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Sample GSM322614 Query DataSets for GSM322614
Status Public on Sep 20, 2008
Title Chondrocytes stimulated with diclofenac treated RASF rep 2
Sample type RNA
 
Source name Human chondrocytes isolated from lateral condyle of femur bone in culture from donor pool 5, stimulated with supernatant from diclofenac treated RASF
Organism Homo sapiens
Characteristics Human chondrocytes cultured in RPMI 1640 medium supplemented with 10% human serum, 100ng/ml amphotericin B, 100U/ml penicillin and 100µg/ml streptomycin for 2 passages, 3D cultured for 14 days in alginate beads in presence of 170µM L-ascorbic acid 2-phosphate and subsequently stimulated for 48h with supernatant of RASF cultured in the same medium and treated with diclofenac for 48h
Treatment protocol Human chondrocytes were isolated by enzymatic digestion (1 U collagenase P, 333 U collagenase II and 33 U hyaluronidase), passage 2 chondrocytes were encapsulated in alginate beads (20 Mio cells/ml in 1,5% (w/v) alginate) and stimulated with conditioned medium of NDSF, RASF or treated RASF
Growth protocol Human chondrocytes obtained from lateral condyle of femur bones of 6 donors post mortem were amplified by cultivation in RPMI 1640 medium supplemented with 10% human serum, 100ng/ml amphotericin B, 100U/ml penicillin and 100µg/ml streptomycin for 2 passages, subsequently 3D cultured in alginate beads for 14 days in presence of 170µM L-ascorbic acid 2-phosphate and finally stimulated with conditioned medium of RASF, NDSF, or treated RASF
Extracted molecule total RNA
Extraction protocol Prior to RNA extraction, alginate beads were solubilized on ice in 55mM sodium citrate, 30mM EDTA and 150mM NaCl and cells were centrifuged at 800g for 5min (4°C). Total RNA isolation was proceeded according to the manufacturer’s protocol. Additionally, a proteinase K and DNase I digestion were performed. Isolation of total RNA was done for the different stimulated donor chondrocytes separately. Afterwards, equal amounts of total RNA from three stimulated donor chondrocytes were pooled, yielding different experimental groups of chondrocytes stimulated with supernatant of NDSF, RASF or treated RASF
Label biotin
Label protocol 2.5 µg of total RNA was amplified and labeled using the GeneChip® one-cycle target labeling and control reagents (Affymetrix)
 
Hybridization protocol 50µg/ml of labeled cRNA was hybridized at 45°C for 16h using the Affymetrix GeneChip hybridization oven. Washing and staining was performed on the Affymetrix Fluidics 450 using the EukGE-WS2v4 protocol.
Scan protocol GeneChips were scanned using the Affymetrix GeneChip Scanner 3000
Description Gene expression data of human chondrocytes stimulated with supernatant of RASF treated for 48h with diclofenac
Data processing Data were processed with GCOS 1.4 (TGT = 150) and subsequently imported and further analyzed in the BioRetis online database (www.bioretis.de). Additionally, expression data was processed with Robust Multi-Array Analysis version 0.4a7 (RMA)
 
Submission date Sep 19, 2008
Last update date Sep 19, 2008
Contact name Thomas Häupl
E-mail(s) thomas.haeupl@charite.de
Phone +49 30 450513293
Organization name Charité
Department Rheumatology
Street address Charitéplatz 1
City Berlin
ZIP/Postal code 10117
Country Germany
 
Platform ID GPL96
Series (1)
GSE12860 Antirheumatic Drug Response in Human Chondrocytes: Potential Molecular Targets to Stimulate Cartilage Regeneration

Data table header descriptions
ID_REF
VALUE RMA log2 transformed signals

Data table
ID_REF VALUE
1007_s_at 7.221074853
1053_at 5.512821719
117_at 5.746202949
121_at 8.253299602
1255_g_at 4.417733996
1294_at 6.532420383
1316_at 5.069663918
1320_at 5.195486498
1405_i_at 5.791311774
1431_at 4.451785639
1438_at 6.01137553
1487_at 6.98856328
1494_f_at 6.271548516
1598_g_at 8.930087225
160020_at 10.66944088
1729_at 6.928768713
177_at 6.13051841
1773_at 5.845617332
179_at 8.557817198
1861_at 5.742920988

Total number of rows: 22283

Table truncated, full table size 496 Kbytes.




Supplementary file Size Download File type/resource
GSM322614.CEL.gz 2.2 Mb (ftp)(http) CEL
GSM322614.EXP.gz 513 b (ftp)(http) EXP
GSM322614_MAS5_data.txt.gz 145.0 Kb (ftp)(http) TXT
Raw data provided as supplementary file
Processed data included within Sample table
Processed data provided as supplementary file

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