Feng Linsen, Ma Jianhua, Ji Haiming, Liu Yichun, Hu Weixing
Department of Neurosurgery, First Affiliated Hospital of Nanjing Medical University, Nanjing, Jiangsu 210029, China.
Department of Neurosurgery, Taixing People's Hospital, Taixing, Jiangsu 225400, China.
Biosci Rep. 2017 Jun 21;37(3). doi: 10.1042/BSR20170019. Print 2017 Jun 30.
The present study intended to investigate the biological effects of miR-330-5p on glioblastoma (GBM) cell proliferation and invasiveness by targeting integrin α5 (ITGA5). The expressions of miR-330-5p and ITGA5 mRNA in GBM cell lines (U87, U251, and U373) and normal brain glial cell line (HEB) were detected using RT-qPCR. Protein expression of ITGA5 was examined using Western blot. The present study used MTT assay, colony formation assay, Transwell assay, wound healing assay, and flow cytometry analysis in order to determine the biological functions of GBM cells (including cell proliferation, invasion, migration, apoptosis, and cell cycle). The present study applied dual-luciferase reporter gene assay to identify the target relationship between miR-330-5p and ITGA5. miR-330-5p was low-expressed in GBM cell lines while ITGA5 was high-expressed compared with HEB. miR-330-5p could directly target ITGA5 as well as suppress its expression in GBM cells. Up-regulation of miR-330-5p and down-regulation of ITGA5 both have an inhibitory effect on cell proliferation, invasion, and migration. Meanwhile, they could also promote GBM cell apoptosis. miR-330-5p could suppress proliferation and invasion of GBM cells through targeting ITGA5.
本研究旨在通过靶向整合素α5（ITGA5）来探究miR-330-5p对胶质母细胞瘤（GBM）细胞增殖和侵袭的生物学作用。采用RT-qPCR检测GBM细胞系（U87、U251和U373）及正常脑胶质细胞系（HEB）中miR-330-5p和ITGA5 mRNA的表达。使用蛋白质印迹法检测ITGA5的蛋白表达。本研究采用MTT法、集落形成试验、Transwell试验、伤口愈合试验及流式细胞术分析来确定GBM细胞的生物学功能（包括细胞增殖、侵袭、迁移、凋亡及细胞周期）。本研究应用双荧光素酶报告基因试验来鉴定miR-330-5p与ITGA5之间的靶向关系。与HEB相比，miR-330-5p在GBM细胞系中低表达，而ITGA5高表达。miR-330-5p可直接靶向ITGA5并抑制其在GBM细胞中的表达。上调miR-330-5p和下调ITGA5均对细胞增殖、侵袭及迁移有抑制作用。同时，它们还可促进GBM细胞凋亡。miR-330-5p可通过靶向ITGA5抑制GBM细胞的增殖和侵袭。
