柔红霉素细胞毒作用对急性淋巴细胞白血病细胞内活性氧水平的影响

Effects of cellular cytotoxicity of daunorubicin on reactive oxygen species levels of acute lymphocyte leukemic cells

目的探讨柔红霉素(DNR)细胞毒作用对儿童急性淋巴细胞白血病(ALL)细胞内活性氧(ROS)水平的影响。方法用Ficoll液从11例ALL患儿骨髓中分离出ALL细胞,用双氢-乙酰乙酸二氯荧光黄(DCFH-DA)作为ROS捕获剂,经流式细胞术检测细胞的固有ROS水平以及DNR50、100、500ng/mL诱导24h后的ROS水平,每次以U937细胞的ROS值作对照。采用MTT法测定各组诱导48h后的细胞相对活力,流式细胞术和Hoechst33342-PI染色法观察细胞凋亡。结果11例患儿的ALL细胞内固有ROS水平与DNRIC50呈负相关(r=-0.700,P=0.016)。ALL患儿细胞内固有ROS水平存在统计学差异(P<0.05)。对照组固有ROS水平为0.260±0.118,DNR50、100、500ng/mL诱导后细胞内ROS水平为0.291±0.132、0.285±0.125和0.257±0.099,DNR500ng/mL组明显低于DNR100ng/mL组(P<0.05)。各浓度DNR诱导后均见典型的凋亡细胞。结论ALL患儿细胞内固有ROS水平存在差异,与DNR细胞毒性正相关。DNR诱导ALL原代细胞后出现ROS水平改变,可能参与细胞的凋亡。

Objective To determine the effects of cellular cytotoxicity of daunorubicin （DNR） on the reactive oxygen species （ROS） levels of acute lymphocyte leukemia （ALL） cells. Methods ALL cells were isolated from bone marrow of 11 children with ALL using Ficoll liquid. With 2＇-7＇-dichlorodihydrofluorescein diacetate （DCFH-DA） as capture agent, ROS basal levels and ROS levels after treatment of ALL cells with DNR （50, 100 and 500 ng/mL）for 24 h were detected by flow cytometry. Relative ROS levels of U937 cells were calculated and served as controls. Cell viabilities were determined by MTT assay after incubation for 48 h, and cell apoptosis was detected by flow cytometry and Hoechst 33342/PI staining. Results There was a negative correlation between basal ROS levels and IC50 values of DNR in these 11 patients with ALL （r = -0. 700, P =0. 016）. There were significant differences in basal ROS levels among patients with ALL（ P 〈0.05）. The basal ROS level of control group was 0. 260 ±0. 118, and the ROS levels after induction by DNR 50, 100 and 500 ng/mL were 0. 291 ±0. 132, 0.285 ±0. 125 and 0. 257 ±0. 099, respectively（P 〈0.05） , with that of DNR 500 ng/mL group significantly lower than that of DNR 100 ng/mL group （ P 〈 0.05）. Typical apoptotic cells were observed after induction by different concentrations of DNR. Conclusion There are differences in the basal ROS levels of ALL cells in patients with ALL, and are positively correlated with DNR cellular cytotoxicity. ROS levels may change after induction by DNR, which may participate in the apoptosis of ALL cells.