Regulation of ACVR1 and ID2 by cell-secreted exosomes during follicle maturation in the mare.
Regulation of ACVR1 and ID2 by cell-secreted exosomes during follicle maturation in the mare.
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马卵泡成熟过程中细胞分泌的外泌体对 ACVR1 和 ID2 的调节。
DOI：
10.1186/1477-7827-12-44
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发表时间：
2014-05-26
期刊：
影响因子：
--
通讯作者：
Bouma GJ
中科院分区：
文献类型：
--
作者：
da Silveira JC;Carnevale EM;Winger QA;Bouma GJ
卵巢卵泡的生长和成熟需要卵泡体细胞和卵母细胞之间进行广泛的交流。近期，一种涉及细胞分泌的被称为外泌体（50 - 150纳米）的囊泡的细胞间通讯被描述，外泌体包含微小RNA（miRNA）和蛋白质，并且已在卵巢卵泡液中被发现。本研究的目的是确定外泌体在卵泡成熟过程中可能发挥的作用。
在发情中期（约35毫米，卵泡成熟诱导前）和排卵前卵泡（卵泡成熟诱导后30 - 34小时）从母马收集卵泡内容物。进行实时聚合酶链反应（PCR）筛选以揭示从发情中期和排卵前卵泡分离出的外泌体微小RNA存在的显著差异，并且根据生物信息学分析，这些外泌体微小RNA预计会靶向转化生长因子β（TGFB）超家族的成员，包括激活素受体1型（ACVR1）和DNA结合抑制因子2（ID2）。培养排卵前卵泡的颗粒细胞，并用从卵泡液中分离出的外泌体进行处理。通过实时PCR和蛋白质印迹法（Western blot）测量信使核糖核酸（mRNA）和蛋白质的变化。
在发情中期和排卵前阶段的颗粒细胞中检测到ACVR1的mRNA和蛋白质，实时PCR分析显示排卵前卵泡的颗粒细胞中ID2（一种ACVR1的靶基因）水平显著降低。暴露于发情中期卵泡卵泡液的外泌体使颗粒细胞中ID2水平降低。此外，从发情中期和排卵前卵泡分离出的外泌体包含ACVR1，并且微小RNA - 27b、微小RNA - 372和微小RNA - 382（ACVR1和ID2的预测调节因子）能够改变排卵前颗粒细胞中ID2的水平。
这些数据表明，从卵泡液中分离出的外泌体能够调节颗粒细胞中TGFB / 骨形态发生蛋白（BMP）信号通路的成员，并可能在调节卵泡成熟过程中发挥作用。
Ovarian follicle growth and maturation requires extensive communication between follicular somatic cells and oocytes. Recently, intercellular cell communication was described involving cell-secreted vesicles called exosomes (50–150 nm), which contain miRNAs and protein, and have been identified in ovarian follicular fluid. The goal of this study was to identify a possible role of exosomes in follicle maturation. Follicle contents were collected from mares at mid-estrous (~35 mm, before induction of follicular maturation) and pre-ovulatory follicles (30–34 h after induction of follicular maturation). A real time PCR screen was conducted to reveal significant differences in the presence of exosomal miRNAs isolated from mid-estrous and pre-ovulatory follicles, and according to bioinformatics analysis these exosomal miRNAs are predicted to target members belonging to the TGFB superfamily, including ACVR1 and ID2. Granulosa cells from pre-ovulatory follicles were cultured and treated with exosomes isolated from follicular fluid. Changes in mRNA and protein were measured by real time PCR and Western blot. ACVR1 mRNA and protein was detected in granulosa cells at mid-estrous and pre-ovulatory stages, and real time PCR analysis revealed significantly lower levels of ID2 (an ACVR1 target gene) in granulosa cells from pre-ovulatory follicles. Exposure to exosomes from follicular fluid of mid-estrous follicles decreased ID2 levels in granulosa cells. Moreover, exosomes isolated from mid-estrous and pre-ovulatory follicles contain ACVR1 and miR-27b, miR-372, and miR-382 (predicted regulators of ACVR1 and ID2) were capable of altering ID2 levels in pre-ovulatory granulosa cells. These data indicate that exosomes isolated from follicular fluid can regulate members of the TGFB/BMP signaling pathway in granulosa cells, and possibly play a role in regulating follicle maturation.
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