High FSH doses during ovarian stimulation protocols for assisted reproductive technologies (ART) are detrimental to ovulatory follicle function and oocyte quality. However, the mechanisms are unclear. In a small ovarian reserve heifer model, excessive FSH doses lead to phenotypic heterogeneity of ovulatory-size follicles, with most follicles displaying signs of premature luteinization, and a range in severity of abnormalities. By performing whole transcriptome analyses of granulosa cells, cumulus cells and oocytes from individual follicles of animals given standard or excessive FSH doses, we identified progressive changes in the transcriptomes of the three cell types, with increasing severity of follicular abnormality with the excessive doses. The granulosa and cumulus cells each diverged progressively from their normal phenotypes and became highly similar to each other in the more severely affected follicles. Pathway analysis indicates a possible dysregulation of the final stages of folliculogenesis, with processes characteristic of ovulation and luteinization occuring concurrently rather than sequentially in the most severely affected follicles. These changes were associated with disruptions in key pathways in granulosa and cumulus cells, which may account for previously reported reduced estradiol production, enhanced progesterone and oxytocin production and diminished ovulation rates. Predicted deficiencies in oocyte survival, stress response and fertilization suggest likely reductions in oocyte health, which could further compromise oocyte quality and ART outcomes.

Ewes heterozygous for combinations of the Inverdale (FecX; I+), Booroola (FecB; B+) and Woodlands (FecX2; W+) mutations have ovulation rates higher than each mutation separately. The aims of the experiments described herein were to examine the ovarian phenotypes in I+B+ and I+B+W+ ewes and to compare these with the appropriate ++ (controls), I+ and BB animals available for this study. The mean ± s.e.m. ovulation rates in the ++ (n = 23), I+ (10), I+B+ (7), I+B+W+ (10) and BB (3) animals were 1.8 ± 0.1, 2.5 ± 0.2, 6.6 ± 1.0, 9.6 ± 0.9 and 9.7 ± 0.9 respectively. The maximum number of granulosa cells per follicle in the ++ and I+ genotypes was accumulated after exceeding 5 mm diameter, whereas in I+B+, I+B+W+ and BB animals, this was achieved when follicles reached >2-3 mm. The number of putative preovulatory follicles, as assessed from those with LH-responsive granulosa cells, 24 h after the induction of luteolysis, was higher (P < 0.01) in the I+B+ and I+B+W+ compared to the ++ and I+ genotypes. The median follicular diameters of these follicles in the ++, I+, I+B+, I+B+W+ and BB genotypes were 6, 5, 3, 3 and 3 mm respectively. The total number of granulosa cells in the putative preovulatory follicles when added together, and total mass of luteal tissue, did not differ between the genotypes. Thus, despite large ovulation rate differences between animals with one or more fecundity genes, the total cell compositions over all preovulatory follicles and corpora lutea, when added together, are similar to that from the one or two such follicles in the wild types.

High FSH doses during ovarian stimulation are detrimental to ovulatory follicle function and decrease live birth rate in cattle and women. However, the mechanism whereby excessive FSH causes ovarian dysfunction is unknown. This study tested the hypothesis that excessive FSH during ovarian stimulation induce premature luteinization of ovulatory-size follicles. Small ovarian reserve heifers were injected twice daily for four days with 70 IU (N = 7 heifers) or 210 IU (N = 6 heifers) Folltropin-V (commercial FSH-enriched preparation of porcine pituitary glands with minor (&lt;1%) LH contamination, cpFSH). Ovulatory-size (≥10 mm) follicles were excised from ovaries after the last cpFSH injection and hormone concentrations in follicular fluid (FF) were determined using ELISA. Luteinization was monitored by assessing cumulus cell-oocyte complex (COC) morphology and measuring concentrations of estradiol (E), progesterone (P) and oxytocin (O) in FF. COC were classified as having compact (cCOC) or expanded (eCOC) cumulus cell layers, and as estrogen-active (EA, E:P in FF ≥1), estrogen-inactive (EI, E:P in FF ≤1 &gt; 0.1) or extreme-estrogen-inactive (EEI, E:P in FF ≤0.1). A high proportion (72%) of ovulatory-size follicles in 210 IU, but not 70 IU, dose heifers displayed eCOC. The high dose also produced higher proportions of EI or EEI follicles which had lower E:P ratio and/or E but higher P and/or O concentrations compared with the 70 IU dose heifers. In conclusion, excessive cpFSH doses during ovarian stimulation may induce premature luteinization of most ovulatory-size follicles in heifers with small ovarian reserves.

When women with small ovarian reserves are subjected to assisted reproductive technologies (ART), high doses of gonadotropins are linked to high oocyte and embryo wastage and low live birth rates. We hypothesized that excessive FSH doses during superovulation are detrimental to ovulatory follicle function in individuals with a small ovarian reserve. To test this hypothesis, heifers with small ovarian reserves were injected twice daily for four days beginning on Day 1 of the estrous cycle with 35 IU, 70 IU, 140 IU, or 210 IU doses of Folltropin-V (FSH). Each heifer (n = 8) was superovulated using a Williams Latin Square Design. During each superovulation regimen, three prostaglandin F2α injections were given at 12 h intervals starting at the 7th FSH injection to regress the newly formed corpus luteum (CL). Human chorionic gonadotropin (hCG) was injected 12 h after the last (8th) FSH injection to induce ovulation. Daily ultrasonography and blood sampling were used to determine number and size of follicles and CL, uterine thickness, and circulating concentrations of estradiol, progesterone, and anti-Müllerian hormone (AMH). The highest doses of FSH did not increase AMH, progesterone, ovulatory-size follicle number, uterine thickness, or number of CL. However, estradiol production and ovulation rate were lower for heifers given high FSH doses compared to lower doses, indicating detrimental effects on ovulatory follicle function.