Review: Roles of follicle-stimulating hormone in preantral folliculogenesis of domestic animals: what can we learn from model species and where do we go from here?

The pituitary gonadotropin FSH is a glycoprotein critical for the development of ovarian follicles. Upon binding to its G protein-coupled membrane receptor located on the granulosa cells of ovarian follicles, FSH elicits a cascade of downstream intracellular responses to promote follicle growth, maturation and steroidogenic activity, leading to the acquisition of meiotic and developmental competence of the enclosed oocyte. The essential role of FSH for proper antral follicle development and fertility is indisputable; over the decades, increasing evidence has also pointed toward survival and growth-promoting effects elicited by FSH in earlier-stage preantral follicles, deeming these follicles FSH-responsive as opposed to the FSH-dependent antral follicles. Transgenic mouse models lacking GnRH1, Fshß or Fshr clearly demonstrate this difference by showing that, morphologically, preantral follicles develop to the secondary stage without FSH signaling; however, exogenous expression or administration of FSH to hormone-deficient mice promotes preantral follicle development, with more pronounced effects seen in earlier stages (i.e., primary follicles). In hypophysectomized sheep, FSH administration also promotes the growth of primary-stage preantral follicles. However, in vivo studies in this area are more challenging to perform in domestic animals compared to rodents, and therefore most of the research to date has been done in vitro. Here, we present the existing evidence for a role of FSH in regulating the growth and survival of preantral follicles from data generated in rodents and domestic animals. We provide an overview of the process of folliculogenesis, FSH synthesis and cellular signaling, and the response to FSH by preantral follicles in vivo and in vitro, as well as interactions between FSH and other molecules to regulate preantral folliculogenesis. The widespread use of FSH in ovarian stimulation programs for assisted reproduction creates a real need for a better understanding of the effects of FSH beyond stimulation of antral follicle growth, and more research in this area could lead to the development of more effective fertility programs. In addition to its importance as an agricultural species, the cow provides a desirable model for humans regarding ovarian stimulation due to similar timing of folliculogenesis and follicle size, as well as similar ovarian architecture. The refinement of minimally invasive methods to allow the study of preantral folliculogenesis in live animals will be critical to understand the short- and long-term effects of FSH in ovarian folliculogenesis.

Isolation of Small Preantral Follicles from the Bovine Ovary Using a Combination of Fragmentation, Homogenization, and Serial Filtration.

Understanding the full process of mammalian folliculogenesis is crucial for improving assisted reproductive technologies in livestock, humans, and endangered species. Research has been mostly limited to antral and large preantral follicles due to difficulty in the isolation of smaller preantral follicles, especially in large mammals such as bovine species. This work presents an efficient approach to retrieve large numbers of small preantral follicles from a single bovine ovary. The cortex of individual bovine ovaries was sliced into 500 µm cubes using a tissue chopper and homogenized for 6 min at 9,000-11,000 rpm using a 10 mm probe. Large debris was separated from the homogenate using a cheese cloth, followed by serial filtration through 300 µm and 40 µm cell strainers. The contents retained in the 40 µm strainer were rinsed into a search dish, where follicles were identified and collected into a drop of medium. The viability of the collected follicles was tested via trypan blue staining. This method enables the isolation of a large number of viable small preantral follicles from a single bovine ovary in approximately 90 min. Importantly, this method is entirely mechanical and avoids the use of enzymes to dissociate the tissue, which may damage the follicles. The follicles obtained using this protocol can be used for downstream applications such as isolation of RNA for RT-qPCR, immunolocalization of specific proteins, and in vitro culture.

A novel sequencing-based vaginal health assay combining self-sampling, HPV detection and genotyping, STI detection, and vaginal microbiome analysis.

The composition of the vaginal microbiome, including both the presence of pathogens involved in sexually transmitted infections (STI) as well as commensal microbiota, has been shown to have important associations for a woman's reproductive and general health. Currently, healthcare providers cannot offer comprehensive vaginal microbiome screening, but are limited to the detection of individual pathogens, such as high-risk human papillomavirus (hrHPV), the predominant cause of cervical cancer. There is no single test on the market that combines HPV, STI, and microbiome screening. Here, we describe a novel inclusive vaginal health assay that combines self-sampling with sequencing-based HPV detection and genotyping, vaginal microbiome analysis, and STI-associated pathogen detection. The assay includes genotyping and detection of 14 hrHPV types, 5 low-risk HPV types (lrHPV), as well as the relative abundance of 31 bacterial taxa of clinical importance, including Lactobacillus, Sneathia, Gardnerella, and 3 pathogens involved in STI, with high sensitivity, specificity, and reproducibility. For each of these taxa, reference ranges were determined in a group of 50 self-reported healthy women. The HPV sequencing portion of the test was evaluated against the digene High-Risk HPV HC2 DNA test. For hrHPV genotyping, agreement was 95.3% with a kappa of 0.804 (601 samples); after removal of samples in which the digene hrHPV probe showed cross-reactivity with lrHPV types, the sensitivity and specificity of the hrHPV genotyping assay were 94.5% and 96.6%, respectively, with a kappa of 0.841. For lrHPV genotyping, agreement was 93.9% with a kappa of 0.788 (148 samples), while sensitivity and specificity were 100% and 92.9%, respectively. This novel assay could be used to complement conventional cervical cancer screening, because its self-sampling format can expand access among women who would otherwise not participate, and because of its additional information about the composition of the vaginal microbiome and the presence of pathogens.

ST segment elevation in lead aVR during exercise testing is associated with LAD stenosis.

PURPOSE: To evaluate, in patients with chest pain, the diagnostic value of ST elevation (STE) in lead aVR during stress testing prior to (99m) Tc-sestamibi scanning correlating ischaemic territory with angiographic findings. METHODS: Consecutive patients attending for (99m) Tc-sestamibi myocardial perfusion imaging (MPI) completed a treadmill protocol. Peak exercise ECGs were coded. STE >or=0.05 mV in lead aVR was considered significant. Gated perfusion images and findings at angiography were assessed. RESULTS: STE in lead aVR occurred in 25% (138/557) of the patients. More patients with STE in aVR had a reversible defect on imaging compared with those who had no STE in aVR (41%, 56/138 vs 27%, 114/419, p=0.003). Defects indicating a left anterior descending artery (LAD) culprit lesion were more common in the STE in aVR group (20%, 27/138 vs 9%, 39/419, p=0.001). There was a trend towards coronary artery stenosis (>70%) in a double vessel distribution involving the LAD in those patients who had STE in aVR compared with those who did not (22%, 8/37 vs 5%, 4/77, p=0.06). Logistic regression analysis demonstrated that STE in aVR (OR 1.36, p=0.233) is not an independent predictor of inducible abnormality when adjusted for STD >0.1 mV (OR 1.69, p=0.026). However, using anterior wall defect as an end-point, STE in aVR (OR 2.77, p=0.008) was a predictor even after adjustment for STD (OR 1.43, p=0.281). CONCLUSION: STE in lead aVR during exercise does not diagnose more inducible abnormalities than STD alone. However, unlike STD, which is not predictive of a territory of ischaemia, STE in aVR may indicate an anterior wall defect.