中华医学杂志
中華醫學雜誌
중화의학잡지
National Medical Journal of China
2015年
4期
299-305
,共7页
王静%许晓军%周睿卿%郭坤元
王靜%許曉軍%週睿卿%郭坤元
왕정%허효군%주예경%곽곤원
伊曲康唑%多柔比星%KG1α细胞%细胞凋亡
伊麯康唑%多柔比星%KG1α細胞%細胞凋亡
이곡강서%다유비성%KG1α세포%세포조망
Itraconazole%Doxorubicin%KG1α cells%Apoptosis
目的 研究伊曲康唑(ITC)单药及联合化疗药多柔比星(ADM)对急性髓系白血病细胞增殖及凋亡的影响.方法 活细胞计数试剂盒(CCK-8)法检测不同浓度ITC、ADM单药及ITC(2、6、15 μmol/L)联合ADM(0.30、0.75 μmol/L)对人急性髓系白血病细胞株KG1α及急性髓系白血病原代细胞的增殖抑制作用;不同浓度ITC持续处理的KG1 α细胞,分别于7、14、21 d观察对集落克隆形成的影响;ADM 0.75 μmol/L、ITC 6 μmol/L及二者联合作用KG1α细胞48 h后,瑞氏染色法观察细胞形态变化,流式细胞术检测细胞凋亡率、线粒体膜电位及细胞周期阻滞情况,Western印迹检测Sonic Hedgehog(Shh)信号通路相关蛋白Shh和胶质瘤相关癌基因同系物1(Gli1)的表达水平.结果 ITC及ADM对KG1α细胞及急性髓系白血病原代细胞均具有增殖抑制作用,并且ITC可降低KG1α克隆形成能力,均呈剂量依赖.Weeb系数检验,ADM0.75 μmol/L与ITC 6 μmol/L两药联合时,细胞实际存活率≤70％细胞预估存活率,有协同效应;联合组与对照组和单药组相比,KG1 α细胞形态变化更为明显.ADM 0.75 μmol/L与ITC 6μmol/L联合诱导KG1α细胞48 h的凋亡率为31.72％±1.58％,均显著高于对照组、ITC单药组和ADM单药组(4.17％±0.74％、4.33％±1.12％、9.53％±1.15％,均P＜0.01).线粒体膜电位检测中,联合组的低红色荧光细胞(P3)比例明显高于对照组、ADM和ITC单药组(18.80％±0.96％比5.00％±0.38％、9.70％±0.43％、7.10％±0.77％,均P＜0.01).ADM 0.75 μmol/L与ITC 6 μmol/L联合组与对照组及单药组比明显阻滞细胞于G2期(P＜0.05).Western印迹结果显示Shh和Gli1的表达水平随着ITC浓度增大而下降.结论 ITC与ADM合用可明显抑制细胞的增殖,增加KG1 α细胞凋亡率、线粒体的损伤及细胞G2期的阻滞;ITC可以抑制Shh信号通路.
目的 研究伊麯康唑(ITC)單藥及聯閤化療藥多柔比星(ADM)對急性髓繫白血病細胞增殖及凋亡的影響.方法 活細胞計數試劑盒(CCK-8)法檢測不同濃度ITC、ADM單藥及ITC(2、6、15 μmol/L)聯閤ADM(0.30、0.75 μmol/L)對人急性髓繫白血病細胞株KG1α及急性髓繫白血病原代細胞的增殖抑製作用;不同濃度ITC持續處理的KG1 α細胞,分彆于7、14、21 d觀察對集落剋隆形成的影響;ADM 0.75 μmol/L、ITC 6 μmol/L及二者聯閤作用KG1α細胞48 h後,瑞氏染色法觀察細胞形態變化,流式細胞術檢測細胞凋亡率、線粒體膜電位及細胞週期阻滯情況,Western印跡檢測Sonic Hedgehog(Shh)信號通路相關蛋白Shh和膠質瘤相關癌基因同繫物1(Gli1)的錶達水平.結果 ITC及ADM對KG1α細胞及急性髓繫白血病原代細胞均具有增殖抑製作用,併且ITC可降低KG1α剋隆形成能力,均呈劑量依賴.Weeb繫數檢驗,ADM0.75 μmol/L與ITC 6 μmol/L兩藥聯閤時,細胞實際存活率≤70％細胞預估存活率,有協同效應;聯閤組與對照組和單藥組相比,KG1 α細胞形態變化更為明顯.ADM 0.75 μmol/L與ITC 6μmol/L聯閤誘導KG1α細胞48 h的凋亡率為31.72％±1.58％,均顯著高于對照組、ITC單藥組和ADM單藥組(4.17％±0.74％、4.33％±1.12％、9.53％±1.15％,均P＜0.01).線粒體膜電位檢測中,聯閤組的低紅色熒光細胞(P3)比例明顯高于對照組、ADM和ITC單藥組(18.80％±0.96％比5.00％±0.38％、9.70％±0.43％、7.10％±0.77％,均P＜0.01).ADM 0.75 μmol/L與ITC 6 μmol/L聯閤組與對照組及單藥組比明顯阻滯細胞于G2期(P＜0.05).Western印跡結果顯示Shh和Gli1的錶達水平隨著ITC濃度增大而下降.結論 ITC與ADM閤用可明顯抑製細胞的增殖,增加KG1 α細胞凋亡率、線粒體的損傷及細胞G2期的阻滯;ITC可以抑製Shh信號通路.
목적 연구이곡강서(ITC)단약급연합화료약다유비성(ADM)대급성수계백혈병세포증식급조망적영향.방법 활세포계수시제합(CCK-8)법검측불동농도ITC、ADM단약급ITC(2、6、15 μmol/L)연합ADM(0.30、0.75 μmol/L)대인급성수계백혈병세포주KG1α급급성수계백혈병원대세포적증식억제작용;불동농도ITC지속처리적KG1 α세포,분별우7、14、21 d관찰대집락극륭형성적영향;ADM 0.75 μmol/L、ITC 6 μmol/L급이자연합작용KG1α세포48 h후,서씨염색법관찰세포형태변화,류식세포술검측세포조망솔、선립체막전위급세포주기조체정황,Western인적검측Sonic Hedgehog(Shh)신호통로상관단백Shh화효질류상관암기인동계물1(Gli1)적표체수평.결과 ITC급ADM대KG1α세포급급성수계백혈병원대세포균구유증식억제작용,병차ITC가강저KG1α극륭형성능력,균정제량의뢰.Weeb계수검험,ADM0.75 μmol/L여ITC 6 μmol/L량약연합시,세포실제존활솔≤70％세포예고존활솔,유협동효응;연합조여대조조화단약조상비,KG1 α세포형태변화경위명현.ADM 0.75 μmol/L여ITC 6μmol/L연합유도KG1α세포48 h적조망솔위31.72％±1.58％,균현저고우대조조、ITC단약조화ADM단약조(4.17％±0.74％、4.33％±1.12％、9.53％±1.15％,균P＜0.01).선립체막전위검측중,연합조적저홍색형광세포(P3)비례명현고우대조조、ADM화ITC단약조(18.80％±0.96％비5.00％±0.38％、9.70％±0.43％、7.10％±0.77％,균P＜0.01).ADM 0.75 μmol/L여ITC 6 μmol/L연합조여대조조급단약조비명현조체세포우G2기(P＜0.05).Western인적결과현시Shh화Gli1적표체수평수착ITC농도증대이하강.결론 ITC여ADM합용가명현억제세포적증식,증가KG1 α세포조망솔、선립체적손상급세포G2기적조체;ITC가이억제Shh신호통로.
Objective To explore the effects of itraconazole (ITC) plus adriamycin (ADM) on proliferation and apoptosis of acute myeloid leukemia cells in vitro.Methods The growth inhibition effects of different concentrations of ITC,ADM or ITC (2,6,15 μmol/L) plus ADM (0.30,0.75 μmol/L) were detected by CCK8 assay in KG1 α and primary adult acute leukemia cells.Different concentrations of ITC for 7,14 and 21 days were applied for observing the effect on colony formation.After 48 h treatments with 6 μmol/L ITC,0.75 μmol/L ADM or ITC (6 μmol/L) plus ADM (0.75 μmol/L),the morphological changes of cells were observed by Wright staining.Flow cytometry was used to detect cell apoptotic rate,mitochondrial membrane potential and cell cycle arrest.And the expression levels of Sonic Hedgehog (Shh) signal pathway-related proteins Shh and glima-associated oncogene homdog1 (Gli1) were determined by Western blot.Results ITC and ADM inhibited the proliferation of KG1 α and primary adult acute leukemia cells and ITC reduced the colony-formation ability of KG1 α cells both in dose-dependent manners.Compared with control or single drug group,the changes of cell morphology were more apparent in combined group.Weeb coefficient test revealed a synergistic effect (D≤70％C) of 0.75 μmol/L ADM plus 6 μmol/L ITC.When KG1α cells were treated with 6 μmol/L ITC plus 0.75 μmol/L ADM,the apoptotic rate was 31.72％ ± 1.58％.And it was significantly higher than that in control,ITC and ADM groups (4.17％ ± 0.74％,4.33％ ± 1.12％,9.53％ ± 1.15％,P ＜0.01).Detection of mitochondrial membrane potential showed that low red-fluorescent cells (P3) of combined group were obviously higher than that in control,ADM and ITC single drug groups (18.80％ ±0.96％ vs 5.00％ ±0.38％,9.70％ ±0.43％,7.10％ ± 0.77％,P ＜0.01).KG1α cells were obviously arrested in G2 phase in combined group and it showed no statistical significance compared with control or single drug group.Western blot showed that the expression levels of Shh and Gli1 decreased with rising concentration of ITC.Conclusions ITC plus ADM can obviously inhibit cell proliferation and increase KG1o cell apoptotic rate,mitochondrial damage and G2 phase retardation.And ITC inhibits the Shh signal pathways.
