Silencing of MGMT with small interference RNA reversed resistance in human BCUN-resistant glioma cell lines.
- Author:
Si-ming XIE
1
;
Mao FANG
;
Hui GUO
;
Xue-yun ZHONG
Author Information
- Publication Type:Journal Article
- MeSH: Blotting, Western; Carmustine; pharmacology; Cell Line, Tumor; Cell Proliferation; drug effects; Drug Resistance, Neoplasm; drug effects; genetics; Glioma; genetics; metabolism; Humans; O(6)-Methylguanine-DNA Methyltransferase; genetics; metabolism; RNA, Small Interfering; Reverse Transcriptase Polymerase Chain Reaction; Sincalide; metabolism
- From: Chinese Medical Journal 2011;124(17):2605-2610
- CountryChina
- Language:English
-
Abstract:
BACKGROUNDOur previous study had cloned two glioma cell lines SWOZ1 and SWOZ2 isolated from parental glioma cell line SWO38. The 1,3-bis(2-chloroethyl)-1-nitrosourea (BCNU) resistance of SWOZ1 was higher than that of SWOZ2. Since O6-methylguanine-DNA methyltransferase (MGMT) was thought to be closely related to BCNU resistance in glioma, this study aimed to explore the function of MGMT in glioma resistant to BCNU.
METHODSA BCNU resistant glioma cell line SWOZ2-BCNU was established. The expression of MGMT was detected in SWOZ1, SWOZ2 and SWOZ2-BCNU. Small interferencing RNA targeting MGMT was used to silence the expression of MGMT in resistant cell lines SWOZ1 and SWOZ2-BCNU. The cytotoxicity of BCNU to these cells was measured using the cell counting kit-8 assay. Statistical analysis was carried out by one-way analysis of variance in statistical package SPSS 13.0.
RESULTSThe resistance of SWOZ1 and SWOZ2-BCNU against BCNU was 4.9-fold and 5.3-fold higher than that of SWOZ2. The results of quantitative RT-PCR and Western blotting confirmed that MGMT was both significantly increased in SWOZ1 and SWOZ2-BCNU compared to SOWZ2. After transfection with small interferencing RNA targeting MGMT, a decreased level of MGMT mRNA expression in SWOZ1 and SWOZ2-BCNU for more than 75% compared to negative control was found and confirmed by Western blotting. As a result, the resistance against BCNU was reversed for about 50% both in the BCNU-resistant cell lines SWOZ1 and SWOZ2-BCNU.
CONCLUSIONSSilencing MGMT with specific small interferencing RNA can reverse the BCNU resistant phenotype in these glioma cell lines. MGMT may play an important role both in intrinsic and acquired BCNU-resistance in glioma.
