141 Effect of different quantities of epidermal growth factor and TCM-199 medium on polar body extrusion of cattle oocytes following in vitro maturation

Epidermal growth factor (EGF) has been shown to stimulate development of cumulus cells with or without supplementation of hormones or serum. As to its role within the oocyte, EGF appears to have a positive impact during IVM in different species. Cattle oocytes mature in vivo at the time of the preovulatory gonadotrophin surge, but can develop in vitro in a suitable medium. Increasing numbers of studies have evaluated the effect of growth factors on nuclear and cytoplasmic development of mammalian oocytes. The objective of this study was to evaluate polar body extrusion following IVM of cattle oocytes in different media quantities of EGF and TCM 199 (500 µL of TCM-199, 400 µL of TCM 199 + 100 µL of EGF, 400 µL of EGF + 100 µL of TCM199, or 250 µL of TCM 199 + 250 µL of EGF). The concentration of EGF medium was 10 µg mL⁻¹ dissolved into 2 mL of M199. Matured cattle ovaries from unknown reproductive status were collected from local slaughterhouse and transported to the laboratory in a thermos flask containing 0.9% saline (Adcock Ingram Critical Care) at 37°C. Oocytes were collected from the ovaries by the aspiration technique, and FSH, LH, and oestradiol) or 400 µL of TCM 199 + 100 µL of EGF or 400 µL of EGF + 100 µL of TCM199, or 250 µL of TCM 199 + 250 µL of EGF all covered with mineral oil. Oocytes were then incubated for 22 h at 38.5°C with 5% CO₂ and 5% O₂ as per their respective groups. After 22 h of IVM, oocyte polar body extrusion was evaluated in all groups with the aid of the Oosight Imaging System (Hamilton-Thorne) connected to an inverted microscope. The total number of oocytes matured per treatment was 260. Data was analysed using the GenStat® program (VSN International). Means of different treatments were separated using Fisher’s protected t-test least significant difference at the 5% level of significance. No difference was recorded for oocyte polar body extrusion rates in 500 µL of TCM-199 (41.7 ± 15.5), 400 µL TCM 199 + 100 µL EGF (35.7 ± 10.1), 400 µL EGF + 100 µL TCM199 (33.2 ± 14.0), and 250 µL TCM 199 + 250 µL EGF (38.8 ± 8.2) media (P > 0.05). In conclusion, the use of TCM 199 and EGF in different quantities did not differ in terms of cattle oocyte maturation rate. Therefore, any of the quantities can be used for maturation of oocytes from cattle. However, a combination of TCM 199 plus EGF is capable of enhancing the maturation of cattle oocytes because it stimulates cumulus expansion of oocytes. Studies using the same quantities of TCM 199 and further supplemented EGF media with hormones and serum should be conducted to improve the maturation rate in cattle oocyte.