自杀基因联合全反式维甲酸治疗对前列腺癌PC-3细胞旁观者效应的影响

Effect of suicide gene combined with all-trans retionic acid on the bystander effect in prostate cancer PC-3 cells

目的观察单纯疱疹病毒胸苷激酶（HSV—TK）／丙氧鸟苷（GCV）系统联合全反式维甲酸（ATRA）对人雄激素非依赖性前列腺癌PC-3细胞旁观者效应的影响。方法噻唑蓝（MTT）比色法检测HSV—TK（Ad—TK）／GCV系统对PC-3细胞的杀伤力。流式细胞仪检查连接蛋白（Cx）43阳性细胞数。结果单纯Ad—TK／GCV系统对PC-3细胞旁观者效应在TK^＋：TK^-细胞混合比例为5：5时明显出现（P〈0．05），而联合ATRA后，在TK^＋：TK^-细胞混合比例为3：7时就明显出现（P〈0．05）。Ad-TK／GCV联合ATRA处理PC-3细胞后，在细胞抑制率均达到约50％时，其TK^＋PC-3细胞混合比例可降低20％。ATRA（10^-6mol／L）处理PC-3细胞1、3、5d后，Cx43阳性细胞率由10．5％分别升高为25．8％、50．8％和62．4％（P〈0．01）。结论ATRA可能通过增强基于Cx43的旁观者效应，从而提高Ad-TK／GCV系统对前列腺癌的杀伤效应。

Objective To probe the effects of Herpes simplex virus thymidine kinase （HSV- TK）/ganciclovir（GCV） system combined with all-trans retinoic acid （ATRA） on the bystander effect （BE） in andrgen-independent cell line PC-3. Methods Multiplicity of infection （MOI） of adenovirus （Ad） to PC-3 cell was examined by flow cytometer （FCM） at the infection rate of 100% and TK gene was identified by using RT-PCR. MTT was used to judge the lethality and BE of Ad-TK/GCV system alone or combined with ATRA on PC-3 cells in vitro. FCM and immunocytochemistry were used to detect positive cells ratio,expression level and the location of Cx43 protein in PC-3 ceils, respectively. Results When MOI was 200,PC-3 cells were infected by Ad stably at the rate of 100. The BE of Ad-TK/GCV on PC-3 cells was poor, but improved after combined with ATRA （ P 〉 0.05 ）. There was synergic effects between ATRA and BE of Ad-TK/GCV （ P 〈 0.05 ）. After PC-3 cells were treated with ATRA, the rate of cells positive for Cx43 was increased significantly, and Cx43 protein was also elevated markedly. Conclusion ATRA might enhance Cx43-based BE, so that Ad-TK/GCV system combined with ATRA could kill PC-3 cells more significantly than Ad-TK/GCV system alone.