沉默LSD1基因对卵巢癌SKOV3细胞系紫杉醇耐药性及生物学行为的影响  

Effect of silent LSD1 gene on taxol resistance and biological behavior of ovarian cancer cell line SKOV3

作　　者：曾敏[1] 张娟 

机构地区：[1]成都市第一人民医院妇产科,四川成都610041 [2]深圳市罗湖区医院病理科

出　　处：《中国妇幼保健》2015年第25期4375-4378,共4页Maternal and Child Health Care of China

基　　金：国家中医药管理局中医临床诊疗技术整理与研究项目课题〔2001ZL48〕

摘　　要：目的探讨LSD1基因对卵巢癌SKOV3细胞系紫杉醇耐药性、增殖和细胞周期的影响。方法采用大剂量间断冲击方法建立人卵巢癌耐药细胞系SKOV3 res TAX。针对LSD1基因m RNA靶向设计小分子干扰RNA(si RNA)沉默片段,构建慢病毒颗粒载体,并转染卵巢癌耐药细胞系SKOV3 res TAX。流式细胞术检测慢病毒转染效率;实时定量聚合链式反应检测转染后LSD1的转录水平;CCK-8试剂盒检测转染后耐药细胞系的增殖能力及紫杉醇耐受能力;流式细胞术检测转染后细胞周期的时相变化;蛋白质免疫印迹法检测LSD1基因、抑癌基因p53和p21、细胞周期依赖性蛋白CDK2、周期蛋白cyclin E及核增殖抗原PCNA的表达。结果通过测定卵巢癌细胞系紫杉醇半抑制浓度,证明耐药性细胞系建立成功。流式细胞术结果显示,慢病毒颗粒载体转染效率均非常高,LSD1沉默慢病毒载体si-LSD1-1能显著降低LSD1的转录水平。转染后耐药细胞系的增殖能力及紫杉醇耐受能力都显著下降,细胞周期被阻滞在G0/G1期。蛋白质印迹结果显示,同si-control组相比,LSD1蛋白表达量下降,同时p53和p21表达上调,CDK2、cyclin E和PCNA表达被显著下调。结论 LSD1基因与卵巢癌SKOV3耐药细胞系的增殖、细胞周期以及紫杉醇耐受能力具有密切关系,可作为紫杉醇等化疗药物联合基因治疗的候选靶点。Objective： To explore the effect of LSD1 gene on taxol resistance,proliferation,and cell cycle of ovarian cancer cell line SKOV3. Methods： Human ovarian cancer cell line SKOV3 res TAX was established by high-dose intermittent shocks. The targetable si RNA for LSD1 gene was synthesized and implanted into lentiviral vector,then the vector was transfected into SKOV3 res TAX. The transfection efficiency was detected by flow cytometry. The LSD1 gene transcription level was detected by real- time quantification polymerase chain reaction. CCK-8 kit was used to detect the proliferation ability and taxol resistance capacity after transfection. The change of cell cycle was measured by flow cytometry. The expressions of LSD1,p53,p21,CDK2,cyclin E,and PCNA were detected by West blot. Results：The resistant cell line was established successfully by measuring the half inhibitory concentration of taxol. Flow cytometry results showed that the lentiviral vector particles had high transfection efficiency. si- LSD1- 1 lentiviral vector significantly reduced the transcription level of LSD1. The proliferation and taxol-resistant abilities significantly decreased after transfection,and cell cycle was arrested in G0 / G1 phase after transfection. Western blot results showed that the expression level of LSD1 protein decreased after transfection compared with si-control group. Meanwhile,p53 and p21 expression levels were up-regulated,CDK2,cyclin E,and PCNA expression levels were significantly downregulated. Conclusion： LSD1 gene is closely correlated with proliferation,cell cycle,and taxol resistance of ovarian cancer cell line SKOV3,which can be used as a candidate target gene of chemotherapeutic therapy combined with gene therapy.

关 键 词：卵巢癌 赖氨酸特异性脱甲基酶1 RNA干扰 紫杉醇 

分 类 号：R711.75[医药卫生—妇产科学]