N-acetylcysteine addition after vitrification improves oocyte mitochondrial polarization status and the quality of embryos derived from vitrified murine oocytes

Abstract

BackgroundVitrification is the safest method to cryopreserve oocytes, however the process alters mitochondrial function resulting from increased reactive oxygen species (ROS) production. Our aim was to alleviate ROS stress in vitrified mice oocytes using N-acetylcysteine (NAC at 1 mM), to improve the oocyte’s developmental competence.ResultsHence, four experimental groups were compared: fresh oocytes (F-C), vitrified oocytes (V-C), NAC addition prior to oocyte vitrification (V-NAC-Pre) and NAC addition after vitrification (V-NAC-Post). V-NAC-Pre and V-NAC-Post exhibited higher levels of mitochondrial polarization compared to vitrified oocytes (36.5 ± 3.1, 37.7 ± 1.3 and 27.2 ± 2.4 measured as the spatial coefficient of variation/oocyte respectively, mean ± SEM; p < 0.05). However, ROS production increased in vitrified oocytes added with NAC compared to the vitrified control (1124.7 ± 102.1 [V-NAC-Pre] and 1063.2 ± 82.1 [V-NAC-Post] vs. 794.6 ± 164.9 [V-C]; arbitrary fluorescence units/oocyte, mean ± SEM; p < 0.05). ATP significantly decreased in V-NAC-Pre compared to V-NAC-Post oocytes (18.5 ± 6.9 vs. 54.2 ± 4.6 fmol/oocyte respectively, mean ± SEM; p < 0.05), and no differences were observed between V-NAC-Post, F-C and V-C groups. Blastocyst rates derived from F-C oocytes was higher than those derived from V-NAC-Pre (90.7 ± 1.8 vs. 79.1 ± 1.8, respectively, mean % ± SEM,; p < 0.05) but similar to those derived from V-NAC-Post (90.7 ± 1.8, mean % ± SEM, p > 0.05). Total blastomere count of blastocysts derived from V-NAC-Post after in vitro fertilization (IVF) was higher than embryos produced from V-C.ConclusionsThe addition of NAC after vitrification improves the quality of vitrified mature murine oocytes while its addition prior to vitrification is advised against.

N-acetyl cysteine restores the fertility of vitrified–warmed mouse oocytes derived through ultrasuperovulation

“…NAC enhances the cryopreservation of mouse oocytes. For instance, after vitrification, 1.0 mM NAC improved mitochondrial activity and vitrified–warmed oocyte maturation [31]. When mouse oocytes were cryopreserved at the germinal vesicle stage, 1.5 mM NAC added to the media for vitrification and in vitro maturation enhanced maturation by inhibiting reactive oxygen species generation [32].…”

Resveratrol enhanced mitochondrial recovery from cryopreservation‐induced damages in oocytes and embryos

“… 27 , 28 Therefore, the effect of antioxidants on oocytes and embryo following vitrification has been extensively studied; supplementation of culture medium of vitrified‐warmed oocytes or embryos with melatonin, glutathione ethyl‐ester, N‐acetyl cysteine, α‐tocopherol, or coenzyme Q reduced ROS levels and improved oocyte viability as well as embryo development in equine, cows, and mice. 6 , 29 , 30 , 31 , 32 In addition, supplementation of vitrified‐warming solution with acetyl‐L‐carnitine, N‐acetyl‐cysteine, and lipoic acid improved the cell number of the blastocysts and fetal development in mice. 33 Furthermore, addition of vitamin E to the culture medium of vitrified‐warming ovarian tissue improved oocyte and subsequent blastocyst stage development.…”

Antioxidant procyanidin B2 protects oocytes against cryoinjuries via mitochondria regulated cortical tension

“…ROS can attack proteins, DNA, cell membrane, and also microtubules, thereby disrupting oocyte structure and function [36,38]. Since vitrified oocytes are under oxidative stress, antioxidants are introduced in conventional cryo-solutions to protect oocytes from cryoinjuries [5,[39][40][41][42]. N-acetylcysteine [40] can improve the quality of mature mouse oocytes after vitrification, melatonin [5] can enhance the efficiency of human oocytes' cryopreservation and resveratrol [39] improves the development of vitrified bovine embryos.…”