Department of Theriogenology and Biotechnology, College of Veterinary Medicine, Seoul National University, Seoul, Korea.
Department of Internal Medicine, College of Medicine, Seoul National University, Seoul, Korea.
J Pineal Res. 2017 Oct;63(3). doi: 10.1111/jpi.12424. Epub 2017 Jun 9.
Melatonin, which is synthesized in the pineal gland and peripheral reproductive organs, has antioxidant properties and regulates physiological processes. It is well known that melatonin affects in vitro maturation (IVM) of oocytes and embryonic development in many species. However, beneficial effects of melatonin on IVM have been explained mainly by indirect antioxidant effects and little information is available on the underlying mechanism by which melatonin directly acts on porcine cumulus oocyte complexes (COCs). Sonic hedgehog (Shh) signaling is important for follicle development, oocyte maturation, and embryo development, and there may be a relationship between melatonin and Shh signaling. To examine this, we designed three groups: (i) control, (ii) melatonin (10 mol/L), and (iii) melatonin with cyclopamine (2 μmol/L; Shh signaling inhibitor). The aim of this study was to investigate the effects of these agents on cumulus expansion, oocyte maturation, embryo development after parthenogenetic activation (PA), gene expression in cumulus cells, oocytes and blastocysts, and protein expression in COCs. Melatonin significantly increased the proportion of COCs exhibiting complete cumulus expansion (degree 4), PA blastocyst formation rates, and total cell numbers, which were inhibited by addition of cyclopamine. Simultaneously, the expression of cumulus expansion-related genes (Ptgs1, Ptgs2, and Has2) and Shh signaling-related genes (Shh, Pthc1, Smo, and Gli1) and proteins (Ptch1, Smo, and Gli1) in cumulus cells was upregulated in the melatonin-treated group, and these effects were also inhibited by cyclopamine. In conclusion, our results suggest that Shh signaling mediates effects of melatonin to improve porcine cumulus expansion and subsequent embryo development.
褪黑素在松果腺和外周生殖器官中合成，具有抗氧化特性，并调节生理过程。众所周知，褪黑素会影响许多物种的卵母细胞体外成熟（IVM）和胚胎发育。然而，褪黑素对 IVM 的有益影响主要通过间接的抗氧化作用来解释，关于褪黑素如何直接作用于猪卵丘卵母细胞复合物（COC）的机制知之甚少。Sonic hedgehog（Shh）信号对于卵泡发育、卵母细胞成熟和胚胎发育很重要，褪黑素和 Shh 信号之间可能存在关系。为了研究这一点，我们设计了三组：（i）对照组，（ii）褪黑素（10 摩尔/L），和（iii）褪黑素加环巴胺（2 微摩尔/L；Shh 信号抑制剂）。本研究旨在探讨这些药物对卵丘扩展、卵母细胞成熟、孤雌激活（PA）后胚胎发育、卵丘细胞、卵母细胞和囊胚的基因表达以及 COC 中的蛋白表达的影响。褪黑素显著增加了完全卵丘扩展（程度 4）、PA 囊胚形成率和总细胞数的 COC 比例，而添加环巴胺则抑制了这些作用。同时，在褪黑素处理组中，卵丘扩展相关基因（Ptgs1、Ptgs2 和 Has2）和 Shh 信号相关基因（Shh、Pthc1、Smo 和 Gli1）以及卵丘细胞中的蛋白（Ptch1、Smo 和 Gli1）的表达上调，而这些作用也被环巴胺抑制。总之，我们的结果表明，Shh 信号介导了褪黑素改善猪卵丘扩展和随后胚胎发育的作用。
