摘要
目的探讨双黄连(SHL)冻干粉诱导急性B淋巴细胞白血病细胞(Nalm6)凋亡的机制。方法采用0、0.025、0.05、0.1、0.2、0.4、0.8 g/L的SHL处理急性白血病细胞株(Nalm6、Jurkat、Molt4、KG1a),CCK-8法检测细胞增殖抑制率,并计算半数抑制浓度(IC50)。Nalm6细胞分为空白对照组(含有10%胎牛血清的RPMI 1640培养基)和SHL0.1、0.2、0.4 g/L组。流式细胞术检测各组细胞周期分布及凋亡情况,Hoechst 33342染色细胞后观察细胞凋亡形态学改变。蛋白免疫印迹法检测各组Nalm6细胞MEK-ERK-c-Myc信号通路蛋白以及凋亡相关蛋白的表达情况。结果SHL对Nalm6的IC50最低,为(0.11±0.01)g/L。细胞周期分析显示,0.1~0.4 g/L SHL处理后,Nalm6的细胞周期分布无明显变化,但随着处理浓度的升高,Nalm6凋亡细胞数增多,总凋亡率升高,t Bid、cleaved Caspase-9、cleavedCaspase-3、cleaved PARP表达升高,MEK/ERK通路相关蛋白p-MEK/MEK、p-ERK/ERK、c-Myc表达下降(均P<0.05)。经凋亡抑制剂Z-VAD-FMK预处理后,SHL对Nalm6细胞的促凋亡作用被抑制。结论SHL可通过抑制MEK-ERK信号通路来诱导Nalm6细胞凋亡。
Objective To explore the mechanism of Shuanghuanglian(SHL)freeze-dried powder induced apoptosis of acute B lymphocytic leukemia cells(Nalm6).Methods SHL was used to treat acute lymphoblastic leukemia cell lines(Nalm6,Jurkat,Molt4 and KGla)at 0,0.025,0.05,0.1,0.2,0.4 and 0.8 g/L.The cell proliferation activity-was detected by the CCK-8 method and the half inhibitory concentration(IC50)was calculated.Nalm6 cells were divided into the blank control group(RPMI 1640 medium containing 10%fetal bovine serum)and the SHL treatment groups(0.1,0.2,and 0.4 g/L).Flow cytometry was used to detect cell cycle distribution and apoptosis in each group.The morphological changes of cell apoptosis were observed after Hoechst 33342 staining.Western blot assay was used to detect the expression levels of MEKERK-c-Myc signaling pathway proteins and apoptosis proteins in Nalm6 cells of each group.Results SHL had the lowest IC50 for Nalm6,which was(0.11±0.01)g/L.After treatment with 0.1-0.4 g/L SHL,flow cytometry analysis showed that the cell cycle distribution of Nalm6 cells had no obvious changes.However,with the SHL concentration increasing,the apoptotic cell number and the total apoptotic rate of Nalm6 increased.In addition,the expression levels of tBid,cleaved Caspase-9,cleaved Caspase-3 and cleaved PARP increased.While the expression levels of MEK/ERK pathway related proteins decreased,including p-MEK/MEK,p-ERK/ERK and c-Myc(all P<0.05).After pretreatment with the apoptosis inhibitor Z-VAD-FMK,the pro-apoptotic effect of SHL on Nalm6 cells was inhibited.Conclusion SHL can induce Nalm6 cell apoptosis by inhibiting the MEK-ERK signaling pathway.
作者
杨友
钟芳芳
黄喆
覃祥
马文哲
刘文君
YANG You;ZHONG Fang-fang;HUANG Zhe;QIN Xiang;MA Wen-zhe;LIU Wen-jun(Department of Pediatrics,the Affiliated Hospital of Southwest Medical University,Luzhou 646000,China;Sichuan Clinical Research Center for Birth Defects;State Key Laboratory of Quality Research in Chinese Medicine,Macao University of Science and Technology)
出处
《天津医药》
CAS
北大核心
2021年第10期1014-1019,共6页
Tianjin Medical Journal
基金
四川省应用基础研究项目(2019YJ0690)
四川省科技厅重点研发项目(2019YFS0531)
北京医卫健康公益基金会(YWJKJJHKYJJ-B20010cs)
核医学与分子影像四川省重点实验室(HYX18004)。
