细胞松弛素B改善冷冻引起的猪卵母细胞皮质颗粒迁移障碍

doi:10.11843/j.issn.0366-6964.2024.05.018

Abstract

Abstract: The purpose of this study was to reveal the interrelationships and effects of cytoskeleton and cortical granule migration during in vitro maturation of frozen porcine GV-stage oocytes. Pig ovary samples were collected from healthy multiparous sows.For each sampling, about 300 Pudong White sows weighing 70-90 kg with good production performance were used to take ovarian samples, the number of ovaries collected was 120-150 per time, and the GV stage oocytes with homogeneous and compact cytoplasmic distribution were selected and randomly divided into 3 groups according to the experimental requirements, and 3 replicates were set up in each group, with at least 30 cells in each group, and cytoskeleton stabilizers, cytochalasin B (CB) were used to treat the frozen porcine oocytes before freezing. Porcine GV oocytes were treated with cytoskeleton stabilizer CB before freezing, and then matured in vitro after freezing and thawing. Oocyte survival, fluorescence co-localization of microfilaments and cortical granules at each time point, degree of spreading of the oval mound, rate of discharge of the first polar body, level of glutathione (GSH), and developmental ability were examined. The result showed that treatment with CB before freezing effectively increased the survival rate of GV stage oocytes after thawing (44.11% vs. 27.91%, P<0.01). During the maturation of fresh oocytes, co-migration of cortical granules with microfilaments was observed. Freezing caused abnormal cortical granule-microfilament migration, and the addition of CB prior to freezing alleviated the freezing-induced migration obstacles, as evidenced by a higher proportion of both microfilaments and cortical granules successfully migrating to the plasma membrane in the CB-treated group, and a better homogeneous distribution of migration to the plasma membrane. After in vitro maturation of frozen oocytes, CB pretreatment was associated with a higher rate of discharge of first polar body ((26.79±2.37)% vs. (8.13±0.30)%, P<0.01), GSH levels (23.12±2.65 vs. 7.27±0.79, P<0.01), cleavage rate after parthenogenetic activation ((20.91±2.84)% vs. (5.64±0.37)%, P<0.05) and blastocyst rate ((5.00±0.03)% vs. (0.41±0.01)%, P<0.05) were significantly increased. The present study showed that the microfilament cytoskeleton has a certain regulatory effect on the migration of intracellular cortical particles from the interior to the plasma membrane during oocyte maturation in vitro, and there exists a certain co-localization relationship between the two, and that the addition of CB incubation before freezing can effectively affect the degree of migration of the cellular microfilament cytoskeleton and cortical particles, mitigate the abnormal distribution of the cytoskeleton induced by freezing, and ease the migration obstruction of cortical particles and microfilaments induced by freezing, thereby enhancing cellular resistance, which is manifested as promoting cytoplasmic maturation, improving the survival rate of frozen oocytes, nuclear maturation rate and embryo development potential in vitro.

Key words: cortical granules, microfilament, vitrification, pig oocyte

CLC Number:

S828.3

LI Wanjun, XU Jiehuan, HE Mengxian, KONG Yuting, ZHANG Defu, DAI Jianjun. Cytochalasin B Alleviates the Migration Disorder of Cortical Particle Caused by Vitrification in Porcine Oocytes[J]. Acta Veterinaria et Zootechnica Sinica, 2024, 55(5): 1999-2010.

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Cytochalasin B Alleviates the Migration Disorder of Cortical Particle Caused by Vitrification in Porcine Oocytes

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