Integrated ultrasensitive metabolomics and single-cell transcriptomics identify crucial regulators of sheep oocyte maturation and early embryo development in vitro,Journal of Advanced Research

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Integrated ultrasensitive metabolomics and single-cell transcriptomics identify crucial regulators of sheep oocyte maturation and early embryo development in vitro
Journal of Advanced Research ( IF 11.4 ) Pub Date : 2024-09-02 , DOI: 10.1016/j.jare.2024.08.040
Bo Pan ₁ , JianPeng Qin ₁ , KunLin Du ₁ , LuYao Zhang ₂ , GongXue Jia ₂ , JiangFeng Ye ₁ , QiuXia Liang ₃ , QiEn Yang ₂ , GuangBin Zhou ₁

Affiliation

Introduction

Developmental competence of oocytes matured in vitro is limited due to a lack of complete understanding of metabolism and metabolic gene expression during oocyte maturation and embryo development. Conventional metabolic analysis requires a large number of samples and is not efficiently applicable in oocytes and early embryos, thereby posing challenges in identifying key metabolites and regulating their in vitro culture system.

Objectives

To enhance the developmental competence of sheep oocytes, this study aimed to identify and supplement essential metabolites that were deficient in the culture systems.

Methods

The metabolic characteristics of oocytes and embryos were determined using ultrasensitive metabolomics analysis on trace samples and single-cell RNA-seq. By conducting integrated analyses of metabolites in cells (oocytes and embryos) and their developmental microenvironment (follicular fluid, oviductal fluid, and in vitro culture systems), we identified key missing metabolites in the in vitro culture systems. In order to assess the impact of these key missing metabolites on oocyte development competence, we performed in vitro culture experiments. Furthermore, omics analyses were employed to elucidate the underlying mechanisms.

Results

Our findings demonstrated that betaine, carnitine and creatine were the key missing metabolites in vitro culture systems and supplementation of betaine and L-carnitine significantly improved the blastocyst formation rate (67.48% and 48.61%). Through in vitro culture experiments and omics analyses, we have discovered that L-carnitine had the potential to promote fatty acid oxidation, reduce lipid content and lipid peroxidation level, and regulate spindle morphological grade through fatty acid degradation pathway. Additionally, betaine may participate in methylation modification and osmotic pressure regulation, thereby potentially improving oocyte maturation and early embryo development in sheep.

Conclusion

Together, these analyses identified key metabolites that promote ovine oocyte maturation and early embryo development, while also providing a new viewpoint to improve clinical applications such as oocyte maturation or embryo culture.

中文翻译：

集成的超灵敏代谢组学和单细胞转录组学可识别绵羊卵母细胞成熟和体外早期胚胎发育的关键调节因子

介绍

由于对卵母细胞成熟和胚胎发育过程中的新陈代谢和代谢基因表达缺乏完全了解，体外成熟的卵母细胞的发育能力受到限制。传统的代谢分析需要大量的样品，不能有效地应用于卵母细胞和早期胚胎，因此对识别关键代谢物和调节其体外培养系统提出了挑战。

目标

为了提高绵羊卵母细胞的发育能力，本研究旨在识别和补充培养系统中缺乏的基本代谢物。

方法

对痕量样品和单细胞 RNA-seq 进行超灵敏代谢组学分析，测定卵母细胞和胚胎的代谢特性。通过对细胞（卵母细胞和胚胎）及其发育微环境（卵泡液、输卵管液和体外培养系统）中的代谢物进行综合分析，我们确定了体外培养系统中关键的缺失代谢物。为了评估这些关键缺失代谢物对卵母细胞发育能力的影响，我们进行了体外培养实验。此外，采用组学分析来阐明潜在机制。

结果

我们的研究结果表明，甜菜碱、肉碱和肌酸是体外培养系统中关键的缺失代谢产物，补充甜菜碱和左旋肉碱显着提高了囊胚形成率（67.48% 和 48.61%）。通过体外培养实验和组学分析，我们发现左旋肉碱具有促进脂肪酸氧化、降低脂质含量和脂质过氧化水平、通过脂肪酸降解途径调节纺锤体形态分级的潜力。此外，甜菜碱可能参与甲基化修饰和渗透压调节，从而有可能改善绵羊的卵母细胞成熟和早期胚胎发育。