circRNA3669通过miR-26a和RCN2激活PI3K/AKT-mTOR与MAPK通路促进山羊子宫内膜上皮细胞增殖

doi:10.1016/j.jia.2023.05.029

Journal of Integrative Agriculture ›› 2024, Vol. 23 ›› Issue (3): 960-974.DOI: 10.1016/j.jia.2023.05.029

circRNA3669通过miR-26a和RCN2激活PI3K/AKT-mTOR与MAPK通路促进山羊子宫内膜上皮细胞增殖

收稿日期:2022-12-05 接受日期:2023-04-13 出版日期:2024-03-20 发布日期:2024-03-03

circRNA3669 promotes goat endometrial epithelial cells proliferation via miR-26a/RCN2 to activate PI3K/AKT-mTOR and MAPK pathways

Xiaorui Liu^1*, Jiuzeng Cui^2*, Mengyao Wei^2*, Xiaofei Wang², Yuexia Liu², Zhongshi Zhu², Min Zhou¹, Gui Ba³, Langda Suo³, Yuxuan Song^2#, Lei Zhang^2#

¹Division of Laboratory Safety and Services, Northwest A&F University, Yangling 712100, China
²College of Animal Science and Technology, Northwest A&F University, Yangling 712100, China
³Institute of Animal Sciences, Tibet Academy of Agricultural and Animal Husbandry Sciences, Lhasa 850009, China

Received:2022-12-05 Accepted:2023-04-13 Online:2024-03-20 Published:2024-03-03
About author:Xiaorui Liu, E-mail: xiaorui_liu925@163.com; #Correspondence Yuxuan Song, E-mail: yuxuan_song2016@163.com; Lei Zhang, E-mail: zhanglei07dongke@163.com *These authors contributed equally to this study.
Supported by:
This study was supported by the China Postdoctoral Science Foundation (2019M653776 and 2020M673516), the Natural Science Basis Research Plan in Shaanxi Province of China (2023-JC-QN-0181), the Shaanxi Livestock and Poultry Breeding Double-chain Fusion Key Proiect, China (2022GD-TSLD-46-0202), and the Natural Science Fundation of Tibet Autonomous Region, China (XZ202101ZR0063G).

摘要/Abstract

摘要：

容受性子宫内膜是胚胎成功植入的必要条件之一，它的形成是一个复杂的动态过程，在这个过程中子宫内膜的形态与功能发生显著改变，其中包括子宫内膜细胞增殖与凋亡。然而，这种分子机制有待进一步研究。目的：本研究旨在探究circRNA3669对山羊子宫内膜上皮细胞的调控作用。方法：利用RT-qPCR、Western blot、双荧光素酶活性检测系统、免疫组化、CCK-8、EdU、细胞凋亡等实验技术，检测circRNA3669在山羊子宫内膜上皮细胞中对miR-26a和网钙蛋白2（RCN2）的调控作用，以及对山羊子宫内膜上皮细胞增殖的影响。结果：在本研究中，circRNA3669在山羊容受期子宫内膜中的表达量显著高于容受前期。功能分析显示， circRNA3669过表达后通过激活PI3K/AKT-mTOR和MAPK信号通路，促进山羊子宫内膜上皮细胞的增殖，从而抑制山羊子宫内膜上皮细胞凋亡。此外，circRNA3669可作为一种竞争性内源性RNA（ceRNA）通过吸附miR-26a上调RCN2在山羊子宫内膜上皮细胞中的表达量。结果显示，RCN2在山羊容受前期子宫内膜中高表达，且β-雌二醇（E2）和孕酮（P4）调控其在山羊子宫内膜上皮细胞中的表达量。RCN2可通过激活PI3K/AKT-mTOR和MAPK信号通路上关键蛋白PI3K、AKT、mTOR、JNK和P38的磷酸化水平，进而促进山羊子宫内膜上皮细胞增殖。结论：circRNA3669作为ceRNA吸附miR-26a，上调RCN2 在山羊子宫内膜上皮细胞中的表达量，并通过激活PI3K/AKT-mTOR和MAPK通路抑制山羊子宫内膜上皮细胞凋亡。本研究在山羊子宫内膜上皮细胞中构建了circRNA3669-miR-26a-RCN2调控网络，为进一步探究circRNA在山羊容受性子宫内膜建立过程中的分子调控机制提供试验依据。

Abstract: The development of receptive endometrium (RE) from pre-receptive endometrium (PE) for successful embryo implantation is a complex dynamic process in which the morphology and physiological states of the endometrial epithelium undergo a series of significant changes, including cell proliferation and apoptosis. However, the molecular mechanisms are not yet fully understood. In this study, a higher circRNA3669 level was observed in PE than in RE of goats. Functional assays revealed that this overexpression promoted the proliferation of goat endometrial epithelial cells (GEECs) by activating the expression of genes related to the PI3K/AKT-mTOR and MAPK pathways, thereby inhibiting apoptosis in vitro. Furthermore, circRNA3669 functioned as a competing endogenous RNA (ceRNA) to upregulate Reticulocalbin-2 (RCN2) expression at the post-transcriptional level by interacting with and downregulating miR-26a in GEECs. In addition, RCN2, which is highly expressed in the PE of goats, was found to be regulated by β-estradiol (E2) and progesterone (P4). Our results demonstrated that RCN2 also affected the key proteins PI3K, AKT, mTOR, JNK, and P38 in the PI3K/AKT-mTOR and MAPK pathways, thereby facilitating GEECs proliferation and suppressing their apoptosis in vitro. Collectively, we constructed a new circRNA3669-miR-26a-RCN2 regulatory network in GEECs, which further provides strong evidence that circRNA could potentially play a crucial regulatory role in the development of RE in goats.

Key words: circRNA3669, RCN2 , miR-26a , goat endometrial epithelial cells (GEECs) , proliferation

. circRNA3669通过miR-26a和RCN2激活PI3K/AKT-mTOR与MAPK通路促进山羊子宫内膜上皮细胞增殖[J]. Journal of Integrative Agriculture, 2024, 23(3): 960-974.

Xiaorui Liu, Jiuzeng Cui, Mengyao Wei, Xiaofei Wang, Yuexia Liu, Zhongshi Zhu, Min Zhou, Gui Ba, Langda Suo, Yuxuan Song, Lei Zhang.

circRNA3669 promotes goat endometrial epithelial cells proliferation via miR-26a/RCN2 to activate PI3K/AKT-mTOR and MAPK pathways [J]. Journal of Integrative Agriculture, 2024, 23(3): 960-974.

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circRNA3669通过miR-26a和RCN2激活PI3K/AKT-mTOR与MAPK通路促进山羊子宫内膜上皮细胞增殖

circRNA3669 promotes goat endometrial epithelial cells proliferation via miR-26a/RCN2 to activate PI3K/AKT-mTOR and MAPK pathways

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