Department of Medical Biotechnology, School of Advanced Technologies in Medicine, Shahid Beheshti University of Medical Sciences, Tehran, Iran.
Cellular and Molecular Biology Research Center, Shahid Beheshti University of Medical Sciences, Tehran, Iran.
Biol Reprod. 2020 Aug 21;103(3):548-559. doi: 10.1093/biolre/ioaa098.
Numerous oocytes are retrieved during in vitro fertilization from patients with polycystic ovary syndrome (PCOS). The poor quality of these oocytes leads to lower fertilization and decreases in cleavage and implantation. MiR-155 is one of the microRNA (miRNA) that is increased in serum and granulosa cells of PCOS patients. In this study, we investigate the effects of miR-155 expression and its target genes on oocyte maturation and embryo development. We used the calcium phosphate protocol to transfect vectors that contained miR-155 or miR-off 155 and alone eGFP into cumulus oophorus complex (COCs) of B6D2F1 female mice for in vitro maturation. Cumulus expansion, nuclear, and cytoplasmic maturation, as well as cleavage rates were determined in groups transfected and compared with the control groups. Quantitative real-time polymerase chain reaction was performed to analyze expression levels of miR-155 and the target genes in the cumulus cells, oocytes, and blastocysts. MiR-155 overexpression in COCs suppressed cumulus expansion, oocyte maturation, and inhibition of endogenous miR-155 by miR-off 155 improved cumulus expansion and oocyte maturation by downregulation and expression increase of the Smad2 and Bcl2 genes. On the other hand, overexpression and downregulation of miR-155 in the COCs led to increase and decrease in cleavage rates by changes in expressions of the Mecp2, Jarid2, and Notch1 genes, respectively (P < 0.05). These results suggested that miR-155 overexpression in granulosa cells of PCOS patients can negatively affect nuclear and cytoplasmic maturation, but this miRNA expression has a positive impact on embryo development.
在多囊卵巢综合征（PCOS）患者的体外受精过程中，会从患者体内获取大量卵子。这些卵子质量较差，导致受精率降低，卵裂和着床减少。miR-155 是一种在 PCOS 患者血清和颗粒细胞中升高的 microRNA（miRNA）之一。在这项研究中，我们研究了 miR-155 表达及其靶基因对卵母细胞成熟和胚胎发育的影响。我们使用磷酸钙转染法将包含 miR-155 或 miR-off 155 和单独 eGFP 的载体转染到 B6D2F1 雌性小鼠的卵丘卵母细胞复合体（COC）中进行体外成熟。在转染组和对照组中，确定卵丘扩张、核和细胞质成熟以及卵裂率，并进行比较。通过定量实时聚合酶链反应分析转染组和对照组中 cumulus 细胞、卵母细胞和囊胚中的 miR-155 表达水平和靶基因表达水平。COC 中 miR-155 的过表达抑制了卵丘扩张、卵母细胞成熟，并通过 miR-off 155 抑制内源性 miR-155，下调和表达增加 Smad2 和 Bcl2 基因，从而促进了卵丘扩张和卵母细胞成熟。另一方面，COC 中 miR-155 的过表达和下调分别通过改变 Mecp2、Jarid2 和 Notch1 基因的表达，导致卵裂率增加和减少（P<0.05）。这些结果表明，PCOS 患者颗粒细胞中 miR-155 的过表达可能会对核和细胞质成熟产生负面影响，但这种 miRNA 表达对胚胎发育有积极影响。
