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华西医学  2010 

铁螯合剂诱导白血病细胞凋亡中caspase-3的变化

, PP. 1683-1685

Keywords: HL-60细胞,去铁胺,脱噬作用,caspase-3

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Abstract:

【】目的探讨铁螯合剂去铁胺(DFO)对诱导白血病细胞HL-60的分子机制。方法2003年7-12月用钙黄绿素(calcein)检测HL-60细胞LIP。台盼蓝活细胞拒染实验进行活细胞计数及细胞存活率测定;光镜形态学观察及流式细胞仪(FCM)等方法检测HL-60细胞凋亡;比色法检测caspase-3(基于pNA标记底物的比色法)活性。结果①不同浓度的DFO作用于HL-60细胞后,随培养时间延长及DFO浓度的增加,动态铁池降低,细胞生存率逐渐下降,凋亡率增加,显示一定的时间剂量依赖性。②HL-60细胞在不同浓度的DFO作用下,caspase-3的活性逐渐升高。50、100μmol/LDFO作用于HL-60细胞24h,caspase-3酶活性升高明显,与对照组相比,有统计学意义(P<0.001);相关分析结果显示,HL-60细胞LIP的改变与caspase-3活性变化呈负相关系(r=-0.887,P<0.05)。结论DFO诱导白血病细胞凋亡的作用可能与螯合细胞内铁,降低细胞LIP,激活caspase-3,最终实施细胞凋亡密切相关。?【Abstract】ObjectiveToobservethechangesofcaspase-3activityduringapoptosisofHL-60cellsinducedbyanirondeferoxamine(DFO).MethodsExponentiallygrowingHL-60cells(1×106/mL)wereusedinthisexperimentfromJuly2003toDecember2003.ThestudygroupsweredividedasfollowsDFOgroup,iron+DFOgroupandcontrolgroup.Theviabilitywasdetectedbytypanblue,apoptosiswasassessedbymorphologicalstudyandflowcytometry(FCM)assay,andthecaspase-3activitywasdetectedbymelorimetry.Theintracellularlabelironpool(LIP)wasmeasuredwithafluorimetricassayusingthemetalsensitiveprobecalcein-AM.Results①WhenHL-60cellswereincubatedwithdifferentconcentrationsofDFO,viabilityassaywaslowerthanthatinthecontrolgroupatthe12th,24thand48thhour(P<0.05).②ThecellsincubatedwithdifferentconcentrationsofDFOshoweddose-timedependenceandwasmuchhigherthanthatinthecontrolgroup(P<0.01).③Thecaspase-3activitywassignificantlyhigherintheapoptoticcellsthanthatinthecontrolcells.ConclusionsTheapoptosisofHL-60cellsinducedbyDFOmaybecorrelatedwiththedecreaseofcellularLIPandactivityofcaspase-3.?

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