The ability to preserve spermatozoa and eggs for a long period of time is critical for retaining fertility, addressing infertility and lengthening the reproductive life span of individuals. Although spermatozoa survive freezing with relative ease, egg (oocyte) preservation is more complex because of the cell's large size, significant water content and fragile cellular skeleton. Current methods to preserve the oocyte are far from optimal, so there is a need to explore novel approaches. This proposal examines what happens when the immature oocyte nucleus, also known as the germinal vesicle, is isolated and dried alone as a potential means of genome preservation. Specifically, we will determine if a desiccated germinal vesicle (from any oocyte collected within the ovary) can be rehydrated and then placed into a fresh (previously enucleated) donor oocyte to reanimate and recover the original maternal genome. The 'reconstructed'oocyte then could be cultured and fertilized in vitro with appropriate spermatozoa to produce a viable embryo of a desired genotype. The general hypothesis is that desiccation and storage at room temperature can preserve the structural and functional integrity of the germinal vesicle. The germinal vesicle will be studied in the domestic cat because (1) this species is an important model for human health and reproduction, and (2) our preliminary data are extensive and highly promising. An important element of the proposed project is compacting the germinal vesicle to a small diameter before drying, which will ensure optimal protection during desiccation or storage while permitting easy transfer by injection into a surrogate oocyte. Our objective is to examine, for the first time, the impact of compaction, desiccation and storage at room temperature on the germinal vesicle, including its ability to reconstitute an oocyte that can undergo meiosis, fertilization and embryo development to term (i.e., production of healthy young). Early data have revealed that (1) in vitro compaction is an excellent protector of the germinal vesicle against stressors, (2) germinal vesicle transfer into a recipient oocyte is successful and (3) desiccation in the presence of sugar safeguards nuclear content. The project challenges existing paradigms about how to preserve female fertility in an area (germinal vesicle preservation and storage) that has never been explored, but shows strong promise. This innovative strategy could circumvent the complications associated with conventional inefficient, damaging and costly freeze-storage of whole oocytes in liquid nitrogen. The required studies also will substantially increase scholarly knowledge about complex interactions between the oocyte's nucleus and cytoplasm, including the mechanisms regulating developmental competence of the germinal vesicle.

Public Health Relevance

(provided by the applicant): As will be described, the drying, storage and reanimation of the oocyte's germinal vesicle represent an enormous potential for preserving human fertility. This approach also could have widespread, practical application to more effective, economical preservation, propagation and management of rare animal models used to study human diseases. The ability to efficiently perpetuate these unique genotypes allows understanding mechanisms to improve human health through translational biomedical research.

Agency
National Institute of Health (NIH)
Institute
Office of The Director, National Institutes of Health (OD)
Type
Research Project (R01)
Project #
5R01OD010948-04
Application #
8456949
Study Section
Special Emphasis Panel (ZRR1-CM-4 (01))
Program Officer
Moro, Manuel H
Project Start
2010-05-01
Project End
2015-02-28
Budget Start
2013-03-01
Budget End
2014-02-28
Support Year
4
Fiscal Year
2013
Total Cost
$352,534
Indirect Cost
$61,270
Name
Smithsonian Institution
Department
Type
DUNS #
089522580
City
Arlington
State
VA
Country
United States
Zip Code
22202