注射用益气复脉(冻干)联合顺铂对肺癌细胞A549增殖的协同作用研究
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摘要
目的观察注射用益气复脉(冻干)(YQFM)与顺铂(Cisplatin,DDP)联合对肺癌细胞株A549增殖抑制的协同效应及作用机制。方法采用CCK-8法检测YQFM、DDP以及两药联用对肺癌细胞株A549的增殖影响,并应用两药相互作用指数评价药物的联合效应;采用荧光探针JC-1来检测线粒体膜电位的变化,Wester Blotting检测凋亡相关蛋白Bax、Cleaved-Caspase 3表达。结果药物作用时间48 h,与DDP等剂量组比较,联合用药(YQFM 16.25 mg/mL+DDP 1.25μg/mL、YQFM 16.25 mg/mL+DDP 2.5μg/mL、YQFM 16.25 mg/mL+DDP 5.00μg/mL)组细胞增殖抑制率显著增加(P<0.05)。与对照组比较,各给药组凋亡率均显著升高(P<0.05);与DDP组比较,联合用药组凋亡率显著升高(P<0.05)。与对照组比较,DDP 5.0、10.0μg/mL组和各联合用药组促凋亡蛋白Bax表达显著增加(P<0.05);DDP质量浓度为5.0、10.0μg/mL时,联合用药组与DDP单独给药组比较,Bax蛋白表达量显著升高(P<0.05)。与对照组比较,各给药组凋亡蛋白Cleaved-caspase 3表达量显著升高(P<0.05);DDP质量浓度为5.0、10.0μg/mL时,联合用药组与DDP单独给药组比较,Cleaved-caspase 3蛋白表达量显著升高(P<0.05)。结论 YQFM联合DDP对A549细胞增殖存在协同抑制作用,机制可能与诱导线粒体凋亡相关。
objective To observe the synergistic effect and mechanism of combined injection of Yiqi Fumai Lyophilized Injection(YQFM) and Cisplatin(DDP) on the proliferation inhibition of A549 lung cancer cell line.Methods The CCK-8 method was used to detect the effect of single DDP and YQFM,and combination of two drugs on the proliferation of lung cancer cell line A549.The changes of mitochondrial membrane potential were detected by fluorescent probe JC-1,which labeled the mitochondrial membrane potential.Western Blot was used to detect the expression of apoptosis-related proteins Bax and Caspase3.Results Compared with DDP equal dose group,the inhibition rate of cell proliferation was significantly increased in YQFM 16.25 mg/mL + DDP 1.25 μg/mL,YQFM 16.25 mg/mL + DDP 2.5 μg/mL,YQFM 16.25 mg/mL + DDP 5.00 μg/mL groups(P<0.05).Compared with the control group,the apoptotic rate of each drug group was significantly higher(P<0.05),and the apoptotic rate of the combined drug group was significantly higher(P<0.05) than that of the DDP group.Compared with control group,the expression of apoptotic protein Bax in DDP 5.0,10.0 μg/mL and combination groups was significantly higher(P<0.05);when DDP concentration was 5.0 and 10.0 μg/mL,the expression of Bax protein in combination group was significantly higher than that in DDP alone group(P<0.05).Compared with the control group,the expression of apoptotic protein Cleaved-caspase 3 was significantly increased in each group(P<0.05).When the concentration of DDP was 5.0 and 10.0 μg/mL,the expression of Cleaved-caspase 3 in the combined group was significantly higher than that in the DDP alone group(P<0.05).Conclusion YQFM combined with DDP could inhibit the proliferation of A549 cells synergistically,and the mechanism might be related to the induction of mitochondrial apoptosis.
objective To observe the synergistic effect and mechanism of combined injection of Yiqi Fumai Lyophilized Injection(YQFM) and Cisplatin(DDP) on the proliferation inhibition of A549 lung cancer cell line.Methods The CCK-8 method was used to detect the effect of single DDP and YQFM,and combination of two drugs on the proliferation of lung cancer cell line A549.The changes of mitochondrial membrane potential were detected by fluorescent probe JC-1,which labeled the mitochondrial membrane potential.Western Blot was used to detect the expression of apoptosis-related proteins Bax and Caspase3.Results Compared with DDP equal dose group,the inhibition rate of cell proliferation was significantly increased in YQFM 16.25 mg/mL + DDP 1.25 μg/mL,YQFM 16.25 mg/mL + DDP 2.5 μg/mL,YQFM 16.25 mg/mL + DDP 5.00 μg/mL groups(P<0.05).Compared with the control group,the apoptotic rate of each drug group was significantly higher(P<0.05),and the apoptotic rate of the combined drug group was significantly higher(P<0.05) than that of the DDP group.Compared with control group,the expression of apoptotic protein Bax in DDP 5.0,10.0 μg/mL and combination groups was significantly higher(P<0.05);when DDP concentration was 5.0 and 10.0 μg/mL,the expression of Bax protein in combination group was significantly higher than that in DDP alone group(P<0.05).Compared with the control group,the expression of apoptotic protein Cleaved-caspase 3 was significantly increased in each group(P<0.05).When the concentration of DDP was 5.0 and 10.0 μg/mL,the expression of Cleaved-caspase 3 in the combined group was significantly higher than that in the DDP alone group(P<0.05).Conclusion YQFM combined with DDP could inhibit the proliferation of A549 cells synergistically,and the mechanism might be related to the induction of mitochondrial apoptosis.
引文
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[13]李小平,郑磊贞,顾建春,等.生脉注射液联合奥沙利铂抗结肠癌肝转移的作用[J].上海交通大学学报:医学版,2008,28(5):552-555.
[14]Gogvadze V,Robertson J D,Enoksson M,et al.Mitochondrial cytochrome release may occur by volume dependent mechanisms not involving permeability transition[J].Biochem J,2004,378(Pt 1):213-217.
[15]Krauskopf A,Eriksson O,Craigen W J,et al.Properties of the permeability transition in VDACI(-/-)mitochondria[J].Biochim Biophys Acta,2006,1757(5/6):590-595.
[2]乔曦.注射用益气复脉在临床中的应用[J].光明中医,2014,29(10):2205-2207.
[3]Wang D G,Wang Z P,Tian B Q,et al.Two hour exposure to sodium butyrate sensitizes bladder cancer to anticancer drugs[J].Int J Urol,2008,15(5):435-441.
[4]鞠爱春,罗瑞芝,秦袖平,等.注射用益气复脉(冻干)药理作用及临床研究进展[J].药物评价研究,2018,41(3):354-364
[5]Lee C H,Kim J H.A review on the medicinal potentials of ginseng and ginsenosides on cardiovascular diseases[J].J Ginseng Res,2014,38(3):161-166.
[6]Lan S,Yi F,Shuang L,et al.Chemical constituents from the fibrous root of Ophiopogon japonicas and their effect on tube formation in human myocardial microvascular endothelial cells[J].Fitoterapia,2013,85:57-63.
[7]Huang Y L,Kou J P,Ma L,et al.Possible mechanism of the anti-inflammatory activity of ruscogenin:Role of intercellular adhesion molecule-1 and nuclear factorkappa B[J].J Pharmacol Sci,2008,108(2):198-205.
[8]徐淑华,刘生友.生脉注射液的药理作用研究进展[J].中国药事,2010,24(4):405-407.
[9]Panossian A,Wikman G.Pharmacology of Schisandra chinensis Bail:An overview of Russian research and uses in medicine[J].J Ethnopharmacol,2008,118(2):183-212.
[10]苗苗,李嵌,刘雅茹.生脉注射液对不同化疗药物的增敏作用[J].中草药,2013,44(7):875-880.
[11]金秀东,徐秋玲,郑学芝,等.复方五味子素B逆转多药耐药性人结肠癌细胞THC-8307[J].医学综述,2011,17(10):1552-1554.
[12]Dalaveris E,Kerenidi T,Katsabeki-Katsafli A,et al.VEGF,TNF-αand 8-isoprostane levels in exhaled breath condensate and serum of patients with lung cancer[J].Lung Cancer,2009,64(2):219-225.
[13]李小平,郑磊贞,顾建春,等.生脉注射液联合奥沙利铂抗结肠癌肝转移的作用[J].上海交通大学学报:医学版,2008,28(5):552-555.
[14]Gogvadze V,Robertson J D,Enoksson M,et al.Mitochondrial cytochrome release may occur by volume dependent mechanisms not involving permeability transition[J].Biochem J,2004,378(Pt 1):213-217.
[15]Krauskopf A,Eriksson O,Craigen W J,et al.Properties of the permeability transition in VDACI(-/-)mitochondria[J].Biochim Biophys Acta,2006,1757(5/6):590-595.
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