Comparison of Gene expression profiling of granulosa cells treated with follicle stimulating hormone or constitutively active protein kinase A. Rattus norvegicus

PKA activation by FSH is essential to transduce FSH-mediated effects on granulosa cell proliferation, differentiation and steroidogenesis. However, It is unknown whether activation of PKA is sufficient to account for the entire program of granulosa cell responses to FSH. We addressed this question by conducting a comprehensive comparative analysis of signaling pathways and gene expression profiles of granulosa cells stimulated with FSH or expressing a constitutively active PKA mutant, PKA-CQR. Overall design: Undifferentiated GCs were isolated from ovaries obtained from 24 day-old female rats and cultured. The following treatments were applied: 1) EGFP lentivirus 24hr (n=2), 2) EGFP lentivirus 24h + FSH (100 ng/ml) for 24h (n=3), 3) PKA-CQR lentivirus 24h (n=3).Total RNA was extracted from the cells 24 hr. later using RNA-Bee solution, frozen at −80 C, and transported to the Genomics and Proteomics Core Laboratory of the University of Pittsburgh to perform microarray analysis with the Affymetrix Rat Genome 230 2.0 array as per the manufacturers instructions. For data analysis, RMA normalized Affymetrix gene expression data were imported into BRB Array Tools (http://linus.nci.nih.gov/BRB-ArrayTools.html) for differential expression (DE) analysis. DE genes were identified using the Student t test (p<0.001) in BRB.