miR-24-3p对猪颗粒细胞雌二醇合成的作用

doi:10.11843/j.issn.0366-6964.2024.01.017

摘要/Abstract

摘要： 旨在探究miR-24-3p对猪颗粒细胞雌二醇合成的影响。本试验收集180日龄健康母猪的卵巢组织，每次试验取20对卵巢进行颗粒细胞的分离培养，将miR-24-3p的mimics及inhibitor转染进颗粒细胞，通过ELISA、RT-qPCR、Western blot、双荧光素酶报告试验等技术探究miR-24-3p对猪颗粒细胞雌二醇合成的作用。结果表明，过表达miR-24-3p可显著促进雌二醇的合成（P<0.01），并加快StAR、CYP19A1和CYP11A1的转录和翻译（P<0.05）；而干扰miR-24-3p则显著抑制雌二醇的合成（P<0.05），并显著下调CYP11A1、CYP19A1的mRNA和蛋白水平（P<0.05）。进一步研究发现，TOP1是miR-24-3p的直接靶基因，过表达miR-24-3p可显著抑制TOP1 的表达（P<0.05），干扰miR-24-3p可显著上调TOP1的表达（P<0.05）。而过表达TOP1则可减弱miR-24-3p对颗粒细胞雌二醇合成的促进作用（P<0.05）。综上所述，miR-24-3p通过靶向TOP1抑制其mRNA和蛋白水平，促进雌二醇合成相关基因的表达水平从而促进颗粒细胞的雌二醇合成。鉴于颗粒细胞的雌二醇合成能力直接影响卵巢卵泡的发育状态，因此本研究为筛选提高母猪繁殖性能的miRNA提供理论依据。

关键词: miR-24-3p, 猪, 颗粒细胞, 雌二醇

Abstract: This study was conducted to investigate the effects of miR-24-3p on estradiol synthesis in porcine granulosa cells. In this experiment, ovarian tissue of 180-day old healthy sows (n=20) was collected, and 20 pairs of ovaries were collected for isolation and culture of granulosa cells in each experiment, and mimics and inhibitor of miR-24-3p were transfected into granulosa cells. The ELISA, RT-qPCR, Western blot, and double luciferase reporting tests were used to explore the effect of miR-24-3p on estradiol synthesis in porcine granulosa cells. The results showed that overexpression of miR-24-3p could significantly promote estradiol synthesis (P<0.01), and up-regulate the transcription and translation of StAR, CYP19A1 and CYP11A1 (P<0.05). Interfering with miR-24-3p significantly inhibited the synthesis of estradiol (P<0.05) and significantly down-regulated the mRNA and protein levels of CYP11A1 and CYP19A1 (P<0.05). Further studies showed that TOP1 was the direct target gene of miR-24-3p, overexpression of miR-24-3p could significantly inhibit the expression of TOP1 (P<0.05), and interference with miR-24-3p could significantly up-regulate the expression of TOP1 (P<0.05). What’s more, overexpression of TOP1 could weaken the promoting effect of miR-24-3p on estradiol synthesis in granulosa cells (P<0.05). In summary, miR-24-3p inhibits mRNA and protein levels of TOP1, and promotes the expression level of genes related to estradiol synthesis, thus promoting estradiol synthesis in granulosus cells by targeting TOP1. Since the estradiol synthesis capacity of granulosa cells directly affects the developmental state of ovarian follicles, this study provides a theoretical basis for screening miRNA that can improve reproductive performance of sows.

Key words: miR-24-3p, pigs, granulosa cells, estradiol

中图分类号:

S828.3

时胜洁, 王立光, 高磊, 蔡传江, 何伟先, 褚瑰燕. miR-24-3p对猪颗粒细胞雌二醇合成的作用[J]. 畜牧兽医学报, 2024, 55(1): 169-178.

SHI Shengjie, WANG Liguang, GAO Lei, CAI Chuanjiang, HE Weixian, CHU Guiyan. Effect of miR-24-3p on Estradiol Synthesis in Porcine Granulosa Cells[J]. Acta Veterinaria et Zootechnica Sinica, 2024, 55(1): 169-178.

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