Abstract
Mounting evidence has revealed that impaired spiral artery remodeling, placental dysfunction, and inadequate trophoblast invasion are closely correlated with the etiology and pathogenesis of pre-eclampsia (PE). Moreover, defective trophoblast invasion may trigger poor maternal–fetal circulation and placental hypoxia, leading to PE. However, the detailed molecular pathology of PE remains unclear. Although circRNAs, as a new type of stable and abundant endogenous noncoding RNA, have been proven to be essential to the pathogenesis of various diseases, their role in PE requires further verification. In this context, it is necessary to unveil the roles of circRNAs in regulating the migration and invasion of extravillous trophoblasts. In this study, using quantitative real-time PCR, we confirmed that hsa_circ_0111277 was upregulated in PE placentas relative to the level in normal pregnancy placentas. In addition, positive correlations between hsa_circ_0111277 expression and PE-related factors (proteinuria level at 24 h and placental weight) were identified by Pearson’s analysis based on the clinical data of 25 PE patients. Moreover, fluorescence in situ hybridization analysis illustrated that circ_0111277 was preferentially localized within the cytoplasm. Mechanistically, circ_0111277 sponged hsa-miR-494-3p in trophoblast cells to attenuate the latter’s repression by regulating HTRA1/Notch-1 expression. In conclusion, trophoblast cell migration and invasion were shown to be promoted and modulated by the hsa_circ_0111277/miR-494-3p/HTRA1/Notch-1 axis, which provides useful insight for exploring a new therapeutic approach for PE.
Highlights
Pre-eclampsia (PE) is a hypertensive disorder of human pregnancy, which manifests as new-onset hypertension and proteinuria after 20 weeks of gestation[1]
The results showed that placental growth factor (PLGF) concentrations were drastically enhanced in both HTR-8/SVneo cells cotransfected with either miR-494 mimics or miR-494 mimics and pLV3-Ctrl compared with the level in the control group, while there was no visible alteration in HTR-8/SVneo cells administered miR-494 mimics and pLV3-High-temperature requirement-A serine peptidase 1 (HTRA1) (Fig. 4f)
Research has confirmed that competing endogenous RNAs play pivotal roles in the molecular pathology of PE15,16
Summary
Hsa_circ_0111277 expression was upregulated in PE placentas To obtain insights into the role of hsa_circ_0111277 in the etiology and pathology of PE, hsa_circ_0111277 was generated by circularizing products in the range of exons 13–17 of the mRNA NM_020318.3 (Fig. 1a). The results of Transwell assays showed that the downregulation of circ_0111277 markedly promoted the invasion and migration of HTR-8/SVneo and JEG-3 cells when compared with those transfected with empty vectors (Fig. 2a, b). To identify whether hsa_circ_0111277 modulates trophoblast cell migration and invasion through the miR-494/HTRA1 pathway, qPCR analysis was performed to examine the alterations of HTRA1 mRNA levels in HTR8/SVneo and JEG-3 cells after co-transfection with miR-494 mimics. Higher Notch-1 protein levels in HTR8/SVneo and JEG-3 cells were elicited by sicirc_0111277-modulated silencing of hsa_circ_0111277, which was in accordance with the above results (Fig. 5b). To directly identify whether HTAR1 overexpression can abolish the increased Notch-1 expression triggered by the silencing of hsa_circ_0111277, western blot assay was performed to detect Notch-1 protein in HTR8/SVneo and JEG-3 cells cotransfected with si-circ_0111277 or si-circ_0111277+ pLV3-HTRA1. The elevated Notch-1 protein level as mentioned above was conspicuously attenuated by HTAR1 overexpression, which was in line with the aforementioned results
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