Effects of Serum‐Free In Vitro Maturation of Bovine Oocytes on Subsequent Embryo Development and Cell Allocation in Two Developmental Stages of Day 7 Blastocysts

Abstract

Maturation of oocytes and the subsequent outcome of the in vitro production (IVP) are affected by the composition of in vitro maturation (IVM) medium. To determine the use of serum interfering with effects of single molecules, we aimed at developing simplified IVM medium. The experimental IVM media were: (1) M199-medium supplemented with hormones and serum (control), (2) as 1 but serum was substituted with fatty acid-free serum albumin (FAFBSA) and (3) M199-medium without hormonal and serum supplementation (M199). The quality of embryos was assessed on day 7 by morphology and cryotolerance, as well as by Terminal deoxynucleotidyl Transferase Biotin-dUTP Nick End Labeling (TUNEL) and differential staining. Results showed that the nuclear maturation was suppressed in M199 group alone. Embryo cleavage and development rates, and the proportion of quality 1 blastocysts were lower in the FAFBSA and M199 groups compared to the control. Differences in the cell allocation of fresh embryos were observed at the blastocyst stage, but not at the expanded blastocyst stage. The control group blastocysts had larger number of cells allocated to the inner cell mass (ICM), and the FAFBSA group blastocysts larger apoptotic cell proportion compared to the blastocysts derived from other groups. After cryopreservation, the reduction of ICM proportion and increase of apoptotic cell proportion of embryos were equal between the experimental groups. In conclusion, exclusion of serum from the IVM media reduces embryo development and may cause perturbations in blastocyst development. Differences in the cell allocation of blastocysts between IVM media may appear only when the developmental stages are taken into account.

Sığır Oositlerinin Kumulus Ekspansiyonu ve Nükleer Maturasyon Üzerine Serum, Gonadotropinler, Epidermal Büyüme Faktörü ve Östradiol 17-Beta’nın Etkileri

“…Cumulus expansion and maturation rates in the present study were comparable to the in studies utilizing EGF, FSH and LH as supplements during in vitro maturation of bovine oocytes 4,11 . This may show that our culture system may support oocyte maturation irrespective of hormonal supplementation because there are culture systems that maturation degree are similar to the level reached in our study 1,16 . When oocytes are removed from antral follicles, they begin spontaneous meiotic maturation, presumably due to release from inhibitory influences of substances in follicular fluid or produced by cumulus cells through gap junctions 13 .…”

“…Stimulatory effect of these hormones on CCE may not necessarily be reflected on nuclear maturation, if one assumes that these hormones act only at the cumulus cell level but not at on the oocytes directly or indirectly. It may also be possible that culture systems in vitro may differ in their response to hormonal effects due to effects of hormonal supplementation to culture media 8,[16][17][18] .…”

“…Some systems require hormonal supplementation while some studies report no beneficial or adverse effects of hormonal supplementation 16,17 in addition to develop serum-free culture systems 18 . This study was designed to evaluate the effects of hormones (FSH, LH and E2) and EGF alone or in combination, in serum-free and serum supplemented TCM-199 medium on cumulus expansion and nuclear maturation in vitro of bovine cumulus-oocyte complexes (COCs) with a view to develop a valid maturation systems responsive to the known factors.…”

Role of PTGS2-generated PGE2 during gonadotrophin-induced bovine oocyte maturation and cumulus cell expansion

“…While $60% of in-vivo matured bovine oocytes reach the blastocyst stage when fertilized and cultured in vitro, less than 40% of oocytes matured in vitro do so. This rate is even lower when serum-free conditions are utilized (Korhonen et al, 2010;Marei et al, 2009), which clearly shows that current conditions for in-vitro oocyte maturation are deficient and require improvement. Achieving this will require a better understanding of the mechanisms regulating the process of oocyte maturation.…”

Efeitos da redução ou substituição do soro fetal bovino por outros compostos na maturação in vitro de oócitos bovinos

“…uma diminuição nas taxas de maturação e/ou desenvolvimento embrionário (Ali et al 2002, Korhonen et al 2010), e estudos relatando taxas satisfatórias (Rusell et al 2006). Recentemente, pesquisas mostraram a possibilidade do uso de suplementos alternativos, como o produto comercial Fluido Embriônico (FE), com resultados satisfatórios nas taxas de maturação oocitária, produção de blastocistos e criotolerância embrionária (Tetzner et al 2010, De Lima et al 2012.…”