【摘要】 目的探讨利用RNAi技术研究靶向抑制宫颈癌细胞株Hela中HPV18E6的可行性。方法根据哺乳动物SiRNA设计原则要求,利用T7RNA聚合酶体外合成针对HPV18E6、E7的SiRNA,用2%的琼脂糖电泳鉴定、脂质体转染Hela细胞,分4组,A组:HPV18E6的SiRNA干预组;B组:HPV18E7的SiRNA干预组;C:HPV18E6、E7的SiRNA干预组;D:基因组文库任意SiRNA为对照组。于转染48、72、96h后,用免疫组化法测定HPV18E6蛋白的表达。结果干预组A和C转染后48、72h,HPV18E6的阳性表达明显减少,以72h为明显,96h后表达无明显减少。而干预B组和对照D组的HPV18E6的阳性表达无明显改变。结论靶向HPV18E6的SiRNA能特异瞬时抑制靶基因E6的表达,该方法将为宫颈癌预防和治疗提供新的策略。更多还原

【Abstract】 【Objective】By RNAi, we investigate whether HPV18 E6 SiRNA targeted Hela cells cervical carcinoma can inhibit the expressions of HPV18 E6 oncoprotein. 【Methods】For SiRNA studies in mammalian cells, we synthesize HPV18 E6 and E7 SiRNAs in vitro using T7 RNA polymerase. The products were checked by running on 2% agarose gel. Then Hela cells were grown in standard conditions and transfecting with lipofectamineTM. The cells were divided into four groups, A group: HPV18 E6 SiRNA; B group: HPV18 E7 SiRNA; C group: HPV18 E6+E7 SiRNA; D group: control. After transfection 48, 72 and 96 hours, we investigated the expressions of HPV18 E6 in Hela cells of different groups by S-P immunohistochemical methods. 【Results】 The expressions of HPV18 E6 in group A and C were significant decreased after transfected 48 hours, 72 hours, especially after transfection 72 hours, but have no difference after transfected 96 hours. In group B and D, the expressions of HPV18 E6 is similar to those before transfected with SiRNA E6 and E7. 【Conclusions】Targeted HPVE6 SiRNA can specific inhibit the expressions of HPV18 E6 oncoprotein and it may be provided a new methods of cervical carcinoma gene therapy.更多还原