关键词: miR-148a, 乳腺癌, 糖酵解, 己糖激酶2

Abstract: In order to investigate the effects of miR-148a and hexokinase 2 (HK2) gene on glycolytic metabolic pathway of human breast cancer cells and their possible mechanisms, real-time fluorescence quantitative PCR (qRT-PCR) was used to detect the expression of miR-148a in various breast cancer cell lines, and then breast cancer cell lines with relatively low expression of miR-148a were selected as research objects. Furthermore, the effect of miR-148a expression on glucose uptake, lactic acid production and cell proliferation index of breast cancer cells was observed to explore the effect of miR-148a on glucose metabolism ability of breast cancer cells. Subsequently, the targeted relationship between miR-148a and HK2 genes was predicted through the online database of TargetScan, and then verified through the double luciferase reporting experiment, Western blot and gene recovery experiment, so as to further clarify the mechanism of miR-148a and HK2 in the glycolytic metabolic pathway of breast cancer cells. Through qRT-PCR, the expression of miR-148a was found to be decreased in various breast cancer cell lines, especially in MDA-MB231 (P<0.000 1). Overexpression of miR-148a significantly decreased glucose uptake, lactic acid production and cell proliferation of MDA-MB231 cells (P<0.01). While inhibition of miR-148a expression significantly increased glucose uptake, lactic acid production and cell proliferation index of MDA-MB231 cells (P<0.01). Predicted by TargetScan online database, miR-148a had partial binding sites with the 3′ untranslated region (3′-UTR) of HK2 gene; and the double luciferase report experiment found that miR-148a was bound to the wild-type HK2 3′-UTR luciferase report vector and not to the mutant HK2 3'-UTR. Western blot analysis showed that overexpression of miR-148a significantly decreased the expression of HK2 protein in MDA-MB231 cells (P<0.000 1), while inhibition of miR-148a promoted the expression of HK2 protein significantly (P<0.05). Gene recovery experiment showed that overexpression of HK2 gene significantly increased glucose uptake, lactic acid production and cell proliferation index of MDA-MB231 breast cancer cells (P<0.01). The overexpressed miR-148a vector and the overexpressed HK2 vector were co-transfected into MDA-MB231 cells, and miR-148a reversed the increasing of glucose uptake and lactic acid production caused by HK2, and inhibited cell proliferation. Therefore, studies suggested that miR-148a could inhibit glycolytic metabolism and cell proliferation of breast cancer cells MDA-MB231 by targeted inhibition of HK2 gene expression.

Key words: miR-148a, breast cancer, glycolysis, hexokinase 2