Qi Jun, Zhang Mei, He Peng-cheng
Department of Hematology, First Clinical Hospital of Xi' an, Jiaotong University, Xi' an.
Zhongguo Zhong Xi Yi Jie He Za Zhi. 2011 Mar;31(3):391-6.
To establish the comparative proteomic profiles of retinoid acid (RA) resistant human acute promyelocytic leukemia (APL) NB4-R1 cells before and after apoptosis induced by realgar (tetra-arsenic tetra-sulfide, As4S4).
First a serial of assays were performed using MTT, transmission electron microscopy, Annexin V FITC/PI double-stain, flow cytometry and confocal laser scanning microscopy to qualitatively and quantitatively observe the in vitro apoptosis inducing effect of realgar on RA-resistant cells. Then the comparative proteomic profile before and after NB4-R1 apoptosis was established using high-resolution two-dimensional electrophoresis system.
The inhibition effect of realgar on NB4-R1 cell growth was dose and time dependent. The 24-h 50% inhibiting concentration (IC50) was 24.06 +/- 0.19 micromol/L, and the 48-h IC50 9.50 +/- 0.13 micromol/L, and 72-h IC50 6.55 +/- 0.03 micromol/L, respectively. 24 h and 48 h were the early and late phase of major NB4-R1 apoptotic cell populations induced by 25 micromol/L realgar respectively. Differential proteomic profiles before and after realgar induced NB4-R1 apoptosis were successfully established. Averagely 1069, 975 and 893 spots could be detected of the untreated group (R0), the 24-h treatment group (R24), and the 48-h treatment group (R48), respectively by ImageMaster 2D Platinum Software. The matching rate between R24 and R0 was 79.94% and that between R48 and R0 69.33%, and that between R24 and R48 71.91%.
Differential proteomic profiles of realgar induced NB4-R1 apoptosis were successfully established for the first time, which provided a basis for comprehensively understanding the signal transduction of realgar induced apoptosis in RA-resistant APL cells, also for screening new bio-markers and drug targets of hematopoietic malignant tumor.
建立维甲酸（RA）耐药的人急性早幼粒细胞白血病（APL）NB4-R1细胞在雄黄（四硫化四砷，As4S4）诱导凋亡前后的比较蛋白质组图谱。
首先采用MTT、透射电子显微镜、Annexin V FITC/PI双染、流式细胞术和共聚焦激光扫描显微镜等一系列实验，定性和定量观察雄黄对RA耐药细胞的体外凋亡诱导作用。然后利用高分辨率二维电泳系统建立NB4-R1细胞凋亡前后的比较蛋白质组图谱。
雄黄对NB4-R1细胞生长的抑制作用呈剂量和时间依赖性。24小时的50%抑制浓度（IC50）为24.06±0.19微摩尔/升，48小时的IC50为9.50±0.13微摩尔/升，72小时的IC50为6.55±0.03微摩尔/升。24小时和48小时分别是25微摩尔/升雄黄诱导NB4-R1细胞凋亡的主要细胞群体的早期和晚期。成功建立了雄黄诱导NB4-R1细胞凋亡前后的差异蛋白质组图谱。通过ImageMaster 2D Platinum软件，未处理组（R0）、24小时处理组（R24）和48小时处理组（R48）分别平均可检测到1069、975和893个斑点。R24与R0之间的匹配率为79.94%，R48与R0之间的匹配率为69.33%，R24与R48之间的匹配率为71.91%。
首次成功建立了雄黄诱导NB4-R1细胞凋亡的差异蛋白质组图谱，为全面了解雄黄诱导RA耐药APL细胞凋亡的信号转导，以及筛选造血恶性肿瘤的新生物标志物和药物靶点提供了依据。
