Evaluación de materiales para el apoyo de la fecundación in vitro

Resum

In recent decades, assisted reproductive techniques (ARTs), have revolutionized the treatment of infertility. Additionally, ARTs represent a significant tool in animal production, improving production efficiency and enhancing animal welfare. However, despite their widespread use, ARTs still face challenges and suboptimal outcomes in terms of the number and quality of embryos produced in vitro compared to those produced in vivo. With the aim of addressing this issue, attempts have been made to mimic the physiological environment of the organ where fertilization naturally occurs, the oviduct. This has been approached from various angles, imitating physiological and physical parameters or adding reproductive fluids as supplements in culture media. Although these approaches have shown improvements in the in vitro embryonic production system, epigenetic, morphological, developmental and viability differences persist between embryos produced in vitro and those produced in vivo. One option to improve this situation has been to construct a device that allows in vitro fertilization (IVF) in the presence of differentiated oviductal cells, reducing differences among the in vivo derived and in vitro produced embryos. In this thesis, we explore, with the future goal of constructing an artificial oviduct, the compatibility of different materials with gametes and embryos, as well as the architecture of the oviduct. In Chapter I, the compatibility of various materials for the construction of an IVF device using 3D printing is investigated. A "Bovine embryo assay" is used to assess embryonic development when these materials have been present during IVF, considering key parameters such as division and blastocyst rates (on days 7 and 8), cell number, the ratio of inner cell mass to trophectoderm, and the index of apoptotic cells. In Chapter II, the compatibility and effect of molybdenum disulfide (MoS2) are studied, which could be used for the functionalization of scaffolds. In turn, to solubilize this material, functionalization with catechin is employed. Furthermore, its impact on porcine sperm during capacitation is evaluated, measuring parameters such as acrosomal integrity, membrane stability, intracellular calcium levels, mitochondrial activity, membrane fluidity, and protein phosphorylation. Ultimately, the effects of these compounds on sperm capacitation are examined through IVF. Finally, in Chapter III, the internal structure of the porcine oviduct is characterized using images obtained through micro-computed tomography (MicroCT). A 3D reconstruction of the oviduct section where fertilization and early stages of development occur, the ampulla-isthmus region, is carried out. In conclusion, this doctoral thesis lays the groundwork for the construction of a new 3D-printed device that can take into account the internal architecture of the oviduct, creating a more realistic model than those built to date.