Date Published: March 11, 2019
Publisher: Public Library of Science
Author(s): Nicholas Hatzirodos, Katja Hummitzsch, Helen F. Irving-Rodgers, James Breen, Viv E. A. Perry, Richard A. Anderson, Raymond J. Rodgers, Gerrit J. Bouma.
Movement and expansion of mesonephric-derived stroma appears to be very important in the development of the ovary. Here, we examined the expression of 24 genes associated with stroma in fetal ovaries during gestation (n = 17; days 58–274) from Bos taurus cattle. RNA was isolated from ovaries for quantitative RT-PCR. Expression of the majority of genes in TGFβ signalling, stromal transcription factors (NR2F2, AR), and some stromal matrix genes (COL1A1, COL3A1 and FBN1, but not FBN3) showed a positive linear increase with gestational age. Expression of genes associated with follicles (INSL3, CYP17A1, CYP11A1 and HSD3B1), was low until mid-gestation and then increased with gestational age. LHCGR showed an unusual bimodal pattern; high levels in the first and last trimesters. RARRES1 and IGFBP3 also increased with gestational age. To relate changes in gene expression in stromal cells with that in non stromal cells during development of the ovary we combined the data on the stromal genes with another 20 genes from non stromal cells published previously and then performed hierarchical clustering analysis. Three major clusters were identified. Cluster 1 genes (GATA4, FBN3, LHCGR, CYP19A1, ESR2, OCT4, DSG2, TGFB1, CCND2, LGR5, NR5A1) were characterised by high expression only in the first trimester. Cluster 2 genes (FSHR, INSL3, HSD3B1, CYP11A1, CYP17A1, AMH, IGFBP3, INHBA) were highly expressed in the third trimester and largely associated with follicle function. Cluster 3 (COL1A1, COL3A1, FBN1, TGFB2 TGFB3, TGFBR2, TGFBR3, LTBP2, LTBP3, LTBP4, TGFB1I1, ALDH1A1, AR, ESR1, NR2F2) had much low expression in the first trimester rising in the second trimester and remaining at that level during the third trimester. Cluster 3 contained members of two pathways, androgen and TGFβ signalling, including a common member of both pathways namely the androgen receptor cofactor TGFβ1 induced transcript 1 protein (TGFB1I1; hic5). GATA4, FBN3 and LHCGR, were highly correlated with each other and were expressed highly in the first trimester during stromal expansion before follicle formation, suggesting that this could be a critical phase in the development of the ovarian stroma.
The mammalian ovary undergoes many different cell processes during its development . The somatic granulosa cells of follicles and the surface epithelial cells, except at the base of the ovary, are derived from a common precursor, the gonadal ridge-epithelial-like (GREL) cell. This precursor cell very likely originates from the surface epithelial cells of the mesonephros and replicates to form the gonadal ridge. The oogonia and oocytes arise from primordial germ cells (PGC) that migrate from as far away as the yolk sac into the mesonephros from where they are incorporated into the gonadal ridge . During their migration, the PGCs start to proliferate—a process which is continued after colonising the gonad. In the developing ovary, the germ cells firstly replicate (oogonia), then subsequently enter meiosis and arrest in the dictyate stage of prophase I (oocytes) until ovulation.
Due to the large number of genes analysed, we will only describe the expression profiles of genes associated with stromal and thecal cells in the following paragraphs.
The transcript abundance in whole ovaries of a number of genes in the developing bovine ovary were measured across gestation. Whilst we focused on 24 genes likely expressed in stroma we also compared these with 20 additional genes associated with germ cells, GREL cells and granulosa cells published previously . We performed hierarchical clustering, and correlation and network analyses, looking for relationships between genes and gestational age. The genes associated into three clusters based upon their expression patterns, and new positive and negative relationships were identified.
The development of the ovary appears to undergo three major phases based upon transcript abundance. The early phase during which the stroma is actively replicating and expanding is characterised by expression of some genes related to PCOS, including FBN3, LHCGR and GATA4. In an intermediate stage the expression of stromal genes suggests that it has matured to express structural collagens and fibrillins associated with adult tissues, and under the control of TGFβ signalling and perhaps androgens. In the third phase the follicles have formed and some have commenced growth. The identification of the first two phases suggests that stroma is clearly dynamic during growth and development of the ovary and abnormalities may impact on the development of later reproductive pathologies.