上皮性卵巢癌论文：转染MiRNA27a模拟物及抑制物对卵巢细胞系的顺铂耐药性的研究.docVIP

上皮性卵巢癌论文：转染MiRNA27a模拟物及抑制物对卵巢细胞系的顺铂耐药性的研究 【中文摘要】探讨采用脂质体2000 (lipofectamin2000)转染miRNA27a (microRNA miRNA)模拟物(mimics)及抑制物(inhabitors)对卵巢癌细胞系的顺铂(DDP)耐药性的研究。方法：本试验采用了小剂量DDP间歇诱导卵巢癌细胞系skov3建立卵巢癌顺铂耐药细胞系skov3/DDP:1.MTT检测细胞耐药系数；2.实时荧光定量PCR (Real-time PCR RT-PCR)检测skov3/DDP及skov3中miRNA27a的表达情况；2lipofectamin2000转染miRNA27a inhabitors、mimics,并用荧光显微镜检测转染效；3.MTT检测细胞增殖情况；4.实时荧光定量PCR检测多耐药基因(Multdrug Resistance Gene MDR1)基因的表达情况；5.流式细胞术检测细胞凋亡；6Western blot检测糖(P-gp)蛋白表达情况。结果：1skov3/DDP相对于skov3耐药系数为5.08；2miRNA27a在skov3/DDP中的表达明显高于skov3,约为2.80倍,有统计学意义；3skov3/DDP与其转染miRNA27a inhabitors, skov3及其转染mimics相比,MDR1的表达无统计学意义；4.转染抑制物的skov3/DDP的P-gp蛋白的表达较skov3/DDP降低了32.90%,转染mimics的skov3的P-gp蛋白的表达较skov3升高了20.71%；5.转染inhibitors及阴性对照(Negative Control NC)的Skov3/DDP在顺铂作用下存活率不同,有统计学意义；6.转染inhibitors的skov3/DDP与转染NC的skov3/DDP相比,前者凋亡率为10.37±2.58%,后者凋亡率为3.12±1.34%,有统计学意义。结论：上皮性卵巢癌耐药可能与miRNA27a的表达上调有关；miRNA27a可能是通过抑制MDR1基因的翻译来发挥作用而不通过抑制MDR1的转录发挥作用的；下调miRNA27a可以逆转卵巢癌细胞系的耐药性,增加其对顺铂的敏感性。 【英文摘要】:to investigate down-regulation and up-regulationm miRNA27a and its relation with drug resistance in human ovarian cancer skov3/DDP.Methods:This test uses a small dose of DDP for intermittenting ovarian cancer cisplatin-resistant cell lines. MTT was used to know the resistant factor and drug sensitivity; real-time PCR was used to detect the expression of miRNA27a in skov3/DDP and skov3. The cells were translated miRNA27a inhibitor, negative control. miRNA27a mimics and negative control by lipofectamin2000. The expression of MDR1 gene in the cells and transltated cells was measured by real-time PCR. cell proliferation analysis was measured by MTT. cell apoptosis analysis was measured by Flow cytometry. p-glycoprotein (p-gp) was measured by western blot.Results:(1) Compared with skov3, resistance coefficient of skov3/DDP is 5.08. (2) miRNA27a was an average of 2.80 -fold higher expression level in skov3/DDP cells than that in skov3 ceils, with a significant difference between the two groups(P0.05), (2) skov3/DDP cells tr