胸苷激酶基因系统联合大蒜素诱导膀胱癌细胞凋亡的研究

The effect of cell killing and apoptosis by human herpes simplex virus-thymidine kinase/ganciclovir system combined with a llitride in BIU87 cells

目的 探讨单纯疱疹病毒胸苷激酶 (HSV TK) /更昔洛韦 (GCV)系统联合大蒜素对膀胱癌细胞的协同杀伤效应及对细胞凋亡的影响。方法 通过脂质体将胸苷激酶 (TK)基因转入膀胱癌BIU87细胞构建BIU87TK细胞,观察GCV(1mg/L)、大蒜素(5mg/L)及GCV+大蒜素(浓度同前)共同作用对BIU87TK细胞形态学、细胞凋亡率及细胞周期变化的影响,检测bcl 2、bax、半胱天冬蛋白酶(caspase 3)表达变化及caspase 3活性变化。结果 药物作用 72h,GCV+大蒜素对BIU87TK细胞的抑制率为 72 50%,高于GCV作用后的 35 00%和大蒜素作用后的 37 00%,二者可发挥协同抑瘤作用; (F=6 46,P<0 05)。GCV+大蒜素联合作用前期凋亡率高于单独作用细胞,细胞周期阻滞在S和G2 期,与单独作用比较阻滞时效提前;二者可抑制bcl 2表达并可促进caspase 3表达及活化。结论 HSV TK/GCV系统联合大蒜素可通过共同诱导凋亡发挥协同作用,抑制膀胱癌细胞生长。细胞周期阻滞,相关凋亡基因表达及活性的变化可能发挥了重要的作用。

Objective To study the kill in g effect of human herpes simplex virus-thymidine kinase/ganciclovir (HSV-TK/GC V) system combined with allitride and the possible apoptosis mechanism in BIU87 cells. Methods The cytotoxicity after combination were estimated by theamine blue te trazolium bromide(MTT). The morphological changes were observed with inverted m icroscope and in-situ cell apoptosis detection kit. Changes of apoptosis rate and cell cycle were assessed by flow cytometry.B-cell lymphoma-2 (bcl-2), b ax, caspase-3 (cysteine aspartate specific proteinase) mRNA changes were detect ed by reverse transcriptase polymerase chain reaction, and caspase-3 activity w as estimated with chromometry. Results For combination group, the cell killing rate was raised to 72.50% to compare with 35.00% of GCV and 37.00% of allitride separately and there was a synergistic effect between these two drugs. The cell apoptosis was induced in all three groups and for the combination group the time of S-phase and G 2 -p hase arrest were earlier than other two groups. Both drugs could inhibit the ex pression of bcl-2 and promote the expression and activity of caspase-3. Conclusions The combination of HSV-TK/GCV system with allitride can inhibit the proliferation of BIU87 cells congenerously through apoptosis, which may be corre lated with S-and G 2-phase arrest, down-regulation of bcl-2 and increased c aspase-3 expression and its activity.