【摘要】：目的研究三氧化二砷(AS_2O_3)對子宮內(nèi)膜癌細(xì)胞和耐藥細(xì)胞的作用及機(jī)制。方法 (1)分別采用MTS法和Annexin V-FITC/PI雙染色流式細(xì)胞術(shù)檢測不同濃度的AS_2O_3在不同作用時(shí)間下對ISK細(xì)胞增殖和凋亡的影響;(2)采用濃度梯度遞增持續(xù)刺激誘導(dǎo)法,體外建立MPA耐藥細(xì)胞后,采用MTS法和Annexin V-FITC/PI雙染色流式細(xì)胞術(shù),檢測AS_2O_3對ISK/MPA增殖和凋亡的影響,并與ISK組對比。(3)Western-blot法檢測AS_2O_3作用后ISK與ISK/MPA細(xì)胞內(nèi)p-AKT、p-ERK1/2及Caspase-3、Bcl-2和Bax蛋白的表達(dá)變化。(4)建立裸鼠皮下移植瘤模型,腹腔注射2 mg/kg的AS_2O_3,觀察裸鼠瘤體的體積變化及毒副作用。結(jié)果 (1)AS_2O_3對ISK細(xì)胞有生長抑制作用,且呈時(shí)間和濃度依賴性;AS_2O_3作用后,ISK的凋亡率呈濃度依賴性升高,48 h細(xì)胞凋亡率大于24 h(P0.05);(2)成功建立人子宮內(nèi)膜癌MPA耐藥細(xì)胞系;AS_2O_3對ISK/MPA細(xì)胞具有生長抑制和促凋亡作用,且呈時(shí)間和濃度依賴性;AS_2O_3對ISK細(xì)胞組的促凋亡作用強(qiáng)于ISK/MPA細(xì)胞組(P0.05),但生長抑制作用比較,差異無統(tǒng)計(jì)學(xué)意義(P0.05);(3)AS_2O_3作用后,ISK及ISK/MPA細(xì)胞內(nèi)p-AKT、p-ERK1/2和Caspase-3表達(dá)降低,Bcl-2表達(dá)降低而Bax表達(dá)升高(P0.05);(4)成功建立裸鼠皮下移植瘤模型,AS_2O_3作用后,裸鼠瘤體體積明顯減小(P0.05)。結(jié)論 AS_2O_3對ISK/MPA細(xì)胞有增殖抑制和促凋亡作用,機(jī)制可能為AS_2O_3可導(dǎo)致Akt、ERK1/2磷酸化水平的降低,抑制P13K/AKT通路和MPAK/ERK通路的激活及在下調(diào)Bcl-2表達(dá)的同時(shí),上調(diào)Bax蛋白的表達(dá),繼而調(diào)節(jié)凋亡相關(guān)蛋白通路分子Caspase-3的表達(dá)發(fā)揮增殖抑制和促凋亡作用。
[Abstract]:Objective to study the effect and mechanism of arsenic trioxide (AS_2O_3) on endometrial cancer cells and drug-resistant cells. Methods (1) the effects of different concentrations of AS_2O_3 on the proliferation and apoptosis of ISK cells were detected by MS assay and Annexin V-FITC/PI double staining flow cytometry. (2) MPA resistant cells were established in vitro by continuous stimulation with increasing concentration gradient. The effects of AS_2O_3 on the proliferation and apoptosis of ISK/MPA were detected by MS method and Annexin V-FITC/PI double staining flow cytometry, and compared with those in ISK group. (3) the effects of ISK and ISK/MPA cells treated with AS_2O_3 were detected by Western-blot assay. (3) ISK and ISK/MPA cells were treated with AS_2O_3. The expression of Bcl-2 and Bax protein was changed. (4) the subcutaneous tumor model of nude mice was established. 2 mg/kg AS_2O_3, was injected intraperitoneally to observe the volume changes and toxic and side effects of the tumors in nude mice. Results (1) AS_2O_3 could inhibit the growth of ISK cells in a time-and concentration-dependent manner, and the apoptosis rate of ISK increased in a concentration-dependent manner, and the apoptosis rate of MPA cells was more than 24 h at 48 h (P 0.05); (2). AS_2O_3 could inhibit the growth and promote apoptosis of ISK/MPA cells in a time-and concentration-dependent manner. The effect of AS_2O_3 on apoptosis in ISK cell group was stronger than that in ISK/MPA cell group (P 0.05), but there was no significant difference in growth inhibition effect between ISK and ISK/MPA cells treated with AS_2O_3 (P < 0.05). The expression of p 鈮，

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