title: Successful cryopreservation of mouse ovaries by vitrification.
abstract: We developed a new method of cryopreservation of whole ovaries by vitrification using DAP213 (2 M dimethyl sulfoxide, 1 M acetamide, and M propylene glycol) as a cryoprotectant. Four-week-old C57BL/6 mice that underwent partial ovariectomy were orthotopically transplanted with cryopreserved or fresh ovaries (experimental or control group) isolated from 10-day-old green fluorescent protein (GFP)-transgenic mice (+/+). GFP-positive pups were similarly obtained from both groups by natural mating or in vitro fertilization (IVF) followed by embryo transfer, indicating that the cryopreserved ovaries by vitrification retain their fecundity. However, a statistically significant difference (P < 0.05) was found between both groups with respect to the following parameters: the number of GFP-positive pups born by natural mating/grafted ovary (0.8 +/- 0.3 for the experimental group versus 2.0 +/- 0.7 for the control group, mean +/- SEM), the number of collected oocytes by superovulation per mouse (7.0 +/- 1.7 for the experimental group versus 22.7 +/- 3.2 for the control group), the percentage of two-cell embryos obtained from GFP-positive oocytes by IVF (38.5% for the experimental group versus 90.0% for the control group). Histologically, normal development of follicles and formation of corpora lutea were observed in frozen-thawed grafts. However, estimated number of follicles decreased in frozen-thawed ovaries compared with fresh ovaries. Taken together, cryopreservation of the ovary by vitrification seems a promising method to preserve ovarian function, but further studies are required to overcome the possible inhibitory effects of this method on the growth of the ovarian graft.
authors.affiliation: Mitsubishi Kagaku Institute of Life Sciences, Minamiooya 11, Machida, Tokyo 194-8511, Japan.
authors.forename: Fujio
authors.initials: F
authors.lastname: Migishima
authors.forename: Rika
authors.initials: R
authors.lastname: Suzuki-Migishima
authors.forename: Si-Young
authors.initials: SY
authors.lastname: Song
authors.forename: Takashi
authors.initials: T
authors.lastname: Kuramochi
authors.forename: Sadahiro
authors.initials: S
authors.lastname: Azuma
authors.forename: Masahiro
authors.initials: M
authors.lastname: Nishijima
authors.forename: Minesuke
authors.initials: M
authors.lastname: Yokoyama
mesh_terms: D000081:Acetamides; D000818:Animals; D000831:Animals, Newborn; D015925:Cryopreservation / Q000379:methods*; D003451:Cryoprotective Agents; D004121:Dimethyl Sulfoxide; D004624:Embryo Transfer; D005260:Female; D005307:Fertilization in Vitro; D049452:Green Fluorescent Proteins; D006651:Histocytochemistry; D008164:Luminescent Proteins / Q000378:metabolism; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008813:Mice, Inbred ICR; D008822:Mice, Transgenic; D008856:Microscopy, Fluorescence; D010053:Ovary / Q000378:metabolism / Q000637:transplantation; D011247:Pregnancy; D011409:Propylene Glycols; D011897:Random Allocation; D013047:Specific Pathogen-Free Organisms
chemical_list: D000081:Acetamides; D003451:Cryoprotective Agents; C093932:DAP213 solution; D008164:Luminescent Proteins; D011409:Propylene Glycols; D049452:Green Fluorescent Proteins; D004121:Dimethyl Sulfoxide