Comment[ArrayExpressAccession]	E-GEOD-48227
MAGE-TAB Version	1.1						
Public Release Date	2013-06-25
Investigation Title	Differential expression of microRNAs in adipose stem cells						
Comment[Submitted Name]	Differential expression of microRNAs in adipose stem cells
Experiment Description	Human adipose derived stem cells (hADSCs) are an attractive multipotent stem cell source with therapeutic applicability in diverse fields for the repair and regeneration of acute and chronically damaged tissues.In the last few years there has been increased interest in hADSCs for tissue engineering applications; however, the molecular mechanisms especially the microRNAs underlying the regulation of hADSCs proliferation and differentiation are not fully understood. To this end, we profiled the global microRNA expression in hADSCs, fibroblasts and endothelial cells by using Exqion mercury LNATM  (Exqion Inc) To identify the microRNAs significantly expressed in human adipose derived stem cells, eight samples were analyzed using Exqion mercury LNATM  (Exqion Inc) including six hADSCs samples and human endothelial and fibroblast cells as controls.						
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	Li	Liu	Jiang	Chen	Shi	Li	Zhou
Person First Name	Min	Xiaoqing	Cizhong	Guangfeng	Xiujuan	Min	Qing
Person Email	minlicq@gmail.com						
Person Affiliation	Tongji University						
Person Address	Life Sciences and Technology, Tongji University, 50 Chifeng Rd., Yangpu District, Shanghai, China						
Person Roles	submitter						
Protocol Name	P-GSE48227-3	P-GSE48227-4	P-GSE48227-1	P-GSE48227-2	P-GSE48227-5		
Protocol Description	The miRCURYM-bM-^DM-" Hy3M-bM-^DM-"/Hy5M-bM-^DM-" Power labeling kit (Exiqon, Vedbaek, Denmark) was used according to the manufacturerM-bM-^@M-^Ys guideline for miRNA labelling. One microgram of each sample was 3'-end-labeled with Hy3TM fluorescent label, using T4 RNA ligase by the following procedure: RNA in 2.0 M-NM-<L of water was combined with 1.0 M-NM-<L of CIP buffer and CIP (Exiqon). The mixture was incubated for 30 min at 37M-0C, and was terminated by incubation for 5 min at 95M-0C. Then 3.0 M-NM-<L of labeling buffer, 1.5 M-NM-<L of fluorescent label (Hy3TM), 2.0 M-NM-<L of DMSO, 2.0 M-NM-<L of labeling enzyme were added into the mixture. The labeling reaction was incubated for 1 h at 16M-0C, and terminated by incubation for 15 min at 65M-0C.	the Hy3TM-labeled samples were hybridized on the miRCURYTM LNA 5th generation Array (Exiqon) according to array manual. The total 25 M-NM-<L mixture from Hy3TM-labeled samples with 25 M-NM-<L hybridization buffer were first denatured for 2 min at 95M-0C, incubated on ice for 2 min and then hybridized to the microarray for 16 M-bM-^@M-^S 20 h at 56M-0C in a 12-Bay Hybridization Systems (Hybridization System - Nimblegen Systems, Inc., Madison, WI, USA), which provides an active mixing action and constant incubation temperature to improve hybridization uniformity and enhance signal. Following hybridization, the slides were achieved, washed several times using Wash buffer kit (Exiqon), and finally dried by centrifugation for 5 min at 400 rpm.	Human umbilical vein endothelial cells and human dermal fibroblasts cells were were cultured in DMEM/F12 medium supplemented with 10% FBS.Adipose tissues obtained from the patients should be washed 3 times by PBS with 1% penicillin/streptomycin and then carefully minced by sterile operation scissors. The minced tissues were then enzymatically dissociated for 45 minutes at 37 M-0C by 0.15% collagenase type I (GIBCO). Then the suspension was then neutralized with isometric culture medium and centrifuged at 500g for 5 minutes. The cell pellet was resuspended in DMEM/F12 medium (GIBCO) supplemented with 10% FBS (GIBCO) or 10% KSR (Invitrogen)at a density of 2x106 cell/mL. Cell cultures were maintained at 37 M-0C in a humidified incubator supplemented with 5% CO2. Passage 3 cells were used for whole human genome microarray.	Total RNA was isolated using TRIzol (Invitrogen) and miRNeasy mini kit (QIAGEN) according to manufacturerM-bM-^@M-^Ys instructions, which efficiently recovered all RNA species, including miRNAs. RNA quality and quantity was measured by using nanodrop spectrophotometer (ND-1000, Nanodrop Technologies) and RNA Integrity was determined by gel electrophoresis	Scanning is performed with the Axon GenePix 4000B microarray scanner. GenePix pro V6.0 is used to read the raw intensity of the image.		
Protocol Type	labelling protocol	hybridization protocol	growth protocol	nucleic acid extraction protocol	array scanning protocol		
Experimental Factor Name	CELL TYPE	AGE					
Experimental Factor Type	cell type	age					
Comment[SecondaryAccession]	GSE48227						
Comment[GEOReleaseDate]	2013-06-25						
Comment[ArrayExpressSubmissionDate]	2013-06-24						
Comment[GEOLastUpdateDate]	2013-06-26						
Comment[AEExperimentType]	transcription profiling by array						
Comment[AdditionalFile:Data1]	GSE48227.txt						
SDRF File	E-GEOD-48227.sdrf.txt