Levator Ani Satellite Cells (LASCs) treated with Nonsteroidal Androgen Receptor Agonist (NARA)

High regenerative capacity of adult skeletal muscle relies on a self-renewing depot of adult stem cells, termed muscle satellite cells (MSCs). A novel MSC line was isolated from the rat levator ani muscle and termed Levator Ani Satellite Cells (LASCs). Androgen, a known mediator of overall body composition and specifically skeletal muscle mass, has been shown to regulate MSCs. The use of non-steroidal androgen receptor agonists (NARA) aims to retain the beneficial influence of androgen on skeletal muscle, while circumventing undesirable cardiovascular and prostate-related side-effects. Primary objectives: 1) Identify biomarkers of satellite cell growth and differentiation, 2) Characterize a novel muscle satellite cell line, 3) Understand the effects of androgen (NARA) on rat satellite cell activation and recruitment. LASCs will be treated with one of the following conditions (N=4 for all groups): 1) 0.2% DMSO, 4 hours; 2) 10 nM NARA, 4 hours; 3) 0.2% DMSO, 48 hours; 4) 10nM NARA, 48 hours RNA was extracted using the PerfectPure RNA isolation kit (5 Prime, Gaithersburg, MD). Five micrograms of total RNA from each sample was labeled and hybridized to Affymetrix Rat Genome 230_2.0 microarrays according to Affymetrix protocols. Each microarray was washed and stained using an Affymetrix Fluidics Station 400 and scanned in an Affymetrix confocal GeneArray scanner. Affymetrix MAS5.0 software was used to scale data to a target intensity of 1500 and calculate transcript abundance (signal). Microarray signal intensities were generated by Affymetrix Expression Console using the MAS5 algorithm with targeted trimmed mean signal set to 1500. The gene chips were checked for quality control by their detection calls, chip pairwise correlation, and principal components analysis (PCA). Three chips (GSM1550754, GSM1550757, and GSM1550758) were identified as outliers and excluded from analysis for obvious low correlation with and deviating on PCA plots from the rest of the chips.