Birth of Healthy Offspring following ICSI in In Vitro-Matured Common Marmoset (Callithrix jacchus) Oocytes

Abstract

Intracytoplasmic sperm injection (ICSI), an important method used to treat male subfertility, is applied in the transgenic technology of sperm-mediated gene transfer. However, no study has described successful generation of offspring using ICSI in the common marmoset, a small non-human primate used as a model for biomedical translational research. In this study, we investigated blastocyst development and the subsequent live offspring stages of marmoset oocytes matured in vitro and fertilized by ICSI. To investigate the optimal timing of performing ICSI, corrected immature oocytes were matured in vitro and ICSI was performed at various time points (1–2 h, 2–4 h, 4–6 h, 6–8 h, and 8–10 h after extrusion of the first polar body (PB)). Matured oocytes were then divided randomly into two groups: one was used for in vitro fertilization (IVF) and the other for ICSI. To investigate in vivo development of embryos followed by ICSI, 6-cell- to 8-cell-stage embryos and blastocysts were nonsurgically transferred into recipient marmosets. Although no significant differences were observed in the fertilization rate of blastocysts among ICSI timing after the first PB extrusion, the blastocyst rate at 1–2 h was lowest among groups at 2–4 h, 4–6 h, 6–8 h, and 8–10 h. Comparing ICSI to IVF, the fertilization rates obtained in ICSI were higher than in IVF (p>0.05). No significant difference was noted in the cleaved blastocyst rate between ICSI and IVF. Following the transfer of 37 ICSI blastocysts, 4 of 20 recipients became pregnant, while with the transfer of 21 6-cell- to 8-cell-stage ICSI embryos, 3 of 8 recipients became pregnant. Four healthy offspring were produced and grew normally. These are the first marmoset offspring produced by ICSI, making it an effective fertilization method for marmosets.

Oocyte‐activating capacity of fresh and frozen–thawed spermatids in the common marmoset (Callithrix jacchus)

“…This short generation turnover might be shortened further if spermatids from immature animals could be used for production of the next generation. In marmosets, the protocol for ICSI has already been established (Takahashi et al, ), and we expect that it can be applied to ROSI with slight modifications. We found that late round spermatids and elongated spermatids could be collected from male marmosets at around 10–12 months of age, although this timing might show individual variations.…”

Single-cell transcriptome analysis of human, marmoset and mouse embryos reveals common and divergent features of preimplantation development

“…Embryos were collected according to established methods using recently developed devices (Takahashi et al, 2014;Thomson et al, 1994). Staging of female marmoset reproductive cycles and embryo collection have been described (Hanazawa et al, 2012).…”

Generation of a Nonhuman Primate Model of Severe Combined Immunodeficiency Using Highly Efficient Genome Editing

“…The mRNA-injected embryos were cultured in ISM-1 medium (Origio) for 4 days at 37 C with 5% CO 2 , 5% O 2 , and 90% N 2 . The live embryos were then noninvasively transferred into the surrogate mothers' uteri as previously described (Marshall et al, 1997;Takahashi et al, 2014). The plasma progesterone levels of the recipients were measured once per week to monitor pregnancy.…”