Objective: To study the effects of knockdown of miR-449a on the proliferation and migration of human breast cancer cell line Michigan Cancer Foundation-7 (MCF-7).
Methods: Using miRNA chip screening of the differential expressions of miRNA in human breast cancer cell MCF-7 and normal breast cells MCF-10A. The inhibitor of miR-449a was synthesized by chemical and detected by real-time PCR after transfection aimed to verify the expression. Cell Counting Kit-8 (CCK-8) assay was used to detect the ability of cell proliferation after transfection with miR-449a inhibitor. Scratch assay was used to detect cell migration of MCF-7, and cell invasion ability was showed by transwell assay; The MCF-7 cell proliferation and migration related proteins, β-catenin and E-cadherin, were detected by Western blot. The potential target gene of miR-449a was predicted by bioinformatics software, and Notch homolog 1 (Notch 1) was proved to be the target gene of miR-449a by luciferase assay.
Results: MCF-7 and MCF-10a cells were collected separately, and miRNA chip results showed that the level of miR-449a in MCF-7 cells was significantly higher than that of MCF-10A. In this study, the cells were divided into mock group, negative control group (NC group) and treatment group, the MCF-7 cells were collected before and after treatment and CCK-8 results showed that knockdown of miR-449a decreased MCF-7 cell proliferation ability significantly. Scratch assay results showed that downregulated miR-449a was related to the decreased metastasis of MCF-7 cells. Transwell results showed that knockdown of miR-449a inhibited the invasion of MCF-7 cells. Western blot showed the expression of β-catenin was decreased and the expression of E-cadherin was increased after knockdown of miR-449a. Luciferase assay showed that miR-449a could significantly decrease the luciferase activity of Notch homolog 1-untranslated region (Notch 1-3'-UTR) plasmid (P<0.01).
Conclusions: Inhibition of miR-449a in breast cancer cell line MCF-7 can significantly inhibit the proliferation and migration of cancer cells, which may be achieved by decreasing the expression of Notch 1 protein.
Keywords: human breast cancer cell line MCF-7; miR-449a; migration; proliferation.