Enhancement of Cytoplasmic Maturation of In Vitro-Matured Pig Oocytes by Mechanical Vibration

Abstract

Abstract. The effects of mechanical vibration during in vitro maturation and/or in vitro culture after artificial activation of pig oocytes on maturation and development were examined. In addition, the optimal conditions were applied to in vitro production of blastocysts derived from miniature pig somatic cell nuclear transfer (SCNT) embryos. Mechanical vibration during in vitro maturation did not affect the rates (60.5 ± 1.9-69.5 ± 2.2%) of oocytes reaching the metaphase-II stage. However, the blastocyst formation rates after activation of oocytes matured with mechanical vibration for 5 sec at intervals of 30-60 min or for 10 sec at intervals of 60 min were significantly (P<0.05) higher than those of oocytes matured without mechanical vibration (25.7 ± 2.0-28.1 ± 2.7% vs. 12.3 ± 1.4% and 25.8 ± 1.8% vs. 15.7 ± 1.9%, respectively). In contrast, mechanical vibration during in vitro culture after activation did not affect the blastocyst formation (11.6 ± 5.2-16.5 ± 3.0%) of oocytes. Mechanical vibration for 5 sec at intervals of 60 min during in vitro maturation of oocytes did not affect fusion (66.8 ± 3.5-72.1 ± 3.1%) with miniature pig somatic cells after enucleation. However, the blastocyst formation rate of SCNT embryos was improved (P<0.05) by mechanically vibrating recipient oocytes for 5 sec at intervals of 60 min during in vitro maturation, regardless of the presence or absence of the same treatment during in vitro culture (17.6 ± 2.5% vs. 9.4 ± 0.9% and 13.0 ± 0.3% vs. 7.4 ± 0.9%, respectively). The results indicated that mechanical vibration enhances the cytoplasmic maturation of in vitro-matured pig oocytes, resulting in improvement of their parthenogenetic development. In addition, it was shown that in vitro maturation of oocytes with mechanical vibration can be applied to efficient production of blastocysts derived from miniature pig SCNT embryos. he low cloning efficiency associated with the development of somatic cell nuclear transfer (SCNT) embryos to offspring remains the major obstacle to use of this technology in various fields of animal science and biomedical applications. Establishment of systems to support efficient production of blastocysts from SCNT embryos in vitro is quite important for basic research to clarify the mechanism controlling the development of SCNT embryos, which will bring about improvement of cloning efficiency. In cattle and pigs, in vitro oocyte maturation systems produce an abundant and stable supply of recipient oocytes for SCNT because immature oocytes can be obtained from slaughtered animals. In vitro-matured oocytes have been commonly used for production of cloned calves [1-3] and pigs [4][5][6][7][8]. The oocyte maturation process is a crucial step for the generation of oocytes capable of being fertilized and undergoing normal embryonic development into blastocysts after in vitro fertilization [9]. Therefore, efficient production of blastocysts from SCNT embryos requires optimization of both recipient oocyte maturation systems and SCNT embryo culture systems.Oocyte ...

Effect of micro-vibration culture system on embryo development

“…Micro-vibration did not influence the maturation of porcine immature oocytes. However, the blastocyst development rates were significantly improved by micro-vibration [21]. It is not known whether micro-vibration has a positive effect on the maturation of human immature oocytes or on the clinical results in IVM.…”

“…Studies of various mammalian embryos culture include this study on micro-vibration culture condition would be useful information for dynamic culture or embryo activation. Based on a study conducted on immature porcine oocytes and embryos, the blastocyst formation rate is significantly increased at a frequency of 20 Hz, 5 s/30 to 60 min duration [21]. Isachenko's group has reported several relevant results.…”

“…In the present study, a micro-vibrator (NSSB-300, Nepa Gene, Ichikawa, Chiba, Japan) was set at a frequency of 42 Hz, 5 s/60 min duration for mouse and human embryo development based on the previous reports [21,22,33]. Micro-vibration was applied to the single-cell zygote stage in the mouse.…”

“…These in vivo mechanical forces are believed to induce cell-to-cell communication, exert beneficial effects by refreshing the fluid surrounding the embryo and eliminate metabolites emitted from embryos [16][17][18][19]. Studies to achieve higher quality embryos by external stimuli using machines (i.e., tilting machines and micro-vibrators) to replicate in vivo mechanical forces have been recently reported [20,21]. Embryonic culture using a micro-vibrator as an external stimulus has been actively investigated [22].…”

Use of Silicone Elastomer-Based Microfluidic Devices and Systems in Reproductive Technologies

“…We developed a tilting embryo culture system (TECS) to move mouse, porcine, and human embryos by fluid motion of the microdroplet, and we have shown that blastocyst development rates of these embryos using TECS improved over those of static controls [85][86][87]. Gentle vibration of porcine and human embryos also appeared to yield improved blastocyst formation rates [88][89][90]. These DCSs can be combined with microwells and microfluidic channels for mammalian embryo cultures.…”