LncRNA-mediated effects of vitrification temperatures and cryoprotectant concentrations on bovine oocyte development following vitrification at the GV stage
LncRNA-mediated effects of vitrification temperatures and cryoprotectant concentrations on bovine oocyte development following vitrification at the GV stage
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LncRNA 介导的玻璃化温度和冷冻保护剂浓度对 GV 阶段玻璃化后牛卵母细胞发育的影响。
DOI：
10.1016/j.theriogenology.2022.03.028
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发表时间：
2022-04-22
期刊：
影响因子：
2.8
通讯作者：
Li, Xiao-Xia
中科院分区：
文献类型：
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作者：
Cai, Meng-Dan;Xu, Zhi-Qian;Li, Xiao-Xia
研究了不同玻璃化冷冻温度（VT）和冷冻保护剂浓度（CPA）对牛卵母细胞萌发泡（GV）期玻璃化冷冻后存活率和长链非编码RNA（lncRNA）表达的影响。本研究结果为改进牛未成熟卵母细胞冷冻保存技术提供了理论依据。收集牛卵丘卵母细胞复合体（COCs），随机分为5组：新鲜卵母细胞（对照组）、液氮玻璃化冷冻组（冷冻组）、冷冻后的卵母细胞（冷冻组）和冷冻后的卵母细胞（冷冻组）（LHe; -269 ℃），5.6 M CPA（LHe 5.6 M），卵母细胞在含6.6 M CPA的LHe中玻璃化冷冻6 M CPA（LN 5.6M）在液氮（LN; -196 ° C）中玻璃化的卵母细胞，以及在具有6.6M CPA（LN 6.6M）的LN中玻璃化的卵母细胞。在4个玻璃化冷冻组中，LHe 5.6 M组的囊胚率最高（13.22%），其次是LHe 6.6 M组（10.19%）和LN 6.6 M组（9.77%），而LN 5.6 M组的囊胚率最低（1.87%）。然后，分析五组中lncRNA的表达。与新鲜组相比，玻璃化冷冻组中总共鉴定了18，271个lncRNA，其中2，158个是差异表达的lncRNA（DEL）（P < 0.05;倍数变化> 2）。共定位（顺式）和共表达（反式）预测显示14个差异表达的靶基因（DETG），对应于17个DEL。基于DEL的分组数据和表达谱，我们证明了不同的VT（-269 ° C与-196 ° C）可以影响MSTRG. 12295. 5、MSTRG. 37123. 1、MSTRG. 37930. 2、MSTRG. 40464. 9、MSTRG. 8869. 3和MSTRG. 26680. 6的表达。这些lncRNA的表达仅在LHe玻璃化（-269 ℃）条件下受CPA影响。MSTRG.35129.6的表达与VT和CPA的暴露相关;而MSTRG. 3578.3、MSTRG. 40576.3、MSTRG. 6723.5、MSTRG. 32862.4、MSTRG. 1184.4、MSTRG. 33110.3、MSTRG. 40454.2、MSTRG. MSTRG. 44732. 4和MSTRG. 6729. 3可能与玻璃化有关。共表达分析表明，MSTRG. 12295. 5、MSTRG. 37930. 2、MSTRG. 40454. 2、MSTRG. 8869. 3和MSTRG. 6723. 5的表达通过调节靶基因的表达影响卵母细胞玻璃化冷冻后的发育。总之，LHe玻璃化冷冻后生物体发育能力的提高可能归因于一些lncRNA表达的变化。我们的研究结果阐明了在不同的VT和CPA下BIO发展的分子机制。(c)2022爱思唯尔公司All rights reserved.
We evaluated the effects of different vitrification temperatures (VTs) and cryoprotective agent concentrations (CPAs) on the viability and expressions of long non-coding RNA (lncRNA) in bovine oocytes following vitrification at the germinal vesicle (GV) stage. Our findings provide a theoretical support for improvement of the cryopreservation technology of bovine immature oocytes (BIOs). Bovine cumulus oocyte complexes (COCs) were collected and randomized into five groups: fresh oocytes (control), oocytes vitrified in liquid helium (LHe; -269 degrees C) with 5.6 M CPAs (LHe 5.6 M), oocytes vitrified in LHe with 6.6 M CPAs (LHe 6.6 M), oocytes vitrified in liquid nitrogen (LN; -196 degrees C) with 5.6 M CPAs (LN 5.6 M), and oocytes vitrified in LN with 6.6 M CPAs (LN 6.6 M). Of the four vitrification groups, the LHe 5.6 M group exhibited the highest blastocyst rate (13.22%), followed by the LHe 6.6 M group (10.19%) and LN 6.6 M group (9.77%), while the LN 5.6 M group had the lowest blastocyst rate (1.87%). Then, lncRNA expressions in the five groups were profiled. A total of 18,271 lncRNAs were identified, of which 2,158 were differentially expressed lncRNAs (DELs) in the vitrified groups, compared to the fresh group (P < 0.05; fold-change > 2). Co-location (cis) and co-expression (trans) prediction revealed 14 differentially expressed target genes (DETGs), which corresponded to 17 DELs. Based on grouping data and expression profiles of the DELs, we demonstrated that different VTs (-269 degrees C vs. -196 degrees C) can affect the expressions of MSTRG.12295.5, MSTRG.37123.1, MSTRG.37930.2, MSTRG.40464.9, MSTRG.8869.3 and MSTRG.26680.6. Expressions of these lncRNAs were affected by CPAs only in the condition of vitrification with LHe (-269 degrees C). Expressions of MSTRG.35129.6 were associated with exposures to both VTs and CPAs; while expressions of MSTRG.3578.3, MSTRG.40576.3, MSTRG.6723.5, MSTRG.32862.4, MSTRG.1184.4, MSTRG.33110.3, MSTRG.40454.2, MSTRG.41073.2, MSTRG.44732.4 and MSTRG.6729.3 might be related to vitrification. Co-expression analysis showed that MSTRG.12295.5, MSTRG.37930.2, MSTRG.40454.2, MSTRG.8869.3 and MSTRG.6723.5 expressions affect oocyte development after vitrification by regulating target gene expressions. Taken together, improvement of the developmental ability of BIOs after LHe vitrification maybe attributed to changes in expressions of some lncRNAs. Our findings elucidate on the molecular mechanisms underlying the development of BIOs under different VTs and CPAs. (c) 2022 Elsevier Inc. All rights reserved.