Su Yan, Xu Jiani, Gao Rufei, Liu Xiaoli, Liu Taihang, Li Cong, Ding Yubin, Chen Xuemei, He Junlin, Liu Xueqing, Li Chunli, Qi Hongbo, Wang Yingxiong
Laboratory of Reproductive Biology, School of Public Health and Management, Chongqing Medical University, Chongqing, China.
Joint International Research Laboratory of Reproduction & Development, Chongqing Medical University, Chongqing, China.
Front Physiol. 2022 Mar 3;13:778116. doi: 10.3389/fphys.2022.778116. eCollection 2022.
Recurrent miscarriage (RM) is associated with numerous clinical factors. However, some RM occurred without specific factors. It has been revealed that some molecules such as hormones, miRNAs, and transcription factors are involved in RM by regulating proliferation, apoptosis, etc. However, the mechanism of RM has yet to be identified clearly. Circular RNAs (circRNAs) are a class of endogenous non-coding RNAs that often act as sponges for miRNAs or binds to proteins involved in biological processes. However, the functional role of circRNAs in the uterine decidua of patients with early RM is still unclear. In this study, we aimed to investigate the mechanisms of circ-CYP24A1 in RM.
The Dual-Luciferase Activity Assay was designed to analyze the bonding between circ-CYP24A1 and miR-224, and miR-224 and prolactin receptor (PRLR) mRNA 3'UTR. hybridization (ISH) and immunohistochemistry (IHC) were used to observe the expression of circ-CYP24A1 and PRLR in the decidua. Rescue experiments were performed to investigate the regulating effects of circ-CYP24A1, miR-224, and PRLR. Western blotting was conducted to test the expression level of PRLR. The proliferation and apoptosis-related markers in Ishikawa cells were analyzed using CCK8, immunofluorescence staining, and the terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL) assay.
In this study, based on the microarray analysis data, we identified a high level of circ-CYP24A1 and PRLR in the decidua of patients with early RM. Based on the bioinformatics prediction, the binding relationship between circ-CYP24A1 and miR-224, as well as miR-224 and PRLR, were verified. Functional experiments demonstrated that circ-CYP24A1 regulated proliferation and apoptosis by binding to and inhibiting miR-224, resulting in increased PRLR expression. Taken together, this study provides new insights into the mechanism of RM.
In this study, we found that circ-CYP24A1 plays a role in RM by impairing the balance of cell proliferation and apoptosis by sponging miR-224, thereby regulating PRLR.
复发性流产（RM）与众多临床因素相关。然而，部分RM发生并无特定因素。已揭示一些分子如激素、微小RNA（miRNA）和转录因子通过调节增殖、凋亡等参与RM过程。然而，RM的机制尚未完全明确。环状RNA（circRNA）是一类内源性非编码RNA，常作为miRNA的海绵或与参与生物过程的蛋白质结合。然而，circRNA在早期RM患者子宫蜕膜中的功能作用仍不清楚。在本研究中，我们旨在探究circ-CYP24A1在RM中的机制。
设计双荧光素酶活性测定法分析circ-CYP24A1与miR-224以及miR-224与催乳素受体（PRLR）mRNA 3'非翻译区（UTR）之间的结合情况。采用原位杂交（ISH）和免疫组织化学（IHC）观察circ-CYP24A1和PRLR在蜕膜中的表达。进行挽救实验以研究circ-CYP24A1、miR-224和PRLR的调节作用。采用蛋白质免疫印迹法检测PRLR的表达水平。使用CCK8、免疫荧光染色和末端脱氧核苷酸转移酶介导的dUTP生物素缺口末端标记（TUNEL）法分析 Ishikawa细胞中增殖和凋亡相关标志物。
在本研究中，基于微阵列分析数据，我们发现早期RM患者蜕膜中circ-CYP24A1和PRLR水平较高。基于生物信息学预测，验证了circ-CYP24A1与miR-224以及miR-224与PRLR之间的结合关系。功能实验表明，circ-CYP24A1通过结合并抑制miR-224来调节增殖和凋亡，从而导致PRLR表达增加。综上所述，本研究为RM的机制提供了新的见解。
在本研究中，我们发现circ-CYP24A1通过充当miR-224的海绵破坏细胞增殖和凋亡的平衡，从而调节PRLR，在RM中发挥作用。
