17α-hydroxylase and the androgen-treated sheep ovary – a model for Polycystic Ovary Syndrome (PCOS)

Abstract

Polycystic Ovary Syndrome (PCOS) is the most common cause of infertility in women of reproductive age, with an estimated five to ten percent of women worldwide affected by the endocrine condition. The aetiology of PCOS remains, for the most part unclear, however, androgens have been implicated in the development of PCOS with hyperandrogenism being one of the most consistent PCOS traits. The products of a complex steroid pathway, androgens, in particular androstenedione and testosterone, are found to be significantly increased in many PCOS patients. The aim of this study was to investigate the activity of a key enzyme in the steroid pathway, 17α-hydroxylase, in the prenatally testosterone treated (T-treatment) sheep ovary and subsequently measure levels of androstenedione present in blood samples taken from the jugular and ovarian veins. In addition to this, a histological study of follicular mapping provided useful insights into the effect of T-treatment on aspects of ovine follicular development. Sheep ovaries and blood samples were collected from pre-pubertal (5 month old) and pubertal (10 month old) animals. Serial sections were examined to study follicle numbers and morphology. A radioactive in situ hybridisation (ISH) was undertaken to determine tissue location and levels of 17α-hydroxylase (17α-OH) mRNA. Androstenedione levels in blood were determined using ELISA. Results showed that while body weight was similar in both T-treated and control groups, T-treatment was associated with an increase in mean ovarian weight in both pre-pubertal and pubertal age groups. T-treated animals experienced some masculinisation of the external genitalia, which has also been reported by other researchers using this model. In pre-pubertal animals, T-treatment was associated with a significant increase in the total number of growing ovarian Type 3-5 follicles within the ovary. When analysed individually, all eight categories of follicles in this study were found to be increased with T-treatment however these proved to be not statistically significant, with the exception of degenerate follicles where a 14-fold increase was seen in T-treated ovaries. Pre-pubertal T-treated ovaries had a significant accumulation of antral follicles ≥4mm, while pubertal T-treated ovaries displayed an increase in follicles ≥1mm. During the isolation of these follicles, it was found that T-treated follicles were more likely to be unhealthy or haemorrhagic than those from control ovaries. Thecal tissue measurements showed that Type 4 and small Type 5 follicles from T-treated ovaries had a significantly larger thecal tissue layer – the site of androstenedione synthesis. In situ hybridisation results showed that expression of 17α-OH mRNA in the theca was significantly increased in Type 4 T-treated follicles and appeared stronger in Type 5 T-treated follicles. Despite 17α-OH being a key enzyme in the production of androstenedione, these results did not translate into a detectable difference in blood levels of androstenedione in both pre-pubertal and pubertal animals. The results of this study suggest that elements of a PCOS phenotype are evident in pre-pubertal, prenatally androgenised sheep. Further study of this model may provide more answers as to the development of PCOS and it’s symptoms and the role that androgens play in the development of the syndrome and the consequent infertility.