摘要
目的死亡效应结构域DNA结合蛋白(death effector domain DNA-binging protein,DEDD)可抑制乳腺癌的生长和转移,但在乳腺癌多药耐药中的作用尚不明确。本研究将探讨DEDD在人乳腺癌多药耐药中的作用及机制。方法通过转染DEDD-shRNA建立稳定敲低DEDD表达的MCF-7人乳腺癌细胞株,MTS实验检测多柔比星、紫杉醇对MCF-7WT(Wild type,野生型)、MCF-7control shRNA和MCF-7DEDD-shRNA细胞活力的影响,Annexin-V和碘化丙啶(propidium iodide,PI)双染后流式细胞仪检测细胞凋亡。蛋白质印迹法检测乳腺癌耐药蛋白(breast cancer resistance protein,BCRP)在药物干预后的蛋白表达水平。结果多柔比星干预后,半数抑制浓度IC50的多因素方差分析结果显示,不同处理(F=89.49,P<0.001)、不同时间(F=50.81,P<0.001)组间比较差异有统计学意义。多柔比星干预24或48h后,MCF-7DEDD-shRNA的半数抑制浓度IC50分别为(0.97±0.15)和(0.62±0.09)μmol/L,明显高于MCF-7 WT(0.46±0.05)和(0.26±0.03)μmol/L和MCF-7control shRNA细胞(0.39±0.05)和(0.25±0.03)μmol/L,P<0.001。紫杉醇干预后,半数抑制浓度IC50的多因素方差分析结果显示,不同处理(F=15.94,P<0.001)、不同时间组间(F=129.70,P<0.001)比较差异有统计学意义。紫杉醇干预24或48h后,MCF-7DEDD-shRNA的半数抑制浓度IC50为(16.74±2.26)和(9.88±1.47)μmol/L,明显高于MCF-7 WT(13.39±1.44)和(7.69±0.92)μmol/L(P=0.001)和MCF-7control shRNA细胞(12.58±1.15)和(7.17±1.12)μmol/L(P<0.001)。多柔比星干预后24h后流式细胞凋亡结果显示,不同处理(F=131.46,P<0.001)、不同浓度组间(F=160.95,P<0.001)比较差异有统计学意义。正常培养状态下,MCF-7control shRNA细胞和MCF-7DEDD-shRNA细胞的凋亡率分别为(14.32±1.47)%和(11.58±1.63)%,而给予0.5μmol/L和1μmol/L多柔比星作用24h后,MCF-7DEDD-shRNA细胞的凋亡率分别为(21.62±1.97)%和(24.39±2.36)%,明显低于MCF-7control shRNA细的(36.26±1.87)%和(38.23±1.46)%,P<0.001。同样,紫杉醇干�
OBJECTIVE Our previous study highlighted that DEDD （death effector domain DNA-binging protein） could inhibit the growth and metastasis of breast cancer. However, the effect of DEDD on multiple drug resistance is still unkown. In this study, we aimed to explore the effects and meschanisms of DEDD on multiple drug resistance of breast cancer. METHODS Stable knockdown DEDD cells were established by transfecting DEDD shRNA into MCF-7 cells. MTS assay was used to evaluate the cell viability of MCF-7 cells （Wild type/WT, MCF-7 control shRNA and DEDD shR- NA） after treated with doxorubicin or paclitaxel for indicated times. Cell apoptosis was measured by flow cytometric anal- ysis after Annexin V/PI double staining. The expressions of BCRP （breast cancer resistance protein） were detected by immunobloting. RESULTS After treatment with doxorubicin, MTS results showed that there were significant differ- ences between the different cells （F=89.49, P（0. 001） or different treatment time （F=50.81, P〈0. 001） groups. Tthe IC±0 values of doxorubicin in MCF7 DEDDshRNA cells were （0.97±0.15） and （0.62±0.09） vmol/L respectively after 24 h or 48 h treatment with doxorubicin, which were significantly higher than that of MCF-7 WT cells [（0.46 ± 0.05） μmol/L, （0.26±.03） μmol/L] and MCF-7 control shRNA cells [（0.39±0.05） μmol/L, （0.25±0.03） μmol/L,P（0. 001]. After treatment with paclitaxel, there were significant differences in IC50 between the different cells （F= 15. 94, P〈 0. 001） or different treatment time （F= 129.70, P（0. 001） groups. The ICs0 valus of paclitaxel in MCF-7 DEDI）shRNA cells were （16.74±2.26） and （9.88±1.47） μmol/L respectively after treatment for 24 h or 48 h, which were signifi- cantly higher than that of MCF-7 WT cells P（13. 39±0. 44 μmol/L, （7.69!0.92） μmol/L,P=0. 001] and MCF-7 con- trol shRNA cells [（12.58±0.15） μmol/L,（7.17±1.12） μmol/L,P（0. 001]. After treatment with doxorubicin, apop- tosis assay showed
出处
《中华肿瘤防治杂志》
CAS
北大核心
2017年第16期1125-1129,1136,共6页
Chinese Journal of Cancer Prevention and Treatment
基金
国家自然科学基金(81202105)
天津市自然科学基金青年项目(13JCQNJC12900)
关键词
DEDD
乳腺癌
多药耐药
乳腺癌耐药蛋白
DEDD
breast cancer
multidrug resistance
breast cancer resistance protein
