澳洲茄碱通过调控Bcl-2/Bax/caspase-3信号通路促进非小细胞肺癌发生凋亡

doi:10.12122/j.issn.1673-4254.2024.06.11

南方医科大学学报 ›› 2024, Vol. 44 ›› Issue (6): 1109-1116.doi: 10.12122/j.issn.1673-4254.2024.06.11

• • 上一篇

澳洲茄碱通过调控Bcl-2/Bax/caspase-3信号通路促进非小细胞肺癌发生凋亡

陈桂玲¹(), 廖晓凤¹, 孙鹏涛¹, 岑欢¹, 舒盛春¹, 李碧晶¹, 黎金华¹^,²^,³()

^1.广州中医药大学第二附属医院，广东广州 510120
^2.广东省第二人民医院，广东广州 510317
^3.南方医科大学生物医学工程学院，广东广州 510515

收稿日期:2024-02-05 出版日期:2024-06-20 发布日期:2024-07-01
通讯作者: 黎金华 E-mail:guilingchen@gzucm.edu.cn;lijinhua0226@163.com
作者简介:陈桂玲，硕士，实习研究员，E-mail: guilingchen@gzucm.edu.cn
基金资助:
国家自然科学基金青年项目(82104456);中国博士后基金资助项目(2020M672745);广州市科技计划项目(202102020205);广东省中医药局中医药科研项目(20244026)

Solasonine promotes apoptosis of non-small cell lung cancer cells by regulating the Bcl-2/Bax/caspase-3 pathway

Guiling CHEN¹(), Xiaofeng LIAO¹, Pengtao SUN¹, Huan CEN¹, Shengchun SHU¹, Bijing LI¹, Jinhua LI¹^,²^,³()

^1.Second Affiliated Hospital of Guangzhou University of Chinese Medicine, Guangzhou 510120, China
^2.Guangdong Second Provincial General Hospital, Guangzhou 510317, China
^3.School of Biomedical Engineering, Southern Medical University, Guangzhou 510515, China

Received:2024-02-05 Online:2024-06-20 Published:2024-07-01
Contact: Jinhua LI E-mail:guilingchen@gzucm.edu.cn;lijinhua0226@163.com
Supported by:
Natural Science Foundation for the Youth (NSFY) of China(82104456)

摘要/Abstract

摘要：

目的探讨龙葵活性成分澳洲茄碱对非小细胞肺癌细胞PC9增殖、凋亡的影响。方法体外培养PC9细胞，设对照组（0 μmol/L）及澳洲茄碱不同剂量组（0、2、5、10、15、20、25 μmol/L），CCK-8试剂盒检测澳洲茄碱对PC9细胞的增殖抑制作用；TMRE检测线粒膜电位；caspase3/7活性试剂盒联合GreenNuc™ Caspase-3/Annexin V-mCherry染色检测caspase-3活性；Annexin V-FITC/PI双染法检测细胞凋亡率；给药处理或者使用PTEN抑制剂后，Western blot检测细胞中相关蛋白的表达量。结果与对照组相比，经澳洲茄碱干预24、48、72 h后，PC9细胞的活力均明显降低（P<0.05）；经澳洲茄碱干预24 h后，细胞线粒体膜电位明显降低，而细胞凋亡比例明显升高（P<0.05）；caspase-3/7活力、活细胞Caspase-3活性及cleaved caspase-3蛋白表达均显著升高（P<0.01）；PI3K和Akt磷酸化水平降低（P<0.05）；而PTEN、Bax蛋白表达上调（P<0.05）；抗凋亡蛋白Bcl-2蛋白的表达下调（P<0.05）。结论澳洲茄碱可通过调控Bcl-2/Bax/caspase-3通路及其上游蛋白活性而抑制PC9细胞增殖，促进其凋亡。

关键词: 澳洲茄碱, 肺癌, Bcl-2/Bax/caspase-3通路, 同源性磷酸酶-张力蛋白, 细胞凋亡

Abstract:

Objective To investigate the effect of solasonine, an active component of Solanum nigrum, on proliferation and apoptosis of non-small cell lung cancer PC9 cells. Methods PC9 cells were treated with 2, 5, 10, 15, 20, or 25 μmol/L solasonine, and the changes in cell proliferation were examined using CCK-8 assay. Tetramethyl rhodamine ethyl ester (TMRE) was used to detect the changes in mitochondrial membrane potential, and caspase-3/7 detection kit and GreenNuc™ caspase-3/Annexin V-mCherry kit for live cell were used to analyze the changes in caspase-3 of the cells. Annexin V-FITC/PI double staining was employed to analyze the apoptosis rate of the cells. The effect of PTEN inhibitors on solasonine-induced cell apoptosis was examined by detecting apoptosis-related protein expressions using Western blotting. Results Solasonine treatment for 24, 48, and 72 h significantly lowered the viability of PC9 cells. The cells treated with solasonine for 24 h showed significantly decreased mitochondrial membrane potential and increased cell apoptosis with enhanced caspase-3/7 and caspase-3 activities and expression of cleaved caspase-3. Solasonine treatment significantly decreased phosphorylation levels of PI3K and Akt, increased the protein expressions of PTEN and Bax, and lowered the expression of Bcl-2 protein in the cells. Conclusion Solasonine inhibits proliferation and induces apoptosis of PC9 cells by regulating the Bcl-2/Bax/caspase-3 pathway and its upstream proteins.

Key words: solasonine, lung cancer, Bcl-2/Bax/caspase-3 signaling pathway, homologous phosphatase tensin protein, apoptosis

陈桂玲, 廖晓凤, 孙鹏涛, 岑欢, 舒盛春, 李碧晶, 黎金华. 澳洲茄碱通过调控Bcl-2/Bax/caspase-3信号通路促进非小细胞肺癌发生凋亡[J]. 南方医科大学学报, 2024, 44(6): 1109-1116.

Guiling CHEN, Xiaofeng LIAO, Pengtao SUN, Huan CEN, Shengchun SHU, Bijing LI, Jinhua LI. Solasonine promotes apoptosis of non-small cell lung cancer cells by regulating the Bcl-2/Bax/caspase-3 pathway[J]. Journal of Southern Medical University, 2024, 44(6): 1109-1116.

图/表 7

图1 澳洲茄碱对PC9细胞增殖的影响

Fig.1 Effect of solasonine on PC9 cell proliferation (Mean±SD, n=3). *P<0.01, **P<0.001 vs Control.

图2 澳洲茄碱对PC9细胞线粒体膜电位的影响

Fig.2 Changes of mitochondrial membrane potential in solasonine-treated PC9 cells detected using TMRE and observed by immunofluorescence microscopy.

图4 澳洲茄碱对PC9细胞cleaved caspase-3蛋白和caspase-3/7酶活性的影响

Fig.4 Western blotting for detecting the expression of cleaved caspase-3 in PC9 cells treated with 5, 10, 15, 20 and 25 μmol/L solasonine for 24 h and quantitative analysis of the protein expression levels using Image J (n=3). A: Protein bands in Western blotting. B, C: Histogram of the protein expression levels. **P<0.01 vs Control.

图5 澳洲茄碱对PC9活细胞内细胞核和细胞膜的染色情况

Fig.5 Activity of caspase-3 in PC9 cells detected using GreenNuc™ caspase-3/Annexin V-mCherry double staining and observed by immunofluorescence microscopy.

图6 澳洲茄碱对PC9细胞Bcl-2和Bax蛋白表达的影响

Fig.6 Western blotting for detecting expressions of Bcl-1 and Bax in PC9 cells treated with 5, 10, 15, 20 and 25 μmol/L solasonine for 24 h and quantitative analysis of the protein expression levels (n=3). *P<0.05, **P<0.01 vs Control.

图7 澳洲茄碱对PI3K、Akt蛋白磷酸化的影响

Fig.7 Western blotting for detecting the expressions of p-PI3K and p-Akt in PC9 cells treated with 20 μmol/L solasonine l for 2, 4, 12 and 24 h and quantitative analysis of the protein expression levels (n=3). *P<0.05, **P<0.01 vs Control.

图8 澳洲茄碱对PC9细胞PTEN蛋白表达的影响

Fig.8 Effect of solasonine on protein expression of PTEN in PC9 cells. n=3, *P<0.05, **P<0.01 vs Control, ▲P<0.05 vs SF1670.

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澳洲茄碱通过调控Bcl-2/Bax/caspase-3信号通路促进非小细胞肺癌发生凋亡

Solasonine promotes apoptosis of non-small cell lung cancer cells by regulating the Bcl-2/Bax/caspase-3 pathway

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