Abstract
Vitrification of matured oocytes is widely adopted in human clinics and animal research laboratories. Cryopreservation of immature oocytes, particularly those at metaphase I (MI), remains a challenge. In the present work, mouse MI oocytes denuded of cumulus cells were vitrified and warmed (V/W) either prior to (V/W-BEFORE-IVM, n = 562) or after (V/W-AFTER-IVM, n = 664) in vitro maturation (IVM). Derivative metaphase II (MII) oocytes were then used for intracytoplasmic sperm injection (ICSI). In the control groups, in vivo matured MII oocytes were used freshly (FRESH-MII, n = 517) or after V/W (MII-V/W, n = 617). In vitro and in vivo developmental competencies were compared among groups. Satisfactory blastocyst rates were achieved in V/W-BEFORE-IVM (27.5%) and V/W-AFTER-IVM (32.4%) groups, albeit as expected still lower than those from fresh-MII (56.1%) or MII-V/W (45.6%) oocytes. Similarly, the term development rates from V/W-BEFORE-IVM and V/W-AFTER-IVM were 12.4% and 16.7% respectively, acceptable but lower than those of the fresh-MII (41.2%) and MII-V/W (23.3%) groups. These data demonstrate that oocytes collected at MI stage are amenable to V/W, which can be performed before or after IVM with acceptable development rates including production of healthy pups. These findings provide useful knowledge to researchers and clinical practitioners for preservation and use of the otherwise discarded MI oocytes.
Highlights
Conventional slow freezing and vitrification are common methods to preserve mammalian gametes and embryos
In metaphase II (MII)-vitrified and warmed (V/W) Group, in vivo matured (IVO) MII oocytes were harvested from the oviducts 15 h after the human chorionic gonadotrophin (hCG) trigger, and were immediately vitrified, stored in liquid nitrogen until they were warmed for intracytoplasmic sperm injection (ICSI) at a later time
In V/W-BEFORE-in vitro maturation (IVM) Group, 818 metaphase I (MI) oocytes were retrieved and vitrified, the MI oocyte survival rate was 92.4% (756/818) post vitrification/warming, and a total of 562 out of 818 starting MI oocytes were matured (68.7%) after 8 h of in vitro culture, significantly lower than that achieved in the V/W-AFTER-IVM Group (P
Summary
Conventional slow freezing and vitrification are common methods to preserve mammalian gametes and embryos. Vitrification has been proven effective on cryopreserving matured female gametes, often leading to minor compromise of the fertility and developmental competency; it is widely adopted in human IVF clinics [1,2,3,4,5,6,7,8,9] and in animal research laboratories [10,11,12]. While vitrification of MII oocytes is reliable, it is noted that a working protocol to vitrify immature oocytes remains to be developed. The effects of vitrification timing on MI oocytes have not been evaluated, and there are no reports on production of live offspring using oocytes V/W at MI stage
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