|
| Status |
Public on Oct 30, 2006 |
| Title |
Glucosamine and IL-1 beta treatment_a |
| Sample type |
RNA |
| |
|
| Source name |
rat chondrocyte cultured in presence of glucosamine and IL-1
|
| Organism |
Rattus norvegicus |
| Characteristics |
Genotype: Rattus norvegicus
Age: 2 months
|
| Treatment protocol |
Articular cartilage was isolated from the femoral heads of male Wistar rats under aseptic conditions (Charles River Laboratories). Chondrocytes were obtained by sequential digestion of the cartilage with pronase and type II collagenase. After filtration to remove tissue debris, the cells were cultured in 75-cm2 flasks in complete Dulbecco’s Modified Eagle Medium (DMEM; supplemented with 10% fetal bovine serum and 1% penicillin-streptomycin at 37°C in a humidified atmosphere containing 5% CO2.
|
| Growth protocol |
Experiments were performed with second-passage cultures whereby the cells from the large cultures were trypsinized, pooled and seeded into twenty 25 cm2 flasks. These were then 1/4 were cultured in presence of 20mM glucosamine for 6 hours then stimulated with IL-1 beta for 14 hours (10ng/ml) (n=4/group).
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Trizol extraction of total RNA was performed according to the manufacturer's instructions.
|
| Label |
streptavidin-phycoerythryn
|
| Label protocol |
Biotinylated cRNA were prepared according to the standard Affymetrix protocol from 3 ug total RNA (Expression Analysis Technical Manual, 2001, Affymetrix).
|
| |
|
| Hybridization protocol |
Following fragmentation, fifteen micrograms of adjusted cRNA from each sample was hybridized for 16 hours at 45°C to Affymetrix GeneChip Rat Genome 230 2.0 arrays.
|
| Scan protocol |
GeneChips were scanned using the Genearray Scanner, Agilent Technologies
|
| Description |
Gene expression data from rat chondrocyte cultured in presence of 20mM glucosamine
|
| Data processing |
Microarray Suite, version 5 (Affymetrix), was used to generate *.cel files, and a computer program (Probe Profiler, ver.1.3.11; Corimbia, Inc.) developed specifically for the GeneChip system (Affymetrix) was used to convert intensity data into quantitative estimates, globally scaled to 100, of gene expression for each probe set.
|
| |
|
| Submission date |
Oct 30, 2006 |
| Last update date |
Oct 30, 2006 |
| Contact name |
jean noel gouze |
| Organization name |
university of florida
|
| Department |
orthopaedics
|
| Lab |
gene therapy
|
| Street address |
1600 SW archer rd
|
| City |
gainesville |
| State/province |
FL |
| ZIP/Postal code |
32610 |
| Country |
USA |
| |
|
| Platform ID |
GPL1355 |
| Series (1) |
| GSE6119 |
Exogenous Glucosamine Globally Protects Chondrocytes from the Arthritogenic Effects of IL-1beta |
|
