Effects of Walnut Green Husk Extract on Proliferation, Apoptosis and EGFR/MAPK Signal Pathway of Breast Cancer MCF-7 Cells

ZHANG Shuowen; LI Dan; HE Jing; DU Zhixing; PANG Jianmin; LI Ruichi; LIU Yongjian; ZHENG Lihua

doi:10.14009/j.issn.1672-2124.2022.06.012

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Effects of Walnut Green Husk Extract on Proliferation, Apoptosis and EGFR/MAPK Signal Pathway of Breast Cancer MCF-7 Cells

更新时间：2022-10-08

- Effects of Walnut Green Husk Extract on Proliferation, Apoptosis and EGFR/MAPK Signal Pathway of Breast Cancer MCF-7 Cells
- Vol. 22, Issue 6, Pages: 684-689(2022)
- 作者机构：
  
  1. 河北医科大学第一医院健康管理中心
  2. 河北医科大学第一医院普外科
- 作者简介：
- 基金信息：
- DOI：10.14009/j.issn.1672-2124.2022.06.012
  CLC： R285
- Published：2022，
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ZHANG Shuowen, LI Dan, HE Jing, et al. Effects of Walnut Green Husk Extract on Proliferation, Apoptosis and EGFR/MAPK Signal Pathway of Breast Cancer MCF-7 Cells. [J]. 22(6):684-689(2022)
DOI：

ZHANG Shuowen, LI Dan, HE Jing, et al. Effects of Walnut Green Husk Extract on Proliferation, Apoptosis and EGFR/MAPK Signal Pathway of Breast Cancer MCF-7 Cells. [J]. 22(6):684-689(2022) DOI： 10.14009/j.issn.1672-2124.2022.06.012.

摘要

目的：探讨核桃青皮提取物对乳腺癌MCF-7细胞增殖、凋亡及表皮生长因子受体(EGFR)/丝裂原活化蛋白激酶(MAPK)信号通路的影响。方法：采用细胞计数试剂盒-8(CCK-8)筛选干预乳腺癌MCF-7细胞的核桃青皮提取物合适浓度。将乳腺癌MCF-7细胞分为空白对照组、阳性对照组、核桃青皮提取物低浓度组、核桃青皮提取物高浓度组和核桃青皮提取物高浓度+EGFR激动剂(NSC228155)组。采用CCK-8和集落形成实验检测各组细胞增殖情况；采用TUNEL法和流式细胞仪检测各组细胞凋亡情况；采用蛋白质印迹法检测各组细胞中增殖、凋亡及EGFR/MAPK信号通路相关蛋白表达水平。结果：当核桃青皮提取物浓度≥20μg/mL时，细胞增殖率显著降低，故选择20、40μg/mL分别为核桃青皮提取物低浓度组、高浓度组的核桃青皮提取物处理浓度。与空白对照组相比，阳性对照组、核桃青皮提取物低浓度组和高浓度组细胞增殖率、细胞集落形成数量，细胞周期蛋白D1(CyclinD1)、增殖细胞核抗原(PCNA)和B细胞淋巴瘤2(Bcl-2)蛋白表达水平，磷酸化EGFR(p-EGFR)/EGFR、磷酸化胞外信号调节激酶(p-ERK)/胞外信号调节激酶(ERK)、磷酸化p38 MAPK(p-p38 MAPK)/p38 MAPK和磷酸化c-Jun氨基端蛋白激酶(p-JNK)/c-Jun氨基端蛋白激酶(JNK)蛋白比值均显著降低；TUNEL阳性细胞率、细胞凋亡率和Bcl-2相关X蛋白(Bax)表达水平均显著升高，差异均有统计学意义(P<0.05)。与核桃青皮提取物低浓度组相比，核桃青皮提取物高浓度组细胞增殖率、细胞集落形成数量，CyclinD1、PCNA和Bcl-2蛋白表达水平，p-EGFR/EGFR、p-ERK/ERK、p-p38 MAPK/p38 MAPK和p-JNK/JNK蛋白比值均降低；TUNEL阳性细胞率、细胞凋亡率和Bax蛋白表达水平均升高，差异均有统计学意义(P<0.05)。核桃青皮提取物高浓度组与阳性对照组上述指标的差异均无统计学意义(P>0.05)。与核桃青皮提取物高浓度组相比，核桃青皮提取物高浓度+NSC228155组细胞增殖率、细胞集落形成数量，CyclinD1、PCNA和Bcl-2蛋白表达水平，p-EGFR/EGFR、p-ERK/ERK、p-p38 MAPK/p38 MAPK和p-JNK/JNK蛋白比值均升高；TUNEL阳性细胞率、细胞凋亡率和Bax蛋白表达水平均降低，差异均有统计学意义(P<0.05)。结论：核桃青皮提取物可能通过抑制EGFR/MAPK信号通路抑制乳腺癌MCF-7细胞增殖，诱导其凋亡。

Abstract

OBJECTIVE: To probe into the effects of walnut green husk extract on proliferation

apoptosis and epidermal growth factor receptor（EGFR）/mitogen activated protein kinase（MAPK） signaling pathway of breast cancer MCF-7 cells. METHODS: Cell counting kit-8（CCK-8） was used to screen the proper concentration of walnut husk extract interfering with breast cancer MCF-7 cells. The breast cancer MCF-7 cells were divided into the blank control group

positive control group

walnut husk husk extract low concentration group

walnut green husk extract high concentration group

walnut green husk extract high concentration + EGFR agonist（NSC228155） group. CCK-8 and colony formation experiment were used to detect cell proliferation in each group. TUNEL method and flow cytometry were used to detect cell apoptosis in each group. Western blot was used to detect the expression levels of proliferation

apoptosis and EGFR/MAPK signaling pathway related proteins of cells in each group. RESULTS: When the concentration of walnut green husk extract was ≥20 μg/mL

the cell proliferation rate decreased significantly

20 μg/mL and 40 μg/mL were selected as the walnut green husk extract low and high concentration treatment

respectively. Compared with the blank control group

the cell proliferation rate

the number of cell colonies

the expression levels of CyclinD1

proliferating cell nuclear antigen（PCNA） and B cell lymphoma-2（Bcl-2） protein

the ratio of phosphorylated-EGFR（p-EGFR）/EGFR

phosphorylated extracellular signal-regulated kinase（p-ERK）/extracellular signal-regulated kinase（ERK）

phosphorylated-p38 MAPK（p-p38 MAPK）/p38 MAPK and phosphorylated c-Jun amino-terminal protein kinase（p-JNK）/c-Jun amino-terminal protein kinase（JNK） protein in the positive control group

the walnut green husk extract low concentration group and high concentration group decreased significantly

and the rate of TUNEL positive cells

the apoptosis rate

the expression level of Bcl-2 related X protein（Bax） increased significantly

with statistically significant differences（P<0.05）. Compared with the walnut green husk extract low concentration group

the cell proliferation rate

the number of cell colonies

the expression levels of CyclinD1

PCNA and Bcl-2 protein

the ratio of p-EGFR/EGFR

p-ERK/ERK

p-p38 MAPK/p38 MAPK and p-JNK/JNK protein in the walnut green husk extract high concentration group decreased

and the rate of TUNEL positive cells

the apoptosis rate

the expression level of Bax protein increased

with statistically significant differences（P<0.05）. There was no significant difference in the above indicators between the walnut green husk extract high concentration group and the positive control group（P>0.05）. Compared with the walnut green husk extract high concentration group

the cell proliferation rate

the number of cell colonies

the expression levels of CyclinD1

PCNA and Bcl-2 protein

the ratio of p-EGFR/EGFR

p-ERK/ERK

p-p38 MAPK/p38 MAPK and p-JNK/JNK protein in the walnut green husk extract high concentration + NSC228155 group increased

and the rate of TUNEL positive cells

the apoptosis rate

the expression level of Bax protein decreased

with statistically significant differences（P<0.05）. CONCLUSIONS: Walnut green husk extract may inhibit the proliferation of breast cancer MCF-7 cells and induce the apoptosis by inhibiting the EGFR/MAPK signaling pathway.

关键词

核桃青皮提取物乳腺癌MCF-7细胞增殖凋亡表皮生长因子受体/丝裂原活化蛋白激酶信号通路

Keywords

Walnut green husk extractBreast cancer MCF-7 cellsProliferationApoptosisEpidermal growth factor receptor/mitogen activated protein kinase signaling pathway

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