Effect of RNA Interference Decreasing MSI1 Expression on Chemosensitivity of Human Glioma Cells
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摘要:目的
探讨RNA干扰沉默MSI1表达对人脑胶质瘤细胞化疗药物敏感度的影响。
方法构建针对MSI1 mRNA的小分子RNA干扰（siRNA）重组质粒pSIREN-MSI1，转染人脑胶质瘤U-87MG细胞；Western blot检测U-87MG细胞内MSI1蛋白的表达；CCK-8法观察U-87MG细胞对化疗药物替莫唑胺（TMZ）敏感度的改变；流式细胞仪（FCM）检测pSIREN-MSI1转染后对U-87MG细胞凋亡的影响。
结果成功构建了重组质粒pSIREN-MSI1，并成功转染U-87MG细胞；Western blot结果显示pSIREN-MSI1抑制U-87MG细胞中MSI1蛋白的表达（P<0.01）；CCK-8结果显示在替莫唑胺作用下，pSIREN-MSI1组U-87MG细胞的存活率明显下降（P<0.01）；FCM结果表明pSIREN-MSI1转染后明显提高U-87MG细胞的凋亡率（P<0.01）。
结论MSI1 siRNA能特异性抑制人脑胶质瘤细胞U-87MG细胞中MSI1的表达，增强人脑胶质瘤U-87MG细胞对化疗药物的敏感度，并促进细胞的凋亡。
Abstract:ObjectiveTo explore the effect of RNA interference decreasing MSI1 gene expression on the chemosensitivity of human glioma cells U-87MG.
MethodsThe design and synthesis of Msi1-specific siRNA and the transfection of U-87MG cells were conducted. The expression of MSI1 in U-87MG cells was detected by Western blot. CCK-8 was employed to observe the sensitivity of U-87MG cells to temozolomide(TMZ). FCM was used to observe the apoptosis rate of U-87MG cells after pSIREN-MSI1 transfection.
ResultsLentiviral vector-mediated MSI1-siRNA was successfully constructed. Immunofluorescence assay demonstrated that the transfection efficiency was above 60%. Western blot analyses demonstrated that pSIREN-MSI1 could significantly inhibit the expression of MSI1 in U-87MG cells (P<0.01); CCK-8 results showed that when U-87MG cells were exposed to temozolomide, the growth inhibiting rate of pSIREN-MSI1 transfected cells was significantly increased, compared with those of blank control group and negative control group (P<0.01). FCM results showed that the apoptosis rate of U-87MG cells were increased after pSIREN-MSI1 transfection(P<0.01).
ConclusionMSI1-siRNA could significantly inhibit the expression of MSI1 in human glioma cells U-87MG. It could induce the apoptosis and enhance the chemosensitivity of U-87MG cells.
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Key words:
- MSI1 /
- RNA interference /
- Glioma /
- Chemosensitivity
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图 1 Western blot检测pSIREN- MSI1转染U-87MG细胞后MSI1蛋白的表达
Figure 1 Expression of MSI1 protein in U-87MG cells after pSIREN-MSI1 transfection detected by Western blot
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图 2 CCK-8法检测不同浓度替莫唑胺作用于各组转染细胞后的细胞存活率
Figure 2 Survival rates of U-87MG cells treated with TMZ in each group detected by CCK-8
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