Zhao Xinyang, Zhang Xudong, Wu Shanshan, Zhang Siwen, Chang Hsun-Ming, Leung Peter C K, Tan Jichun
Center of Reproductive Medicine, Department of Obstetrics and Gynecology, Shengjing Hospital of China Medical University, No.39 Huaxiang Road, Tiexi District, Shenyang, Liaoning, 110022, China; Key Laboratory of Reproductive Dysfunction Disease and Fertility Remodeling of Liaoning Province, China.
Center of Reproductive Medicine, Department of Obstetrics and Gynecology, Shengjing Hospital of China Medical University, No.39 Huaxiang Road, Tiexi District, Shenyang, Liaoning, 110022, China; Key Laboratory of Reproductive Dysfunction Disease and Fertility Remodeling of Liaoning Province, China.
Theriogenology. 2025 Oct 1;245:117518. doi: 10.1016/j.theriogenology.2025.117518. Epub 2025 May 31.
The effective utilization of immature oocytes holds promise for improving conception rates in women with low ovarian reserve. The oocyte microenvironment, comprising the extracellular matrix and intercellular communication with somatic cells, is crucial for supporting oocyte maturation. This study hypothesizes that a three-dimensional (3D) culture system, constructed with hydrogels modified by laminin-mimetic peptides-a major component of the extracellular matrix-and granulosa cells (GCs) from mature mouse oocytes, can replicate the biomechanical microenvironment required for oocyte maturation, thereby promoting the development of immature mouse oocytes. To validate this hypothesis, we initially assessed the physical properties of hydrogels and determined the optimal concentration for in vitro oocyte culture. A total of 304 germinal vesicle (GV) phase mouse oocytes were retrieved and randomly allocated to two-dimensional (2D), 3D, and 3D-GCs culture groups. The 3D-GCs group exhibited the highest rate of first polar body extrusion (83.1 ± 5.0 %). Oxidative stress levels and mitochondrial membrane potential were evaluated using DCFH-DA and JC-1 staining, confirming that 3D culture significantly reduced oxidative stress and enhanced mitochondrial activity. Cytoplasmic maturation, assessed by cortical granule distribution and endoplasmic reticulum organization, further highlighted the superiority of the 3D-GCs system. The 3D-GCs group demonstrated 87.0 % of oocytes with grade III cortical granule distribution and 84.6 % with mature endoplasmic reticulum, significantly surpassing the other groups (p < 0.05). In conclusion, the 3D-GCs culture system effectively supports synchronized nuclear and cytoplasmic maturation in immature oocytes. This approach provides a promising platform for improving the developmental competence of oocytes, potentially benefiting women with diminished ovarian reserve.
有效利用未成熟卵母细胞有望提高卵巢储备功能低下女性的受孕率。卵母细胞微环境由细胞外基质和与体细胞的细胞间通讯组成，对支持卵母细胞成熟至关重要。本研究假设，用层粘连蛋白模拟肽（细胞外基质的主要成分）修饰的水凝胶和来自成熟小鼠卵母细胞的颗粒细胞构建的三维（3D）培养系统，可以复制卵母细胞成熟所需的生物力学微环境，从而促进未成熟小鼠卵母细胞的发育。为了验证这一假设，我们首先评估了水凝胶的物理性质，并确定了体外卵母细胞培养的最佳浓度。总共收集了304个生发泡（GV）期小鼠卵母细胞，并随机分配到二维（2D）、3D和3D-颗粒细胞培养组。3D-颗粒细胞组的第一极体排出率最高（83.1±5.0%）。使用DCFH-DA和JC-1染色评估氧化应激水平和线粒体膜电位，证实3D培养显著降低了氧化应激并增强了线粒体活性。通过皮质颗粒分布和内质网组织评估的细胞质成熟进一步突出了3D-颗粒细胞系统的优势。3D-颗粒细胞组显示87.0%的卵母细胞皮质颗粒分布为III级，84.6%的卵母细胞内质网成熟，显著超过其他组（p<0.05）。总之，3D-颗粒细胞培养系统有效地支持未成熟卵母细胞的核和细胞质同步成熟。这种方法为提高卵母细胞的发育能力提供了一个有前景的平台，可能使卵巢储备功能减退的女性受益。
