Oocyte lipid accumulation in cattle: effects of in vitro maturation variations

Author(s)

Luana Alves

• Document type: Doctoral Thesis
• Press: Pirassununga.
• Institution: Universidade de São Paulo (USP). Faculdade de Zootecnica e Engenharia de Alimentos (FZE/BT)
• Defense date: 2024-05-10

Examining board members

Felipe Perecin; Tiago Henrique Camara de Bem; Lindsay Unno Gimenes; Juliano Coelho da Silveira; Edson Guimarães Lo Turco

• Advisor: Felipe Perecin

Abstract

Reproductive success depends on oocyte competence, which can be negatively affected when these gametes are subjected to an in vitro environment. One of the commonly observed adverse effects is the excessive accumulation of lipids during maturation, resulting in loss of quality, metabolism changes, lower tolerance to cryopreservation and pregnancy rate reduction. The components of in vitro maturation medium (IVM) and the transport of biomolecules between cumulus- oocyte cells via transzonal projections (TZPs) and small extracellular vesicles (EVs) are shown to be crucial in this mechanism. Thus, our hypothesis is that the composition of the IVM medium and the communication between cumulus and oocyte cells modulate intraoocyte lipid accumulation, as well as the expression of related genes in bovine cumulus-oocyte complexes (COCs). In Experiment 1 we investigated the effects of presence or absence of small fetal calf serum (FCS) EVs in IVM medium regarding intraoocyte lipid accumulation after 9 h of IVM through EVs characterization by presence, size and concentration, and its effect on oocytes lipid droplets accumulation and maturation rate. We confirmed the presence of FCS small EVs with average size of 144nm and concentration of 1,15¹¹ particles/mL. Presence or absence of FCS small EVs showed no effect on lipid droplets and maturation rate. In Experiment 2 we assessed the influence of prologued communication within COC mediated by TZPs regarding intraoocyte lipid accumulation during IVM validating a pre-IVM (PIVM) system for this experimental model, as well as intraoocyte lipid accumulation in COCs that were submitted to pre-IVM and were matured in conventional medium (MC) and modified medium (MF). Oocytes lipid droplets accumulation and maturation rate were assessed, along with transcript patterns and specific genes in cumulus cells (CCs) after 9 h IVM (RNASeq Sequencing). There was no difference on lipid droplets comparing all experimental groups, however we identified a tendency of higher lipid accumulation on PIVM oocytes. Maturation rates had no difference. RNASeq results showed effect of PIVM presence or absence on transcripts patterns identifying 83 differentially expressed genes (DEGs): the Regulation of Actin Cytoskeleton Pathway genes in PIVM group; and genes that regulates protein processing in endoplasmic reticulum, estrogen signaling, TGF-beta signaling and lipid metabolism in IVM group. Analyzing specific genes by transcripts per million value (TPM), we found SCD and ATF4 genes increased in the PMIV group; and ACACA, FABP3 and HSPA5 genes increased in the MIV group. We conclude that the presence/absence of SFB EVs is not a determining factor for oocyte lipid accumulation nor for altering oocyte development during IVM. Nonetheless, the modulation of communication in the COC through PIVM proved to be a promising model to understand the mechanism by which the oocyte accumulates more lipids in in vitro environment, once effects were observed on specific transcripts genes in CCs with a slight change in oocytes phenotype. Studies regarding lipolysis and lipogenesis related genes in oocytes that have undergone PIMV are interesting as a next step, as oocytes that have undergone PIMV may be suppressing the expression of these genes in response to higher lipid droplets in their cytoplasm. (AU)

• FAPESP's process: 21/09123-4 - Oocyte lipid accumulation in cattle: effects of variations in the in vitro maturation system
• Grantee: Luana Alves
• Support Opportunities: Scholarships in Brazil - Doctorate