Abstract
To investigate the role of SIRT1/autophagy pathway in mediating the regulatory effect of lncRNA SOX2OT on 5-fluorouracil (5-FU) resistance in cholangiocarcinoma cells. HCCC-9810 cells were used to construct a 5-FU-resistant cell model (HCCC-9810/5-FU cells), and the expression levels of lncRNA SOX2OT and SIRT1 mRNA and the protein expressions of SIRT1, Beclin1, LC3 and P62 were detected with qRT-PCR and Western blotting. The effects of transfection with a SOX2OT mimic on drug resistance and cell migration of HCCC-9810/5-FU cells were detected using CCK-8 assay and wound healing assay, and the changes in expressions of SOX2OT, SIRT1, Beclin1, LC3 and P62 were detected. Rescue experiment was performed by co-transfection of HCCC-9810/5-FU cells with both a SOX2OT-overexpressing plasmid and si-SIRT1 to confirm the role of SIRT1 in SOX2OT-mediated regulation of 5-FU resistance. A RNA pulldown assay was used to verify the targeted binding between SOX2OT and SIRT1. The proliferation of HCCC-9810 cells was significantly inhibited after treatment with different concentrations of 5-FU (P < 0.05). The 5-FU-resistant cells showed significantly increased protein expressions of SIRT1, Beclin1 and p62, an increased LC3 Ⅱ/LC3 Ⅰ ratio, and enhanced expressions of SIRT1 mRNA and SOX2OT (P < 0.05). Transfection of the resistant cells with SOX2OT mimic significantly enhanced cell migration and increased the protein expressions of SIRT1, Beclin1 and p62, the LC3Ⅱ/LC3Ⅰ ratio, and expression levels of SIRT1 mRNA and SOX2OT (P < 0.05), and these changes were obviously attenuated by SIRT1 knockdown, which also resulted in lowered 5-FU resistance of the cells without significantly affecting the expression level of SOX2OT (P > 0.05). RNA pulldown assay suggested that SOX2OT could directly bind to SIRT1. LncRNA SOX2OT enhances 5-FU resistance in HCCC-9810 cells by promoting autophagy through up-regulating SIRT1 expression.
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Schedule a callMore From: Nan fang yi ke da xue xue bao = Journal of Southern Medical University
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