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Archive > Volume 27 Issue 6 > 2020,27(6):622-628. DOI:10.3872/j.issn.1007-385X.2020.06.005 Prev Next
Basic Research
miR-125a-5p promotes the resistance to gefitinib in non-small cell lung carcinoma cells by targeting APAF1
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Abstract:
Objective: To investigate the role of miR-125a-5p in inducing the gefitinib (Gef)-resistance of non-small cell lung carcinoma (NSCLC) cells and its possible mechanism. Methods: Human NSCLC drug-resistant cell line A549/GR and NSCLC cell line A549 were chosen for this study. miR-125a-5p mimic, miR-125a-5p inhibitor, pcDNA3.1-APAF1 and empty vector pcDNA3.1 were transfected into A549/GR cells. The expression level of miR-125a-5p in cell lines was detected by qPCR. MTT, Transwell and Flow cytometry were used to detect the effects of Gef on proliferation, migration and apoptosis of cell lines, respectively. The targeting relationship between miR-125a-5p and APAF1 (apoptotic peptidase activating factor 1) was verified by Dual-luciferase reporter gene system. In addition, the expression of APAF1 protein in A549/GR cells was detected by Western blotting. The expression levels of caspase-3 and caspase-9 were assessed by colorimetry. Results: Expression level of miR-125a-5p was upregulated significantly in Gefresistant A549/GR cells (P<0.01). And the influences of Gef onA549/GR cells were enhanced by knockdown of miR-125a-5p, including inhibiting cell proliferation and migration (all P<0.05) and inducing apoptosis (P<0.01). Dual luciferase reporter gene assay confirmed that miR-125a-5p targeted APAF1 and negatively regulated its expression. Furthermore, by targetedly downregulating APAF1,miR-125a-5p alleviated the inhibition of proliferation and migration (all P<0.05) and promotion of apoptosis (P<0.05) of A549/GR cells caused by Gef, and attenuated Gef-induced upregulation of apoptosis-related proteins caspase-3 and caspase-9 (all P<0.05).Conclusion: miR-125a-5p promotes Gef-resistance of A549/GR cells, and the underlying mechanisms are promotion of proliferation,migration and inhibition of apoptosis of non-small cell lung cancer cells by targeting APAF1.
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