Cellular and Molecular Research Center, Ahvaz Jundishapur University of Medical Sciences, Ahvaz, Iran.
Department of Anatomical Sciences, Faculty of Medicine, Ahvaz Jundishapur University of Medical Sciences, Ahvaz, Iran.
JBRA Assist Reprod. 2024 Jun 1;28(2):289-294. doi: 10.5935/1518-0557.20240019.
Today, researchers have succeeded in achieving oocyte-like cells through the in vitro differentiation of stem cells. MicroRNAs are key regulators of oocyte development. In this study we decided to evaluate the expression pattern of microRNA-21, microRNA-15a, and microRNA-372 in oocyte-like cells, to determine the maturation stage of oocyte-like cells.
Human follicular fluid samples were collected and centrifuged, and their cells were divided into 3 groups; day 7 as control group, days 14 and 21. During this period, the cells were evaluated for their morphological appearance and viability by inverted microscopy. RNA isolation was performed and cDNA was reversely transcribed by specific stem-loop RT primers. Real-time RT-PCR was used to detect microRNA expression.
The relative expression of microRNA-21 and microRNA-15a on day 21 was significantly down-regulated compared to the control group (day 7), but microRNA-372 did not show a significant difference. Also, on day 14 compared to the control group (day 7), microRNA-21 did not show a significant difference; but microRNA-15a and microRNA-372 were significantly down-regulated. MicroRNA-21 and microRNA-15a on day 21 compared to day 14 revealed down-regulated levels, but microRNA-372 revealed up-regulated levels.
Our results showed significant decreases in the expression of microRNA-21 and microRNA-15a in oocyte-like cells, as well as in oocytes, which may lead to cytoplasmic maturation, germinal vesicle break down and the completion of meiosis І. In addition, down-regulation expression of microRNA-372 maybe a confirmation that mesenchymal stem cells have differentiated into germ cells, and these cells were differentiated into oocyte-like cells.
如今，研究人员已经成功地通过体外干细胞分化获得卵母细胞样细胞。microRNA 是卵母细胞发育的关键调节因子。在这项研究中，我们决定评估 microRNA-21、microRNA-15a 和 microRNA-372 在卵母细胞样细胞中的表达模式，以确定卵母细胞样细胞的成熟阶段。
收集人卵泡液样本并离心，将其细胞分为 3 组；第 7 天作为对照组，第 14 天和第 21 天。在此期间，通过倒置显微镜评估细胞的形态外观和活力。通过特定的茎环 RT 引物进行 RNA 分离和 cDNA 反转录。使用实时 RT-PCR 检测 microRNA 表达。
与对照组（第 7 天）相比，第 21 天 microRNA-21 和 microRNA-15a 的相对表达明显下调，但 microRNA-372 没有明显差异。此外，与对照组（第 7 天）相比，第 14 天 microRNA-21 没有明显差异；但 microRNA-15a 和 microRNA-372 明显下调。第 21 天 microRNA-21 和 microRNA-15a 与第 14 天相比呈现下调水平，但 microRNA-372 呈现上调水平。
我们的结果表明，卵母细胞样细胞和卵母细胞中 microRNA-21 和 microRNA-15a 的表达明显下降，可能导致细胞质成熟、生发泡破裂和减数分裂Ⅰ的完成。此外，microRNA-372 的下调表达可能证实间充质干细胞已分化为生殖细胞，这些细胞分化为卵母细胞样细胞。
