CircFBXW4 regulates human trophoblast cell proliferation and invasion via targeting miR-324-3p/TJP1 axis in recurrent spontaneous abortion

Introduction: Increasing evidence has shown that circular RNAs (circRNAs) play vital roles in embryonic devel-opment. However, the function of circRNAs in recurrent spontaneous abortion (RSA) is largely unknown. This study aimed to investigate the expression profile of human circRNAs and their functional mechanisms in regu-lating RSA. Methods: The profiles of circRNAs in placental villus tissues from women with RSA and healthy pregnancy with induced abortion were investigated using RNA-sequencing and bioinformatics. Nine circRNAs were verified in the 50 placental villus samples. RNase R digestion, actinomycin D treatment, and fluorescence in situ hybridi-zation were performed to characterize circFBXW4. Furthermore, direct binding of circFBXW4 to miR-324-3p was confirmed by dual-luciferase reporter assay. The roles of circFBXW4 were determined by loss-and gain-of-function assays including cell proliferation, invasion, and apoptosis using CCK8 kit, transwell migration assay, and TUNEL kit in vitro, respectively. Results: A total of 417 aberrantly expressed circRNAs was detected. circFBXW4, a circRNA significantly up-regulated in the RSA group, was further evaluated. circFBXW4 showed higher stability than FBXW4 mRNA and was localized in the cytoplasm and nucleus in HTR-8/SVneo cells. MiR-324-3p was lowly expressed in the RSA group, and directly regulated circFBXW4 and TJP1 expression in a targeted manner. Overexpression and knockdown of circFBXW4 and miR-324-3p mimic/inhibitor could increase or decrease HTR-8/SVneo cell pro-liferation and invasion. circFBXW4 regulated TJP1 expression, cell proliferation, and invasion by sponging miR-324-3p. Discussion: The circFBXW4/miR-324-3p/TJP1 axis is involved in the occurrence and progression of RSA and may be a promising therapeutic target in RSA.

Automated summary

This study identified the critical role of circFBXW4/miR-324-3p/TJP1 axis in the occurrence and progression of RSA, suggesting a promising therapeutic target for RSA treatment.