RNA干扰hTERT基因对人恶性黑素瘤A375细胞迁移和侵袭的影响.pdf

摘要 目的：使用特异性siRNA 靶向干扰黑素瘤A375 细胞中hTERT 基因的表达后，观察对黑素瘤细胞迁 移和侵袭能力的影响，从而探索hTERT 基因在黑素瘤发病过程中的作用。 方法：培养黑素瘤A375 细胞，分别设置空白对照组、阴性对照组和干扰组。干扰组的黑素瘤A375 细胞转染课题组前期筛选出的特异性hTERT siRNA ，阴性对照组则使用阴性对照siRNA 对细胞进行 转染，空白对照组细胞不做处理。Real-Time PCR 和 Western-blot 实验分别检测各组细胞中hTERT mRNA 和蛋白的表达水平，之后使用细胞划痕实验和Transwell 侵袭实验检测各组细胞的迁移和侵袭 能力。 结果：干扰组中使用特异性hTERT siRNA 靶向干扰hTERT 基因可明显下调hTERT 基因mRNA 和 蛋白的表达，与阴性对照组和空白对照组相比差异具有统计学意义（P ＜0.05 ）。细胞划痕实验和 Transwell 侵袭实验结果显示，当黑素瘤A375 细胞中hTERT 基因表达沉默后，细胞的迁移和侵袭能 力也出现明显下降（P ＜0.05）。 结论：特异性hTERT siRNA 可以有效沉默黑素瘤A375 细胞中hTERT 基因的表达，并抑制恶性黑素 瘤A375 细胞的迁移和侵袭能力。 关键词：黑色素瘤；小干扰RNA ；人端粒酶逆转录酶；侵袭；迁移 I Abstract Objective: To target the interference of hTERT gene expression in melanoma A375 cells with specific siRNA and observe the effect on the migration and invasion ability of melanoma cells, in order to explore the role of hTERT gene in the pathogenesis of melanoma. Methods: Culture melanoma A375 cells ，and set up blank control group, negative control group and interference group respectively. The interference group used specific hTERT siRNAs screened by the research group to transfect melanoma A375 cells, and the negative control group used negative control siRNA to transfect the cells. The blank control group cells were not treated. Real-Time PCR and western experiments were used to detect hTERT mRNA and protein expression levels in each group of cells. Cell scratch and Transwell experiments were used to detect the cell migration and invasion ability of each group before and after interference. Results: Targeted interference with hTERT gene using specific hTERT siRNA can significantly down-regulate the expression of hTERT gene mRNA and protein,