苯甲醇对红细胞冻干前海藻糖负载红细胞影响研究 (1)

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1、蟾蜍灵在诱导HL60细胞凋亡过程中对Bcl2和PKC的影响作者：田昕作者单位：中国医科大学附属第一医院肿瘤内科，沈阳110001【摘要】为了研究蟾蜍灵在诱导HL60细胞凋亡过程中对Bcl2表达与裂解和磷酸化的作用，及对蛋白激酶C（PKC）活性与PKCs亚细胞定位的影响，分别采用台盼蓝拒染法检测细胞生存率，细胞染色法观察凋亡形态，流式细胞术分析细胞周期，琼脂糖凝胶电泳检测凋亡DNA片段化，［γ32P］同位素掺入法测定PKC活性，Westernblot分析Bcl2及PKC蛋白表达。结果表明：①蟾蜍

2、灵抑制HL60细胞增殖，24、48及72小时的IC50分别为(25.8±2.1)、(8.0±1.2)及(2.3±0.3)nmol/L;②50nmol/L蟾蜍灵作用24小时可诱导HL60细胞凋亡;③50nmol/L蟾蜍灵处理HL60细胞6-24小时，Bcl2蛋白表达水平明显下调，并出现23kD的裂解片段，磷酸化水平逐渐降低;④1-100nmol/L蟾蜍灵分别作用30分钟，对PKC总活性无影响，但可促使PKCβⅡ膜转位。结论：蟾蜍灵诱导HL60细胞凋亡可能与Bcl2表达降低、裂解及脱磷酸化有关。【

3、关键词】白血病蟾蜍灵凋亡Bcl2蛋白激酶CEffectofBufalinInducingApoptosisonBcl2andPKCinHL60CellsTIANXin，WANGPingPing，LIUYunPeng，HOUKeZuo，JINBo，LUOYing，QUXiuJuanDepartmentofOncology，TheFirstAffiliatedHospital，ChinaMedicalUniversity，Shenyang110001，ChinaAbstractPrevious

4、studyrevealedthatbufalincaninhibitproliferation,andinduceapoptosisinsomehumancancercelllines.However,themechanismofitsanticancereffecthasnotbeenfullyunderstood.ThepresentstudywasdesignedtoinvestigatetheeffectsofbufalininducedapoptosisonBcl2andPKCinhuman

5、leukemicHL60cells.Thecellviabilitywasdeterminedbytrypanbluedyeexclusion.Theapoptosiswasdetectedbymorphology,flowcytometryandDNAagarosegelelectrophoresis.TheexpressionsofBcl2andPKCwereanalyzedbyWesternblot,andactivityofPKCwasassayedby［γ32P］isotopeincorp

6、orationmethod.Theresultsshowedasfollows:(1)proliferationofHL60cellswasinhibitedbybufalinandtheIC50at24,48,72hourswere(25.8±2.1),(8.0±1.2)and(2.3±0.3)nmol/L,respectively.(2)apoptosisofHL60cellswasinducedwhenthecellsweretreatedwithbufalinatconcentrationof

7、50nmol/Lfor24hours.(3)comparedwithcontrol,treatmentwithbufalinatconcentrationof50nmol/Lfor6-24hoursresultedindownregulationofproteinexpression,decreaseofphosphorylation,andcleavageofBcl2,simultaneously.(4)theactivityoftotalPKCwasunchangedwhenHL60cellswe

8、reexposedto1-100nmol/Lbufalinfor30minutes,butPKCβⅡunderwenttranslocationfromcytosoltomembrane.Itisconcludedthatapoptosisinducedbybufalinisassociatedwithdownregulationofproteinexpression,dephosphorylation,andcleavage