Comment[ArrayExpressAccession] E-GEOD-56976 MAGE-TAB Version 1.1 Public Release Date 2014-04-23 Investigation Title Development of gene expression signatures for ANP in HUVEC Comment[Submitted Name] Development of gene expression signatures for ANP in HUVEC Experiment Description Analysis of HUVEC treated with ANP. ANP is a cardiac hormone, binding to the guanylate cyclase-A (GC-A) receptor with a major role in cardiovascular homeostatic mechanisms. Results provide insight into the molecular mechanisms of ANP in vascular endothelial cell. ANP induced gene expression in HUVEC was measured at 0, 1 and 6 hours. Four independent experiments were performed at each time (0, 1 or 6 hours) for each experiment. Term Source Name ArrayExpress EFO Term Source File http://www.ebi.ac.uk/arrayexpress/ http://www.ebi.ac.uk/efo/efo.owl Person Last Name Nojiri Hasegawa Person First Name Takashi Takeshi Person Email nojirit@thoracic.med.osaka-u.ac.jp Person Affiliation National Cerebral and Cardiovascular Center Research Institute Person Phone +81-6-6833-5012 Person Address Biochemistry, National Cerebral and Cardiovascular Center Research Institute, 5-7-1, Fujishirodai, Suita, Osaka, Japan Person Roles submitter Protocol Name P-GSE56976-1 P-GSE56976-5 P-GSE56976-6 P-GSE56976-2 P-GSE56976-3 P-GSE56976-4 P-GSE56976-7 Protocol Description Intensities were extracted with Agilent Feature Extraction software ver.10.7.3.1. Data analyses were conducted with Agilent GeneSpring GX software ver.12.5. ID_REF = VALUE = Normalized signal intensity Cy3 labeled cRNA was prepared from 200 ng RNA using the Agilent Low Input Quick Amp Labeling Kit according to the manufacturer's instructions, followed by Qiagen RNAeasy column purification. Dye incorporation and cRNA yield were checked with the NanoDrop ND-1000 Spectrophotometer. 1.65ug of Cy3-labelled cRNA (specific activity >10.0 pmol Cy3/ug cRNA) was fragmented at 60C for 30 minutes in a reaction volume of 55ul containing 25x Agilent GE fragmentation buffer and 10x AgilentGE blocking agent following the manufacturers instructions. On completion of the fragmentation reaction, 55ul of 2x Agilent GE hybridization buffer was added to the fragmentation mixture and hybridized to Agilent Whole Human Genome Oligo Microarrays (G4112A) for 17 hours at 65C in a rotating Agilent hybridization oven. After hybridization, microarrays were washed 1 minute at room temperature with GE Wash Buffer 1 (Agilent) and 1 minute with 37C GE Wash buffer 2 (Agilent). HUVECs are incubated with 0.1ŒºM ANP or vehicle for 0, 1 and 6 hours. HUVEC was purchased from Lonza (Walkersville, MD, USA), then maintained in EGM-2 according to the manufacturer's instructions at 37 ∫C in a humidified incubator with 5% CO2. RNA are extracted from HUVEC using the Qiagen RNeasy Mini Kit. Slides were scanned immediately after washing on the Agilent DNA Microarray Scanner (G2505B) using one color scan setting for 4x44k array slides (Scan resolution 5µm, Dye channel is set to Green and Green PMT is set to 100%). Protocol Type normalization data transformation protocol labelling protocol hybridization protocol sample treatment protocol growth protocol nucleic acid extraction protocol array scanning protocol Experimental Factor Name compound dose time Experimental Factor Type compound dose time Comment[SecondaryAccession] GSE56976 Comment[GEOReleaseDate] 2014-04-23 Comment[ArrayExpressSubmissionDate] 2014-04-22 Comment[GEOLastUpdateDate] 2014-04-23 Comment[AEExperimentType] transcription profiling by array Pubmed ID 25775533 Publication DOI 10.1073/pnas.1417273112 Publication Title Atrial natriuretic peptide prevents cancer metastasis through vascular endothelial cells Publication Author List Nojiri T, Hosoda H, Tokudome T, Miura K, Ishikane S, Otani K, Kishimoto I, Shintani Y, Inoue M, Kimura T, Sawabata N, Minami M, Nakagiri T, Funaki S, Takeuchi Y, Maeda H, Kidoya H, Kiyonari H, Shioi G, Arai Y, Hasegawa T, Takakura N, Hori M, Ohno Y, Miyazato M, Mochizuki N, Okumura M, Kangawa K SDRF File E-GEOD-56976.sdrf.txt