the Korean Society of Plant Pathology Conference
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    • 2003.10a
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    • pp.122-123
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    • 2003
  • Apple stem grooving virus (ASGV) belongs to Capillovirus and infects pome fruits. Transmission mode of ASGV is known by grafting and mechanical inoculation into susceptible hosts, not by any other natural vectors. But we have observed the spread of ASGV in the field without mechanical inoculation or grafting. Transmission seems to be occurred from tree-to-tree and tree-to-susceptible herbaceous plants along but not across ditches in the field. In order to ascertain this possibility, various fungi were isolated and cultured from ASGV-infected plants and 69 isolates were characterized. By means of RNA dot-blot hybridization and PCR analysis, 3 isolates were sorted out for further studies. The isolates were identified to Tataromyces sp. and belonged to Phenicillium by morphological characteristics and molecular markers. As an experimental host, 10 kidney beans (Phaseolus vulgaris) were screened and Kyunggi-5 was selected for virus amplification and symptom development. Kyunggj-5 infected by fungi which seemed to carry ASGV showed the typical disease symptoms and viral coat protein genes were detected from all tested plants. To confirm the Koch's rule, fungi cultured from inoculation origins of kidney bean were grown on PDA media and re-inoculated to hosts. The fungi isolated from inoculation origins induced the typical disease symptoms on hosts. However virus free fungi did not induce any symptom on the experimental hosts. This bioassay showed that these typical symptoms were caused by virus, not fungi.

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Cloning and Expression Analysis of a Novel Mouse Zinc Finger Protein Gene Znf313 Abundantly Expressed in Testis

  • Li, Na;Sun, Huaqin;Wu, Qiaqing;Tao, Dachang;Zhang, Sizhong;Ma, Yongxin
    • BMB Reports
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    • v.40 no.2
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    • pp.270-276
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    • 2007
  • We have cloned a novel mouse zinc finger protein gene Znf313 by rapid amplification of cDNA ends (RACE) according to the homologue of human ZNF313 gene. The cDNA is 2,163 base pairs (bp) in length and encodes a 229 amino acids (aa) protein with a $C_3HC_4$ ring finger domain and three $C_2H_2$ domains. 89% and 93% nucleotide (nt) and aa sequence identity is observed with its human homologue. Revealed by Northern blot and RT-PCR, full mRNA consists of 2.16 kb and widely expresses in tissues as a single transcript, most abundantly in heart, liver, kidney and testis. The expression of Znf313 in testis is detected in all development stages. Western blot analysis also reveals that Znf313 is expressed in the tissues. Immunohistochemical staining and subcellular localization demonstrate that Znf313 is expressed both in the cytoplasm and nucleus whereas predominantly localized in the nucleus. Present data suggests that Znf313 gene might play a fundamental role in gene transcription and regulation in organism and relates to spermatogenesis.

Molecular Differentiation of Schistosoma japonicum and Schistosoma mekongi by Real-Time PCR with High Resolution Melting Analysis

  • Kongklieng, Amornmas;Kaewkong, Worasak;Intapan, Pewpan M.;Sanpool, Oranuch;Janwan, Penchom;Thanchomnang, Tongjit;Lulitanond, Viraphong;Sri-Aroon, Pusadee;Limpanont, Yanin;Maleewong, Wanchai
    • Parasites, Hosts and Diseases
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    • v.51 no.6
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    • pp.651-656
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    • 2013
  • Human schistosomiasis caused by Schistosoma japonicum and Schistosoma mekongi is a chronic and debilitating helminthic disease still prevalent in several countries of Asia. Due to morphological similarities of cercariae and eggs of these 2 species, microscopic differentiation is difficult. High resolution melting (HRM) real-time PCR is developed as an alternative tool for the detection and differentiation of these 2 species. A primer pair was designed for targeting the 18S ribosomal RNA gene to generate PCR products of 156 base pairs for both species. The melting points of S. japonicum and S. mekongi PCR products were $84.5{\pm}0.07^{\circ}C$ and $85.7{\pm}0.07^{\circ}C$, respectively. The method permits amplification from a single cercaria or an egg. The HRM real-time PCR is a rapid and simple tool for differentiation of S. japonicum and S. mekongi in the intermediate and final hosts.

Identification of Sex-specific Expression Markers in the Giant Tiger Shrimp (Penaeus monodon)

  • Khamnamtong, Bavornlak;Thumrungtanakit, Supaporn;Klinbunga, Sirawut;Aoki, Takashi;Hirono, Ikuo;Menasveta, Piamsak
    • BMB Reports
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    • v.39 no.1
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    • pp.37-45
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    • 2006
  • Bulked segregant analysis (BSA) and AFLP were used for isolation of genomic sex determination markers in Penaeus monodon. A total of 256 primer combinations were tested against 6-10 bulked genomic DNA of P. monodon. Five and one candidate female- and male-specific AFLP fragments were identified. Female-specific fragments were cloned and further characterized. SCAR markers derived from FE10M9520, FE10M10725.1, FE10M10725.2 and FE14M16340 provided the positive amplification product in both male and female P. monodon. Further analysis of these markers using SSCP and genome walk analysis indicated that they were not sex-linked. In addition, sex-specific (or differential) expression markers in ovaries and testes of P. monodon were analyzed by RAP-PCR (150 primer combinations). Twenty-one and fourteen RAP-PCR fragments specifically/differentially expressed in ovaries and testes of P. monodon were successfully cloned and sequenced. Expression patterns of 25 transcripts were tested against the first stranded cDNA of ovaries and testes of 3-month-old and broodstock-sized P. monodon (N = 5 and N = 7 - 10 for females and N = 4 and N = 5 - 7 for males, respectively). Five (FI-4, FI-44, FIII-4, FIII-39 and FIII-58) and two (M457-A01 and MII-51) derived RAP-PCR markers revealed female- and male-specific expression patterns in P. monodon. Surprisingly, MII-5 originally found in testes showed a higher expression level in ovaries than did testes of juvenile shrimps but a temporal female-specific pattern in P. monodon adults.

Significance of Expression of Human METCAM/MUC18 in Nasopharyngeal Carcinomas and Metastatic Lesions

  • Lin, Jin-Ching;Chiang, Cheng-Feng;Wang, Shur-Wern;Wang, Wen-Yi;Kwan, Po-Cheung;Wu, Guang-Jer
    • Asian Pacific Journal of Cancer Prevention
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    • v.15 no.1
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    • pp.245-252
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    • 2014
  • Human METCAM/MUC18, a cell adhesion molecule (CAM) in the immunoglobulin-like gene super family, plays a dual role in the progression of several epithelium cancers; however, its role in the nasopharyngeal carcinoma (NPC) remains unclear. To initiate the study we determined human METCAM/MUC18 expression in tissue samples of normal nasopharynx (NP), NPCs, and metastatic lesions, and in two established NPC cell lines. Immunoblotting analysis was used for the determination in lysates of frozen tissues, and immunohistochemistry (IHC) for expression in formalin-fixed, paraffin-embedded tissue sections of 7 normal nasopharynx specimens, 94 NPC tissue specimens, and 3 metastatic lesions. Human METCAM/MUC18 was expressed in 100% of the normal NP, not expressed in 73% of NPC specimens (or expressed at very low levels in only about 27% of NPC specimens), and expressed again in all of the metastatic lesions. The level of human METCAM/MUC18 expression in NPC tissues was about one fifth of that in the normal NP and metastatic lesions. The low level of human METCAM/MUC18 expression in NPC specimens was confirmed by a weak signal of RT-PCR amplification of the mRNA. Low expression levels of human METCAM/MUC18 in NPC tissues were also reflected in the seven established NPC cell lines. These findings provided the first evidence that diminished expression of human METCAM/MUC18 is an indicator for the emergence of NPC, but increased expression then occurs with metastatic progression, suggesting that huMETCAM/MUC18, perhaps similar to TGF-${\beta}$, may be a tumor suppressor, but a metastasis promoter for NPC.

Comparative Pathogenicity of Fusarium graminearum Isolates from Wheat Kernels in Korea

  • Shin, Sanghyun;Son, Jae-Han;Park, Jong-Chul;Kim, Kyeong-Hoon;Yoon, Young-mi;Cheong, Young-Keun;Kim, Kyong-Ho;Hyun, Jong-Nae;Park, Chul Soo;Dill-Macky, Ruth;Kang, Chon-Sik
    • The Plant Pathology Journal
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    • v.34 no.5
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    • pp.347-355
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    • 2018
  • Fusarium head blight (FHB) caused by Fusarium species is a major disease of wheat and barley around the world. FHB causes yield reductions and contamination of grains with trichothecene mycotoxins including; nivalenol (NIV), deoxynivalenol (DON), 3-acetyldeoxynivalenol (3-ADON), and 15-acetyldeoxynivalenol (15-ADON). The objectives of this study were to identify strains of F. graminearum isolated in Korea from 2012-harvested wheat grain and to test the pathogenicity of these NIV- and DON-producing isolates. Three hundred and four samples of wheat grain, harvested in 2012 in Chungnam, Chungbuk, Gyeongnam, Jeonbuk, Jeonnam, and Gangwon provinces were collected. We recovered 44 isolates from the 304 samples, based on the PCR amplification of internal transcribed spacer (ITS) rRNA region and sequencing. Our findings indicate that F. asiaticum was the predominant (95% of all isolates) species in Korea. We recovered both F. asiaticum and F. graminearum from samples collected in Chungnam province. Of the 44 isolates recovered, 36 isolates had a NIV genotype while 8 isolates belonged to the DON genotype (3-ADON and 15-ADON). In order to characterize the pathogenicity of the strains collected, disease severity was assessed visually on various greenhouse-grown wheat cultivars inoculated using both NIV- and DON-producing isolates. Our results suggest that Korean F. graminearum isolates from wheat belong to F. asiaticum producing NIV, and both F. graminearum and F. asiaticum are not significantly different on virulence in wheat cultivars.

Comorbid Gastric Adenocarcinoma and Gastric and Duodenal Strongyloides stercoralis Infection: A Case Report

  • Seo, An Na;Goo, Youn-Kyoung;Chung, Dong-Il;Hong, Yeonchul;Kwon, Ohkyoung;Bae, Han-Ik
    • Parasites, Hosts and Diseases
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    • v.53 no.1
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    • pp.95-99
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    • 2015
  • Strongyloides stercoralis can cause systemic infection, termed strongyloidiasis, and gastrointestinal ulcer disease in immunocompromised patients. However, to our knowledge, there are no reported cases of comorbid gastric adenocarcinoma and S. stercoralis infection. Here, we report a case of an 81-year-old Korean man who presented with S. stercoralis infection coexisting with early gastric adenocarcinoma (T1aN0M0). S. stercoralis eggs, rhabditiform larvae, and adult females were observed in normal gastric and duodenal crypts. They were also observed in atypical glands representative of adenocarcinoma and adenoma. Preliminary laboratory tests revealed mild neutrophilic and eosinophilic leukocytosis. A routine stool test failed to detect rhabditiform larvae in the patient's fecal sample; however, S. stercoralis was identified by PCR amplification and 18S rRNA sequencing using genomic DNA extracted from formalin-fixed paraffin-embedded tissues. Postoperatively, the patient had a persistent fever and was treated with albendazole for 7 days, which alleviated the fever. The patient was followed-up by monitoring and laboratory testing for 4 months postoperatively, and no abnormalities were observed thus far. The fact that S. stercoralis infection may be fatal in immunocompromised patients should be kept in mind when assessing high-risk patients.

Induction of Apoptosis and Cell Cycle Arrest by Dorema Glabrum Root Extracts in a Gastric Adenocarcinoma (AGS) Cell Line

  • Jafari, Naser;Zargar, Seyed Jalal;Yassa, Narguess;Delnavazi, Mohammad Reza
    • Asian Pacific Journal of Cancer Prevention
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    • v.17 no.12
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    • pp.5189-5193
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    • 2016
  • Objective: Dorema glabrum Fisch. & C.A. Mey is a perennial plant that has several curative properties. Anti-proliferative activity of seeds of this plant has been demonstrated in a mouse fibrosarcoma cell line. The aim of the present study was to evaluate cytotoxicity of D. glabrum root extracts in a human gastric adenocarcinoma (AGS) cell line and explore mechanisms of apoptosis induction, cell cycle arrest and altered gene expression in cancer cells. Materials and Methods: The MTT assay was used to evaluate IC50 values, EB/AO staining to analyze the mode of cell death, and flow cytometry to assess the cell cycle. Quantitative real-time polymerase chain reaction (qRT-PCR) amplification was performed with apoptosis and cell cycle-related gene primers, for cyclin D1, c-myc, survivin, VEGF, Bcl-2, Bax, and caspase-3 to determine alteration of gene expression. Results: Our results showed that n-hexane and chloroform extracts had greatest toxic effects on gastric cancer cells with IC50 values of $6.4{\mu}g/ml$ and $4.6{\mu}g/ml$, respectively, after 72 h. Cell cycle analysis revealed that the population of treated cells in the G1 phase was increased in comparison to controls. Cellular morphological changes indicated induction of apoptosis. In addition, mRNA expression levels of Bax and caspase-3 were increased, and of bcl-2 survivin, VEGF, c-myc and cyclin D1 were decreased. Conclusion: Our study results suggest that D. glabrum has cytotoxic effects on AGS cells, characterized by enhanced apoptosis, reduced cell viability and arrest of cell cycling.