The present study was conducted to investigate the effects of attachment of cumulus cells to porcine oocytes during the process of maturation and fertilization on the nuclear maturation, fertilization and subsequent development after in vitro fertilization (IVF). In the first experiment, the cumulus cells were removed from cumulus-oocyte complexes (COCs) at 0, 24 and 42 h after the onset of maturation culture and were then cultured until reaching 42 h of cultivation. In the second experiment, COCs were denuded as described in the first experiment, then fertilized and cultured for 7 days. As a control, cumulus cells were allowed to maintain attachment to the oocytes until the end of IVF. The proportion of oocytes reaching metaphase II significantly increased with the delay in the removal treatment of cumulus cells. The proportion of normal fertilization gradually increased with delay in the removal treatment of cumulus cells from COCs until the end of IVF. However, no significant difference in the proportion of normal fertilization was found between the 42-h and control groups. The removal treatment of cumulus cells in the 0- and 24-h group significantly (p < 0.05) decreased the proportion of cleaved embryos when compared with the control, and none of them developed to the blastocyst stage. The proportion of development to the blastocyst stage was significantly higher (p < 0.05) in the control group than in the 42-h group (18.1% vs 12.4%; p < 0.05). The present study indicates that the attachment of cumulus cells to the oocyte during maturation and fertilization is important to support oocyte nuclear maturation, fertilization and subsequent embryo development. Particularly, the attachment of cumulus cells to the oocyte during IVF promotes embryonic development.
The present study was conducted to determine the effects of hexoses on in vitro maturation, fertilization, and development of porcine oocytes. In the first experiment, porcine oocytes were matured in modified North Carolina State University (NCSU)-37 medium supplemented with 0.6 mM cysteine, 1 mM dibutyryl cyclic AMP (dbc AMP), 10 IU/mL equine chorionic gonadotrophin, 10 IU/mL human chorionic gonadotrophin, 50 μ/mL gentamycin (Sigma-Aldrich, Tokyo, Japan), 10% (v/v) porcine follicular fluid, and various hexoses (glucose, fructose, and galactose) at various concentrations of 0 (control), 2.5, 5.5, and 10 mM. They were subsequently cultured in the maturation medium without hormones and dbcAMP for an additional 22 h. Fertilization was performed according to Kikuchi et al. (2002 Biol. Reprod. 66, 1033–1041); 15 oocytes were co-incubated with 1 million frozen thawed sperm/mL in fertilization medium for 5 h. Supplementation of either glucose (2.5 or 5.5 mM) or fructose (5.5 mM) in the maturation medium significantly increased the percentages of maturation to metaphase II (68.5%, 79.4%, and 70.2%, respectively) and monospermic fertilization of oocytes (55.0%, 64.5%, and 58.9%, respectively), as compared with control group (metaphase II: 52.8%; monospermic: 42.7%; P < 0.05). Supplementation of galactose had no effect on the meiotic maturation and monospermic fertilization of oocytes. In the second experiment, presumptive zygotes were cultured in modified NCSU-37 supplemented with 4 mg/mL BSA, 0.17 mM sodium pyruvate, 2.73 mM sodium lactate, and 50 μg/mL gentamycin. The cleaved embryos were collected at Day 3 after in vitro fertilization and then cultured for a further 4 days in modified NCSU-37 medium supplemented with 5.5 mM of glucose, fructose, or galactose. The percentages of blastocyst formation, calculated from cleaved embryos, were significantly higher in the glucose and fructose groups (20.4% and 18.4%, respectively) than in the galactose group (12.9%) and the non-supplemented control group (9.2%). Although fructose supplementation did not accelerate blastocyst formation, it did significantly increase the mean cell number of blastocysts (48.0 ± 2.9 vs. 38.6 ± 1.8) and reduced the index of DNA-fragmented nuclei in the blastocysts stained by the TUNEL method (7.6 ± 0.9 vs. 11.8 ± 0.9), as compared with glucose supplementation. In the present study, although all hexoses were used through the glycolysis pathway, supplementation of galactose in the maturation and embryo culture medium had no beneficial effect on development. In contrast, both glucose and fructose enhanced development with an optimal concentration of 5.5 mM. Replacement of glucose with fructose did not accelerate blastocyst formation but did enhance embryo quality in terms of increasing cell number and decreasing the number of fragmented nuclei.
scite is a Brooklyn-based organization that helps researchers better discover and understand research articles through Smart Citations–citations that display the context of the citation and describe whether the article provides supporting or contrasting evidence. scite is used by students and researchers from around the world and is funded in part by the National Science Foundation and the National Institute on Drug Abuse of the National Institutes of Health.
hi@scite.ai
10624 S. Eastern Ave., Ste. A-614
Henderson, NV 89052, USA
Copyright © 2024 scite LLC. All rights reserved.
Made with 💙 for researchers
Part of the Research Solutions Family.