赖氨藤黄酸对乳腺癌MCF-7细胞增殖和凋亡的影响及其作用机制

doi:10.13481/j.1671-587x.20150308

摘要/Abstract

摘要：

目的: 研究赖氨藤黄酸(GAL)对乳腺癌MCF-7细胞增殖和凋亡的影响并探讨其作用机制,为GAL 抗乳腺癌的临床研究和应用提供理论依据。方法: 选取处于对数生长期的MCF-7细胞,并随机分为空白对照组、不同剂量 (0.25、0.50、1.00、2.00、4.00和8.00 μmol·L^-1)GAL组和不同剂量(0.25、0.50、1.00、2.00、4.00和8.00μmol·L^-1)藤黄酸组,采用MTT法检测各组MCF-7细胞48 h增殖活性。选取对数生长期的MCF-7细胞,2 μmol·L^-1 GAL作用MCF-7细胞不同时间(0、6、12、24、36、48和60 h),采用MTT法检测作用不同时间后各组MCF-7细胞增殖活性。流式细胞术和Hoechst 33258染色检测各组MCF-7细胞24 h时凋亡情况,采用Western blotting 法检测各组MCF-7细胞24 h时凋亡相关蛋白的表达水平。结果: 细胞培养24 h 后,不同剂量GAL组MCF-7细胞存活率低于空白对照组(P<0.05),随剂量增加和作用时间延长细胞存活率下降(P<0.05)。流式细胞术和Hoechst 33258染色,与空白对照组比较,不同剂量GAL组MCF-7细胞凋亡率明显升高(P<0.05),且呈剂量依赖性。Western blotting检测,与空白对照组比较,不同剂量GAL组细胞沉默信息调节因子1(SIRT1)蛋白表达水平明显降低 (P<0.05),而caspase-3切割片段蛋白的表达水平明显高于对照组 (P<0.05)。结论: GAL 通过抑制SIRT1蛋白表达水平和上调caspase-3切割片段蛋白的表达水平诱导乳腺癌MCF-7细胞凋亡。

关键词: 赖氨藤黄酸, 乳腺肿瘤, 细胞凋亡, 沉默信息调节因子1

Abstract:

Objective To investigate the effect of gambogic acid lysinate (GAL) on the proliferation and apoptosis of human breast cancer MCF-7 cells and its mechanism, and to provide theoretical foundation for clinical trial and use of GAL against breast cancer. Methods The human breast cancer MCF-7 cells in logarithm growth phase were selected and randomly divided into blank control group, different doses (0.25, 0.50, 1.00, 2.00, 4.00, 8.00 μmol·L^-1) of GAL groups, and different doses(0.25, 0.50, 1.00, 2.00, 4.00, 8.00 μmol·L^-1) of gambogic acid groups.After the MCF-7 cells were treated with GAL or gambogic acid for 48 h, MTT assay was used to detect the proliferation activities of MCF-7 cells.After the MCF-7 cells in logarithm growth phase were treated with 2 μmol·L^-1 GAL for different time (0, 6, 12, 24, 36, 48, 60 h), MTT assay was also used to detect the proliferation activities of MCF-7 cells in various groups.The flow cytometry method and Hoechst 33258 staining were used to analyze the apoptosis of MCF-7 cells in various groups. The expression levels of apoptosis-associated proteins were detected by Western blotting method. Results After 48 h cell culture, the cell viabilities in GAL groups were lower than those in control group (P< 0.05) and the cell viability was inhibited in a dose-dependent manner (P<0.05).After the MCF-7 cells were treated by 2 μmol·L^-1 GAL for different time, the cell proliferation was inhibited in a time-dependent manner (P<0.05).The flow cytometry method and Hoechst 33258 staining Results showed that the apoptotic rates of MCF-7 cells in GAL groups were higher than those in control group (P<0.05).The expressions levels of SIRT1 in GAL groups were significantly lower than that in control group (P<0.05), while the expression amounts of cleaved caspase-3 were higher than that in control group (P<0.05). Conclusion GAL can induce the apoptosis of breast cancer MCF-7 cells by inhibiting the expression level of SIRT1 and up-regulating the expression amount of cleaved caspase-3.

Key words: gambogic acid lysinate, breast neoplasms, apoptosis, silent information regulator 1

中图分类号:

R737.9

魏洁, 李开济, 魏静波, 赵毓芳, 闫丰, 吕翠平, 甄永占. 赖氨藤黄酸对乳腺癌MCF-7细胞增殖和凋亡的影响及其作用机制[J]. 吉林大学学报(医学版), 2015, 41(03): 476-480.

WEI Jie, LI Kaiji, WEI Jingbo, ZHAO Yufang, YAN Feng, LYU Cuiping, ZHEN Yongzhan. Influence of gambogic acid lysinate in proliferation and apoptosis in breast cancer MCF-7 cells and its mechanism[J]. Journal of Jilin University Medicine Edition, 2015, 41(03): 476-480.

参考文献

[1] Amiri-Kordestani L, Blumenthal GM, Xu QC, et al. FDA approval: ado-trastuzumab emtansine for the treatment of patients with HER2-positive metastatic breast cancer [J].Clin Cancer Res, 2014.Epub ahead of print.

[2] Wang H, Vo T, Hajar A, et al.Multiple mechanisms underlying acquired resistance to taxanes in selected docetaxel-resistant MCF-7 breast cancer cells [J].BMC Cancer, 2014, 14:37.

[3] Wang X, Chen W.Gambogic acid is a novel anti-cancer agent that inhibits cell proliferation, angiogenesis and metastasis [J].Anticancer Agents Med Chem, 2012, 12(8):994-1000.

[4] Cascão R, Vidal B, Raquel H, et al.Potent anti-inflammatory and antiproliferative effects of gambogic acid in a rat model of antigen-induced arthritis [J].Mediators Inflamm, 2014, 2014:195327.

[5] Wen J, Pei H, Wang X, et al.Gambogic acid exhibits anti-psoriatic efficacy through inhibition of angiogenesis and inflammation [J].J Dermatol Sci, 2014, 74(3):242-250.

[6] Duan D, Zhang B, Yao J, et al.Gambogic acid induces apoptosis in hepatocellular carcinoma SMMC-7721 cells by targeting cytosolic thioredoxin reductase [J].Free Radic Biol Med, 2014, 69:15-25.

[7] DeSantis C, Ma J, Bryan L, et al.Breast cancer statistics, 2013 [J].CA Cancer J Clin, 2014, 64(1):52-62.

[8] Fountzilas G, Dafni U, Papadimitriou C, et al.Dose-dense sequential adjuvntchemotherapy followed, as indicated, by trastuzumab for one year in patients with early breast cancer: first report at 5-year median follow-up of a Hellenic Cooperative Oncology Grouprandomized phase Ⅲ trial [J].BMC Cancer, 2014, 14(1):515.

[9] Proietti S, Cucina A, Dobrowolny G, et al.Melatonin down-regulates MDM2 gene expression and enhances p53 acetylation in MCF-7 cells [J].J Pineal Res, 2014, 57(1):120-129.

[10] Hwang BJ, Madabushi A, Jin J, et al.Histone/protein deacetylase SIRT1 is an anticancer therapeutic target [J].Am J Cancer Res, 2014, 4(3):211-221.

[11] Yuan H, Su L, Chen WY.The emerging and diverse roles of sirtuins in cancer: a clinical perspective [J].Oncol Targets Ther, 2013, 6:1399-1416.

[12] Thompson KJ, Humphries JR, Niemeyer DJ, et al.The effect of alcohol on Sirt1 expression and function in animal and human models of hepatocellular carcinoma (HCC) [J].Adv Exp Med Biol, 2015, 815:361-373.

[13] Chen IC, Chiang WF, Huang HH, , et al.Role of SIRT1 in regulation of epithelial-to-mesenchymal transition in oral squamous cell carcinoma metastasis [J].Mol Cancer, 2014, 13(1):254.

[14] Ye D, Shuhendler AJ, Pandit P, et al.Caspase-responsive smart gadolinium-based contrast agent for magnetic resonance imaging of drug-induced apoptosis [J].Chem Sci, 2014, 4(10):3845-3852.

[15] Ji YB, Yu L.N-Butanol extract of capparis spinosa L.induces apoptosis primarily through a mitochondrial pathway involving mPTP open, cytochrome C release and Caspase activation [J].Asian Pac J Cancer Prev, 2014, 15(21):9153-9157.