Developmental competence of gametes reconstructed by germinal vesicle transplantation from fresh and cryopreserved bovine oocytes

Objective

To evaluate the use of fresh or frozen bovine oocytes as an animal model for reconstructing artificial gametes by germinal vesicle transplantation (GVT), to study nucleocytoplasmic interaction and define clinical procedures for ooplasm donation in humans.

Design

Prospective experimental study.

Setting

University-based experimental laboratory.

Animal(s)

Bovine oocytes from slaughterhouse ovaries.

Intervention(s)

A total of 446 gametes were reconstructed from fresh immature oocytes; nuclear and cytoplasmic competencies were analyzed through the assessment of meiotic progression and cytoskeleton reorganization; embryonic developmental capability was evaluated after parthenogenetic activation of metaphase II (MII) reconstructed oocytes. Furthermore, the distribution of mitochondria in karyoplast and cytoplast in grafted oocytes was studied. Finally, meiotic and developmental competencies were determined in 199 gametes reconstructed from vitrified immature oocytes.

Main Outcome Measure(s)

Maturational and developmental rate of reconstructed oocytes, cytoskeleton organization, and mitochondrial distribution.

Result(s)

Gametes reconstructed from either fresh or cryopreserved immature oocytes showed similar meiotic competence (41.6% vs. 37.7%, respectively). All reconstituted oocytes that reached MII displayed a normal distribution of cytoskeletal elements. Embryonic developmental capability was higher in oocytes derived from fresh than from cryopreserved gametes (30.8% vs. 8.1%, respectively). Finally, oocyte centrifugation was effective in obtaining karyoplasts with <5% of mitochondria.

Conclusion(s)

Cows can provide a suitable organism model to develop GVT technique in both research and clinical settings as well as in fertility preservation programs.