Investigation Title	Transcription profiling of pig coronary arterial endothelial cells by multiple passaging in vitro	
Comment[Submitted Name]	To study the genomic changes in senescent porcine coronary arterial endothelial cells by multiple passaging in vitro	
Experimental Design	unknown_experiment_design_type	transcription profiling by array	
Experimental Design Term Source REF		EFO	
Comment[AEMIAMESCORE]	4	
Comment[ArrayExpressReleaseDate]	2008-06-16	
Comment[SecondaryAccession]	GSE8974	
Comment[ArrayExpressAccession]	E-GEOD-8974	
Comment[MAGETAB TimeStamp_Version]	2010-08-09 12:05:27 Last Changed Rev: 13058	
Experimental Factor Name	
Experimental Factor Type	
Experimental Factor Term Source REF	
Person Last Name	Lee	
Person First Name	Mary	
Person Mid Initials	
Person Email	leemary@hkucc.hku.hk	
Person Phone	
Person Fax	
Person Address	L2, Dept of Pharmacology, The University of Hong Kong, 21 Sassoon Road , Hong Kong , 852, Hong Kong, China	
Person Affiliation	The University of Hong Kong	
Person Roles	submitter	
Person Roles Term Source REF	The MGED Ontology	
Quality Control Type	
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Replicate Type	
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Normalization Type	
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Date of Experiment	
Public Release Date	2008-06-16	
PubMed ID	
Publication DOI	
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Publication Title	
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Publication Status Term Source REF	
Experiment Description	Nitric oxide helps to prevent endothelial dysfunction and senescence. This study aimed to define genomic and proteomic changes in cultured porcine senescent endothelial cells and their resemblance with those observed in regenerated endothelial cells. Senescent endothelial cells were produced by passaging primary porcine coronary arterial endothelial cells until passage four. The protein presence of endothelial nitric oxide synthase, cyclic GMP levels [basal and during stimulation (bradykinin and A23187)], were reduced. The mRNA expression level was measured by microarray assays. Genes related to oxidative stress [superoxide dismutase (MnSOD), glutathione peroxidase 3, glutathione S-transferase M1] were downregulated, extracellular matrix components (type III collagen, thrombospondin 1 and 3, transforming growth factor ?) upregulated and nuclear factor kappa B (NF?B)-signaling pathway [IkappaB, TNF receptor-associated factor 1 and 5 (TRAF1 and 5)] activated in senescent cells. The differential gene expression of MnSOD and TRAF5 was confirmed at the protein level by Western blotting and biochemical assay (MnSOD). The basal and stimulated (by tumor necrosis factor-?)?levels of NF?B were augmented as demonstrated by electrophoretic mobility shift assay. In summary, cultured senescent endothelial cells exhibit reduced nitric oxide production, and decreased antioxidative, proliferative capacities, augmented expression of extracellular matrix components and activation of NF?B. These molecular changes do not exactly mimick those occurring during endothelial regeneration in vivo. Experiment Overall Design: Totally 8 samples were analyzed with 4 replicates each for control (cells at passage one) and test (cells at passage four) samples.	
Protocol Name	P-G8974-1	P-G8974-2	P-G8974-4	P-G8974-3	Affymetrix:Protocol:Hybridization-Unknown	P-AFFY-6	Affymetrix:Protocol:ExpressionStat	
Protocol Type	grow	specified_biomaterial_action	nucleic_acid_extraction	labeling	hybridization	feature_extraction	bioassay_data_transformation	
Protocol Description	porcine coronary arterial endothelial cells were brought to primary culture. Cultured endothelial cells for 1 weeks produced cells at passage one (control cells). Serial passaging was performed on a weekly basis until cells at passage four (senescent cells) were reached (test cells).	Cultured endothelial cells for 1 weeks produced cells at passage one (control cells). Serial passaging was performed on a weekly basis until cells at passage four (senescent cells) were reached (test cells).  Both control and test cells at basal condition were collected for total RNA extraction for microarray analysis.	Total RNA was extracted with Rneasy mini kit (Qiagen) performed according to the manufacturer's instructions.	Biotinylated cRNA were prepared according to the standard Affymetrix protocol from 1ug total RNA.	Title: Affymetrix Generic Hybridization. Description: 	Title: Affymetrix CEL analysis. Description: 	Title: Affymetrix CHP Analysis (ExpressionStat). Description: 	
Protocol Parameters	
Protocol Hardware	
Protocol Software					MicroArraySuite 5.0	MicroArraySuite 5.0	MicroArraySuite 5.0	
Protocol Contact	
Protocol Term Source REF					mo		The MGED Ontology	
SDRF File	E-GEOD-8974.sdrf.txt	
Term Source Name	ncbitax		The MGED Ontology	ArrayExpress	EFO	The MGED Ontology	mo	
Term Source File	http://www.ncbi.nlm.nih.gov/Taxonomy/taxonomyhome.html		http://mged.sourceforge.net/ontologies/MGEDontology.php	http://www.ebi.ac.uk/arrayexpress	http://www.ebi.ac.uk/efo/	http://mged.sourceforge.net/ontologies/MGEDontology.php	http://mged.sourceforge.net/ontologies/MGEDontology.php	
Term Source Version