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Sample GSM1108303 Query DataSets for GSM1108303
Status Public on May 21, 2018
Title HMVEC-L_TNF-α-induced premature senescent_rep2
Sample type RNA
 
Source name HMVEC-L_PD < 5; treated with 20ng/ml TNF-α_replicate 2
Organism Homo sapiens
Characteristics cell type: TNF-α-induced premature senescent HMVEC-L
treatment: TNFalpha
tissue: pulmonary microvascular
Treatment protocol TNF-α (Sigma-Aldrich, USA) was used to induce premature senescence in HMVEC-L. The cells were subjected to fifteen successive sub-lethal treatments of 20 ng/ml TNF-α in EGM-2MV containing 5% FBS, with one treatment per day for fifteen consecutive days. Control cultures were maintained in parallel without TNF-α.
Growth protocol HMVEC-L was purchased from Lonza (Walkersville, MD, USA) and maintained in Microvascular Endothelial Cell Growth Medium-2 (EGM-2MV) (Lonza), at 37oC in a humidified atmosphere of 95% air/5% CO2. Young and RS cells used in experiments are cells with PD < 5 and PD > 20 respectively.
Extracted molecule total RNA
Extraction protocol Total RNAs were isolated from cells using miRNeasy mini kit (Qiagen) according to manufacturer’s protocol. RNA was quantified using Nanophotometer (Implen GmbH, Germany). RNA integrity was accessed using Agilent 2100 Bioanalyzer (Agilent Technologies Inc., Palo Alto, CA, USA).
Label Cy3
Label protocol Total RNAs (50ng) were labeled with Cyanine 3-CTP using Low Input Quick Amp Labeling Kit (Agilent Technologies Inc., USA) according to manufacturer’s protocol, followed by RNAeasy mini column purification (Qiagen, Germany). Dye incorporation and cRNA yield were checked with the Nanophotometer (Implen GmbH, Germany).
 
Hybridization protocol 1.65 μg of Cy3-labelled cRNA (specific activity > 6.0 pmol Cy3/μg cRNA) was fragmented at 60°C for 30 minutes in a reaction volume of 55μl containing 25x Agilent fragmentation buffer and 10x Agilent blocking agent following the manufacturers instructions. On completion of the fragmentation reaction, 55 μl of 2x Agilent hybridization buffer was added to the fragmentation mixture and hybridized to Agilent Whole Human Genome Microarrays (G4112F) for 17 hours at 65°C in a rotating Agilent hybridization oven. After hybridization, microarrays were washed 1 minute with moderate stirring at room temperature with GE Wash Buffer 1 (Agilent) and 1 minute with moderate stirring at 37°C GE Wash buffer 2 (Agilent).
Scan protocol Slides were scanned immediately after washing on the Agilent Microarray Scanner C using scan one color setting for 4x44k array slide (Scan Area 61x21.6 mm, Scan resolution 5μm, Dye channel is set to Green and output is 20 bit TIFF).
Description Gene expression of TNF-α-induced premature senescent HMVEC-L
Data processing The scanned images were analyzed with Feature Extraction Software 10.5.1.1 (Agilent) using protocol: GE1_105_Dec08 and Grid: 014850_D_20070820.
 
Submission date Mar 27, 2013
Last update date May 21, 2018
Contact name Pooi-Fong Wong
Organization name University of malaya
Department Pharmacology
Street address Faculty of Medicine
City KUALA LUMPUR
ZIP/Postal code 50603
Country Malaysia
 
Platform ID GPL4133
Series (2)
GSE45540 Gene signature of young, replicative and TNF-α-induced premature senescent of human microvascular endothelial cells-lung (HMVEC-L)
GSE45541 Young, replicative and TNF-a-induced premature senescent of human microvascular endothelial cells-lung (HMVEC-L)

Data table header descriptions
ID_REF
VALUE Agilent Feature Extraction default gProcessedSignal

Data table
ID_REF VALUE
1 4.87E+04
2 1.97E+00
3 4.16E+00
4 1.97E+00
5 1.97E+00
6 1.97E+00
7 1.97E+00
8 1.97E+00
9 1.97E+00
10 1.98E+00
11 1.98E+00
12 3.55E+01
13 3.76E+00
14 6.88E+01
15 1.54E+01
16 5.86E+03
17 3.41E+01
18 8.26E+01
19 2.29E+04
20 1.19E+01

Total number of rows: 45015

Table truncated, full table size 648 Kbytes.




Supplementary file Size Download File type/resource
GSM1108303_US91203660_251485045074_S01_GE1_105_Dec08_1_2.txt.gz 2.2 Mb (ftp)(http) TXT
Processed data included within Sample table
Processed data provided as supplementary file

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