参术抗白血病细胞作用的研究

目的 :探讨参术对白血病细胞有无杀伤或诱导凋亡及分化作用 ,从细胞和分子水平探讨其作用机理 ,为寻找一种选择性杀伤白血病细胞、高效低毒的诱导凋亡及分化的中药方剂提供实验依据。方法 :a.通过MTT试验、台盼蓝拒染检测活细胞数 ,观察不同浓度参术作用后 HL-60细胞增殖的改变 ;b.通过细胞形态学、流式细胞术、DNA琼脂糖凝胶电泳进行细胞凋亡的检测和观察 ;c.以药物维甲酸为阳性对照组 ,采用6.2 5 mg/m L浓度参术作用 HL-60细胞 72 h,通过细胞形态学观察、硝基四氮唑蓝 ( NBT)还原实验、细胞吞墨实验来观察参术作用后 HL-60细胞分化情况。结果 :a.在 6.2 5～ 5 0 mg/m L浓度范围内 ,参术对 HL-60细胞的增殖有抑制作用并呈剂量、时间依赖关系 ,半数抑制浓度约为 2 5 .0 mg/m L;b.典型的细胞形态学改变、DNA琼脂糖凝胶电泳条形梯带的出现和流式细胞仪检测亚 G1 峰的检出等证实参术能诱导白血病细胞凋亡 ;c.NBT还原实验和细胞吞墨实验 :在参术 (浓度为 6.2 5 mg/m L)作用 72 h后 ,NBT阳性反应率和细胞吞墨率分别为5 0 .8%和 5 6.0 % ,明显高于对照组 (分别为 7.8%和 8.0 % ) P <0 .0 5。全反式维甲酸 ( 1 0 - 6 mol/L)作用组 NBT阳性反应率和细胞吞墨率分别为 5 5 .3 %和 69.0 % ,稍高于参术作用组 ,但无

THE STUDY ON SHENZHU ANTI-LEUKEMIA CELL ACTION

Objective: To give the experimental evidence for a high efficiency-low toxicity traditional Chinese medicine which possess the characteristic of selectively killing the leukemia cells. To examine Shenzhu's functional mechanism at both cellular level and molecular level, the in vitro experiment is designed to test whether Shenzhu can kill leukemia cells, induce apoptosis and differentiation of leukemia cells. Methods: (1)MTT assay and trypan blue resisting staining method are used to find out Shenzhu's inhibition on HL-60 cell (a human leukemia cell line). (2)Morphological analysis, flow cytometry (FCM) assay and electrophoresis assay are used to examine cell apoptosis. (3)Using retinoic acid as the positive control, morphological analysis, NBT reduction assay and ink phagocytizing assay are used to examine HL-60 cell differentiation after treating HL-60 cell with Shenzhu(6.25 mg/mL) for 72 hours. Results: (1)HL-60 cells' proliferation was inhibited by Shenzhu concentrations from 6.25 mg/mL to 50 mg/mL. This effect shows dosage and time-dependent relation. IC 50 is 25.0 mg/mL. (2)Typical morphological changes were found in morphological analysis; the sub-G 1 peak was found in FCM analysis and the typical ladder was found in DNA agarose gel electrophoresis analysis. These results showed Shenzhu's definite effect on HL-60 cells' apoptosis. (3)In NBT reduction assay and ink phagocytizing assay, after treating HL-60 cell with Shenzhu(6.25 mg/mL) for 72 hours. The NBT positive reacting rate and in phagocytizing rate were 50.8% and 56.0% respectively, which were much higher than the negative control group, and statistical analysis showed significant difference between the two groups. The rates of the two assays in the retinoic acid group(the positive control group) is 55.3% and 69.0%, which were slightly higher than the Shenzhu group, but statistical analysis showed no significant difference between them. Conclusion: Shenzhu can inhibit HL-60 cells' proliferation and this effect shows dosage and time-dependent relation. Shenzhu can induce apoptosis and it can interfere the cell-cycle phase in HL-60 cell and cause cell remain in G 1 phase. Shenzhu can induce HL-60 cells' differentiation, and the effect is similar with retinoic acid.