Journal of Applied Biosciences (J. Appl. Biosci.) [ISSN 1997 - 5902]
Volume 24: 1520 - 1534. Published December 7, 2009.
Effect of hormones, culture media and oocyte quality on in vitro maturation of Egyptian Sheep oocytes
I.M. Farag* , S.M. Girgis*^ , W.K.B.Khalil*, N.H.A. Hassan**, A.A.M.Sakr *, S.M. Abd Allah ***, N. I. Ali*
*Cell Biology Department, National Research Centre, 12622- Dokki, Cairo, Egypt.
**Zoology Department, Faculty of Science, Ain Shams University, Cairo, Egypt.
***Theriogenology Department., Faculty of Veterinary Medicine, Benisuef University Benisuef, Egypt.
ABSTRACT
Objective: The present study was carried out to investigate the role of adding hormone combinations (PMSG+HCG+E2) alone with or without fetal bovine serum (FBS) to the culture media (TCM-199 or RPMI-1640) on the in vitro maturation (IVM) of sheep oocytes. The effects of type of culture media and oocyte quality were also investigated.
Methodology and results: Sheep ovaries were collected form local slaughterhouse. Cumulus oocyte complexes COCs and cumulus denuded oocytes [CDOs] were recovered from visible antral follicles (2-6mm) by aspiration method. The collected oocytes were matured in culture media for 26-29h at 39ْC under 5% CO2 in air and 95% humidity. The results indicated that the addition of hormones combined with FBS to the basic medium (TCM-199) significantly (P<0.05) improved the IVM of COCs as compared to the control (medium alone) (41.25 vs. 3.50, respectively). Supplementing TCM – 199 with hormones alone (PMSG+HCG+E2) insignificantly increased the IVM of COCs compared to the control (14.75 vs. 3.50, respectively). However, supplementing TCM – 199 medium with hormones alone or hormones combined with FBS did not improve the IVM of denuded oocytes. On the other hand, the addition of hormones combined with FBS to RPMI medium significantly (P<0.05) improved the IVM of COCs as compared to the control (medium alone) (7.22 vs. 4.69, respectively). However, the addition of hormones alone to RPMI medium did not improve the IVM of COCs. Supplementing RPMI medium with hormones alone significantly (P<0.05) improved the IVM of denuded oocytes as compared to the control group (3.30 vs. 0.00, respectively). In contrast, the addition of hormones plus FBS did not improve the IVM of denuded oocytes. Concerning, the effect of type of culture media on the IVM of sheep oocytes, the results showed that the proportion of oocytes (COCs or denuded oocytes) reaching MII significantly increased (P<0.05) in the group that were matured in TCM-199 medium compared to RPMI medium (19.84 vs. 3.98; or 2.50 vs. 1.07, respectively). The effect of oocye quality on IVM showed that COCs had higher maturation rates compared to denuded oocytes in either TCM-199 (P<0.01) supplemented groups (23.98 vs. 2.57%, respectively) or in RPMI supplemented groups (2.59 vs. 1.36%, respectively).
Conclusion and application of findings: The present study demonstrated that the addition of hormone combinations (PMSG + hCG + E2) with FBS to culture media (TCM- 199 or RPMI- 1640) could significantly improve the IVM of sheep oocytes especially COCs). TCM – 199 medium is more effective for in vitro maturation of sheep oocytes than RPMI- 1640 medium. For optimum oocyte nuclear maturation, the use of COCs is recommended.
Key words: Hormones, culture media, oocyte quality, sheep,cytogenetics, nuclear maturation.
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