天津医药

天津医药 ›› 2021, Vol. 49 ›› Issue (10): 1014-1019.doi: 10.11958/20210189

• 细胞与分子生物学 • 上一篇    下一篇

双黄连冻干粉通过抑制MEK-ERK信号通路诱导急性淋巴细胞白血病Nalm6细胞凋亡

杨友，钟芳芳，黄喆，覃祥，马文哲，刘文君   

  1. 1西南医科大学附属医院儿科（邮编646000）；2四川省出生缺陷临床医学研究中心；3澳门科技大学中药质量研究国家重点实 验室
  • 收稿日期:2021-01-25 修回日期:2021-07-20 出版日期:2021-10-15 发布日期:2021-10-15
  • 通讯作者: 刘文君 E-mail:lwjlyfy@qq.com
  • 基金资助:
    四川省应用基础研究项目;四川省科技厅重点研发项目;西南医科大学;西南医科大学附属医院;泸州市科技厅

Shuanghuanglian freeze-dried powder induced apoptosis of acute B cell lymphocytic leukemia cells (Nalm6) through inhibiting the MEK-ERK signaling pathway

YANG You, ZHONG Fang-fang, HUANG Zhe, QIN Xiang, MA Wen-zhe, LIU Wen-jun #br#   

  1. 1 Department of Pediatrics, the Affiliated Hospital of Southwest Medical University, Luzhou 646000, China; 2 Sichuan Clinical
    Research Center for Birth Defects; 3 State Key Laboratory of Quality Research in Chinese Medicine,
    Macau University of Science and Technology
  • Received:2021-01-25 Revised:2021-07-20 Published:2021-10-15 Online:2021-10-15
  • Contact: Wen-Jun Liu E-mail:lwjlyfy@qq.com

PDF (PC)

2987

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SHL 0.1、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，tBid、cleaved Caspase-9、cleaved Caspase-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。

关键词: 白血病, 淋巴样, 白血病, 髓样, 细胞系, 肿瘤, 双黄连注射剂, 细胞凋亡, MAP激酶信号系统, Nalm6细胞

Abstract: 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 KG1a) 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 IC 50 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.

Key words: leukemia, lymphoid, leukemia, myeloid, cell line, tumor, Shuanghuanglian injection, apoptosis, MAP kinase signaling system, Nalm6 cell