醋酸甲羟孕酮增强顺铂对人卵巢癌耐药细胞抗癌作用的研究

Enhancement of the antitumor effect of cisplatin against the drug-resistant human ovarian cancer cells CoC1/cDDP by medroxyprogesterone acetate in vivo.

目的:观察醋酸甲羟孕酮(MPA)对人卵巢癌CoC1/cDDP细胞移植瘤的生长抑制作用及对DDP的耐药逆转作用,并分析其作用机制。方法:建立人卵巢癌裸鼠皮下移植瘤模型,随机分为4组,每组5只。(1)对照组:腹腔注射等体积生理盐水;(2)DDP治疗组:每只每次腹腔注射DDP 3mg/kg。(3)MPA治疗组:每只每次灌胃30mg/kg;(4)联合治疗组:每只每次灌胃MPA 30mg/kg,1h后每只每次腹腔注射DDP 3mg/kg,每隔2天给药1次,共4次,于治疗第1、5、10、15、20天分别测瘤体积,20天后处死裸鼠,完整剥出瘤组织,称瘤重,计算抑瘤率;通过流式细胞仪检测亚G1期细胞及AnnexinV+/PI-细胞鉴定细胞凋亡,分析移植瘤细胞周期;用半定量RT-PCR法检测移植细胞组织Survivin-ΔEx3、caspase-3、P21WAF1/CIP1及GST-π4基因mRNA表达。结果:(1)MPA组、MPA+DDP组治疗第10天起皮下移植瘤体积明显小于DDP组和对照组,且进行性缩小(P<0.01);抑瘤率分别为51.63%、62.21%,均明显大于DDP组的6.84%(P<0.01),并且两组间有显著差异(P<0.01);(2)流式细胞仪分析显示,移植瘤出现亚G1期峰及AnnexinV+/PI-细胞均证实MPA能诱导CoC1/cDDP细胞凋亡,并显著高于对照组及DDP组,并出现G1期阻滞;与DDP合用除出现G1期阻滞外又出现G2/M期阻滞,S期明显减少,亚G1期细胞及AnnexinV+/PI-细胞进一步上升;(3)半定量RT-PCR检测显示,MPA组Survivin-ΔEx3、GST-πmRNA下调,而P21WAF1/CIP1、caspase-3 mRNA上调,与DDP合用后,对Survivin-ΔEx3、P21WAF1/CIP1及GST-πmRNA的表达无协调作用,而对caspase-3 mRNA有协同上调作用。结论:MPA通过阻滞G0/G1期明显抑制了CoC1/cDDP移植瘤生长,并有很强的致凋亡作用,同时下调GST-πmRNA,从而逆转对顺铂耐药。

Objective：To investigate the enhancement of antitumor effect of cisplatin against human ovarian cancer xenografts in nude mice by medroxyprogesterone acetate. Methods： Mouse models bearing xenografted cisplatin-resistant ovarian carcinoma were established, which were then divided equally into four groups consisting of three treatment groups [ a DDP- treated group（ n = 5 ） received 3mg/kg DDP, a MPA-treated group （ n = 5 ） received 30mg/kg MPA via gastrointestinal gavage and a MPA plus DDP-treated group] and a control group. Four groups were treated one time every two days for four times. The tumor volumes were measured every five days. After treatment, the mice were sacrificed. The tumor xenografts were removed and weighed. Apoptosis was confirmed by sub G1 and AnnexinV ＋/PI - cells content , the cells cycle were analyzed with flow cytometry, surviving-△Ex3, caspase-3, P21^ WAF1/CIP1 and GST-π genes were detected by semi-quantitative RT-PCR. Results： （ 1 ） Both MPA plus DDP and MPA had significant inhibition effect on the growth of the xenografted tumors, especially in the former group;（2） The G0/G1 phase fraction of the xenografted tumors was increased and the CoC1/ cDDP cells apoptosis was confirmed by sub-G1 phase and Annexin V/PI;the G2/M phase was increased and S-phase was decreased besides G0/G1 increasion and sub-G1 phase and Annexin V ＋/PI - cell was progressionly increased after treatment of MPA plus DDP ; （3） The expression of surviving-△Ex3 and GST-π mRNA were down-regulated, but the expression of P21^WAF1/CIP1 and caspase-3 mRNA were up-regulated after treatment of MPA. MPA was not in cooperation with DDP in the expression of surviving-△Ex3 ,P21^WAF1/CIP1 and GST-π mRNA,but in coopera- tion with DDP in the expression of caspase-3 mRNA. Conclusion：MPA can significantly suppress the proliferation and reverse the drug-resistance of the CoC1/cDDP of xenografted tumors to cisplatin by regulating the cell cycle, apoptotic and GST-π genes.