Effects of cumulus cells, different cryoprotectants, various maturation stages and preincubation before insemination on developmental capacity of frozen-thawed bovine oocytes

To improve freezability of bovine follicular oocytes, it is necessary to minimize injury to the oocytes caused by freezing and the toxicity of cryoprotectants. The maturing ability of frozen-thawed follicular oocytes with or without cumulus complexes was tested. The proportion of frozen-thawed follicular oocytes reaching the metaphasc II (M II) stage after in vitro maturation of 24 h was significantly (P < 0.05) higher in cumulus oocyte complexes (COCs; 44%) than in denuded oocytes (30%). Oocytes were cultured for 0, 6, 12, 18 or 24 h then frozen-thawed with 1,2-propanediol (PROH) or dimethyl sulfoxide (DMSO), and cultured for 24, 18, 12, 6 or 0 h respectively. In PROH, 24:0 (67%) showed significantly (P < 0.05) higher maturation rate than 0:24 (38%), 6:18 (41%). In DMSO, 18:6 (72%) and 24:0 (61%) showed significantly (P < 0.05) higher maturation rate than 0:24 (30%), 6:18 (33%) and 12:12 (44%). In case of 18:6, DMSO (72%) showed significant (P < 0.05) higher maturation rate than PROH (52%), however in case of 0:24, 6:18, 12:12 and 24:0, there was no significant (P < 0.05) difference in the maturation rate between PROH and DMSO. The proportion of embryos developed to > or = 2 cell, > or = 8 cell, morula and blastocyst in 18:6 DMSO (35, 10, 3 and 0%) and 24:0 PROH (38, 12, 5 and 0%) was significantly (P < 0.05) lower than that of fresh oocytes (67, 38, 31 and 16%). There was no significant (P < 0.05) difference in the rate of embryos that developed to > or = 2 cells, > or = 8 cells, morulae and blastocysts between PROH and DMSO. When the frozen oocytes were grouped as rewarming culture (21:2 PROH) and control (24:0 PROH), there was no significant (P < 0.05) difference in the rate of embryos that developed to > or = 2 cells, > or = 8 cells, morulae and blastocysts between 24:0 PROH (42, 24, 11 and 1%) and 21:2 PROH (51, 29, 16 and 4%) but 21:2 PROH showed slightly higher developmental capacity than 24:0 PROH. Transferable blastocysts (4%) were obtained in 21:2 PROH when the frozen-thawed follicular oocytcs were fertilized and cultured for 8 to 9 d.