酸浆苦素B对人非小细胞肺癌细胞增殖、迁移及凋亡的影响
|
摘要
目的探讨酸浆苦素B(PB)对非小细胞肺癌(NSCLC)A549细胞增殖、迁移和凋亡的影响,初步探讨其作用机制。方法细胞分为不同浓度(2.5、5、10、20、40μmol/L)PB组、对照组、空白组,实时无标记细胞分析技术(RTCA)、CCK-8法检测PB对A549细胞增殖抑制率变化,Transwell检测细胞迁移,流式细胞仪检测PB对A549细胞的凋亡及周期的影响,Western blotting检测PB对相关蛋白表达的影响。结果 2.5、5、10、20、40μmol/L PB可抑制A549细胞增殖,且呈剂量依赖性;Transwell结果显示,10、20μmol/L PB组较对照组的细胞迁移能力下降;10、20μmol/L PB处理后,A549细胞的凋亡率升高,呈剂量依赖性。与对照组相比,10、20μmol/L PB组G_2/M期细胞比例增高、G_0/G_1期比例减少。Western blotting检测结果显示,与对照组相比,10、20μmol/L PB组使Akt蛋白表达水平下降,而Caspase-9和PTEN蛋白表达水平上升。结论 PB可抑制A549细胞的增殖,诱导细胞凋亡且细胞周期阻滞,这可能与调控细胞周期相关蛋白表达有关。
Objective To investigate the effects of Physalin B(PB)on proliferation,migration and apoptosis of A549cells in non-small cell lung cancer(NSCLC),and preliminarily explore its mechanism.Methods The A549 cells w ere divided into different concentrations(2.5,5,10,20,40μmol/L)of PB groups,control group and blank group.The inhibition rate of PB on A549 cells proliferation w as detected by real-time cell analyzer(RTCA)and CCK-8 assay,cell migration w as examined by transw ell chamber assay,the apoptosis and cycle of PB on A549 cells w ere detected by flow cytometry and the effect of PB on the expression of related proteins w as detected by Western blotting.Results PB w ith2.5,5,10,20 and 40μmol/L significantly inhibited the proliferation of A549 cells in a dose-dependent manner.The cell migration ability of 10 and 20μmol/L PB groups w as significantly low er than that of control group.After treatment w ith 10 and 20μmol/L PB,the apoptosis rate of A549 cells increased in a dose-dependent manner.Compared w ith control group,the proportion of G_2/M phase cells increased and the proportion of G_0/G_1phase cells decreased in 10 and 20μmol/L PB groups.Compared with the control group,the expression level of Akt protein decreased while the expression levels of Caspase-9 and PTEN increased in 10 and 20μmol/L PB groups.Conclusion PB can inhibit the proliferation of A549 cells,induce cell apoptosis and cell cycle arrest,w hich may be related to the regulation of cell cycle related protein expression.
Objective To investigate the effects of Physalin B(PB)on proliferation,migration and apoptosis of A549cells in non-small cell lung cancer(NSCLC),and preliminarily explore its mechanism.Methods The A549 cells w ere divided into different concentrations(2.5,5,10,20,40μmol/L)of PB groups,control group and blank group.The inhibition rate of PB on A549 cells proliferation w as detected by real-time cell analyzer(RTCA)and CCK-8 assay,cell migration w as examined by transw ell chamber assay,the apoptosis and cycle of PB on A549 cells w ere detected by flow cytometry and the effect of PB on the expression of related proteins w as detected by Western blotting.Results PB w ith2.5,5,10,20 and 40μmol/L significantly inhibited the proliferation of A549 cells in a dose-dependent manner.The cell migration ability of 10 and 20μmol/L PB groups w as significantly low er than that of control group.After treatment w ith 10 and 20μmol/L PB,the apoptosis rate of A549 cells increased in a dose-dependent manner.Compared w ith control group,the proportion of G_2/M phase cells increased and the proportion of G_0/G_1phase cells decreased in 10 and 20μmol/L PB groups.Compared with the control group,the expression level of Akt protein decreased while the expression levels of Caspase-9 and PTEN increased in 10 and 20μmol/L PB groups.Conclusion PB can inhibit the proliferation of A549 cells,induce cell apoptosis and cell cycle arrest,w hich may be related to the regulation of cell cycle related protein expression.
引文
[1]Jemal A, Siegel R, Xu JQ, et al. Cancer statistics, 2010[J]. CA Cancer J Clin, 2010, 60(5): 277-300.
[2]Chen WQ, Zheng RS, Zeng HM, et al. Epidemiology of lung cancer in China[J]. Thorac Cancer, 2015, 6(2): 209-215.
[3]Verdecchia A, Francisci S, Brenner H, et al. Recent cancer survival in Europe: a 2000-02 period analysis of EUROCARE-4 data[J]. Lancet Oncol, 2007, 8(9): 784-796.
[4]Bravo LE, García LS, Collazos PA. Cancer survival in Cali, Colombia: a population-based study, 1995-2004[J]. Colomb Med, 2014, 45(3): 110-116.
[5]J?nne PA, Johnson BE. Effect of epidermal growth factor receptor tyrosine kinase domain mutations on the outcome of patients with non-small cell lung cancer treated with epidermal growth factor receptor tyrosine kinase inhibitors[J]. Clin Cancer Res, 2006, 12(14 Pt 2): 4416s-4420s.
[6]杨燕军, 陈梅果, 胡玲, 等. 苦蘵的化学成分研究[J]. 中国药学杂志, 2013, 48(20): 1715-1718.YANG Yanjun, CHEN Meiguo, HU Ling, et al. Chemical constituents of whole plant of physalis angulata L[J]. Chinese Pharmaceutical Journal, 2013, 48(20): 1715-1718.
[7]Cragg GM, Newman DJ. Plants as a source of anti-cancer agents[J]. J Ethnopharmacol, 2005, 100(1/2): 72-79.
[8]Makino B, Ohya J, Yamamura H, et al. Cytotoxic activity of physalins possessing modified skeletal structures against HeLa cells[J]. Pharmazie, 2003, 58(1): 70-71.
[9]Wu SJ, Ng LT, Chen CH, et al. Antihepatoma activity of Physalis angulata and P. peruviana extracts and their effects on apoptosis in human Hep G2 cells[J]. Life Sci, 2004, 74(16): 2061-2073.
[10]Hsieh WT, Huang KY, Lin HY, et al. Physalis angulata induced G2/M phase arrest in human breast cancer cells[J]. Food Chem Toxicol, 2006, 44(7): 974-983.
[11]Sá MS, de Menezes MN, Krettli AU, et al. Antimalarial activity of physalins B, D, F, and G[J]. J Nat Prod, 2011, 74(10): 2269-2272.
[12]Jacobo-Herrera NJ, Bremner P, Marquez N, et al. Physalins from Witheringia solanacea as modulators of the NF-kappaB cascade[J]. J Nat Prod, 2006, 69(3): 328-331.
[13]Vandenberghe I, Créancier L, Vispé S, et al. Physalin B, a novel inhibitor of the ubiquitin-proteasome pathway, triggers NOXA-associated apoptosis[J]. Biochem Pharmacol, 2008, 76(4): 453-462.
[14]Hsu CC, Wu YC, Farh L, et al. Physalin B from Physalis angulata triggers the NOXA-related apoptosis pathway of human melanoma A375 cells[J]. Food Chem Toxicol, 2012, 50(3/4): 619-624.
[15]Ye YC, Tang XY, Sun ZZ, et al. Upregulated WDR26 serves as a scaffold to coordinate PI3K/ AKT pathway-driven breast cancer cell growth, migration, and invasion[J]. Oncotarget, 2016, 7(14): 17854-17869.
[16]Cai J, Xu LJ, Tang HJ, et al. The role of the PTEN/PI3K/Akt pathway on prognosis in epithelial ovarian cancer: a meta-analysis[J]. Oncologist, 2014, 19(5): 528-535.
[17]Chen JZ, O'Donoghue A, Deng YF, et al. The effect of lycopene on the PI3K/Akt signalling pathway in prostate cancer[J]. Anticancer Agents Med Chem, 2014, 14(6): 800-805.
[18]Khan KH, Yap TA, Yan L, et al. Targeting the PI3K-AKT-mTOR signaling network in cancer[J]. Chin J Cancer, 2013, 32(5): 253-265.
[19]Carnero A, Blanco-Aparicio C, Renner O, et al. The PTEN/PI3K/AKT signalling pathway in cancer, therapeutic implications[J]. Curr Cancer Drug Targets, 2008, 8(3): 187-198.
[20]Jiang BH, Liu LZ. PI3K/PTEN signaling in tumorigenesis and angiogenesis[J]. Biochim Biophys Acta, 2008, 1784(1): 150-158.
[21]Oda K, Stokoe D, Taketani Y, et al. High frequency of coexistent mutations of PIK3CA and PTEN genes in endometrial carcinoma[J]. Cancer Res, 2005, 65(23): 10669-10673.
[22]Jiang BH, Zheng JZ, Aoki M, et al. Phosphatidylinositol 3-kinase signaling mediates angiogenesis and expression of vascular endothelial growth factor in endothelial cells[J]. Proc Natl Acad Sci U S A, 2000, 97(4): 1749-1753.
[23]Jeong YJ, Choi Y, Shin JM, et al. Melittin suppresses EGF-induced cell motility and invasion by inhibiting PI3K/Akt/mTOR signaling pathway in breast cancer cells[J]. Food Chem Toxicol, 2014, 68: 218-225. doi:10.1016/j.fct.2014.03.022.
[24]Zhang H, Zhao B, Huang C, et al. Melittin restores PTEN expression by down-regulating HDAC2 in human hepatocelluar carcinoma HepG2 cells[J]. PLoS One, 2014, 9(5): e95520. doi:10.1371/journal.pone.0095520.
[2]Chen WQ, Zheng RS, Zeng HM, et al. Epidemiology of lung cancer in China[J]. Thorac Cancer, 2015, 6(2): 209-215.
[3]Verdecchia A, Francisci S, Brenner H, et al. Recent cancer survival in Europe: a 2000-02 period analysis of EUROCARE-4 data[J]. Lancet Oncol, 2007, 8(9): 784-796.
[4]Bravo LE, García LS, Collazos PA. Cancer survival in Cali, Colombia: a population-based study, 1995-2004[J]. Colomb Med, 2014, 45(3): 110-116.
[5]J?nne PA, Johnson BE. Effect of epidermal growth factor receptor tyrosine kinase domain mutations on the outcome of patients with non-small cell lung cancer treated with epidermal growth factor receptor tyrosine kinase inhibitors[J]. Clin Cancer Res, 2006, 12(14 Pt 2): 4416s-4420s.
[6]杨燕军, 陈梅果, 胡玲, 等. 苦蘵的化学成分研究[J]. 中国药学杂志, 2013, 48(20): 1715-1718.YANG Yanjun, CHEN Meiguo, HU Ling, et al. Chemical constituents of whole plant of physalis angulata L[J]. Chinese Pharmaceutical Journal, 2013, 48(20): 1715-1718.
[7]Cragg GM, Newman DJ. Plants as a source of anti-cancer agents[J]. J Ethnopharmacol, 2005, 100(1/2): 72-79.
[8]Makino B, Ohya J, Yamamura H, et al. Cytotoxic activity of physalins possessing modified skeletal structures against HeLa cells[J]. Pharmazie, 2003, 58(1): 70-71.
[9]Wu SJ, Ng LT, Chen CH, et al. Antihepatoma activity of Physalis angulata and P. peruviana extracts and their effects on apoptosis in human Hep G2 cells[J]. Life Sci, 2004, 74(16): 2061-2073.
[10]Hsieh WT, Huang KY, Lin HY, et al. Physalis angulata induced G2/M phase arrest in human breast cancer cells[J]. Food Chem Toxicol, 2006, 44(7): 974-983.
[11]Sá MS, de Menezes MN, Krettli AU, et al. Antimalarial activity of physalins B, D, F, and G[J]. J Nat Prod, 2011, 74(10): 2269-2272.
[12]Jacobo-Herrera NJ, Bremner P, Marquez N, et al. Physalins from Witheringia solanacea as modulators of the NF-kappaB cascade[J]. J Nat Prod, 2006, 69(3): 328-331.
[13]Vandenberghe I, Créancier L, Vispé S, et al. Physalin B, a novel inhibitor of the ubiquitin-proteasome pathway, triggers NOXA-associated apoptosis[J]. Biochem Pharmacol, 2008, 76(4): 453-462.
[14]Hsu CC, Wu YC, Farh L, et al. Physalin B from Physalis angulata triggers the NOXA-related apoptosis pathway of human melanoma A375 cells[J]. Food Chem Toxicol, 2012, 50(3/4): 619-624.
[15]Ye YC, Tang XY, Sun ZZ, et al. Upregulated WDR26 serves as a scaffold to coordinate PI3K/ AKT pathway-driven breast cancer cell growth, migration, and invasion[J]. Oncotarget, 2016, 7(14): 17854-17869.
[16]Cai J, Xu LJ, Tang HJ, et al. The role of the PTEN/PI3K/Akt pathway on prognosis in epithelial ovarian cancer: a meta-analysis[J]. Oncologist, 2014, 19(5): 528-535.
[17]Chen JZ, O'Donoghue A, Deng YF, et al. The effect of lycopene on the PI3K/Akt signalling pathway in prostate cancer[J]. Anticancer Agents Med Chem, 2014, 14(6): 800-805.
[18]Khan KH, Yap TA, Yan L, et al. Targeting the PI3K-AKT-mTOR signaling network in cancer[J]. Chin J Cancer, 2013, 32(5): 253-265.
[19]Carnero A, Blanco-Aparicio C, Renner O, et al. The PTEN/PI3K/AKT signalling pathway in cancer, therapeutic implications[J]. Curr Cancer Drug Targets, 2008, 8(3): 187-198.
[20]Jiang BH, Liu LZ. PI3K/PTEN signaling in tumorigenesis and angiogenesis[J]. Biochim Biophys Acta, 2008, 1784(1): 150-158.
[21]Oda K, Stokoe D, Taketani Y, et al. High frequency of coexistent mutations of PIK3CA and PTEN genes in endometrial carcinoma[J]. Cancer Res, 2005, 65(23): 10669-10673.
[22]Jiang BH, Zheng JZ, Aoki M, et al. Phosphatidylinositol 3-kinase signaling mediates angiogenesis and expression of vascular endothelial growth factor in endothelial cells[J]. Proc Natl Acad Sci U S A, 2000, 97(4): 1749-1753.
[23]Jeong YJ, Choi Y, Shin JM, et al. Melittin suppresses EGF-induced cell motility and invasion by inhibiting PI3K/Akt/mTOR signaling pathway in breast cancer cells[J]. Food Chem Toxicol, 2014, 68: 218-225. doi:10.1016/j.fct.2014.03.022.
[24]Zhang H, Zhao B, Huang C, et al. Melittin restores PTEN expression by down-regulating HDAC2 in human hepatocelluar carcinoma HepG2 cells[J]. PLoS One, 2014, 9(5): e95520. doi:10.1371/journal.pone.0095520.