Downregulation of miRNA⁃449b is an important factor to promote proliferation and invasion of glioma cells

Abstract

Objective To investigate the expression of miRNA⁃449b in gliomas and its effect on cell proliferation and invasion. Methods The expression level of miRNA⁃449b was detected by locked⁃oligonucleotide⁃probe in situ hybridization (ISH) in 60 human glioma specimens of different WHO grades and 10 non⁃tumoral control brain tissues. miRNA⁃449b was quantified by Stem⁃loop quantitation real⁃time polymerase chain reaction (qRT⁃PCR) with U6 as the internal control in U251, LN229 and UC2 cells. EdU staining, MTS method, flow cytometry assay (FCM), Phalloidin staining and Transwell assays were used to evaluate the effect of miRNA⁃449b on the proliferation and invasion of U251 and LN229 cells. Results The expression of miRNA⁃449b was decreased in gliomas compared with that in the non⁃tumoral control brain tissues (P < 0.001, for all), and was significantly declined with the elevation of glioma grades (P < 0.001, for all). miRNA⁃449b level was significantly lower in U251 and LN229 cells than that in UC2 cells (P = 0.001, 0.002). The relative expression of miRNA⁃449b in U251⁃miRNA⁃449b group was higher (P < 0.001), while EdU⁃positive rates (P = 0.029) and cell proliferation (P = 0.004, 0.001, 0.002) was lower than U251⁃Scr group; the relative expression of miRNA⁃449b in LN229⁃miRNA⁃449b group was higher (P < 0.001), while EdU⁃positive rates (P = 0.032) and cell proliferation (P = 0.003, 0.001, 0.004) was lower than LN229⁃Scr group. The percentages of each phase G 0 /G 1 in U251⁃miRNA⁃449b group and LN229⁃miRNA⁃449b group were higher than those in U251 ⁃ Scr group (P < 0.001) and LN229 ⁃ Scr group (P = 0.018) respectively, cell proliferation were lower than U251⁃Scr group (P = 0.002) and LN229⁃Scr group (P = 0.005). F⁃actin was centralized mostly on cell cortex or lamellae in the groups of U251⁃Scr and LN229⁃Scr, whereas diffusely distributed in the groups of U251⁃miRNA⁃449b and LN229⁃miRNA⁃449b. Conclusions miRNA⁃449b is an important tumor⁃suppressive miRNA and its downregulation may be an important cause leading to tumorigenesis of glioblastoma, which could be used as a valuable biomarker to assess the malignant degree of gliomas. Furthermore, the exogenous miRNA⁃449b could effectively inhibit the proliferation and invasion abilities of glioma cells, confirming it has potential value in malignant gliomas therapy.

DOI：10.3969/j.issn.1672⁃6731.2019.11.007