Gene expression profile in human cumulus cells stimulated with purified FSH (hMG) or recombinant FSH (recFSH) combined with gonadotropin-releasing hormone agonist (GnRH-a) or antagonist (GnRH-ant)

Abstract

In order to evaluate the impact of controlled ovarian stimulation (COS) protocols on cumulus cells, we studied their messenger RNA (mRNA) expression profile variations in patients undergoing long (GnRH-a) and short (GnRH-ant) controlled ovarian stimulation (COS) protocols using hMG or recFSH. The mRNA expression profiles of cumulus cells from individual patients were evaluated on DNA microarray chips. Gene expression levels and differential intensities between the groups were analyzed with bioinformatics tools. Cumulus cells of patients (n = 18) referred for an intracytoplasmic sperm injection (ICSI), stimulated with hMG or recFSH in a short GnRH-ant or long GnRH-a protocol, were mechanically removed on the day of egg collection for mRNA extraction. Pools of cumulus cells from a single patient were analyzed on Affymetrix™ HG-U133 plus 2.0 GeneChip microarrays. The Affymetrix™ GeneChip Operating Software 1.2 (GCOS) was used to evaluate the signal intensity for the 30000 unique human known or predicted genes. Sample groups were compared with the Significance Analysis of Microarrays (SAM) software to identify genes displaying restricted expression profiles. Principal component analysis on the genes with the highest average deviation across all samples revealed important variations between all patients, including those of a same COS group. When comparing hMG vs. recFSH, across all samples or within the long GnRH-a and short GnRH-ant groups, only trends in gene expression variation were observed. Yet, genes were found to be strictly restricted to hMG or recFSH COS protocols. Differential expression was more important for the GnRH-a vs. GnRH-ant comparison: under hMG stimulation, 52 genes are up-regulated (fold change >3, False Discovery Rate 0%) with the short GnRH-ant protocol whereas none is up-regulated by the long protocol. Similar results but with different genes were found for the same comparison between the recFSH groups. Exclusive genes were also found in the long GnRH-a and short GnRH-ant groups. Despite important patient to patient variations, we identified human genes differentially expressed when comparing hMG vs. recFSH or GnRH-a vs. GnRH-ant protocols. This will further the identification and understanding of the gene markers induced by the COS that are pertinent for the IVF outcome, thus permitting to better monitor the COS protocols for each patient and to improve the IVF practice.