Dados do Trabalho

Title

Next-generation of Oocyte In Vitro Maturation: Potential 3D Matrix using Bovine Model

Objective

Despite advances in assisted reproductive technologies (ART), current conditions in human medicine and animal production desired standards. Abnormalities in oocytes matured under in vitro maturation (IVM) conditions can negatively affect embryonic development. In comparison, oocytes matured in vivo produce more and better embryos with differences in morphology, metabolism, and gene expression than in vitro embryos. The culture environment has a significant impact on bovine blastocyst development. Traditionally, IVM has relied on two-dimensional (2D) culture systems that cannot fully reflect the complexity of the in vivo environment. Consequently, oocytes matured in 2D systems may be of lower quality, resulting in a lower rate of successful embryo development. In contrast, three-dimensional (3D) culture systems provide a more biomimetic environment by using matrices or scaffolds that closely resemble the extracellular matrix in vivo. These matrices support interactions between oocyte and cumulus cells, promote a more natural cell-cell communication, and provide a microenvironment that better resembles the ovarian follicle. In this study, we present an innovative 3D-IVM approach using a custom 3D matrix to enhance oocyte maturation by preventing cumulus-oocyte complex (COC) flattening and preserving its structural and functional integrity. We investigated meiotic progression in oocytes and blastocyst yield using this 3D matrix.

Methods

COCs were collected from cattle ovaries at a slaughterhouse (n= 50 COCs/group; with 10 replicates) and subjected to either the new 3D-IVM system or a control group using 2D-IVM for 24 hours in a basic medium. The maturation medium consisted of TCM199 with bicarbonate and Earle's salts, supplemented with recombinant follicle-stimulating hormone (FSH - 0.1 IU/mL), sodium pyruvate (22 µg/mL), amikacin (75 µg/mL), and bovine serum albumin without fatty acids (BSA; 4 mg/mL). After the IVM, matured COCs were denuded and the meiotic progress was examined under a microscope to determine the meiotic stage (Hoechst 33342 protocol). In a second experiment, the matured COCs were subjected to regular in vitro fertilization (IVF) and in vitro culture (IVC).

Results

In our 3D-IVM system, COCs did not adhere to the surface of the plate, enhancing their exchange surface area with the culture medium. Although the meiotic progress among 2D-IVM and 3D-IVM was not affected by the new system (P= 0.76; 86.45 ± 2.65 and 85.77 ± 2.08, respectively) the results suggest a brief reduction in degenerated oocytes using our 3D-IVM (P= 0.06; 2.30 ± 0.56; 0.81 ± 0.54, for 2D-IVM and 3D-IVM, respectively). Furthermore, our system was able to improve the blastocyst yield (P= 0.04; 37.20 ± 1.99) compared to the 2D system (27.73 ± 3.56), which may have a beneficial effect on early embryonic development.

Conclusion

The proposed 3D-IVM approach represents a groundbreaking and highly promising advancement in the field of assisted reproduction. 3D maturation offers a paradigm shift by providing a more biologically relevant and physiologically accurate microenvironment for oocyte development. As research and technology in the field advance, the adoption of 3D-IVM has the potential to revolutionize reproductive technologies, leading to increased success rates and improved fertility treatments in both human medicine and animal reproduction programs. This study was financially supported by CAPES and FAPESP- Pipe (nº 2022/06068-5).

Keywords

oocyte maturation, three-dimensional (3D) culture, blastocyst yield, fertility treatments.