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目的通过合成靶向分化抗原簇44(CD44)基因的siRNA片段,下调CD44在脑胶质瘤细胞U251中的表达,揭示其在胶质瘤增殖、迁移和侵袭中所起的作用。方法根据CD44基因序列设计并合成的siRNA片段转染U251细胞,利用荧光实时定量(qRT-PCR)检测细胞中CD44基因mRNA水平的变化;蛋白印迹实验(Western blot)检测CD44基因蛋白水平的变化;四甲基偶氮唑蓝染色(methyhhiazolyl tetrazolium,MTT)法检测U251细胞增殖;单细胞划痕愈合实验、Transwell小室侵袭实验检测U251细胞的迁移与侵袭能力变化。结果体外合成的CD44-siRNA能有效抑制U251细胞中CD44基因在mRNA水平与蛋白水平的表达(P<0.05),并抑制细胞的增殖(P<0.05)和迁移侵袭能力(P<0.05)。结论 CD44-siRNA能够有效抑制U251脑胶质瘤细胞的增殖及其迁移侵袭力,为以CD44为靶点的肿瘤基因治疗奠定了基础。
OBJECTIVE: To investigate the role of CD44 in the proliferation, migration and invasion of glioma by down-regulating the expression of CD44 in the glioma cell line U251 by synthesizing siRNA fragments targeting differentiated antigen cluster 44 (CD44) gene. Methods U251 cells were transfected with siRNA fragments designed and synthesized according to the sequence of CD44 gene. The expression of CD44 mRNA was detected by qRT-PCR. The protein level of CD44 was detected by Western blot. Methyhhiazolyl tetrazolium (MTT) assay was used to detect the proliferation of U251 cells. Single cell wound healing assay and Transwell chamber invasion assay were used to detect the migration and invasion of U251 cells. Results The CD44-siRNA synthesized in vitro could effectively inhibit the expression of CD44 mRNA and protein in U251 cells (P <0.05) and inhibit the proliferation (P <0.05) and invasion (P <0.05). Conclusion CD44-siRNA can effectively inhibit the proliferation and migration of U251 glioma cells and lay a foundation for the gene therapy of CD44.