Use of human gametes obtained from anonymous donors for the production of human embryonic stem cell lines.
OBJECTIVE
To investigate the use of donated gametes for the production of human embryonic stem cell lines.


METHODS
Basic research study.


METHODS
Assisted Reproductive Technology (ART) program at an academic institution.


METHODS
Consenting oocyte and sperm donors.


METHODS
None.


METHODS
Oocytes were aspirated from oocyte donors (n = 12) and inseminated with frozen-thawed donor (n = 2) sperm followed by culture of embryos to day 5 or 6 in sequential media. The inner cell masses of expanded blastocysts were isolated using immunosurgery and cultured for 4-11 days on irradiated primary mouse embryonic fibroblasts (PMEFs). Viable cell colonies were passed every 7-10 days onto fresh PMEFs in the presence of leukemia inhibitory factor (0.1 microg/mL) and evaluated for appropriate cell surface markers.


RESULTS
Immunosurgery of 40 blastocysts resulted in the culture of 18 inner cell masses, which have produced three cell lines. One of these cell lines has been shown to stain positive for alkaline phosphatase and stage-specific embryonic antigen (SSEA)-4 and negative for SSEA-1, express telomerase activity, and produce hCG when allowed to differentiate.


CONCLUSIONS
These findings demonstrate that the future production of human embryonic stem cell lines for therapeutic use is possible with the use of donated gametes. Many ethical issues were considered before the initiation of this study, and it was our goal to ensure that both oocyte and sperm donors understood the nature and purpose of the research before their participating in the study.
Lanzendorf
S E
SE
Technology Development Center, Jones Institute for Reproductive Medicine, Eastern Virginia Medical School, Norfolk, Virginia 23507, USA. lanzense@evms.edu
Boyd
C A
CA
Wright
D L
DL
Muasher
S
S
Oehninger
S
S
Hodgen
G D
GD
D000469:Alkaline Phosphatase / Q000378:metabolism; D001755:Blastocyst / Q000166:cytology; D002454:Cell Differentiation / Q000502:physiology; D002460:Cell Line / Q000378:metabolism; D006063:Chorionic Gonadotropin / Q000378:metabolism; D004622:Embryo, Mammalian / Q000166:cytology*; D005260:Female; D006028:Glycosphingolipids / Q000378:metabolism; D006801:Humans; D007158:Immunologic Techniques; D016256:Lewis X Antigen / Q000378:metabolism; D008297:Male; D018587:Oocyte Donation; D009865:Oocytes; D013094:Spermatozoa; D055746:Stage-Specific Embryonic Antigens; D013234:Stem Cells / Q000166:cytology*; D019098:Telomerase / Q000378:metabolism; D014019:Tissue Donors
D006063:Chorionic Gonadotropin; D006028:Glycosphingolipids; D016256:Lewis X Antigen; D055746:Stage-Specific Embryonic Antigens; C066584:stage-specific embryonic antigen-4; D019098:Telomerase; D000469:Alkaline Phosphatase