Identification of differential gene expression in in vitro FSH treated pig granulosa cells using suppression subtractive hybridization

In order to find new regulated transcripts, however rare, we have used a Suppression Subtractive Hybridization approach (SSH) on pig granulosa cells in primary culture treated or not with FSH. Two SSH libraries were generated and 76 clones were sequenced after selection by differential screening. Sixty four different sequences were identified, including 3 novel sequences. Experiments demonstrated the presence of 25 regulated transcripts.A gene ontology analysis of these 25 genes revealed (1) catalytic; (2) transport; (3) signal transducer; (4) binding; (5) anti-oxidant and (6) structural activities. These findings may deepen our understanding of FSH's effects. Particularly, they suggest that FSH is involved in the modulation of peroxidase activity and remodelling of chromatin.The development of ovarian follicles leading to ovulation requires endocrine regulation by the gonadotropins FSH and LH as the main actors. The complex regulatory network also includes steroids and peptides (e.g. growth factors, inhibins) acting via the autocrine and paracrine pathways. Recent studies have highlighted the importance of FSH in ovarian follicle maturation: in FSH-deficient mice the folliculogenesis is blocked prior to antral formation [1,2]. In order to obtain functional gametes, granulosa cell (GC) communication with the oocyte also seems essential [2,3]. GCs constitute an important compartment in the mammalian ovarian follicle, contributing to follicle development. They actively participate in the endocrine function of the ovaries by secreting oestradiol or progesterone under FSH stimulation. Besides their functional importance, GCs have been intensively studied for their convenient isolation. In murine, porcine, and bovine species they constitute a well-standardized model for the in vitro study of GC function, including hormonal regulation.Even if much data has been accumulated on the action of gonadotropins on GCs, the entire spectrum of genes regulated by FSH is not known. B