Qu Ping, Zhou Fen, Tan Ling-Fang, Wang Zhong-Jian, Wang Mei-Li, Jin Run-Ming, Han Juan
Department of Pediatrics, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, Hubei Province, China.
Department of Pediatrics, Union Hospital, Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430022, Hubei Province, China,E-mail:
Zhongguo Shi Yan Xue Ye Xue Za Zhi. 2020 Oct;28(5):1451-1458. doi: 10.19746/j.cnki.issn.1009-2137.2020.05.004.
To investigate the proliferation inhibition and pro-apoptotic effect of Huaier aqueous extract combined with routine chemotherapeutic drugs including Vincristine (VCR), Daunorubicin (DNR), L-aspartase (L-Asp) on human acute lymphoblastic leukemia cell lines Nalm-6 and Sup-B15.
Nalm-6 and Sup-B15 cell lines were treated with different concentrations of Huaier aqueous extract and chemotherapeutics including VCR, DNR, L-Asp alone or in combination for 48 h, and the growth inhibitory effect and IC values (the half maximal inhibitory concentration) were detected by CCK-8. Jin's formula was used to estimated the synergistic effect of these combinations. Apoptosis rates of Nalm-6 and Sup-B15 cells and expression of apoptosis-related proteins BAX, BCL-2, cleaved Caspase-3 were determined by flow cytometry and Western blot respectivcly.
Huaier aqueous extract, VCR, DNR and L-Asp had inhibition effect on Nalm-6 and Sup-B15 cell lines. The inhibition rate of Huaier aqueous extract combined with VCR, DNR and L-Asp were all higher than those of each dug alone (P＜0.05) and the combination index (q) was between 0.85 and 1.15 or greater than 1.15. The two kinds of drugs showed had additive or synergistic effects. The results of flow cytometry showed that the cell apoptosis rates in combined treatment group were higher than those of each drug alone (P＜0.05). The results of Western blot revealed that Huaier aqueous extract and VCR all decreased protein expression of BCL-2 (P＜0.05) and increase protein expression of BAX (P＜0.05) and cleaved Caspase-3 (P＜0.05) in Nalm-6 and Sup-B15 cells. Compared with Huaier aqueous extract or VCR alone, the effect of two drug combination were more significant. DNR down-regulated protein expression of BCL-2 (P＜0.05) and up-regulated cleaved Caspase-3 (P＜0.05). However, it had no effect on the expression of BAX in Nalm-6 and Sup-B15 cells. When it was combined with Huaier aqueous extract, the expression of cleaved Caspase-3 and BCL-2 showed more significant changes. The expression of BAX in combined treated group did not show significant difference, compared with group treated with Huaier aqueous extract in Nalm-6 and Sup-B15 cells. L-Asp did not show significant effect on the three apoptosis-related proteins and there was no significant difference between the combination group and the Huaier aqueous extract group.
the combination of Huaier aqueous extract and VCR, DNR, L-Asp shows additive or synergistic effects on human acute lymphoblastic leukemia cell lines Nalm-6 and Sup-B15.
探讨槐耳水提取物联合长春新碱（VCR）、柔红霉素（DNR）、L-天冬酰胺酶（L-Asp）等常规化疗药物对人急性淋巴细胞白血病细胞系Nalm-6和Sup-B15的增殖抑制及促凋亡作用。
用不同浓度的槐耳水提取物及化疗药物VCR、DNR、L-Asp单独或联合处理Nalm-6和Sup-B15细胞系48 h，采用CCK-8法检测其生长抑制作用及IC值（半数最大抑制浓度）。用金氏公式计算这些联合用药的协同效应。分别采用流式细胞术和蛋白质免疫印迹法检测Nalm-6和Sup-Bl5细胞的凋亡率及凋亡相关蛋白BAX、BCL-2、裂解型Caspase-3的表达。
槐耳水提取物、VCR、DNR和L-Asp对Nalm-6和Sup-B15细胞系均有抑制作用。槐耳水提取物联合VCR、DNR和L-Asp的抑制率均高于各单药组（P＜0.05），联合指数（q）在0.85至1.15之间或大于1.15。两种药物联合显示有相加或协同作用。流式细胞术结果显示，联合治疗组细胞凋亡率高于各单药组（P＜0.05）。蛋白质免疫印迹结果显示，槐耳水提取物和VCR均可降低Nalm-6和Sup-B15细胞中BCL-2蛋白表达（P＜0.05），增加BAX蛋白表达（P＜0.05）和裂解型Caspase-3蛋白表达（P＜0.05）。与槐耳水提取物或VCR单药组相比，两药联合效果更显著。DNR下调BCL-2蛋白表达（P＜0.05），上调裂解型Caspase-3蛋白表达（P＜0.05）。但对Nalm-6和Sup-B15细胞中BAX表达无影响。与槐耳水提取物联合时，裂解型Caspase-3和BCL-2的表达变化更显著。在Nalm-6和Sup-B15细胞中，联合治疗组BAX表达与槐耳水提取物组相比无显著差异。L-Asp对三种凋亡相关蛋白无显著影响，联合组与槐耳水提取物组无显著差异。
槐耳水提取物联合VCR、DNR、L-Asp对人急性淋巴细胞白血病细胞系Nalm-6和Sup-B15显示相加或协同作用。
