【摘 要】
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目的 目的:探讨长链非编码RNA H19(lncRNA H19)在紫杉醇耐药的三阴性乳腺癌(Triplenegative breast cancer,TNBC)细胞中的作用方法 方法:1.利用实时定量PCR,扩增编码lncRNA H19 的cDNA,并使用特异性lncRNA H19 发夹shRNA 敲除lncRNA H19,然后将它们插入慢病毒pHBLV-U6-ZsGreen-Puro 质粒中并收
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目的 目的:探讨长链非编码RNA H19(lncRNA H19)在紫杉醇耐药的三阴性乳腺癌(Triplenegative breast cancer,TNBC)细胞中的作用方法 方法:1.利用实时定量PCR,扩增编码lncRNA H19 的cDNA,并使用特异性lncRNA H19 发夹shRNA 敲除lncRNA H19,然后将它们插入慢病毒pHBLV-U6-ZsGreen-Puro 质粒中并收集慢病毒颗粒以感染TNBC 细胞(包括MDA-MB-453,MDA-MB-157,MDA-MB-231 细胞系),使lncRNA H19 在MDAMB-157 和MDA-MB-231 细胞系中稳定表达;2.MTT 法检测显示MDA-MB-157_H19、MDA-MB-231_H19 细胞与对照组细胞对紫杉醇的耐药性;3.通过凋亡分析、蛋白质印迹法及免疫组化,经过紫杉醇处理,分析MDA-MB-231/PTX_shH19 细胞与MDA-MB-231/PTX_shNC 细胞的凋亡率,以及Caspase-3 抑制剂Z-DEVD-FMK 在紫杉醇诱导细胞凋亡中的作用;4.观察敲除lncRNA H19 裸鼠中紫杉醇耐药的TNBC 肿瘤生长,以及紫杉醇处理后MDA-MB-231/PTX_shH19 和MDA-MB-231/PTX_shNC 异种移植物的肿瘤体积和重量.
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