Candidate genes for polycystic ovary syndrome are regulated by TGF beta in the bovine foetal ovary

STUDY QUESTION: Could changes in transforming growth factor beta (TGF beta) signalling during foetal ovary development alter the expression of polycystic ovary syndrome (PCOS) candidate genes leading to a predisposition to PCOS? SUMMARY ANSWER: TGF beta signalling molecules are dynamically expressed during foetal ovary development and TGF beta 1 inhibits expression of the androgen receptor (AR) and 7 (INSR, C8H9orf3, RAD50, ERBB3, NEIL2, IRF1 and ZBTB16) of the 25 PCOS candidate genes in foetal ovarian fibroblasts in vitro, whilst increasing expression of the AR cofactor TGF beta-induced transcript 1 (TGFBIII or Hic5). WHAT IS KNOWN ALREADY: The ovarian stroma arises from the mesonephros during foetal ovary development. Changes in the morphology of the ovarian stroma are cardinal features of PCOS. The ovary is more fibrous and has more tunica and cortical and subcortical stroma. It is not known why this is and when this arises. PCOS has a foetal origin and perhaps ovarian stroma development is altered during foetal life to determine the formation of a polycystic ovary later in life. PCOS also has a genetic origin with 19 loci containing 25 PCOS candidate genes. In many adult tissues, TGF beta is known to stimulate fibroblast replication and collagen deposition in stroma, though it has the opposite effect in the non-scaring foetal tissues. Our previous studies showed that TGF beta signalling molecules [TGF beta s and their receptors, latent TGF beta binding proteins (LTBPs) and fibrillins, which are extracellular matrix proteins that bind LTBPs] are expressed in foetal ovaries. Also, we previously showed that TGF beta 1 inhibited expression of AR and 3 PCOS candidate genes (INSR, C8H9orf3 and RAD50) and stimulated expression of TGFBIII in cultured foetal ovarian fibroblasts. STUDY DESIGN, SIZE, DURATION: We used Bos taurus for this study as we can ethically collect foetal ovaries from across the full 9-month gestational period. Foetal ovaries (62-276 days, n- 19) from across gestation were collected from pregnant B. taurus cows for RNA-sequencing (RNA-seq) analyses. Foetal ovaries from B. taurus cows were collected (160-198 days, n 6) for culture of ovarian fibroblasts. PARTICIPANTS/MATERIALS, SETTING, METHODS: RNA-seq transcriptome profiling was performed on foetal ovaries and the data on genes involved in TGF beta signalling were extracted. Cells were dispersed from foetal ovaries and fibroblasts cultured and treated with TGF beta 1. The effects of TGF beta regulation on the remaining eight PCOS candidate genes not previously studied (ERBB3, MAPRE1, FDFT1, NEIL2, ARL14EP, PLGRKT, IRF1 and ZBTB16) were examined. MAIN RESULTS AND THE ROLE OF CHANCE: Many TGF beta signalling molecules are expressed in the foetal ovary, and for most, their expression levels increased accross gestation (LTBP1/2/3/4, FBN1, TGFB2/3, TGFBR2/3 and TGFB1I1), while a few decreased (FBN3, TGFBR3L, TGFBI and TGFB1) and others remained relatively constant (TGFBRAP1, TGFBR1 and FBN2). TGF beta 1 significantly decreased expression of PCOS candidate genes ERBB3, NEIL2, IRF1 and ZBTB16 in cultured foetal ovarian fibroblasts. LARGE SCALE DATA: The FASTQ files, normalized data and experimental information have been deposited in the Gene Expression Omnibus (GEO) accessible by accession number GSE178450. LIMITATIONS, REASONS FOR CAUTION: Regulation of PCOS candidate genes by TGF beta was carried out in vitro and further studies in vivo are required. This study was carried out in bovine where foetal ovaries from across all of the 9-month gestational period were available, unlike in the human where it is not ethically possible to obtain ovaries from the second half of gestation. WIDER IMPLICATIONS OF THE FINDINGS: From our current and previous results we speculate that inhibition of TGF beta signalling in the foetal ovary is likely to (i) increase androgen sensitivity by enhancing expression of AR, (ii) increase stromal activity by stimulating expression of COL1A1 and COL3A1 and (iii) increase the expression of 7 of the 25 PCOS candidate genes. Thus inhibition of TGF beta signalling could be part of the aetiology of PCOS or at least the aetiology of polycystic ovaries.