|
| Status |
Public on Dec 01, 2015 |
| Title |
Palmitate rep1 vs. Control rep1 |
| Sample type |
RNA |
| |
|
| Channel 1 |
| Source name |
LHCN-M2 mytoubes
|
| Organism |
Homo sapiens |
| Characteristics |
treatment: 0.5 mM Palmitate
cell line: LHCN-M2
differentiation: 10 days post-differentiation induction
tissue: skeletal muscle
|
| Treatment protocol |
Sodium salts of palmitic and oleic acids were prepared in deionized water containing 1.2 equivalents of NaOH at 70ºC, until an optically clear dispersion was obtained. The fatty acid salt solution was added to cell culture media containing fatty acid-free bovine serum albumin (BSA) with continuous agitation. The fatty acid: BSA molar ratio were 5:1.
|
| Growth protocol |
Myoblasts were grown in DMEM/MEM199 (4:1) added with 15% fetal bovine serum (FBS), 16 mM Hepes, 0.03 μg/ml zinc sulfate, 1.4 μg/ml vitamin B12, 0.055 µg/ml dexamethasone , 0.025 μg/ml human hepatocyte growth factor and 2.5 ng/ml fibroblast growth factor. In confluent cultures, myoblast fusion was stimulated by incubating cells in the DMEM/MEM199 media supplemented with 0.005% FBS, 20 mM Hepes, 10 µg/ml insulin, 0.1 mg/ml apotransferrin and 0.055 µg/ml dexamethasone during 2 days. To further induce cell differentiation, media was then switched to DMEM/MEM199 supplemented with 0.005% FBS, 20 mM Hepes and 0.055 µg/ml dexamethasone, and the culture was continued for 8 additional days.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted from cultured LHCN-M2 myotubes with RNeasy kit (Qiagen, Valencia, CA, USA), with the same DNase treatment than in the RNeasy fibrous tissue kit, and homogenized with a Polytron.
|
| Label |
Cy5
|
| Label protocol |
Low Input Quick Amp Labeling Protocol v 6.5 (Agilent)
|
| |
|
| Channel 2 |
| Source name |
LHCN-M2 mytoubes
|
| Organism |
Homo sapiens |
| Characteristics |
treatment: Control
cell line: LHCN-M2
differentiation: 10 days post-differentiation induction
tissue: skeletal muscle
|
| Treatment protocol |
Sodium salts of palmitic and oleic acids were prepared in deionized water containing 1.2 equivalents of NaOH at 70ºC, until an optically clear dispersion was obtained. The fatty acid salt solution was added to cell culture media containing fatty acid-free bovine serum albumin (BSA) with continuous agitation. The fatty acid: BSA molar ratio were 5:1.
|
| Growth protocol |
Myoblasts were grown in DMEM/MEM199 (4:1) added with 15% fetal bovine serum (FBS), 16 mM Hepes, 0.03 μg/ml zinc sulfate, 1.4 μg/ml vitamin B12, 0.055 µg/ml dexamethasone , 0.025 μg/ml human hepatocyte growth factor and 2.5 ng/ml fibroblast growth factor. In confluent cultures, myoblast fusion was stimulated by incubating cells in the DMEM/MEM199 media supplemented with 0.005% FBS, 20 mM Hepes, 10 µg/ml insulin, 0.1 mg/ml apotransferrin and 0.055 µg/ml dexamethasone during 2 days. To further induce cell differentiation, media was then switched to DMEM/MEM199 supplemented with 0.005% FBS, 20 mM Hepes and 0.055 µg/ml dexamethasone, and the culture was continued for 8 additional days.
|
| Extracted molecule |
total RNA |
| Extraction protocol |
Total RNA was extracted from cultured LHCN-M2 myotubes with RNeasy kit (Qiagen, Valencia, CA, USA), with the same DNase treatment than in the RNeasy fibrous tissue kit, and homogenized with a Polytron.
|
| Label |
Cy3
|
| Label protocol |
Low Input Quick Amp Labeling Protocol v 6.5 (Agilent)
|
| |
|
| |
| Hybridization protocol |
Oligoarray control targets and hybridization buffer (Agilent In Situ Hybridization Kit Plus) were added, and samples were applied to microarrays enclosed in Agilent SureHyb-enabled hybridization chambers. After hybridization, slides were washed sequential
|
| Scan protocol |
Scanned on an Agilent G2505C scanner.
Images were quantified using Agilent Feature Extraction Software (version 10.7.3.1).
|
| Description |
Zhu, C. H et al. 2007. Cellular senescence in human myoblasts is overcome by human telomerase reverse transcriptase and cyclin-dependent kinase 4: consequences in aging muscle and therapeutic strategies for muscular dystrophies. Aging Cell 6: 515-523.
|
| Data processing |
Agilent Feature Extraction Software (v 10.7.3.1) was used for background subtraction and LOWESS normalization.
|
| |
|
| Submission date |
Dec 09, 2013 |
| Last update date |
Dec 01, 2015 |
| Contact name |
Anna Maria Gómez Foix |
| E-mail(s) |
agomezfoix@ub.edu
|
| Phone |
34-93-4039283
|
| Organization name |
Universitat de Barcelona
|
| Department |
Bioquímica i Biologia Molecular
|
| Lab |
Enginyeria Cel.lular i Teràpia Gènica
|
| Street address |
Diagonal 643
|
| City |
Barcelona |
| ZIP/Postal code |
08028 |
| Country |
Spain |
| |
|
| Platform ID |
GPL13607 |
| Series (1) |
| GSE53116 |
GENE TARGETS OF PALMITATE, OLEATE AND THEIR MIXTURE IN CULTURED LHCN-M2 MYOTUBES |
|
