论文部分内容阅读
目的研究三氧化二砷(AS_2O_3)对子宫内膜癌细胞和耐药细胞的作用及机制。方法 (1)分别采用MTS法和Annexin V-FITC/PI双染色流式细胞术检测不同浓度的AS_2O_3在不同作用时间下对ISK细胞增殖和凋亡的影响;(2)采用浓度梯度递增持续刺激诱导法,体外建立MPA耐药细胞后,采用MTS法和Annexin V-FITC/PI双染色流式细胞术,检测AS_2O_3对ISK/MPA增殖和凋亡的影响,并与ISK组对比。(3)Western-blot法检测AS_2O_3作用后ISK与ISK/MPA细胞内p-AKT、p-ERK1/2及Caspase-3、Bcl-2和Bax蛋白的表达变化。(4)建立裸鼠皮下移植瘤模型,腹腔注射2 mg/kg的AS_2O_3,观察裸鼠瘤体的体积变化及毒副作用。结果 (1)AS_2O_3对ISK细胞有生长抑制作用,且呈时间和浓度依赖性;AS_2O_3作用后,ISK的凋亡率呈浓度依赖性升高,48 h细胞凋亡率大于24 h(P<0.05);(2)成功建立人子宫内膜癌MPA耐药细胞系;AS_2O_3对ISK/MPA细胞具有生长抑制和促凋亡作用,且呈时间和浓度依赖性;AS_2O_3对ISK细胞组的促凋亡作用强于ISK/MPA细胞组(P<0.05),但生长抑制作用比较,差异无统计学意义(P>0.05);(3)AS_2O_3作用后,ISK及ISK/MPA细胞内p-AKT、p-ERK1/2和Caspase-3表达降低,Bcl-2表达降低而Bax表达升高(P<0.05);(4)成功建立裸鼠皮下移植瘤模型,AS_2O_3作用后,裸鼠瘤体体积明显减小(P<0.05)。结论 AS_2O_3对ISK/MPA细胞有增殖抑制和促凋亡作用,机制可能为AS_2O_3可导致Akt、ERK1/2磷酸化水平的降低,抑制P13K/AKT通路和MPAK/ERK通路的激活及在下调Bcl-2表达的同时,上调Bax蛋白的表达,继而调节凋亡相关蛋白通路分子Caspase-3的表达发挥增殖抑制和促凋亡作用。
Objective To study the effect and mechanism of arsenic trioxide (AS_2O_3) on endometrial carcinoma cells and drug-resistant cells. Methods (1) MTS and Annexin V-FITC / PI double staining were used to detect the effects of different concentrations of AS_2O_3 on the proliferation and apoptosis of ISK cells at different time points. (2) After establishment of MPA resistant cells in vitro, MTS and Annexin V-FITC / PI double staining were used to detect the effect of AS_2O_3 on the proliferation and apoptosis of ISK / MPA cells and compared with ISK group. (3) The expressions of p-AKT, p-ERK1 / 2, Caspase-3, Bcl-2 and Bax in ISK and ISK / MPA cells were detected by Western-blot. (4) A subcutaneous xenograft tumor model was established in nude mice. Intraperitoneal injection of 2 mg / kg AS_2O_3 was used to observe the volume changes and side effects of the tumor in nude mice. RESULTS: AS_2O_3 inhibited the growth of ISK cells in a time and concentration-dependent manner. The apoptosis rate of ISK cells increased in a concentration-dependent manner after AS_2O_3 treatment for 48 h, and the apoptosis rate was greater than 24 h (P <0.05) ); (2) MPA resistant cell line of human endometrial cancer was established successfully; AS_2O_3 could inhibit growth and induce apoptosis of ISK / MPA cells in a time and concentration dependent manner; AS_2O_3 promoted the apoptosis of ISK cells (P <0.05). (3) After AS_2O_3 treatment, the expression of p-AKT and p-AKT in ISK and ISK / MPA cells was significantly higher than that in ISK / MPA cells The expression of ERK1 / 2 and Caspase-3 decreased, the expression of Bcl-2 decreased and Bax increased (P <0.05). (4) The model of subcutaneous xenograft in nude mice was successfully established. Small (P <0.05). Conclusion AS_2O_3 can inhibit the proliferation and induce the apoptosis of ISK / MPA cells. The possible mechanism is that AS_2O_3 can reduce the phosphorylation of Akt and ERK1 / 2, inhibit the activation of P13K / AKT pathway and MPAK / ERK pathway and decrease the expression of Bcl- 2 expression at the same time, up-regulate the expression of Bax protein, which in turn regulates apoptosis-related protein pathway expression of Caspase-3 to play a role in proliferation inhibition and apoptosis.