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Sample GSM1976989 Query DataSets for GSM1976989
Status Public on Feb 01, 2016
Title AV281
Sample type RNA
 
Channel 1
Source name Reference sample
Organism Homo sapiens
Characteristics type: Reference sample
Treatment protocol NA
Growth protocol Human primary CAMs and ATMs had previously been generated from patients undergoing surgery for gastric or esophagal cancer. Normal myofibroblasts from both parts (antrum, corpus) of healthy stomach and esophagus had been generated from deceased transplant donors. The patients, and the myofibroblasts obtained from them have previously been reported (Czepan M et al., Pflugers Archiv : European journal of physiology 463: 459-475, 2012; Holmberg C et al., Carcinogenesis 33: 1553-1562, 2012). This work had been approved by the Ethics Committee of the University of Szeged, Hungary. Myofibroblasts were cultured as previously described and were used between passages 3 and 10 (Holmberg C et al., Carcinogenesis 33: 1553-1562, 2012). Tumor and adjacent tissues were characterized using the TNM classification (Supplementary Methods, available at Carcinogenesis Online -Holmberg. C., et al. Carcinogenesis ; 33(8): 1553–1562, 2012) for gastric cancer (Sobin L.H., et al. Wiley-Blackwell; Chichester,2009).
Extracted molecule total RNA
Extraction protocol Myofibroblasts (1.5 million) were cultured overnight in 75cm dishes and total RNA was extracted using miRNeasy according the manufacturer’s instructions (Qiagen, Crawley, UK)
Label Hy5
Label protocol Total RNA and reference samples were labelled with Hy3TM and Hy5TM fluorescence labels respectively (Exiqon Services, Vedbaek, Denmark).
 
Channel 2
Source name Gastric
Organism Homo sapiens
Characteristics type: NTM-C
Treatment protocol NA
Growth protocol Human primary CAMs and ATMs had previously been generated from patients undergoing surgery for gastric or esophagal cancer. Normal myofibroblasts from both parts (antrum, corpus) of healthy stomach and esophagus had been generated from deceased transplant donors. The patients, and the myofibroblasts obtained from them have previously been reported (Czepan M et al., Pflugers Archiv : European journal of physiology 463: 459-475, 2012; Holmberg C et al., Carcinogenesis 33: 1553-1562, 2012). This work had been approved by the Ethics Committee of the University of Szeged, Hungary. Myofibroblasts were cultured as previously described and were used between passages 3 and 10 (Holmberg C et al., Carcinogenesis 33: 1553-1562, 2012). Tumor and adjacent tissues were characterized using the TNM classification (Supplementary Methods, available at Carcinogenesis Online -Holmberg. C., et al. Carcinogenesis ; 33(8): 1553–1562, 2012) for gastric cancer (Sobin L.H., et al. Wiley-Blackwell; Chichester,2009).
Extracted molecule total RNA
Extraction protocol Myofibroblasts (1.5 million) were cultured overnight in 75cm dishes and total RNA was extracted using miRNeasy according the manufacturer’s instructions (Qiagen, Crawley, UK)
Label Hy3
Label protocol Total RNA and reference samples were labelled with Hy3TM and Hy5TM fluorescence labels respectively (Exiqon Services, Vedbaek, Denmark).
 
 
Hybridization protocol The Hy3TM labelled samples and Hy5TM labelled reference RNA samples were mixed pairwise and hybridised to the miRCURY TM LNA Array ver5 that contains capture probes targeting all miRNAs for human, mouse or rat registered in the miRBase version 14.0
Scan protocol Microarrays were scanned using an Agilent G2565BA Microarray Scanner System (Agilent Technologies, Santa Clara, CA, USA). Image analysis was carried out using ImaGene 8.0 software (BioDiscovery, Hawthorne, CA, USA).
Description Slide 34
Total RNA and reference samples were labelled with Hy3TM and Hy5TM fluorescence labels respectively (Exiqon Services, Vedbaek, Denmark)
Cy3 File: 1_Exiqon_14172274_S01.txt
Cy5 File: 0_Exiqon_14172274_S01.txt
Data processing Log2 transformed median Hy3/Hy5 ratios were calculated using median signals on capture probe replicates and the average log2 median ratios across samples taken. The difference in log2 median ratio (LMR) between sample groups was calculated and subjected to Student t-test (two tailed) or ANOVA using Bonferroni correction. The miRNA data resource was accessed at www.mirbase.org. Principal component analysis (PCA) and hierarchical clustering were performed using DNA Chip Analyser (dChip0 software, www.dchip.org). Validated target miRNAs were obtained from TarBase (Vergoulis T et al.,; Nucleic acids research 40: D222-229, 2012), miRecords (Xiao F et al.,; Nucleic acids research 37: D105-110, 2009) and MetaCore databases (GeneGo, ST Joseph, MI, USA). Network enrichment analysis of miRNA targets was performed using MetaCore and statistically significant networks and pathways were identified at p<0.05 with 5% FDR. Predicted targets of miR-181d were retrieved from TargetScan release 5.1 (Friedman RC et al.,; Genome Res 19: 92-105, 2009) using a context score below -0.2 for high efficacy targets. Gene expression profiles of gastric myofibroblasts, previously carried out using GeneChip Human Genome U133 Plus 2.0 array and deposited at http://0-www-ncbi-nlm-nih-gov.brum.beds.ac.uk/geo/query/acc.cgi?acc=GSE44740 (Balabanova S et al., Carcinogenesis 35: 1798-1806, 2014) were used to determine the abundance of potential miRNA regulators and target transcripts.
Signals were background corrected (i.e. Normexp with offset value 10) and normalised using the globally weighted scatterplot smoothing (LOWESS) regression algorithm.
 
Submission date Dec 21, 2015
Last update date Feb 01, 2016
Contact name Liyi Wang
E-mail(s) liyi@liverpool.ac.uk
Organization name University of Liverpool
Department Department of Physiology
Lab Green block, Sherrington Building
Street address Crown Street
City Liverpool
State/province Merseyside
ZIP/Postal code L693BX
Country United Kingdom
 
Platform ID GPL17107
Series (2)
GSE76218 miRNA analysis in myofibroblats derived from normal stomach, both antrum (A) and corpus (C) separately and gastric cancers
GSE76221 miRNA profiling of oesophageal and gastric myofibroblast in health and disease

Supplementary file Size Download File type/resource
GSM1976989_0_Exiqon_14172274_S01.txt.gz 812.4 Kb (ftp)(http) TXT
GSM1976989_1_Exiqon_14172274_S01.txt.gz 901.9 Kb (ftp)(http) TXT
Processed data are available on Series record

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