四种砷拮抗剂对三氧化二砷诱导的三种粒系白血病细胞凋亡和端粒酶活性的减量调节作用(英文)

目的:研究巯基供体N-乙酰半胱氨酸(NAC)和二巯丁二钠(NDMS)、抗氧化剂过氧化氢酶(CAT)和Ca~(2 )清除刘(Quin 2)对三氧化二砷诱导的三种粒系白血病细胞凋亡和端粒酶活性改变的调控作用。方法:用流式细胞仪和PCR ELISA法分别检测NAC、NDMS、CAT或Quin 2与三氧化二砷共同作用于三种粒系白血病细胞后其凋亡和端粒酶活性的变 化。结果:三氧化二砷0.6、2.7和8.1μmol/L可分别诱导急性早幼粒细胞白血病细胞株NB4,慢性粒细胞白血病细胞株K562,急性粒细胞白血病细胞株HL-60细胞发生40％-60％的凋亡,同时下调三种细胞的端粒酶活性。NAC 4mmol/L,NDMS 200μmol/L,CAT 80kU/L,Quin 220μmol/L不同程度抑制这种凋亡作用。NAC和CAT既可独立降低三种细胞的端粒酶活性,也可促进三氧化二砷对端粒酶的下调作用,而Quin 2可抑制K562和HL-60细胞中的这种下调作用。结论:三氧化二砷诱导的三种细胞的凋亡过程涉及了疏基失活、自由基的改变、细胞内Ca~(2 )浓度改变及端粒酶活性下降。NAC、NDMS、CAT及Quin 2可不同程度拮抗三氧化二砷对三种细胞的作用。     

菊苣酸诱导人白血病细胞HL-60凋亡作用及机制研究

目的观察菊苣酸诱导人急性髓系白血病细胞株HL-60凋亡的作用并初步探讨其机制。方法培养HL-60细胞,分别给予终浓度为10¹00μmol·L-1的菊苣酸48 h,检测细胞活性和凋亡、Caspase-3活性以及Bcl-2蛋白表达水平。结果终浓度为10100μmol·L-1的菊苣酸48 h,检测细胞活性和凋亡、Caspase-3活性以及Bcl-2蛋白表达水平。结果终浓度为10¹00μmol·L-1的菊苣酸能呈浓度依赖性降低HL-60细胞增殖活性和增加凋亡,增强Caspase-3活性和下调Bcl-2蛋白表达。结论菊苣酸具有抑制白血病细胞株HL-60增殖活性和诱导其凋亡作用,其机制与降低抗凋亡蛋白Bcl-2表达及增强Caspase-3活性有关。     

大蒜素对HL-60细胞端粒酶活性的影响

目的 研究大蒜素对人早幼白血病HL-60细胞端粒酶活性的影响。方法 不同浓度的大蒜素作用HL-60细胞后,MTT法测细胞的生长和增殖情况,TRAP-PCR-ELISA法研究细胞端粒酶活性的变化。结果 大蒜素能明显地抑制HL-60细胞的生长,且呈时间、浓度依赖性。不同浓度的大蒜素能下调HL-60细胞的端粒酶活性,且同样呈时间、浓度依赖性。结论 大蒜素可抑制HL-60细胞的端粒酶活性,可能是其诱导HL-60细胞凋亡的重要机制之一。     

砷诱导的肝癌细胞凋亡及端粒酶活性改变

目的：研究三氧化二砷（As2O3)对肝癌细胞凋亡的诱导作用及其机制。方法：1μmol/L As2O3处理培养的人肝癌HCC－9204细胞48h，用形态学观察、细胞超微结构分析和流式细胞仪检测等方法鉴定细胞凋亡。用噻唑蓝（MTT）比色试验观察蛋白质全盛抑制剂放线菌素D对As2O3凋亡诱导作用的影响。用TRAP－PCR－ELISA法检测As2O3处理前后肝癌细胞端粒酶活性的变化情况。结果：1μmol/L As2O3作用48h后，HCC－9204细胞出现了典型的凋亡形态学改变和超微结构变化；流式细胞仪检测到大量Annexin-阳性、PI－阴性细胞、放线菌素D可明显拮抗As2O3的凋亡诱导作用。细胞凋亡发生后端粒酶活性显著降低，A450/A630值从2．874降为0．767。结论：As2O3可以有效诱导肝癌细胞凋亡，端粒酶活性降低可能是其中的机制之一。     

紫杉醇体外诱导人胃癌细胞凋亡及对端粒酶活性的影响

目的　研究紫杉醇对人胃癌细胞诱导凋亡作用及端粒酶活性变化。方法　 0 .0 0 1～ 1μmol/L的紫杉醇处理SGC 790 1细胞后 ,用MTT法测定胃癌细胞的生长抑制率 ,通过形态学观察及流式细胞术检测细胞凋亡率 ,半定量TRAP -银染法测定端粒酶活性变化。结果　不同浓度的紫杉醇对胃癌细胞均有明显抑制作用 ,且呈时间依赖性及剂量依赖性 ,光镜及流式细胞术分析表明 ,0 .0 1μmol/L的紫杉醇处理后 2 4h ,细胞即出现明显的凋亡形态特征及凋亡峰 ,对端粒酶活性的同步检测结果显示 ,紫杉醇在诱导细胞凋亡的同时伴随端粒酶活性下调 ,且随紫杉醇浓度增大 ,抑制作用逐渐增强 ,2 4h酶活性即显著受抑制 ,72h变为阴性。结论　紫杉醇对胃癌细胞具有明显抑制作用 ,诱导细胞凋亡并抑制端粒酶活性可能是其发挥抗癌作用的机制之一 ,端粒酶可作为肿瘤化疗的敏感性指标。     

MEK抑制剂联合三氧化二砷对髓系白血病细胞凋亡的研究

目的:研究MEK抑制剂PD98059联合三氧化二砷(As2O3)对髓系白血病细胞凋亡的影响及其作用机制。方法:将PD98059、As2O3单独或联合作用于髓系白血病细胞系HL-60、K562细胞,用AnnexinV-FITC法检测细胞凋亡,用流式细胞术检测Bcl-2、Caspapse-3表达。结果:联合组与单用组相比,细胞凋亡率明显增高。Bcl-2在HL-60、K562细胞均高水平表达。As2O3明显抑制HL-60细胞Bcl-2表达,对K562细胞Bcl-2无明显抑制作用。单用PD98059、As2O3及两药合用在诱导HL-60、K562细胞凋亡过程中,活化caspapse-3均明显上升,两药合用较单用PD98059或As2O3活化caspapse-3明显升高。结论:PD98059联合As2O3同时抑制ERK/MAPK和Bcl-2,激活Caspase酶,对HL-60细胞有协同促凋亡效应。两药联合同时靶向作用ERK/MAPK和BCR/ABL,活化Caspase酶,协同诱导K562细胞凋亡。PD98059可增强As2O3对髓系白血病细胞的凋亡诱导作用。     

三氧化二砷对人肺癌A549细胞凋亡及耐药基因表达的影响

目的研究三氧化二砷（As2O3）对人肺癌细胞株的诱导凋亡作用及对耐药基因LRP表达的影响。方法应用人肺腺癌A549细胞株,运用体外细胞培养法、MTT法、流式细胞术检测As2O3对人肺癌细胞株抑制及诱导凋亡作用;用逆转录-聚合酶链反应（RT-PCR）方法检测LRP mRNA的表达。结果As2O3对人肺癌A549细胞具有抑制作用,其抑制率呈时间-剂量依赖关系。不同浓度的As2O3均可诱导凋亡。1．0μmol／L、3．0μmol／L的As2O3可下调LRPmRNA的表达。结论As2O3具有抑制肿瘤细胞生长及诱导细胞凋亡作用,其机制与下调LRPmRNA表达有密切关系。     

熊果酸诱导慢性粒细胞白血病细胞凋亡过程中活性氧的生物学功能探讨

目的研究熊果酸对慢性粒细胞白血病细胞株K562的凋亡及其过程中活性氧自由基的作用。方法不同浓度UA处理后用显微镜观察K562细胞的形态学变化;用Annexin V/PI双染色法检测凋亡;用DCFH-DA法检测活性氧自由基的水平。结果 20μmol/L浓度组可见细胞变形、坏死,40μmol/L浓度组可见细胞变形、核固缩、核碎裂和胞膜突起等现象。10、20、40μmol/L作用48h后K562细胞凋亡呈剂量依赖关系,凋亡率分别为21.66%、40.55%、70.21%,随着UA浓度的增高细胞凋亡率也增高,差异具有统计学意义(P0.01)。熊果酸能显著增高K562细胞内的活性氧水平,且活性氧水平随着熊果酸的浓度增加而升高。平均荧光强度由48.96升至78.23,差异具有统计学意义(P0.01)。结论熊果酸可诱导慢性粒细胞白血病凋亡,且活性氧自由基的水平在凋亡诱导过程中起了重要作用。     

苦参碱类生物碱抗粒细胞和单核细胞性白血病的药理作用研究进展

苦参碱类生物碱对人红白血病K562细胞、TF-1细胞、急性早幼粒细胞白血病HL-60细胞、NB4细胞、急性单核细胞白血病THP-1细胞、白血病KG1a干细胞、慢性粒细胞白血病细胞和辐射引起的骨髓造血畸变细胞等均有抑制增殖并诱导分化和凋亡的作用。其诱导分化和凋亡及抑制增殖的作用机制是多方面的,与下调原癌基因c-myc、β-链蛋白、生存素表达和端粒酶活性以及上调半胱天冬酶活性和白血病细胞表面分化抗原CD11b、CD15表达有关。苦参碱类生物碱上调白血病细胞表面自然杀伤细胞活化受体NKG2D的配体表达,能提高白血病细胞对自然杀伤细胞的敏感性。综述苦参碱类生物碱抗粒细胞和单核细胞性白血病的药理作用文献,并对其研究进展做了分析。     

辛伐他汀对K562细胞端粒酶活性及人端粒酶逆转录酶表达的影响

目的探讨辛伐他汀治疗慢性粒细胞白血病可能的作用机制。方法辛伐他汀10和20μmol.L-1与K562细胞作用48或72 h后,用流式细胞术AnnexinⅤ-FITC/PI双染法检测细胞凋亡百分率;提取细胞端粒酶,用PCR-ELISA检测端粒酶活性;实时荧光定量PCR检测人端粒酶逆转录酶(hTERT),c-myc和bcl2-mRNA表达。结果辛伐他汀10和20μmo.lL-1与K562细胞作用48 h后,细胞凋亡率分别为(6.24±0.18)%和(9.41±0.22)%,与对照组(1.88±0.14)%比较明显增加;作用72 h后细胞凋亡率分别为(12.41±0.32)%和(19.08±0.26)%,与对照组(4.20±0.19)%比较明显增加。辛伐他汀10和20μmo.lL-1与K562细胞作用48或72 h后,端粒酶活性,hTERT,c-myc和bcl-2 mRNA表达明显低于对照组。结论辛伐他汀可使K562细胞端粒酶活性降低,其机制可能与下调hTERT mRNA表达有关。     

Inhibition of mitogen-activated protein kinase kinase enhances apoptosis induced by arsenic trioxide in human breast cancer MCF-7 cells

Arsenic trioxide (As2O3) has recently been used to treat acute promyelocytic leukaemia and has activity in vitro against several solid tumour cell lines where the induction of differentiation and apoptosis are the prime effects. The mechanism of As2O3-induced cell death has yet to be clarified, especially in solid cancers. In the present study, the human breast cancer cell line MCF-7 was examined as a cellular model for As2O3 treatment. The involvement of extracellular signal-regulated kinase (ERK), p38 and c-Jun N-terminal kinase (JNK) was investigated in As2O3-induced cell death. 3. It was found that As2O3 activates the prosurvival mitogen-activated protein kinase kinase (MEK)/ERK pathway in MCF-7 cells, which, conversely, may compromise the efficacy of As2O3. Hence, a combination treatment of As2O3 and MEK inhibitors was investigated to determine whether this treatment could lead to enhanced growth inhibition and apoptosis in MCF-7 cells. 4. Inhibition of MEK/ERK with the pharmacological inhibitors U0126 (10 micromol/L) or PD98059 (20 micromol/L) together with As2O3 (2 and 5 micromol/L) resulted in a significant enhancement of growth inhibition in breast cancer MCF-7 cells as determined by the 3-(4,5-dimethyl-2 thiazoyl)-2,5-diphenyl-2H-tetrazolium bromide assay and [Methyl-3H]-thymidine incorporation. Furthermore, the results demonstrated that combined treatment with As2O3 and the MEK1/2 inhibitor U0126 could augment breast cancer MCF-7 cell apoptosis approximately twofold compared with the effects of the two drugs alone, as determined by Hoechst 33258 or annexin V/propidium iodide (PI) staining and flow cytometry. 5. In addition, As2O3 activated p38 in a dose-dependent manner, but had no effect on JNK1/2. Treatment with a p38 inhibitor did not prevent As2O3-induced apoptosis. 6. In conclusion, the results of the present study showed that enhanced apoptosis is detected in breast cancer MCF-7 cells in the presence of As2O3 and an MEK inhibitor, which may be a new promising adjuvant to current breast cancer treatments.     

三氧化二砷对HL-60细胞凋亡及其端粒酶hTERT-mRNA表达影响的实验研究

目的　探讨不同浓度的三氧化二砷 (As2 O3)对急性早幼粒细胞白血病细胞株HL 60细胞端粒酶亚单位hTERT mRNA表达及其诱导凋亡的作用。方法　不同浓度的As2 O3与HL 60细胞株共培养 48h ,流式细胞术鉴定HL 60细胞凋亡 ;RT PCR法检测HL 60细胞端粒酶亚单位hTERTmRNA的表达。结果　在 0 1、1 0和 5 0 μmol·L-13种浓度的As2 O3作用下 ,HL 60细胞凋亡率分别为 2 2 8%± 0 40 %、2 5 5 %± 0 2 2 %和 47 5 7%± 2 79% ;端粒酶亚单位hTERT mRNA表达的相对值分别为 73 97%、63 70 %和 2 9 0 4%。细胞凋亡率在 5 0 μmol·L-1与 0 1和 1 0μmol·L-1浓度组间差异有显著性 (P <0 0 1) ;端粒酶亚单位表达的差异也有显著性 (P <0 0 5 )。As2 O3诱导HL 60细胞凋亡效应与其抑制端粒酶亚单位hTERTmRNA的表达相关。结论　As2 O3对HL 60细胞具有凋亡诱导效应 ,且对其端粒酶亚单位hTERTmRNA的表达具有抑制作用 ,两者均呈现浓度依赖效应。As2 O3可能通过抑制HL 60细胞端粒酶亚单位hTERTmRNA的表达而诱导肿瘤细胞凋亡     

As2S2诱导K562细胞凋亡及其机制

目的:探索As_2S_2对K562细胞的作用及其机制.方法:As_2S_2对K562细胞的生长抑制作用用细胞计数法;细胞凋亡的检测用流式细胞分析、基因组DNA电泳、细胞形态学观察等方法;Western-blot方法用于蛋白表达的检测;基因表达的变化用半定量RT-PCR方法.结果:As_2S_2浓度在1-5μmol/L作用24-72 h即可抑制K562细胞生长,大于3μmol/L时可诱导K562细胞凋亡.As_2S_2能降低K562细胞中Bcr-Abl蛋白水平及 c-abl和 Bcr-Abl PTK活性,但不调变bcr-abl基因表达水平.As_2S_2也能诱导慢性粒细胞性白血病(CML)患者单个核细胞凋亡,且Ph~ 单个核细胞比Ph~- 单个核细胞对As_2S_2诱导的凋亡更敏感.结论:As_2S_2可通过降低Bcr-Abl蛋白含量而诱导CML细胞凋亡.As_2S_2可能为治疗CML的有效药物.     

Diethyldithiocarbamate-induced cytotoxicity and apoptosis in leukemia cell lines

Diethyldithiocarbamate (DDTC) has been shown to induce cytotoxicity in several different systems. We examined whether the DDTC-induced cytotoxicity was via apoptosis, or in relation to intracellular glutathione (GSH) in various murine and human leukemia cell lines. The cells most sensitive to DDTC-induced cytotoxicity were P388 lymphoid neoplasma cells and NALM-6, a B cell line of acute lymphocytic leukemia (ALL). The next level of susceptible cells included J774.1, having a macrophage function, HL-60 premyelocytic leukemia cells, MOLT-4, an acute lymphoblastic leukemia cell, and Jurkat, a T-cell leukemia. U937 (expressing many monocyte-like characteristics), K562 erythroleukemia and K562/DXR (a multidrug-resistant clone derived from K562) were almost unaffected by DDTC. P388 was also highly susceptible to H(2)O(2), a most useful exogenous reactive oxygen species generator, and was lower in intracellular total GSH content than other leukemia cells. DDTC-induced cytotoxicity was closely related to intracellular GSH, but the level of cellular GSH did not always correlate with H(2)O(2)-induced cytotoxicity in this experiment. K562 had a higher intracellular total GSH content and showed lower susceptibility to DDTC and H(2)O(2), but with the combination of DDTC and DL-buthionine-(S,R)-sulfoximine (BSO), cytotoxicity increased significantly. The ratio of GSH/GSSG in P388 was reduced by DDTC or H(2)O(2). H(2)O(2)-induced cytotoxicity was completely blocked by catalase (CAT), while it was enhanced by superoxide dismutase (SOD). CAT or SOD did not affect DDTC-induced cytotoxicity. N-Acetylcysteine (NAC: 1 mM), a vanguard substance of GSH, and aurintricarboxylic acid (ATA: 100 microM), an endonuclease inhibitor, ameliorated DDTC-induced cytotoxicity and apoptosis. In conclusion, we suggest that DDTC-induced cytotoxicity was via an oxidative shift in the intracellular redox state, and accompanied the activation of endonuclease through apoptosis in leukemia cell lines.     

喜树碱诱导人白血病HL-60细胞凋亡过程中端粒酶的调控(英文)

目的:研究在喜树碱诱导人白血病HL—60细胞凋亡过程中端粒酶的调节变化规律.方法:用MTT法测定药物对细胞存活率的影响;用琼脂糖电泳及流式细胞术检测和定量凋亡的发生;用以PCR为基础的TRAP法测定端粒酶活力;逆转录PCR检测凋亡过程中bcl-2及端粒酶亚基hTR、hEST2/hTERT和TLPl/TPl的基因表达水平的变化.结果:端粒酶活力伴随喜树碱诱导HL-60细胞凋亡的发生而逐渐降低,在此过程中端粒酶各亚基的mRNA水平无可见性变化,而bcl-2的基因表达水平则相应下调.结论:端粒酶活力的下调和喜树碱诱导的HL-60凋亡密切相关,端粒酶活力的阻断并非发生在其亚基基因转录水平,bcl-2对端粒酶活力的调节也不是通过影响端粒酶亚基的转录水平来实现的.     

Cytotoxic and apoptogenic effect of Swietenia mahagoni (L.) Jacq. leaf extract in human leukemic cell lines U937, K562 and HL-60

The apoptogenic activity of Swietenia mahagoni leaf extract (SMLE) was investigated against three human leukemic cell lines – U937, K562 and HL-60. SMLE inhibited cell growth and metabolic activity of the leukemic cells and showed characteristic features of apoptosis. Flow-cytometric analysis showed that SMLE arrested U937 and K562 cell populations in the G2-M phase and the HL-60 cell population in the G1 phase of cell cycle. SMLE induced apoptosis was found to be mediated through mitochondrial intrinsic pathway involving the release of cytochrome c into the cytosol and activation of caspase-9 and caspase-3. Two flavonoids, catechin and quercetin-3-O-glucoside, isolated from SMLE, were found to inhibit the growth and metabolic activity of U937, K562 and HL-60 cells at much lower concentrations thus indicating that these two flavonoids might be the active ingredients responsible for the anti-leukemic activity of SMLE.     

Emodin induces apoptosis in human promyeloleukemic HL-60 cells accompanied by activation of caspase 3 cascade but independent of reactive oxygen species production

Emodin (1,3,8-trihydroxy-6-methylanthraquinone) is an active constituent of Rheum palmatum, and showed inhibitory activity on lipopolysaccharide-induced NO production in our previous study. However, the apoptosis-inducing activity of emodin has remained undefined. Among three structurally related anthraquinones, including emodin, physcion, and chrysophanol, emodin showed the most potent cytotoxic effects on HL-60 cells, accompanied by the dose- and time-dependent appearance of characteristics of apoptosis including an increase in DNA ladder intensity, morphological changes, appearance of apoptotic bodies, and an increase in hypodiploid cells. Emodin at apoptosis-inducing concentrations causes rapid and transient induction of caspase 3/CPP32 activity, but not caspase 1 activity, according to cleavage of caspase 3 substrates poly(ADP-ribose) polymerase and D4-GDI proteins, the appearance of cleaved caspase 3 fragments being detected in emodin- but not physcion- or chrysophanol-treated HL-60 cells. A decrease in the anti-apoptotic protein, Mcl-1, was detected in emodin-treated HL-60 cells, whereas other Bcl-2 family proteins including Bax, Bcl-2, Bcl-XL, and Bad remained unchanged. The caspase 3 inhibitor, Ac-DEVD-CHO, but not the caspase 1 inhibitor, Ac-YVAD-CHO, attenuated emodin-induced DNA ladders, associated with the blockage of PARP and D4-GDI cleavage. Free radical scavenging agents including NAC, catalase, SOD, ALL, DPI, L-NAME and PDTC showed no preventive effect on emodin-induced apoptotic responses, whereas NAC, CAT and PDTC prevented HL-60 cells from ROS (H(2)O(2))-induced apoptosis through inhibition of caspase 3 cascades. Induction of catalase, but not SOD, activity was detected in emodin-treated HL-60 cells by in gel activity assays, and H(2)O(2)-induced intracellular peroxide level was significantly reduced by prior treatment of emodin in HL-60 cells. Our experiments provide evidence that emodin is an effective apoptosis inducer in HL-60 cells through activation of the caspase 3 cascade, but that it is independent of ROS production.