Abstract
Successful production of transgenic pigs requires oocytes with a high developmental competence. However, cumulus–oocyte complexes (COCs) obtained from antral follicles have a heterogeneous morphology. COCs can be classified into one of two classes: class I, with five or more layers of cumulus cells; and class II, with one or two layers of cumulus cells. Activator [e.g., epidermal growth factor (EGF)] or inhibitors (e.g., wortmannin and U0126) are added to modulate kinases in oocytes during meiosis. In the present study, we investigated the effects of kinase modulation on nuclear and cytoplasmic maturation in COCs. Class I COCs showed a significantly higher developmental competence than class II COCs. Moreover, the expression of two kinases, AKT and ERK, differed between class I and class II COCs during in vitro maturation (IVM). Initially, inhibition of the PI3K/AKT signaling pathway in class I COCs during early IVM (0–22 h) decreased developmental parameters, such as blastocyst formation rate, blastomere number, and cell survival. Conversely, EGF-mediated AKT activation in class II COCs enhanced developmental capacity. Regarding the MAPK signaling pathway, inhibition of ERK by U0126 in class II COCs during early IVM impaired developmental competence. However, transient treatment with U0126 in class II COCs increased oocyte maturation and AKT activity, improving embryonic development. Additionally, western blotting showed that inhibition of ERK activity negatively regulated the AKT signaling pathway, indicative of a relationship between AKT and MAPK signaling in the process underlying meiotic progression in pigs. These findings may help increase the developmental competence and utilization rate of pig COCs with regard to the production of transgenic pigs and improve our understanding of kinase-associated meiosis events.
Highlights
Transgenic pigs are regarded as potentially good animal models for biomedical research
To examine the maturation competence and subsequent embryonic development according to cumulus–oocyte complexes (COCs) morphology, PA-derived mature oocytes from class I and class II COCs were cultured in IVC medium for 6 days, and developmental parameters were scored on Day 6
The present study was the first to demonstrate the distinct activities of two signaling pathways, PI3K/AKT and mitogen-activated protein kinase (MAPK), between class I and II COCs in the pig after 20 h during in vitro maturation (IVM)
Summary
Transgenic pigs are regarded as potentially good animal models for biomedical research. To produce transgenic pigs, porcine cumulus–oocyte complexes (COCs) with high developmental competence are needed because of the low development rate of in vitro-produced (IVP) blastocysts compared with embryos produced in vivo. Many laboratories have cultured COCs of heterogonous morphology in vitro to produce mature oocytes upon fertilization or parthenogenesis (PA) [1, 2]. Many researchers have cultured COCs to produce IVP embryos according to the number of cumulus cell layers [3, 4]. Researchers have focused on enhancing developmental competence via treatment with supplements, such as growth factors, hormones, and inhibitors, to alter the activity of proteins in COCs [7, 8]. There is limited information on the improvement of maturation competence via the regulation of proteins in oocytes according to differences in COC morphology
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