Expressions of miR-124* and laminin-8 in gliomas and their interactive regulation
10.3760/cma.j.issn.1671-8925.2014.08.002
- VernacularTitle:miR-124*和laminin-8β1链在脑胶质瘤中的表达及相互调控作用
- Author:
Guohui LU
1
;
Chunmei JIANG
;
Jian DUAN
;
Dongwei ZHOU
;
Tao HONG
;
Shizhong ZHANG
Author Information
1. 南昌大学第一附属医院神经外科
- Keywords:
Glioma;
MiR-124*;
Laminin-8;
Regulation
- From:
Chinese Journal of Neuromedicine
2014;13(8):763-767
- CountryChina
- Language:Chinese
-
Abstract:
Objective To explore the expressions ofmiR-124* and laminin-8 in different grades of human gliomas,and analyze the potential regulatory relationship between them.Methods Thirty human glioma specimens of different stages and two healthy brain tissues,collected in our hospital from January 2011 to December 2012,were used in our study; Western blotting was performed to detect the laminin-8 and miR-124* protein expressions; plasmid vectors carried psiCHECKTM-2-wild-type (WT) laminin β1 chain 3 'untranslated region (3'UTR) and psiCHECKTM-2-mutant (MUT) laminin β1 chain 3'UTR were established,and they were,respectively,co-transfected with the miR-124* mimics,negative control ofmiRNA mimics,miR-124* inhibitors,negative control ofmiRNA inhibitors into the U87 cells;besides,cotransfection with psiCHECKTM-2 into U87 cells was used as blank control group; the fluorescence intensity of each group was determined by dual-luciferase assay 48 h after the cotransfection.Dual-luciferase assay was developed to verify the regulatory effects ofmiR-124* on laminin-8.The nu/nu nude mice were given subcutaneous injection of U87 cells to establish giloma animal models; the expression of β1 chain-containing laminin-8 protein was detected in gliomas by Western blotting; the expression of vascular endothelial cell marker CD31 was observed by immunohistochemical staining.Results Western blotting showed high expressions of laminin-8 protein in high-grade gliomas (WHO grade Ⅲ/ⅣV),but low expressions in low-grade gliomas (WHO grade Ⅰ/Ⅱ); real-time fluorescent quantitative PCR showed that the expression of miR-124* in high-grade gliomas was significantly lower than that in low-grade gliomas.Dual-luciferase assay confirmed that fluorescence intensity of U87 cells carried psiCHECKTM-2-WT laminin β1 chain 3'UTR and miR-124* mimics (2.13±0.25) was significantly weaker than that of cells carried psiCHECKTM-2-WT laminin β1 chain 3'UTR and negative control of miRNA mimics (2.71±0.08,P＜0.05); that of those carried psiCHECKTM-2-MUT laminin β1 chain 3'UTR and miR-124* inhibitors (3.18 ±0.22) was significantly stronger than those carried,psiCHECKTM-2-MUT laminin β1 chain 3'UTR and negative control of miRNA inhibitors (2.70±0.17,P＜0.05).The animal model results indicated the laminin-8 and CD31 expressions in miR-124* treatment group were obviously lower than those in control group.Conclusion MiR-124*,which can affect the malignant degrees of human gliomas through playing its negative regulation role in laminin-8 expression,maybe a target gene for the diagnosis and therapy of gliomas.
