CAG方案清除T细胞急性淋巴细胞白血病细胞株A3细胞作用机制的研究

Mechanism of CAG Regimen Eliminating T-cell Acute Lymphoblastic Leukemia A3 Cell Line

本研究主要探讨CAG方案清除T细胞性急性淋巴细胞白血病(T-ALL)细胞株A3细胞的作用机制及G-CSF/G-CSFR配体受体系统在清除过程中的作用。以单克隆抗体结合流式细胞术测定A3细胞表面G-CSFR表达率;以A3细胞为体外实验模型,分组加入不同浓度的G-CSF作用48小时后收集细胞,碘化丙锭细胞核染色法分析细胞周期变化;用Cell Counting Kit(CCK-8)检测不同浓度阿糖胞苷(Ara-C)和G-CSF组合作用48小时后对细胞生长抑制率的影响;用AnnexinV试剂盒结合流式细胞术检测Ara-C、阿克拉霉素(ACR)及G-CSF联合作用48小时后细胞凋亡水平。结果表明:T-ALL细胞株A3细胞表面高表达G-CSFR(94.2%);G-CSF作用48小时后S期A3细胞比例在一定浓度范围内随G-CSF浓度的增加而升高;CCK-8检测显示Ara-C+G-CSF组较单药Ara-C组抑制细胞生长更明显(Ara-C浓度10-5mmol/L和10-6mmol/L时p<0.05);凋亡检测显示Ara-C+ACR+G-CSF组细胞早期凋亡率高达38%,显著高于Ara-C+ACR组。结论:T-ALL细胞株A3细胞表面高表达G-CSFR;G-CSF/G-CSFR配体受体系统在CAG方案清除A3细胞的过程中与化疗药物具有协同作用,即通过动员G0期细胞进入细胞增殖周期使细胞对化疗药物的敏感性增强、导致细胞凋亡,从而清除细胞。诱导凋亡是CAG方案清除T-ALL细胞株A3细胞的机制之一。

The purpose of this study was to explore the mechanism of CAG regimen eliminating T-cell acute lymphoblastic leukemia （T-ALL） A3 cell line and evaluate the role of G-CSF/G-CSFR system in this process. The expression levels of G-CSFR on the A3 cell membrane were detected by flow cytometry. Cell cycle changes of A3 cells treated with different concentrations of G-CSF for 48 hours were examined by propidium iodide staining method. The inhibition and apoptosis rates of A3 cells treated with various combinations of G-CSF, cytarabine （ Ara-C）, aclarubicin （ACR） were analyzed by Cell Counting Kit and AnnexinV Kit, respectively. The results indicated that the expression level of G-CSFR on A3 cells was 94.2%. The proportion of A3 cells in S-phase rised concomitantly with the increasing of G-CSF concentrations within 0 - 20 ng/mL After incubation with Ara-C and G-CSF for 48 hours, A3 ceils were inhibited more obviously compared with incubation with Ara-C alone （p 〈 0.05, Ara-C 10^- 5 mol/L and 10^- 6 mol/L）. After incubation with Ara-C, ACR and G-CSF for 48 hours, the apoptotic rate of A3 cells was much more than that after incubation with Ara-C and ACR. It is concluded that the expression level of G-CSFR on A3 cells is high, G-CSF/G- CSFR system has a synergetic effect on eliminating A3 cells when administrated simultaneously with chemical agents. This effect is caused through driving more cells from GO phase into S phase, increasing sensitivity of A3 cells to chemical drugs and inducing cell apoptosis which may be one of the mechanisms of CAG regimen eliminating A3 cells.