脑肿瘤二级康复预防:阳离子脂质体介导HSV-TK/ACV系统对人脑胶质瘤细胞增殖活性的影响(英文)

PhaseⅡrehabilitation/prevention of brain tumor:effect of the HSV-TK/ACV syst em mediated by cationic lipsome on the proliferative activity of human glioma cells

背景:基因治疗是国内外对于脑胶质瘤生物学治疗的研究热点。目的:应用已克隆并构建的真核表达载体pCR3-TK,探讨HSV-TK/ACV系统对人脑胶质瘤细胞抑制生长和杀伤作用研究。设计:以细胞为研究对象的实验研究。单位:一所大学医院神经外科、肿瘤内科。对象:实验于2004-01/04在哈尔滨兽医研究所生物技术国家重点实验室完成。所用的真核表达载体pCR3-TK由作者构建,TJ905细胞株由天津神经病学研究所浦佩玉教授惠赠。选择未转染和转染空载体的细胞作为对照组。方法:用阳离子脂质体Lipofectamine将pCR3-Uni及含HSV-TK基因的真核表达载体pCR3-TK转染至人脑胶质瘤细胞株TJ905中,筛选出阳性克隆,阳性细胞克隆给予ACV(50mg/L),72h后,收集玻片,进行AgNOR染色。主要观察指标:对未转染和转染不同载体的TJ905细胞ACV作用后银染颗粒进行记数。结果:转染HSV-TK基因的细胞,在给予ACV后,细胞增殖活性明显降低,转染pCR3-Uni和pCR3-TK细胞克隆的AgNOR颗粒数分别为14.33和6.67(P<0.01)。结论:AgNOR计数是一种操作简便、检测细胞增殖活性的方法,为研究HSV-TK/ACV系统的抗肿瘤机制提供帮助。

BACKGROUND:Gene therapy is a popular topic in domestic and overseas studies on biological therapy for brain tumor. OBJECTIVE:By using a newly constructed eukaryotic expression vector of pCR3 T K,the effect of the HSV TK/ACV system on the proliferative activity of human gl ioma cells was investigated. DESIGN:Experimental study based on cells. SETTING:Department of neurosurgery and department of oncology in a university hospital. MATERIALS:The study was conducted at the National Key Laboratory of Veterinary Biotechnology of Harbin Veterinary Research Institute from January to April in 2004.The eukaryotic expression vector of pCR3 TK was constructed by the author. The TJ905 strain was a gift from professor Pu Pei yu,who worked in the Neurolo gy Institute of Tianjin city.The nontransfected cells and the cells transfected with pCR3 Uni vector were set as controls. METHODS:By using Lipofectamine(a cationic liposome),the pCR3 Uni vector and t he recombinant pCR3 TK plasmid(inserted with HSV TK gene) were transfected int o the human glioma cell strain TJ905.Then the positive clones were picked out a nd were given ACV(50 mg/L).Totally 72 hours later,the cover slips were collected and silver staining for nucleolus organizer regions(AgNORs) was performed. MAIN OUTCOME MEASURES:After the ACV treatment and AgNORs staining, the numbers of silver stained granules in TJ905 cells with or without transfections were c ounted respectively. RESULTS:In those cells transfected with HSV TK gene,after ACV treatment,a sig nificant decreasing in proliferative activity could be observed,and the average numbers of the silver stained granules in cells transfected with pCR3 Uni or p CR3 TK were 14.33 and 6.67 respectively(P< 0.01). CONCLUSION:As an easy to operate method, AgNOR counting is helpful for the s tudies on the proliferative activity of cells and the investigations into the po tential anti tumor mechanism of the HSV TK/ACV system.