Comment[ArrayExpressAccession]	E-GEOD-49858
MAGE-TAB Version	1.1					
Public Release Date	2013-08-14
Investigation Title	Gene expression of SCP-1 cells after incubation with conditioned medium of the breast carcinoma cell line MCF-7.					
Comment[Submitted Name]	Gene expression of SCP-1 cells after incubation with conditioned medium of the breast carcinoma cell line MCF-7.
Experiment Description	Disseminated tumor cells (DTCs) in the bone marrow can be detected in patients with solid tumors early on in disease progression. Via interaction with mesenchymal stromal cells (MSCs) these tumor cells may interfere with hematopoiesis. Using appropriate co-culture models, we investigated whether DTCs can change the bone marrow microenvironment by modulating MSC function with a special emphasis on their chemoattractive activity towards hematopoietic stem and progenitor cells (HSPCs). Human bone marrow derived MSCs as well as an immortalised MSC line (SCP-1) were co-cultured with MCF-7, MDA-MB231 breast carcinoma or MCF-10A non-malignant breast epithelial cells or their conditioned medium. Gene expression analysis of SCP-1 cells cultured with MCF-7 conditioned medium revealed SDF-1/CXCL12 as one of the significantly downregulated genes. Both tumor cell lines caused an inhibited SDF-1 promoter activity in SCP-1 cells, whereby MCF-7 medium decreased it to 77% and MDA-MB231 to 47%. Moreover, the SDF-1 mRNA and protein levels were significantly reduced. As a functional consequence of lower SDF-1 levels, we detected a decreased trans-well migration potential of CD34+ HSPC to MSC/tumor cell co-cultures or conditioned medium. The specificity of this chemokine mediated effect was confirmed by blocking studies with the CXCR4 antagonist AMD3100. Downregulation of SP1 transcription factor and increased miR23a levels in MSCs after contact with tumor cell medium as well as an enhanced TGFb1 expression were identified as potential molecular regulators of SDF-1 activity in MSCs. We propose an additional mechanism by which tumor cells affect the niche environment of HSPCs and therefore negatively impact hematopoiesis. Gene expression of human immortalized mesenchymal stromal cells (SCP-1) was investigated after incubation with conditioned medium of the breast carcinoma cell line MCF-7 for 24, 48, and 72 hours. Three independent experiments were performed at each time.					
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	Wobus	Wobus	BornhM-CM-$user			
Person First Name	Manja	Manja	Martin			
Person Email	manja.wobus@uniklinikum-dresden.de					
Person Affiliation	University Hospital Dresden					
Person Address	Hematology/Oncology, University Hospital Dresden, Fetscherstr. 74, Dresden, Germany					
Person Roles	submitter					
Protocol Name	P-GSE49858-1	P-GSE49858-4	P-GSE49858-5	P-GSE49858-2	P-GSE49858-3	P-GSE49858-6
Protocol Description	The Agilent Feature Extraction Software (FES 10.5.1.1 ) was used to read out and process the microarray image files. ID_REF =  VALUE = median normalized signal intensities	1 M-5g of each RNA was used as template to produce Cy3- labeled cRNA. The RNA samples were amplified and labeled using the Agilent Quick Amp Labeling Kit (Agilent Technologies) following the manufacturerM-bM-^@M-^Ys protocol.	Cy3 labeled cRNAs were hybridized overnight (17 hours, 65M-0C) to an Agilent Whole Human Genome Oligo Microarrays (44K) using AgilentM-bM-^@M-^Ys recommended hybridization chamber and oven.Finally, microarrays were washed once with wash buffer 1 for 1 min at RT followed by a second wash with wash buffer 2 for 1 min at 37M-0C and a final wash step with acetonitrile for 30 sec at RT.	Human immortalized mesenchymal stromal cells (SCP-1) were incubated with conditioned medium of the breast carcinoma cell line MCF-7 for 24, 48, and 72 hours at 37 M-:C in a humidified incubator with 5% CO2.	RNA was prepared using standard RNA extraction protocols (Trizol). Quality was checked via the Agilent 2100 Bioanalyzer platform.	Fluorescence signals of the hybridized Agilent Microarrays were detected using AgilentM-bM-^@M-^Ys Microarray Scanner System (G2505C, Agilent Technologies, Palo Alto, USA).
Protocol Type	normalization data transformation protocol	labelling protocol	hybridization protocol	sample treatment protocol	nucleic acid extraction protocol	array scanning protocol
Experimental Factor Name	SAMPLE DAY	CO-CULTURED MEDIUM				
Experimental Factor Type	sample day	co-cultured medium				
Comment[SecondaryAccession]	GSE49858					
Comment[GEOReleaseDate]	2013-08-14					
Comment[ArrayExpressSubmissionDate]	2013-08-13					
Comment[GEOLastUpdateDate]	2013-08-14					
Comment[AEExperimentType]	transcription profiling by array					
SDRF File	E-GEOD-49858.sdrf.txt