论文部分内容阅读
目的探讨热休克蛋白90(HSP90)抑制剂17-丙烯胺基-17-去甲氧格尔德霉素(17-AAG)联合紫杉醇对人未分化甲状腺癌FRO细胞增殖和凋亡的影响。方法 1采用四甲基偶氮唑盐微量酶反应比色法(MTT比色法)测定不同浓度(17-AAG:0.312 5、0.625 0、1.250 0、2.500 0及5.000 0μmol/L;紫杉醇:0.001 0、0.010 0、0.100 0及1.000 0μmol/L)、不同时间(24、48及72 h)17-AAG、紫杉醇单药和联合处理(17-AAG:0.625 0μmol/L,紫杉醇:0.001 0、0.010 0、0.100 0及1.000 0μmol/L)后FRO细胞的增殖抑制率。2采用流式细胞仪检测17-AAG、紫杉醇单药及联合处理24 h后(17-AAG:0.625 0μmol/L、紫杉醇:0.100 0μmol/L;联合用药:17-AAG的浓度为0.625 0μmol/L,紫杉醇的浓度为0.100 0μmol/L)FRO细胞的细胞周期变化及凋亡率。3采用胱天蛋白酶-3(Caspase-3)和Caspase-9检测试剂盒检测17-AAG、紫杉醇单药及联合处理24 h后(17-AAG:0.625 0μmol/L、紫杉醇:0.100 0μmol/L;联合用药:17-AAG的浓度为0.625 0μmol/L,紫杉醇的浓度为0.100 0μmol/L)后FRO细胞中的Caspase-3和Caspase-9活性。空白对照组均不加任何药物,只加培养液。结果 1同时点空白对照组、各剂量17-AAG组/紫杉醇组/17-AAG联合紫杉醇组的增殖抑制率随浓度升高而逐渐升高,任2组比较差异均有统计学意义(P<0.05);各剂量17-AAG组/紫杉醇组/17-AAG联合紫杉醇组的增殖抑制率在24、28及72 h逐渐增高,任2组比较差异均有统计学意义(P<0.05);同时点同浓度情况下,17-AAG联合紫杉醇组的增殖抑制率均高于单独用药组(P<0.05)。各时点17-AAG与紫杉醇联合的q值均大于1.15,两者之间呈协同作用。2 17-AAG组、紫杉醇组及17-AAG联合紫杉醇组FRO细胞的凋亡率均明显高于空白对照组(P<0.05),且17-AAG联合紫杉醇组FRO细胞的凋亡率高于17-AAG组和紫杉醇组(P<0.05)。3 17-AAG组、紫杉醇组及17-AAG联合紫杉醇组FRO细胞的Caspase-3和Caspase-9活性均高于空白对照组(P<0.05);且17-AAG联合紫杉醇组细胞的Caspase-3和Caspase-9活性均高于17-AAG组和紫杉醇组相应指标(P<0.05)。结论 17-AAG和紫杉醇均可明显抑制FRO细胞的增殖并诱导细胞凋亡,联合用药有一定的协同效应,呈剂量依赖关系。
Objective To investigate the effects of heat shock protein 90 (HSP90) inhibitor 17-acrylamido-17-demethoxygeldanamycin (17-AAG) and paclitaxel on the proliferation and apoptosis of human undifferentiated thyroid cancer cell line FRO. Method 1 MTT colorimetry was used to determine the effect of different concentrations of 17-AAG: 0.312 5,0.625 0,1.250 0,2.500 0 and 5.000 0μmol / L; paclitaxel 0.001 (17-AAG: 0.625 0μmol / L, paclitaxel: 0.001 0,0.010) at different times (24,48 and 72 h) at different time points (0, 0.010 0, 0.100 0 and 1.000 μmol / L) 0, 0.100 0 and 1.000 0μmol / L). (2) After 17-AAG, paclitaxel alone and 24 h treatment with 17-AAG: 0.625 0 μmol / L and paclitaxel: 0.100 0 μmol / L by flow cytometry, the concentration of 17-AAG was 0.625 0 μmol / , Paclitaxel concentration 0.100 0μmol / L) FRO cell cycle and apoptosis rate. 3 Caspase-3 and Caspase-9 detection kit for 17-AAG, paclitaxel monotherapy and combined treatment for 24 h (17-AAG: 0.625 0μmol / L, paclitaxel: 0.100 0μmol / L; Caspase-3 and Caspase-9 activity in FRO cells after co-administration of 0.625 0 μmol / L 17-AAG and 0.100 0 μmol / L paclitaxel. Blank control group without any drugs, only add medium. Results 1 At the same time point blank control group, the proliferation inhibition rate of 17-AAG group / paclitaxel group / 17-AAG combined with paclitaxel group increased gradually with the increase of concentration, and the difference was statistically significant (P < 0.05). The proliferation inhibition rate of 17-AAG group / paclitaxel group / 17-AAG combined with paclitaxel group increased gradually at 24, 28 and 72 hours, and there was significant difference between any two groups (P <0.05) At the same concentration, 17-AAG combined with paclitaxel showed higher proliferation inhibition rate than that of the single drug alone group (P <0.05). At each time point 17-AAG combined with paclitaxel q values ?? were greater than 1.15, the synergy between the two. The apoptosis rate of FRO cells in 17-AAG group, paclitaxel group and 17-AAG combined with paclitaxel group was significantly higher than that in blank control group (P <0.05), and the apoptosis rate of FRO cells in 17-AAG combined with paclitaxel group was higher than 17 -AAG group and paclitaxel group (P <0.05). The Caspase-3 and Caspase-9 activities of FRO cells in 17-AAG group, paclitaxel group and 17-AAG combined with paclitaxel group were significantly higher than those in blank control group (P <0.05). Caspase-3 And Caspase-9 activity were higher than 17-AAG group and paclitaxel group corresponding indicators (P <0.05). Conclusions Both 17-AAG and paclitaxel can significantly inhibit the proliferation of FRO cells and induce apoptosis. The combination of 17-AAG and paclitaxel has a synergistic effect in a dose-dependent manner.