腺病毒介导单纯疱疹病毒胸苷激酶基因/更昔洛韦系统杀伤膀胱癌细胞的研究

目的：探讨单纯疱疹病毒胸苷激酶基因／更昔洛韦（HSV－tk/GCV)系统杀伤膀胱癌细胞的作用。方法；采用人巨细胞病毒早期蛋白启动子驱动HSV－tk基因重组腺病毒（Ad)载体，体外转染HTB9和Scan BER细胞，MTT法测定杀伤率，PCR检测Ad-tk转染细胞中tk基因及腺病毒E1区基因。结果：GCV对转染的癌细胞有杀伤作用，感染复数（MOI）100时，10mg/L GCV杀伤80％的tk^ 细胞，而对未转染的细胞无杀伤作用。不同MOI的Ad-tk转染两种细胞，其毒性与Ad-tk MOI正相关，MOI100时，15mg/L GCV杀伤100％的细胞，无GCV组未受到抑制。PCR证实Ad-tk转染细胞有tk基因，腺病毒E1区基因阴性。Annexin V法证实杀伤效应与凋亡有关。结论：HSV-tk/GCV系统是一种有效，安全治疗膀胱癌的新方法。

Study of adenoviral-mediated gene therapy for bladder cancer cell

Objective To study the killing effects of adenovirus-mediated defective adenovirus carring the HSV-tk gene(Ad-tk ) in bladder cancer cells.Methods Ad-tk was amplified on a large scale in293 cells.The purified recombinant adenovirus had the advanta ges of higly efficient gene transduction and high titer.In vitro experiments,the HTB9 cells and ScanBER cells transferred by tk gene were killed in the presence of GCV treatment.Results When MOI was 100,10mg/L GCV killed 80% tk cells.In addition,Ad tk~ density was related with toxicity of cells.15mg/L GCV killed 100% tumor cells.PCR proved that tk gene had been transferred into cells successfully.Fluorescent staining confirmed that HSV-tk/GCV system could induce apoptosis of bladder cancer cells .C onclusion Adenovirus-mediated HSV-tk/GCV system might be used as a n apromising suicide gene therapy for bladder carcinoma.