The Possible Involvement of miR-371a-5p Regulating XIAP in the Pathogenesis of Recurrent Pregnancy Loss

Abstract

Apoptosis is an interactive and dynamic biological process involved in all phases of embryogenesis. If apoptosis dominates the trophoblast cell growth process, it will result in adverse pregnancy outcomes. X-linked inhibitor of apoptosis protein (XIAP) is a potent caspase inhibitor and an important barrier to apoptotic cell death. MicroRNAs involve in posttranscriptional gene expression regulation and apoptosis. Online sequence alignment analysis showed that there was a putative binding site of miR-371a-5p on the 3′-untranslated region (UTR) of XIAP. Thirty chorionic villi samples were collected to examine the expression of miR-371a-5p and XIAP. The dual-luciferase reporter assay was applied to determine the relationship between miR-371a-5p and XIAP by human placental choriocarcinoma cells (JEG-3) cells in vitro. After 48-hour transfection of mimics and inhibitor by JEG-3 cells in vitro, Western blotting was used to, respectively, detect the protein expression levels of XIAP and caspase-3. Flow cytometry was used to validate the apoptosis ratio of transfection. The expression of miR-371a-5p and XIAP in recurrent pregnancy loss was greatly decreased. The results from the luciferase reporter assay strongly suggested binding of the XIAP 3′-UTR by miR-371a-5p. Apoptosis percentage of miR-371a-5p mimic was significantly greater than that of normal control. However, apoptosis percentage of miR-371a-5p inhibitor was significantly lower than that of normal control. A significant decrease in luciferase activity was observed in miR-371a-5p mimics-transfected JEG-3 cells compared with controls. These findings provide the evidence that miR-371a-5p is one of the regulating factors according to apoptosis pathway of XIAP-caspase-3 and may be involved in the pathogenesis of recurrent pregnancy loss.