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• W4281555319 abstract "Oxidative stress is a major problem in in vitro handling of gametes and embryos. In this study, the effects of Verbascoside (VB), a bioactive polyphenol with known antioxidant activity, added during In Vitro Maturation (IVM) of equine oocytes on blastocyst quality were analyzed after conventional or mercury-free piezo-assisted intracytoplasmic sperm injection (ICSI). Cumulus-oocyte complexes recovered from the ovaries of slaughtered mares underwent overnight holding and IVM in the presence of 0 (control) or 100nM VB. Fresh sperm of fertile stallions was subjected to swim-up procedure and used to microinject matured oocytes by conventional (Dell'Aquila et al. Biology of Reproduction 2003;68:2065-72) or piezo-assisted ICSI (Martino et al. Reproduction Fertility and Development 2019;31:1862-73). Embryos were in vitro cultured in DMEM/F-12 with 10% FCS for 9 days under 5%O2, 5%CO2, 90%N2 atmosphere. Blastocyst quality was assessed for bioenergetic/oxidative status by confocal laser scanning microscopy after labeling mitochondria (mt) with MitoTracker Orange CMTM Ros and reactive oxygen species (ROS) with 2’,7’-dichlorodihydrofluorescein diacetate (Marzano et al. Molecular Reproduction and Development 2019;86:1388-1404). Then, blastocysts were stained with Hoechst 33258 for total cell numberevaluation and confirmed when showing more than 64 nuclei with organization of an outer trophoblast layer. Proportions of blastocyst rates after ICSI were compared between VB-exposed group and controls by the Chi-square-test, whereas blastocyst total cell number, ROS levels and mt/ROS colocalization were compared by the Student's T-test. Values with P<0.05 were considered significantly different. VB addition during IVM did not influence oocyte maturation rate (31/55, 56% vs 38/76, 50% for VB-exposed and CTRL, respectively). After both conventional and piezo-assisted ICSI, VB had no effects on blastocyst formation rate (4/21, 19% vs 3/24, 12.5%; P>0.05 and 3/10, 30% vs 2/14, 14%; P>0.05) and total cell number (187.3±104.7 vs 143.3±63.2, P>0.05; and 247.3±109.8 vs 465±17.0, P>0.05). Instead, after both microinjection procedures, VB significantly reduced ROS levels (29.7±5.1 vs 36.1±4.2, P<0.001; and 22.8±8.9 vs 30.1±4.4, P<0.01) and increased mt/ROS colocalization (0.63±0.04 vs 0.61±0.03, P<0.001; 0.79±0.06 vs 0.74±0.12, P<0.001). These findings indicate long-term beneficial antioxidant effects of in vitro exposure to nanomolar VB concentrations during oocyte IVM on equine blastocyst quality. Acknowledgments: funded by the Italian Ministry of Health: Project GR-2011-02351396." @default.
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• W4281555319 date "2022-06-01" @default.
• W4281555319 modified "2023-09-29" @default.
• W4281555319 title "Verbascoside addition during IVM improves equine blastocyst redox status after conventional and mercury-free piezo-assisted ICSI" @default.
• W4281555319 doi "https://doi.org/10.1016/j.jevs.2022.103973" @default.
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