Production of dromedary (Camelus dromedarius) embryos by IVM and IVF and co-culture with oviductal or granulosa cells.
The general objective of this work was to produce dromedary embryos from cumulus-oocyte complexes (COCs) that were matured, fertilized and co-cultured in vitro. A total of 1598 COCs were recovered from 457 ovaries; 1308 were deemed suitable for IVM and were cultured at 38.5 degrees C, 5% CO2, and >95% humidity for 36 h in TCM-199 supplemented with 10% heat-treated fetal calf serum (FCS), 10 ng/ml epidermal growth factor (EGF), 1 microg/ml FSH, and 500 microM cysteamine. Matured COCs (n = 88) were denuded, fixed, and stained to determine nuclear status; 63% (56/88) had reached metaphase II (MII) at 36 h. Overall, 1135 COCs were inseminated with ejaculated fresh semen (0.5 x 10(6)spermatozoa/ml in modified TALP-solution). Inseminated oocytes (n = 155) were examined for evidence of fertilization; 68% (106/155) were penetrated by spermatozoa, including 52% (55/106) with two pronuclei and 34% (36/106) with polyspermy. Inseminated, denuded oocytes (n = 819) were co-cultured with dromedary oviductal epithelial or granulosa cells in TCM-199 supplemented with 10% heat-treated FCS. Although the rate of first cleavage (two to eight cells) was similar for the two co-culture systems (32 versus 33%, respectively), more embryos (two-cell to blastocyst stage) were obtained from oocytes co-cultured with oviductal versus granulosa cells (61 versus 45%; P < 0.05). The proportions of fertilized oocytes developing to the early morula stage were 19% (80/417) and 12% (48/402) for oocytes co-cultured for 7 days with oviductal or granulosa cells, respectively (P > 0.05). However, development to the blastocyst stage (10% of fertilized oocytes) occurred only in oocytes co-cultured with oviductal cells. In conclusion, dromedary embryos were produced in vitro using abattoir-derived oocytes, fresh (ejaculated) semen, and oviductal cell co-culture.
Veterinary Research Center, Abu Dhabi, United Arab Emirates.
Hadj
H
Khatir
Abdelhaq
A
Anouassi
Ahmed
A
Tibary
D000818:Animals; D002162:Camelus / Q000196:embryology* / Q000502:physiology; D002478:Cells, Cultured; D002970:Cleavage Stage, Ovum / Q000187:drug effects / Q000502:physiology; D018920:Coculture Techniques / Q000662:veterinary; D003538:Cystamine / Q000494:pharmacology; D004815:Epidermal Growth Factor / Q000494:pharmacology; D005187:Fallopian Tubes / Q000166:cytology; D005260:Female; D005307:Fertilization in Vitro / Q000662:veterinary*; D005640:Follicle Stimulating Hormone / Q000494:pharmacology; D006107:Granulosa Cells / Q000166:cytology; D008297:Male; D009865:Oocytes / Q000187:drug effects / Q000254:growth & development / Q000502:physiology*; D006080:Ovarian Follicle / Q000166:cytology / Q000502:physiology; D012661:Semen / Q000502:physiology
D004815:Epidermal Growth Factor; D005640:Follicle Stimulating Hormone; D003538:Cystamine