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.


METHODS
Prospective experimental study.


METHODS
University-based experimental laboratory.


METHODS
Bovine oocytes from slaughterhouse ovaries.


METHODS
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.


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


RESULTS
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.


CONCLUSIONS
Cows can provide a suitable organism model to develop GVT technique in both research and clinical settings as well as in fertility preservation programs.
Franciosi
Federica
F
Division of Veterinary Anatomy and Histology, Department of Animal Sciences, Faculty of Veterinary Medicine, University of Milan, Milan, Italy.
Perazzoli
Federica
F
Lodde
Valentina
V
Modina
Silvia C
SC
Luciano
Alberto M
AM
D000818:Animals; D001755:Blastocyst / Q000502:physiology; D002417:Cattle; D002467:Cell Nucleus / Q000502:physiology; D015925:Cryopreservation; D054885:Cumulus Cells / Q000502:physiology; D003593:Cytoplasm / Q000502:physiology; D022162:Cytoplasmic Vesicles / Q000502:physiology*; D003599:Cytoskeleton / Q000502:physiology; D046149:Embryo Culture Techniques; D005260:Female; D005307:Fertilization in Vitro; D008540:Meiosis; D008928:Mitochondria / Q000502:physiology; D023421:Models, Animal; D053652:Nuclear Transfer Techniques; D009865:Oocytes / Q000502:physiology*; D010312:Parthenogenesis; D027724:Reproductive Techniques, Assisted; D014015:Tissue Banks