MiR-222-3p inhibits formation of medulloblastoma stem-like cells by targeting Notch2/c-myc signaling pathway

Background: Medulloblastoma (MB) is an embryonal tumor of the cerebellum, which commonly occurs in childhood. Herein, we investigated the effects of miR-222-3p on the formation of MB stem-like cells via the Notch2/c-Myc pathway.

Methods: Quantitative Real-Time PCR (qRT-PCR) or western blotting was performed to determine the expression of miR-222-3p and Notch2, c-Myc, proliferating cell nuclear antigen (PCNA), and Caspase-3. Luciferase reporter gene, RNA immunoprecipitation (RIP), and RNA pull-down assay were applied to confirm the interaction between miR-222-3p and Notch2. Cell growth was examined by Cell Counting Kit-8. Cell cycle distribution and the number of stem cell marker CD133⁺ cells were examined using flow cytometry. The sphere formation assay was performed.

Results: miR-222-3p expression was decreased and Notch2 expression was increased in human medulloblastoma cells. miR-222-3p overexpression inhibited cell viability and the sphere formation, induced cell cycle arrest, decreased the number of CD133⁺ cells, and up-regulated Caspase-3 expression and down-regulated PCNA, Notch2, and c-Myc expression. However, Notch2 overexpression counteracted these effects of miR-222-3p overexpression. Simultaneous overexpression of Notch2 and miR-222-3p increased the c-Myc promoter-luciferase activity which was decreased by miR-222-3p overexpression. Luciferase reporter gene, RIP, and RNA pull-down assay revealed that miR-222-3p targeted Notch2.

Conclusion: MiR-222-3p suppressed cell viability, altered cell cycle distribution, and inhibited the formation of MB stem-like cells via the Notch2/c-Myc pathway.