CircSETD3 (hsa_circ_0000567) inhibits proliferation and induces apoptosis in cholangiocarcinoma cells via downregulation of microRNA-421 expression

Abstract

ABSTRACT Cholangiocarcinoma (CCA) is a fatal tumor associated with chronic inflammation. Circular RNAs (circRNAs) have been evidenced to be involved in tumorigenesis and tumor progression. This study aimed to explore the effects and potential molecular mechanism of circSETD3 in CCA progression. Levels of CircSETD3 and microRNA (miR)-421 in CCA tissue and cell lines were measured using quantitative real-time polymerase-chain reaction (qRT-PCR). A direct target of miR-421 was predicted using TargetScan and further confirmed by a dual-luciferase reporter assay. Cell proliferation and apoptosis were measured using MTT (3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide) assay and flow cytometry, respectively. The activity of caspase-3 was also examined using caspase-3 activity detection kits. Moreover, the levels of B-cell lymphoma-2 modifying factor (BMF), B-cell lymphoma 2 (BCL2), and Bcl-2-associated X protein (BAX) in TFK1 cells were assessed using qRT-PCR and western blot analysis. We found that circSETD3 was downregulated, while miR-421 was upregulated in CCA tissues and cell lines. CircSETD3 negatively regulated miR-421 levels in TFK1 cells. Functional assays revealed that circSETD3-plasmid inhibited cell proliferation, induced apoptosis, promoted caspase-3 activity, enhanced Bax and cleaved-Caspase 3 expression, and reduced Bcl-2 levels, and these effects were reversed by miR-421 mimic. Meanwhile, similar results were observed in miR-421 inhibitor-transfected TFK1 cells, and these results were abolished by BMF-siRNA. BMF, a direct target of miR-421, was downregulated in CCA tissues and cell lines. These findings demonstrate that circSETD3 inhibits proliferation and induces apoptosis in CCA cells by regulating the miR-421/BMF axis, indicating its potential as a promising candidate for CCA therapy.