siRNA干扰MTMG对脑胶质瘤BCNU耐药的影响 |
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引用本文: | 谢思明,钟雪云. siRNA干扰MTMG对脑胶质瘤BCNU耐药的影响[J]. 中华医学杂志(英文版), 2010, 123(5) |
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作者姓名: | 谢思明 钟雪云 |
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作者单位: | 黄埔大道西601号暨南大学医学院1037,Jinan University |
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基金项目: | The National Natural Science Foundation of China (General Program, Key Program, Major Research Plan),China Postdoctoral Science Foundation |
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摘 要: | 目的:探讨MGMT在脑胶质瘤BCNU耐药中的作用。方法:前期研究中我们从脑胶质瘤细胞系SWO38中克隆出两株亚系SWOZ1和SWOZ2,其中SWOZ1对BCNU的耐药性强于SWOZ2。我们以SWOZ2细胞系为对象诱导出BCNU耐药的细胞系SWOZ2-BCNU,并检测SWOZ1、SWOZ2和SWOZ2-BCNU这三株细胞系中MGMT的表达情况。同时构建MGMT的小分子干扰RNA(siRNA)序列,利用脂质体法转染耐药细胞系SWOZ1和SWOZ2-BCNU,以SWOZ2为对照。通过荧光定量RT-PCR和Western blot法检测MGMT干扰后mRNA和蛋白质的表达水平,并通过CCK-8法检测耐药性的变化情况。结果:与SWOZ2相比,SWOZ1和SWOZ2-BCNU对BCNU的耐药性分别增加了4.9倍和5.3倍,同时在SWOZ1和SWOZ2-BCNU中MGMT的表达高于SWOZ2。经siRNA后,MGMT在SWOZ1和SOWZ2-BCNU的mRNA和蛋白质表达水平均显著下降(大于75%)。CCK-8法结果表明经siRNA干扰后SWOZ1和SWOZ2-BCNU对BCNU的耐药性均下降50%。结论:我们的结果证明MGMT经siRNA后能部分逆转BCNU耐药细胞SWOZ1和SWOZ2-BCNU对BCNU的耐药性,提示MGMT在脑胶质瘤BCNU耐药中起着重要的作用。
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关 键 词: | 脑胶质瘤;MGMT;BCNU耐药;小分子干扰RNA |
Silencing of MGMT with small interference RNA induced reversal of BCNU resistance in human BCNU-resistant glioma cell lines |
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Abstract: | Background: To explore the function of MGMT in glioma resistant against BCNU.Methods: Our previous study had cloned two glioma cell lines SWOZ1 and SWOZ2 isolated from parental glioma cell line SWO38. The BCNU resistance of SWOZ1 were stronger than that of SWOZ2. In this study, a BCNU resistant glioma cell line SWOZ2-BCNU was established. The expression of MGMT was detected in SWOZ1, SWOZ2 and SWOZ2-BCNU. SiRNA targeting MGMT was used to silence the expression of MGMT in resistant cell lines SWOZ1 and SWOZ2-BCNU. The expression level of MGMT mRNA and protein was detected by quantitative RT-PCR and western blot. The cytotoxicity of BCNU to these cell lines was measured using the cell counting kit assay.Results: The resistance of SWOZ1 and SWOZ2-BCNU against BCNU was 4.9-fold and 5.3-fold stronger than that of SWOZ2. The results of quantitative RT-PCR and western blot confirmed that MGMT was both significantly increased in SWOZ1 and SWOZ2-BCNU compared to SOWZ2. After transfection with siRNA 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 blot. As a result, the resistance against BCNU were reversed for about 50% both in the BCNU- resistant cell lines SWOZ1 and SWOZ2-BCNU.Conclusion: Our data demonstrated silencing MGMT with specific siRNA can reverse the BCNU resistant phenotype in these glioma cell lines, which indicate that MGMT may play an important role both in intrinsic and acquired BCNU-resistance in glioma. |
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Keywords: | Glioma MGMT SiRNA Drug resistance BCNU |
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