CAJ | 학술논문

目的探讨蛋白酶体抑制剂硼替佐米单用及与三氧化二砷或高三尖杉酯碱联合作用对髓系白血病细胞株HL-60的凋亡诱导作用.方法 MTT法、Hoechst33342染色分别观察细胞增殖抑制及细胞凋亡情况,Western印迹检测bcl-2 、Caspase-9、Caspase-3、PARP蛋白表达.结果硼替佐米、三氧化二砷、高三尖杉酯碱单独作用时均对HL-60细胞有明显的增殖抑制作用及凋亡诱导作用；三氧化二砷或高三尖杉酯碱与硼替佐米联合对HL-60细胞的增殖抑制作用比3种药物单独作用增强(均P＜0.05)；形态学观察显示药物联合作用后凋亡诱导作用也比单药作用增强.Western印迹法检测显示15 μmol/L三氧化二砷单独作用细胞后,Caspase-9、Caspase-3、PARP均出现裂解片段,bcl-2蛋白表达水平下降；30 nmol/L高三尖杉酯碱单药作用后PARP出现裂解片段、bcl-2蛋白表达水平下降,但是Caspase-9、Caspase-3的表达与对照组比较无改变；联合用药后相关蛋白的改变与细胞凋亡相平行.结论高三尖杉酯碱或三氧化二砷与硼替佐米联合后对细胞的凋亡诱导有相加作用.三氧化二砷与硼替佐米的凋亡诱导相加作用与二者能共同抑制Caspase信号途径及bcl-2蛋白的表达有关,高三尖杉酯碱和硼替佐米的相加作用与二者共同抑制bcl-2蛋白表达及促进PARP裂解活化有关.
목적 탐토단백매체억제제붕체좌미단용급여삼양화이신혹고삼첨삼지감연합작용대수계백혈병세포주HL-60적조망유도작용.방법 MTT법、Hoechst33342염색분별관찰세포증식억제급세포조망정황,Western인적검측bcl-2 、Caspase-9、Caspase-3、PARP단백표체.결과 붕체좌미、삼양화이신、고삼첨삼지감단독작용시균대HL-60세포유명현적증식억제작용급조망유도작용；삼양화이신혹고삼첨삼지감여붕체좌미연합대HL-60세포적증식억제작용비3충약물단독작용증강(균P＜0.05)；형태학관찰현시약물연합작용후조망유도작용야비단약작용증강.Western인적법검측현시15 μmol/L삼양화이신단독작용세포후,Caspase-9、Caspase-3、PARP균출현렬해편단,bcl-2단백표체수평하강；30 nmol/L고삼첨삼지감단약작용후PARP출현렬해편단、bcl-2단백표체수평하강,단시Caspase-9、Caspase-3적표체여대조조비교무개변；연합용약후상관단백적개변여세포조망상평행.결론 고삼첨삼지감혹삼양화이신여붕체좌미연합후대세포적조망유도유상가작용.삼양화이신여붕체좌미적조망유도상가작용여이자능공동억제Caspase신호도경급bcl-2단백적표체유관,고삼첨삼지감화붕체좌미적상가작용여이자공동억제bcl-2단백표체급촉진PARP렬해활화유관.
Objective To explore the apoptosis effect induced by bortezomib combined with homoharringtonine or arsenious acid in HL-60 cell line and the mechanism.Methods Cell' s apoptosis was demonstrated by MTT assay and Hoechst33342 staining.Expression of bcl-2,Caspase-9,Caspase-3 and PARP protein was detected by Western blot.Results HL-60 cells' apoptosis could be induced by bortezomib,homoharringtonine and arsenious acid respectively.Proliferation inhibition of HL-60 cells could be enhanced significantly when treated by bortezomib combined with homoharringtonine or arsenious acid compared with treated by any of the three drugs alone (P ＜ 0.05).At the same time morphology shows the apoptosis induced by drugs combined is more obviously than by one drug.Western blot showed bcl-2 protein was down-regulated and Caspase-9,Caspase-3 and PARP proteins were all cleaved activation when cells were treated by 15 μmol/ L arsenious acid alone,but only cleaved activation of PARP and down-regulation of bcl-2 protein be detected when cells were treated with 30 nmol/L homoharringtonine alone,expression of Caspase-9 and Caspase-3 had no change compared with the control.The changes of associated proteins were paralleled with the cell' s apoptosis when treated with combined drugs.Conclusion HL-60 cells' apoptosis effect is inhanced significantly when bortezomib combined with homoharringtonine or arsenious acid.Arsenious acid and bortezomib can inhibit caspase signaling pathway and down-regulate the expression of bcl-2 protein together,but homoharringtonine and bortezomib can only down-regulate the expression of bcl-2 protein and induce cleaved activation of PARP together.