|Application:||Gene expression microarray analysis|
|Number of samples:||12|
|Release date:||Sep 29 2011|
|Last update date:||Oct 29 2018|
|Diseases:||Muscular Dystrophies, Muscular Dystrophy, Facioscapulohumeral, Chronobiology Disorders|
|Dataset link||Expression profiling FSHD vs. control myoblasts and myotubes|
For the microarray, two normal-control myoblast-myotube sample pairs and one disease-control pair (sporadic inclusion body myositis; IBM), which are previously described samples GSM443914_CM33j_ Undiff_06042009_, GSM443913_CM33j_Diff_06042009_, GSM443916_CM01154_002_Undiff, GSM443915_CM01154_002_Diff_06012009, GSM443918_CM01201_001_Undiff_06012009, GSM443917_CM01201_001_Diff_06012009, were compared to three myoblast-myotube sample pairs generated from moderately affected skeletal muscle from patients with molecularly confirmed FSHD. Myoblasts were harvested at about 70% confluency, and myotubes were harvested at maximal myotube formation, 5 or 7 days of differentiation. The control myoblast and myotube biological replicates 1-3 were from a normal-control 23 year-old male (quadriceps biopsy), a normal-control 27 year-old female (quadriceps biopsy), and a 74 year-old female (deltoid biopsy), respectively. The FSHD myoblast biological replicates 1-3 were from a 45 year-old female (quadriceps biopsy), 22 year-old female (orthopedic scapular fixation rhomboid tissue), and 13 year-old male (quadriceps biopsy) and contained six, five, and three copies of the D4Z4 repeat unit in their disease-linked arrays, respectively. The FSHD myotube biological replicates 1-3 were from the same batch of cells as for the FSHD myoblasts with the exception of biological replicate 3, which was from a 14 year-old female (deltoid biopsy, 2 copies of the disease-linked D4Z4 repeat unit). For qRT-PCR validation, 4 - 8 FSHD myoblast or myotube samples from different patients with a molecularly confirmed diagnosis were compared to 4 - 8 analogous control samples from different unaffected individuals. Most of these were from quadriceps biopsies, and all but one had not been used on the microarray. Methods for generation of myoblast cell strains, propagation of myoblasts, and induction of differentiation by serum limitation were previously described (http://genome.ucsc.edu/ENCODE/protocols/cell/human/HSMM_HSMMtube_Crawford_protocol.pdf). Each batch of myoblasts and myotubes was checked by immunocytochemical staining for the quality of the cell preparation. The myoblast preparations for the microarray or qRT-PCR validation had >85% desmin-positive cells as determined with an antibody preparation confirmed to have selectivity for myogenic cells (Labvision/Thermo Scientific) using DAPI counterstaining. The myotube preparations had >70% of the nuclei in desmin-positive cells containing more than 2 nuclei per cell. Five of the six preparations for the microarray analysis (all but FM1795) were also immunostained with a monoclonal antibody to myosin heavy chain (MF20, a gift of Stephen D. Hauschka) and had >70% of the nuclei in multinucleated myosin heavy chain-positive cells.