蛇床子素对TRAIL诱导白血病HL-60细胞凋亡的作用及其相关机制研究
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摘要
目的:探讨蛇床子素(Osthole)对肿瘤坏死因子相关凋亡诱导配体(TRAIL)诱导白血病HL-60细胞凋亡的影响及其可能机制。方法:采用MTT法检测不同浓度Osthole和TRAIL单独及联合应用对HL-60细胞增殖的影响。选择低于半数抑制浓度(IC50)的100μmol/L的Osthole和40 ng/ml TRAIL单独或联合处理细胞,通过流式细胞术测定HL-60细胞凋亡和细胞线粒体膜电位(MMP)的变化,RT-PCR法检测BCL-2、BAX和DR5 mRNA的表达,用分光光度法检测Caspase-3、-8、-9活性变化。结果:100μmol/L Osthole和40 ng/ml TRAIL联合处理HL-60细胞48 h,HL-60细胞的凋亡率达(33.9±2.7)%,较单用Osthole、TRAIL均显著提高(P<0.05),同时2者联合应用能增强降低MMP和BCL-2/BAX表达比值的效果,提高DR5的表达和Caspase-3、-8、-9活性。结论:Osthole能增强TRAIL诱导HL-60细胞凋亡的敏感性,其机制可能与其激活线粒体凋亡途径和上调DR5的表达密切相关。
Objective: To explore the effects of Osthole on apoptosis of HL-60 cells induced by tumor necrosis factor related apoptosis inducing ligand( TRAIL) and its possible mechanism. Methods: The proliferative inhibition of HL-60 cells treated with different concentrations of Osthole,TRAIL alone and Osthole combined with TRAIL was measured by MTT assay. The HL-60 cells were treated with Osthole,TRAIL alone and Osthole combined with TRAIL at the concentration < IC50 value,i. e. 100 μmol/L for Osthole and 40 ng/ml for TRAIL. Apoptosis and mitochondrial membrane potential( MMP) of HL-60 cells were detected by flow cytometry; the mRNA expression of BCL-2,BAX and DR5 was determined by RT-PCR; and the levels of Caspase-3,-8,-9 activity were detected by spectrophotometry.Results: The combined treatment( 100 μmol/L Osthole + 40 ng/ml TRAIL) of HL-60 cells for 48 h induced an apoptotic rate of( 33. 9 ± 2. 7) %,which was significantly higher than that of cells treated with Osthole or TRAIL alone( P < 0. 05); at the same time,the combined treatment promoted the decrease of MMP and the expression rate of BCL-2/BAX,and potentiated the expression of DR5 and Caspase-3,-8,-9 activity. Conclusion: Osthole can sensitize HL-60 cells to TRAIL-induced apoptosis,which may be related with the activation of mitochondrial pathways and up-regulation of DR5.
Objective: To explore the effects of Osthole on apoptosis of HL-60 cells induced by tumor necrosis factor related apoptosis inducing ligand( TRAIL) and its possible mechanism. Methods: The proliferative inhibition of HL-60 cells treated with different concentrations of Osthole,TRAIL alone and Osthole combined with TRAIL was measured by MTT assay. The HL-60 cells were treated with Osthole,TRAIL alone and Osthole combined with TRAIL at the concentration < IC50 value,i. e. 100 μmol/L for Osthole and 40 ng/ml for TRAIL. Apoptosis and mitochondrial membrane potential( MMP) of HL-60 cells were detected by flow cytometry; the mRNA expression of BCL-2,BAX and DR5 was determined by RT-PCR; and the levels of Caspase-3,-8,-9 activity were detected by spectrophotometry.Results: The combined treatment( 100 μmol/L Osthole + 40 ng/ml TRAIL) of HL-60 cells for 48 h induced an apoptotic rate of( 33. 9 ± 2. 7) %,which was significantly higher than that of cells treated with Osthole or TRAIL alone( P < 0. 05); at the same time,the combined treatment promoted the decrease of MMP and the expression rate of BCL-2/BAX,and potentiated the expression of DR5 and Caspase-3,-8,-9 activity. Conclusion: Osthole can sensitize HL-60 cells to TRAIL-induced apoptosis,which may be related with the activation of mitochondrial pathways and up-regulation of DR5.
引文
1 Park MH,Kim SY,Kim YJ,et al.ALS2CR7(CDK15)attenuates TRAIL induced apoptosis by inducing phosphorylation of survivin Thr34.Biochem Biophys Res Commun,2014;450(1):129-134.
2 Cao W,Li X,Zheng S,et al.Selenocysteine derivative overcomes TRAIL resistance in melanoma cells:evidence for ROS-dependent synergism and signaling crosstalk.Oncotarget,2014;5(17):7431-7445.
3赵素容,段海峰,张配,等.2-DG增强TRAIL诱导白血病细胞HL-60凋亡的作用.中国实验血液学杂志,2013;21(2):351-355.
4胡荣,张嵘,李迎春,等.硼替佐米增加HL-60细胞对TRAIL的敏感性.中国实验血液学杂志,2010;18(3):617-620.
5王绩英,赵雪强,王昌明,等.三氧化二砷逆转A549细胞对TRAIL耐药的研究.重庆医学,2012;41(7):676-682.
6徐燕,廖建华,吴龙火.蛇床子素的生物学活性与作用机制研究进展.湖北农业科学,2013;52(18):4313-4318.
7李慧.蛇床子素及其衍生物抗肿瘤作用机制研究进展.中药药理与临床,2015;31(3):208-214.
8 Lin K,Gao Z,Shang B,et al.Osthole suppresses the proliferation and accelerates the apoptosis of human glioma cells via the upregulation of microRNA-16 and downregulation of MMP-9.Mol Med Rep,2015;12(3):4592-4597.
9方柯,袁小芬,廖琼,等.厚朴酚对HL-60细胞增殖和凋亡的作用及分子机制研究.中国实验血液学杂志,2016;24(2):388-393.
10 Dimberg LY,Anderson CK,Camidge R,et al.On the TRAIL to successful cancer therapy?Predicting and counteracting resistance against TRAIL-based therapeutics.Oncogene,2013;32(11):1341-1350.
11 Trivedi R,Maurya R,Mishra DP.Medicarpin,a legume phytoalexin sensitizes myeloid leukemia cells to TRAIL-induced apoptosis through the induction of DR5 and activation of the ROS-JNK-CHOPpathway.Cell Death Dis,2014;5:e1465.
12齐玲,赵东海,王玮瑶,等.Caspase-8和Bcl-2在脑肿瘤干细胞肿瘤坏死因子相关凋亡诱导配体耐药中的作用研究.中国药学杂志,2015;50(2):152-156.
13于有江,彭建明,叶记林,等.蛇床子素对宫颈癌Hela细胞凋亡的作用研究.重庆医学,2017;46(7):883-885.
14叶记林,刘永春,于有江,等.TRAIL诱导Hela细胞凋亡的线粒体信号通路.基础医学与临床,2011;31(10):1120-1123.
15 Quast SA,Berger A,Pl9tz M,et al.Sensitization of melanoma cells for TRAIL-induced apoptosis by activation of mitochondrial pathways via Bax.Eur J Cell Biol,2014;93(1-2):42-48.
16 Leanza L,Venturini E,Kadow S,et al.Targeting a mitochondrial potassium channel to fight cancer.Cell Calcium,2015;58(1):131-138.
17 James MA,Seibel WL,Kupert E,et al.A novel,soluble compound,C25,sensitizes to TRAIL-induced apoptosis through upregulation of DR5 expression.Anticancer Drugs,2015;26(5):518-530.
18 Azijli K,Weyhenmeyer B,Peters GJ,et al.Non-canonical kinase signaling by the death ligand TRAIL in cancer cells:discord in the death receptor family.Cell Death Differ,2013;20(7):858-868.
2 Cao W,Li X,Zheng S,et al.Selenocysteine derivative overcomes TRAIL resistance in melanoma cells:evidence for ROS-dependent synergism and signaling crosstalk.Oncotarget,2014;5(17):7431-7445.
3赵素容,段海峰,张配,等.2-DG增强TRAIL诱导白血病细胞HL-60凋亡的作用.中国实验血液学杂志,2013;21(2):351-355.
4胡荣,张嵘,李迎春,等.硼替佐米增加HL-60细胞对TRAIL的敏感性.中国实验血液学杂志,2010;18(3):617-620.
5王绩英,赵雪强,王昌明,等.三氧化二砷逆转A549细胞对TRAIL耐药的研究.重庆医学,2012;41(7):676-682.
6徐燕,廖建华,吴龙火.蛇床子素的生物学活性与作用机制研究进展.湖北农业科学,2013;52(18):4313-4318.
7李慧.蛇床子素及其衍生物抗肿瘤作用机制研究进展.中药药理与临床,2015;31(3):208-214.
8 Lin K,Gao Z,Shang B,et al.Osthole suppresses the proliferation and accelerates the apoptosis of human glioma cells via the upregulation of microRNA-16 and downregulation of MMP-9.Mol Med Rep,2015;12(3):4592-4597.
9方柯,袁小芬,廖琼,等.厚朴酚对HL-60细胞增殖和凋亡的作用及分子机制研究.中国实验血液学杂志,2016;24(2):388-393.
10 Dimberg LY,Anderson CK,Camidge R,et al.On the TRAIL to successful cancer therapy?Predicting and counteracting resistance against TRAIL-based therapeutics.Oncogene,2013;32(11):1341-1350.
11 Trivedi R,Maurya R,Mishra DP.Medicarpin,a legume phytoalexin sensitizes myeloid leukemia cells to TRAIL-induced apoptosis through the induction of DR5 and activation of the ROS-JNK-CHOPpathway.Cell Death Dis,2014;5:e1465.
12齐玲,赵东海,王玮瑶,等.Caspase-8和Bcl-2在脑肿瘤干细胞肿瘤坏死因子相关凋亡诱导配体耐药中的作用研究.中国药学杂志,2015;50(2):152-156.
13于有江,彭建明,叶记林,等.蛇床子素对宫颈癌Hela细胞凋亡的作用研究.重庆医学,2017;46(7):883-885.
14叶记林,刘永春,于有江,等.TRAIL诱导Hela细胞凋亡的线粒体信号通路.基础医学与临床,2011;31(10):1120-1123.
15 Quast SA,Berger A,Pl9tz M,et al.Sensitization of melanoma cells for TRAIL-induced apoptosis by activation of mitochondrial pathways via Bax.Eur J Cell Biol,2014;93(1-2):42-48.
16 Leanza L,Venturini E,Kadow S,et al.Targeting a mitochondrial potassium channel to fight cancer.Cell Calcium,2015;58(1):131-138.
17 James MA,Seibel WL,Kupert E,et al.A novel,soluble compound,C25,sensitizes to TRAIL-induced apoptosis through upregulation of DR5 expression.Anticancer Drugs,2015;26(5):518-530.
18 Azijli K,Weyhenmeyer B,Peters GJ,et al.Non-canonical kinase signaling by the death ligand TRAIL in cancer cells:discord in the death receptor family.Cell Death Differ,2013;20(7):858-868.