This study was conducted to explore the effect of energy substrates in the culture medium during in vitro maturation of bovine oocytes. A modified TCM199 medium (M-7528) was used to mature bovine oocytes in vitro. Oocytes were supplemented with different pyruvate (0.1, 0.2, 0.4 mM) and glucose (1.5, 5.6, 20.0 mM) concentrations for 48 hours at 38.5 °C under 5% CO2 atmosphere with 95% humidity. Their maturity was checked at 24 and 48 hours. After 48 hours, the denuded oocytes were stained with fluorescent dye JC-1 and avidin-FITC. Fluorescent dye JC-1 is a membrane permeable to the cell and would indicates membrane activity or its organization. Fluorescence intensity of avidin-FITC determination using corrected total cell fluorescence (CTCF) expressed oxidative stress level. There is a significant contribution of energy substrates towards oocyte maturation. Pyruvate at 0.2 mM produced mature oocytes with a diameter of ≥ 120 μm, promoted oocytes maturation to metaphase II (MII) stage faster and reduced cell’s oxidative stress levels. In comparison, 5.6 mM glucose is the optimum concentration for glucose to reduce cell stress level. Unfortunately, this concentration only produced mature oocytes with a small diameter of up to 116 μm. All changes were significant at the level of p < 0.05. As a conclusion, pyruvate at 0.2 mM is the optimum concentration for in vitro maturation after taking cell’s stress level into consideration.

Apoptosis is a programed cell death that is vital for tissue homeostasis. However, embryo apoptosis had been known to be related to embryo fragmentation which should be avoided in in vitro fertilization (IVF). The purpose of this study was to evaluate the relationship of embryo apoptosis with the grade of immature oocytes and cleavage stage of in vitro produced (IVP) cattle embryos. This study consisted of 345 oocytes collected through ovary slicing. Immature oocytes were graded as A, B and C. This grading was based on cumulus cell thickness and compactness. All oocytes then underwent an in vitro maturation (IVM) procedure. An IVF was done 24 h after IVM culture. Prior to staining, stage of cleaved embryos was determined and classified as either 2, 4, 8 or > 8-cell embryo stage. Apoptosis status of cleaved IVP embryos was determined by using annexin V-FITC staining technique at 48 and 72 h post insemination (hpi). Apoptosis status for each embryo was classified as either early or late. The result showed that there was no significant difference (p > 0.05) of apoptosis status among grade A, B and C embryos. All grades of oocytes showed embryo apoptosis where 1.5% late apoptosis for grade A, 4.5% and 10.4% of early and late apoptosis for grade B and grade C. Early apoptosis was not seen in grade A embryo. We also noted no significant difference (p > 0.05) of apoptosis status between 2, 4, 8 and >8-cell embryo stage. Early apoptosis was also not seen in >8-cell stage. Even though there were no differences in apoptosis expression between the three classes, the cleavage rate of grade A oocytes was significantly higher (p < 0.01) than grade B and grade C. In conclusion, the apoptosis expression in the embryo can occur regardless of the oocyte quality and the cleavage stage of the embryo produced. (C) 2015 The Authors. Production and hosting by Elsevier B.V. on behalf of King Saud University.