Abstract
This study was conducted to investigate the effects of sericin on meiotic maturation, hydrogen peroxide (H 2 O 2 ) concentration and deoxyribonucleic acid (DNA) fragmentation in buffalo oocytes. Oocytes were matured in vitro in tissue culture medium (TCM199) with in vitro maturation (IVM) groups under several conditions, namely without bovine serum albumin (BSA), (+BSA), and 0.025%, 0.05%, 0.1% and 0.25% (w/v) sericin. The results showed that supplementation of the maturation medium with 0.05% sericin significantly increased the rate of oocytes that reach metaphase II compared with other groups, except for the 0.025% sericintreated group. Intracellular H 2 O 2 concentrations in oocytes of all groups were measured using 2',7'dichlorodihydrofluorescein diacetate (DCHFDA). The concentration of H 2 O 2 in matured oocytes treated with 0.05% sericin was lower than in other groups. DNA fragmentation and the nuclear status of oocytes were examined using the terminal deoxyribonucleotidyl transferase-mediated dUTP digoxigenin nick endlabelling (TUNEL) method. The total proportion of TUNELpositive oocytes at the MII stage was lower in the 0.05% sericin group. The results indicate that addition of 0.05% sericin to the maturation medium may improve nuclear maturation and may inhibit DNA fragmentation in oocytes by decreasing H2O2 concentrations. Keywords: antioxidant, apoptosis, in vitro maturation, reactive oxygen species
Highlights
In buffalo, the efficiency of in vitro embryo production (IVEP) is still low (Neglia et al, 2003; Drost, 2007)
Each ovary was sliced with a scalpel blade to release the cumulus-oocyte complexes in phosphate buffered saline (PBS) supplemented with 0.3% bovine serum albumin (BSA) (Sigma-Aldrich) and antibiotics
The metaphase II (MII) rate of oocytes that were cultured in 0.025% sericin was significantly higher than that of oocytes cultured in (BSA) (P ≤0.05), the rate was not significantly different from oocytes in the (+BSA) group
Summary
The efficiency of in vitro embryo production (IVEP) is still low (Neglia et al, 2003; Drost, 2007). Only 20% of buffalo oocytes that are fertilized in vitro form blastocysts (Gasparrini, 2002; Nandi et al, 2002; Manjunatha et al, 2008) This inefficiency in buffalo IVEP could be associated with the quality of the oocytes at the beginning of the maturation process, oocytes were selected based on the compaction of cumulus cells and the homogeneity of ooplasm (Chauhan et al, 1998). The oxygen tension in in vitro culture systems (≈ 20%) is often higher than in the female reproductive tract (3% to 9%) (Rocha-Frigoni et al, 2016) This condition is believed to possibly induce the generation of reactive oxygen species (ROS), such as superoxide anions, hydroxyl radicals and hydrogen peroxide (H2O2) (Morado et al, 2009).
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