Avaliação histológica de folículos pré-antrais inclusos em tecido ovariano equino e uso de metodologias ativas no ensino superior

Abstract

The equine species has been an attractive experimental model for comparative research on folliculogenesis in humans and domestic animals due to several similarities in follicular dynamics. In this regard, obtaining high-quality equine ovarian tissue is crucial to ensure the effectiveness of ovarian manipulation techniques. Therefore, it is essential to perform a histological analysis that preserves the morphological integrity of the tissue by establishing an appropriate fixation protocol to ensure the preservation of pre-antral follicles as faithfully as possible to the original fresh ovarian tissue, mitigating damage. Thus, this thesis aimed to improve fixation protocols to preserve the morphology of pre-antral follicles within equine ovarian tissue. Additionally, active teaching methodologies were developed to foster research skills among undergraduate veterinary students. In the first study, ovaries (n = 6) from young mares subjected to ovariectomy were fragmented (3 x 3 x 1 mm; 20 fragments/ovary). Equine ovarian tissue fragments were subjected to fixation with 70% ethanol for a short period (ST-EtOH, 45 minutes) in comparison to two classic fixatives [Carnoy's solution (CAR) and 4% paraformaldehyde (PFA)] at different fixation times (6 hours, 12 hours). After standard histological processing, the obtained slides were assessed for morphology (normal or abnormal), follicular developmental stage, densities of follicular and stromal cells, as well as nuclear diameters of follicles and oocytes within equine ovarian tissue. As a result, it was demonstrated that Carnoy's solution better preserved the cellular morphology of equine ovarian tissue. It was observed that 4% paraformaldehyde is a suitable fixative for morphological evaluation of pre-antral follicles, and it has the advantage of being compatible with immunohistochemical technique, compared to the other fixatives. Additionally, the STEtOH fixative

proved to be a good alternative when a quick histological evaluation is required. Furthermore, in the second study, it was possible to assess the involvement of undergraduate veterinary students in research, utilizing active teaching methodologies aimed at equipping students with essential skills for their education, build scientific knowledge, and stimulating critical thinking. The experience of using active teaching methodologies contributed to the development of a second study focused on equine ovarian tissue manipulation procedures and provided students with a broader perspective on the practical and scientific applications in this field.