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目的:检测flavopiridol联合泰素对人卵巢癌细胞系SKOV3、裸鼠人卵巢癌皮下及腹腔移植瘤模型的治疗作用。方法:应用CCK-8法、流式细胞术和原位细胞凋亡检测法(TUNEL)检测flavopiridol和泰素作用后卵巢癌细胞系SKOV3的细胞存活率和凋亡率;应用逆转录-多聚酶链反应(RT-PCR)检测作用前后细胞中cyclinD1的表达:应用ELISA法检测细胞中活性caspase-3的表达。建立裸鼠人卵巢癌皮下及腹腔移植瘤模型,观测flavopiridol和泰素治疗前后裸鼠肿瘤体积的变化并计算抑瘤率,用TUNEL和免疫组化法分别检测作用前后肿瘤组织中的凋亡情况和微血管密度(MVD)值。结果:flavopiri-dol(300nmol/L)或泰素(1μmol/L)单独应用24h均能显著降低细胞存活率,增强caspase-3活性,诱导细胞凋亡,并下调细胞中cyclinD1的表达。Taxol先作用24h再用flavopiridol作用24h的联合用药对二者的上述作用产生显著的协同效应:联合用药后细胞存活率为(5.2±0.8)%,凋亡率为(51.10±2.52)%,caspase-3活性相对值为0.602±0.008,cy-clinD1相对表达水平为0.264±0.076。Flavopiridol和泰素单独应用均能显著抑制皮下移植瘤生长(抑瘤率分别为84.5%和85.7%),并降低肿瘤组织MVD值(分别为12.4±4.7vs 35.2±10.3和15.2±2.9 vs 35.2±10.3)。二者联合应用抑瘤作用稍差(抑瘤率68.9%),抑制MVD作用(23.0±5.1 vs 35.2±10.3)也不及单药治疗。结论:Flavopiridol和泰素均能通过致凋亡作用显著抑制卵巢癌细胞及移植瘤生长,二者联合应用对体外培养的卵巢癌细胞具有协同杀伤作用,但体内治疗中无协同抑瘤作用。
OBJECTIVE: To investigate the therapeutic effect of flavopiridol combined with taxol on human ovarian cancer cell line SKOV3 and human ovarian cancer subcutaneously and intraperitoneally transplanted tumor model in nude mice. Methods: The cell viability and apoptosis rate of ovarian cancer cell line SKOV3 after flavopiridol and paclitaxel treatment were detected by CCK-8, flow cytometry and in situ cell apoptosis assay (TUNEL). Reverse transcription-polymerase chain reaction The expression of cyclinD1 was detected by RT-PCR. The expression of active caspase-3 in cells was detected by ELISA. The subcutaneous and abdominal xenograft models of human ovarian cancer were established. The changes of tumor volume in nude mice before and after treatment with flavopiridol and paclitaxel were observed and the tumor inhibition rates were calculated. TUNEL and immunohistochemical methods were used to detect the apoptosis in tumor tissues And microvessel density (MVD) values. RESULTS: Both flavopiri-dol (300 nmol / L) and paclitaxel (1 μmol / L) alone could significantly decrease cell viability, increase caspase-3 activity, induce apoptosis and down-regulate the expression of cyclinD1 in cells. The synergistic effect of Taxol for 24 h followed by flavopiridol for 24 h resulted in a significant synergistic effect on the above effects: cell survival rate (5.2 ± 0.8)%, apoptosis rate (51.10 ± 2.52)%, caspase -3 activity relative value of 0.602 ± 0.008, cy-clinD1 relative expression level of 0.264 ± 0.076. Flavopiridol and paclitaxel alone significantly inhibited the growth of subcutaneous xenografts (tumor inhibition rates were 84.5% and 85.7%, respectively) and decreased MVD in tumor tissues (12.4 ± 4.7 vs 35.2 ± 10.3 and 15.2 ± 2.9 vs 35.2 ± 10.3). The combination of the two had a poorer anti-tumor effect (inhibition rate of 68.9%) and no effect on MVD (23.0 ± 5.1 vs 35.2 ± 10.3) compared with monotherapy. CONCLUSION: Both Flavopiridol and Taxol can significantly inhibit the growth of ovarian cancer cells and xenografts by inducing apoptosis. The combination of the two drugs has synergistic cytotoxic effect on ovarian cancer cells cultured in vitro. However, there is no synergistic anti-tumor effect in vivo.