Chaetoglobosins E诱导乳腺癌MCF-7细胞凋亡作用机制研究

于大永; 高鸿雁; 曹鹤; 卢轩; 史丽颖

doi:10.6054/j.jscnun.2022010

Chaetoglobosins E诱导乳腺癌MCF-7细胞凋亡作用机制研究

doi: 10.6054/j.jscnun.2022010

大连大学生命科学与技术学院，大连 116622

基金项目:

辽宁省科学技术计划项目 2019-ZD-0564

大连大学优秀青年科研创新创业团队项目 XQN202004

详细信息

通讯作者:
史丽颖, Email: shiliying@dlu.edu.cn

中图分类号: R966
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- 被引次数: 0
出版历程
- 收稿日期: 2021-03-03
- 刊出日期: 2022-02-25

The Effect of Chaetoglobosins E on the Proliferation and Apoptosis of MCF-7 Cells and the Related Mechanisms

The School of Life Science and Technology, Dalian University, Dalian 116622, China

摘要

摘要: 为探究Chaetoglobosins E (ChE)对肿瘤细胞增殖和凋亡的影响，体外培养人乳腺癌MCF-7细胞、人膀胱癌T-24细胞、人黑色素瘤C8161细胞、人白血病U937细胞，用不同浓度的ChE分别作用于4种细胞24 h或48 h，MTT法检测4种肿瘤细胞的增殖情况；为进一步研究其作用机制，Hoechst 33342染色观察MCF-7经ChE处理后细胞形态的变化，流式细胞术检测MCF-7经ChE处理后细胞凋亡、周期、活性氧以及线粒体膜电位的变化情况，Western Blot法检测MCF-7细胞中凋亡相关蛋白的表达情况。结果显示：ChE对MCF-7、T-24、C8161和U937细胞增殖均有抑制作用，且均呈现出时间和剂量依赖性，4种肿瘤细胞中，ChE对MCF-7增殖的抑制效果最强，24、48 h的IC₅₀分别为82.04±7.01、49.87±2.28 μmol/L；Hoechst 33342染色发现，随着ChE浓度的升高，凋亡的MCF-7细胞数逐渐增多，细胞凋亡特征显著，细胞核的体积缩小，细胞核裂解并伴有凋亡小体；通过流式细胞术发现，MCF-7细胞经ChE处理后, 细胞凋亡增加、细胞周期改变、活性氧增加以及线粒体膜电位降低；Western Blot实验发现，Bid、Caspase 3蛋白的表达量降低，Cleaved Caspase 3、Bax蛋白与Bcl-2蛋白表达量的比值增加。综上所述，ChE诱导的MCF-7细胞凋亡与Caspase依赖性线粒体途径有关。
- Chaetoglobosins E /
- MCF-7 /
- 细胞增殖 /
- 凋亡 /
- 线粒体途径
Abstract: To explore the effect of Chaetoglobosins E (ChE) on tumor cell proliferation and apoptosis, human breast cancer MCF-7 cells, human bladder cancer T-24 cells, human melanoma C8161 cells and human leukemia U937 cells were treated with ChE of different concentrations for 24 h or 48 h. The MTT method was used to detect the proliferation of 4 kinds of tumor cells. To further study its mechanism of action, the Hoechst 33342 staining was used to observe the changes in the morphology of MCF-7 cells treated with ChE. Flow cytometry was used to detect the changes of apoptosis, cycle, reactive oxygen species and mitochondrial membrane potential of MCF-7 cells treated with ChE. The Western Blot method was used to detect the expression of apoptosis-related proteins in MCF-7 cells. The results showed that ChE inhibited the proliferation of MCF-7, T-24, C8161 and U937 cells in a time- and dose-dependent manner. Among the four tumor cells, ChE had the strongest inhibitory effect on the proliferation of MCF-7 with IC₅₀ of 82.04±7.01 μmol/L and 49.87±2.28 μmol/L at 24 h and 48 h, respectively. The Hoechst 33342 staining found that the number of apoptotic MCF-7 cells gradually increased, with significant characteristics of cell apoptosis and the volume of the nucleus was reduced, with the nucleus lysed and accompanied by apoptotic bodies with the increase of ChE concentration. The flow cytometry found that, after MCF-7 cells were treated with ChE, the number of apoptosis cells increased, the cell cycle was changed, the reactive oxygen species increased, and the mitochondrial membrane potential decreased. The Western Blot experiment found that the expression of Bid and Caspase 3 protein decreased and cleaved Caspase 3 and the ratio of Bax protein to Bcl-2 protein expression increased. To sum up, ChE-induced apoptosis of MCF-7 cells is related to the Caspase-dependent mitochondrial pathway.
- Chaetoglobosins E /
- MCF-7 /
- cell proliferation /
- apoptosis /
- mitochondrial pathway

HTML全文

图 1 Chaetoglobosins E结构式

Figure 1. The structure of Chaetoglobosins E

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图 2 不同浓度ChE对4种肿瘤细胞24、48 h增殖的影响(x±s, n=3)

注：与空白对照组相比，*表示P < 0.05, **表示P < 0.01，下图同。

Figure 2. The effects of different concentrations of ChE on the 24 h or 48 h proliferation of four kinds of tumor cells(x±s, n=3)

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图 3 不同浓度ChE对MCF-7细胞处理6 h后的细胞形态(×400)

Figure 3. The morphology of MCF-7 cells treated with different concentrations of ChE for 6 h (×400)

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图 4 不同浓度ChE处理MCF-7细胞12 h后的凋亡情况

注：A图中的左下象限代表活细胞，右下象限代表早期凋亡细胞，右上象限代表晚期凋亡细胞与死细胞，左上象限代表误差。

Figure 4. The apoptosis of MCF-7 cells treated with ChE for 12 h

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图 5 不同浓度ChE处理MCF-7细胞4 h后细胞周期的变化

Figure 5. The cell cycle changes of MCF-7 cells after ChE treatment for 4 h

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图 6 不同浓度ChE处理MCF-7细胞2 h后细胞内ROS的变化

注：A图中虚线表示该组细胞平均荧光强度。

Figure 6. The ROS production changes of MCF-7 cells after ChE treatment for 2 h

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图 7 不同浓度ChE处理MCF-7细胞3 h后细胞线粒体膜电位损失率的变化

注：A图中按对照组数据确定好固定荧光强度值后，统计低于该荧光强度的总细胞数所占全部细胞百分比。

Figure 7. The loss of MMP changes of MCF-7 cells after ChE treatment for 3 h

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图 8 不同浓度ChE处理MCF-7细胞6 h后细胞蛋白表达量的变化

Figure 8. The protein expression changes of MCF-7 cells after ChE treatment for 6 h

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表 1 ChE对4种肿瘤细胞的IC₅₀

Table 1. The IC₅₀ of ChE on four kinds of tumor cells μmol/L

处理时间/h	T24	C8161	MCF-7	U937
24	120.70±9.88	99.05±9.97	82.04±7.01	86.64±8.61
48	100.50±9.67	89.63±7.43	49.87±2.28	62.03±4.03

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