MiR-181a-5p facilitates proliferation, invasion, and glycolysis of breast cancer through NDRG2-mediated activation of PTEN/AKT pathway

Abstract

Dysregulation of microRNAs (miRNAs) is associated with the occurrence and development of breast cancer. In this research, we explored the involvement of miR-181a-5p in the progression of breast cancer and investigated potential molecular mechanisms. Firstly, the miR-181a-5p and N-myc downstream-regulated gene (NDRG) 2 expression was detected by real-time quantitative polymerase chain reaction. Cellular processes were assessed using Cell Counting Kit 8, Bromodeoxyuridine, colony formation and transwell assays. HK2, PKM2 and LDHA activities were assessed by ELISA. The combination between miR-181a-5p was assessed by dual-luciferase reporter assay and RNA pull-down assay. The results indicated that miR-181a-5p levels were upregulated and NDRG2 levels were downregulated in breast cancer, leading to poor prognosis. Silencing of miR-181a-5p inhibited cell proliferation, invasion, glycolysis, and xenograft tumor growth, while enhanced miR-181a-5p got the opposite results. Furthermore, NDRG2 acts as a target of miR-181a-5p. Knockout of NDRG2 facilitated biological behaviors and meanwhile enhanced phosphorylation (p)-PTEN and p-AKT levels. Rescue experiments showed that restoring NDRG2 abolished the effects caused by miR-181a-5p in breast cancer cells. In conclusion, miR-181a-5p facilitated tumor progression through NDRG2-induced activation of PTEN/AKT signaling pathway of breast cancer, suggesting that focusing on miR-181a-5p may provide new insight for breast cancer therapy. Abbreviations Brdu: Bromodeoxyuridine; CCK-8: Cell Counting Kit-8; miRNA: microRNAs; mut: mutant; RT-qPCR: real-time quantitative polymerase chain reaction; UTR: untranslated region; WT: wild-type

lncRNA LINC01315 promotes malignancy of triple-negative breast cancer and predicts poor outcomes by modulating microRNA-876-5p/GRK5

“…Cell cultures were seeded at 5 × 10 4 cells/well into the 96-well plates and starved for 12 h. Cells were supplied with a completed growth medium and incubated for 0, 24, 48, and 72 h. Then, 20 μL MTT solution (Sigma) was mixed with the cultured cells and incubated for another 4 h. The crystal was dissolved by 150 μL DMSO (Sigma) and the absorbance at 490 nm of each well was measured with a Microplate Reader [ 20 , 21 ].…”

“…Cell cultures were placed in the upper chamber (coated with Matrigel in invasion assay) of the 24-well transwell plates (Corning) and supplied with an FBS-free culture medium. The completed medium was placed in the bottom chamber and incubated at 37°C for 48 h. Then, cells on the upper surface were removed, and the migrated and invaded cells were counted with a microscope (magnification ×200) [ 20 , 21 ].…”

miR-125a attenuates the malignant biological behaviors of cervical squamous cell carcinoma cells through Rad51

“…SiHa and HCC-0214 cells were seeded in 96-well plates and transfected with oligonucleotides in accordance with the experimental aims. The cells in each group were continuously cultured at 37°C, and 10 μL of CCK-8 reagent (Dojindo, Japan) was added at 0, 24, and 48 h. After incubation for another 2 h at 37°C, the absorbance of each cell group was measured with a microplate reader [ 24 ].…”

“…After washing with PBS, the CSCC cells were fixed with 4% paraformaldehyde for 20 min and stained with 0.2% crystal violet for 5 min. After washing with water, the 6-well plate was dried, scanned and photographed under a microscope [ 24 ].…”

“…The invaded cells were fixed with 4% formaldehyde and stained with 0.2% crystal violet. The invaded cells were observed and counted under a microscope [ 24 ].…”