miR-96 plays an oncogenic role by targeting FOXO1 in osteosarcoma

Objective: To explore the difference in miR-96 expression in human osteosarcoma tissue and cells, as well as its molecular mechanisms in regulating the malignant phenotype of Osteosarcoma. Methods: MiR-96 expression in human osteosarcoma tissue and cells was detected using quantitative reverse transcription polymerase chain reaction (qRT-PCR), and the correlation of miR-96 expression with the clinicopathological features of patients was analyzed. MiR-96 inhibitor was transfected in vitro into human osteosarcoma cells MG-63, and changes in miR-96 expression in the cells after transfection were detected using qRT-PCR. Meanwhile, changes in cell proliferation and apoptosis after transfection were determined using CCK-8 assay and flow cytometry. Changes in PCDA and caspase-3 protein expression after transfection were detected using Western blotting. Bioinformatics was used to predict the targeting genes of miR-96. Double luciferase reporter gene assay combined with qRT-PCR and Western blotting were used to validate the targeting regulation of miR-96 on FOXO1. In addition, the effect of miR-96 on osteosarcoma xenograft in the nude mouse was detected through the nude mouse subcutaneous tumorigenicity assay. Results: MiR-96 showed aberrantly high expression in human osteosarcoma cells MG-63, Saos-2 and U2OS, as well as osteosarcoma tissue. Further statistical analysis suggested that miR-96 expression in osteosarcoma tissue was closely related to the Enneking stage and distant metastasis. FOXO1 is a direct target gene of miR-96. Moreover, miR-96 expression level was remarkably reduced in MG-63 cells after transfection with miR-96 inhibitor, while both FOXO1 mRNA and protein expression levels were notably increased, cell proliferation capacity and PCDA protein expression level were markedly lowered, and cell apoptosis rate and caspase-3 protein expression level were evidently enhanced. Furthermore, nude mouse osteosarcoma xenograft growth results revealed that xenograft in MG-63 cells transfected with miR-96 inhibitor was outstandingly reduced in terms of volume and weight. Conclusion: MiR-96 shows aberrantly high expression in osteosarcoma tissue and cells. Down-regulating miR-96 expression can apparently suppress osteosarcoma cell proliferation and tumorigenic ability, and enhance its apoptosis, the mechanism of which may be related to the target regulation of FOXO1.