【作者】 吴永忠；

【导师】 李少林；

【作者基本信息】 重庆医科大学 ， 药理学， 2003， 博士

【摘要】 第一部分目的：探讨c-erbB-2与c-raf-1基因反义寡核苷酸联合转染对人卵巢癌SKOV3细胞增殖的抑制作用及其分子机制。 方法：实验分成7组：正常对照组、c-erbB-2正义治疗组、c-raf-1正义治疗组、c-erbB-2反义治疗组、c-raf-1反义治疗组、全剂量联合治疗组和半剂量联合治疗组。脂质体转染后不同时间分别进行MTT试验、集落形成试验、细胞周期检测、电镜检测、荧光显微镜检测、RT-PCR、蛋白质荧光强度测定，以观察不同条件处理后各组细胞的增殖、凋亡、细胞周期、mRNA及蛋白质表达有无差别。 结果：就全剂量联合治疗组和半剂量联合治疗组而言，转染72h的OD值分别为0.151（p﹤0.005）和0.073（p﹤0.005），增殖抑制率分别达到了88.7%和94.5%，克隆形成率分别为19.4%（p﹤0.01）和12.8%（p﹤0.01），凋亡率分别为24.3%（p﹤0.05）和31.5%（p﹤0.01），明显地下调c-erbB-2与c-raf-1基因mRNA及其蛋白质的表达水平。 结论：在卵巢癌的基因治疗中，c-erbB-2与c-raf-1基因反义寡核苷酸联合治疗优于单反义治疗，半剂量联合又优于全剂量联合治疗。关键词：SKOV3细胞; 反义寡核苷酸; 联合转染; 卵巢癌更多还原

【Abstract】 PART ONEOBJECTIVE: To investigate the proliferating inhibition and the mechanisms of c-erbB-2 and c-raf-1 genes antisense oligodeoxynucleotides combined transfection in the human ovarian carcinoma SKOV3 cell lines. METHODS: There were 7 groups in our study, normal control group, c-erbB-2 sense experimental group, c-raf-1 sense experimental group, c-erbB-2 antisense experimental group, c-raf-1 antisense experimental group, whole-dose experimental group, half-dose experimental group. In the different time after liposome-mediated transfection, the cell proliferation, apoptosis, mRNA level, protein expressing level and cell cycle were observed by MTT assay, flow cytometry, RT-PCR, electronic microscope, fluorescent microscope and cloning test. RESULTS: According to the results of whole-dose experimental group, half-dose experimental group, the OD-value were 0.151(p﹤0.005) and 0.073(p﹤0.005) transfected<WP=12>after 72h, the inhibitory rate of proliferation were 88.7% and 94.5%, the cloning efficiency were 19.4%（p﹤0.01）and 12.8%（p﹤0.01）, the apoptosis rate were 24.3%（p﹤0.05）and 31.5%（p﹤0.01） and the level of mRNA and protein were decreased statistically in both c-erbB-2 and c-raf-1,respectively. CONCLUSIONS: These results suggested that c-erbB-2 and c-raf-1 combined ASODN is superior to single ASODN, and the half-dose group is more efficient than the whole-dose group in the treatment of ovarian carcinoma.KEY WORDS: SKOV3 cell lines; antisense oligodeoxynucleotides; combined transfection; ovarian carcinoma更多还原