国际外科学杂志
國際外科學雜誌
국제외과학잡지
INTERNATIONAL JOURNAL OF SURGERY
2013年
5期
310-314,封3
,共6页
夏咸军%马芳%张丽军%刘保池
夏鹹軍%馬芳%張麗軍%劉保池
하함군%마방%장려군%류보지
热休克蛋白90%17-二甲胺乙胺基-17-去甲氧基格尔德霉素%细胞增生%细胞凋亡
熱休剋蛋白90%17-二甲胺乙胺基-17-去甲氧基格爾德黴素%細胞增生%細胞凋亡
열휴극단백90%17-이갑알을알기-17-거갑양기격이덕매소%세포증생%세포조망
Heat shock protein 90%17-DMAG%Cell proliferation%Apoptosis
目的 观察17-二甲胺乙胺基-17-去甲氧基格尔德霉素(17-dimethylaminoethylamino-17-demethoxygeldanamyein,17-DMAG)对人结肠癌HT-29细胞增生抑制及诱导其凋亡的作用.方法 用CCK-8检测17-DMAG对结肠癌HT-29细胞增生的影响；DAPI染色在倒置荧光显微镜下观察17-DMAG作用于HT-29细胞24 h后细胞核的形态；AnnexinV-FITC/PI双染法流式细胞检测细胞凋亡率；免疫印迹实验分析Caspase-3蛋白表达水平变化.结果 17-DMAG呈时间-剂量-依赖性抑制HT-29细胞增生.0.1μmol/L、0.25μmol/L 、0.5μmol/L、1.0μmol/L、2.5μmol/L和0.5μmol/L作用于HT-29细胞24 h后,细胞增生抑制率分别为(14.36±0.95)％、(22.17±1.15)％、(28.45±1.16)％、(35.04±1.58)％、(46.85±2.44)％和(57.19±2.06)％；作用48h后,细胞增生抑制率分别为(20.80±1.17)％、(27.55±0.65)％、(33.33±1.23)％、(46.20±4.76)％、(55.45±4.47)％和(61.75±2.72)％；作用72h,细胞增生抑制率分别为(29.62±2.27)％、(39.19±1.74)％、(44.29±2.00)％、(50.66±2.17)％、(58.84±3.18)％和(70.74±2.65)％.DAPI染色倒置荧光显微镜观察0.25μmol/L、0.5μmol/L、1.0μmol/L与2.5μmol/L的17-DMAG作用HT-29细胞24h,均可见到细胞凋亡的改变.AnnexinV-FITC双染法检测结果显示,正常对照组HT-29细胞24h的自然总凋亡率(早期+晚期)为(2.72±0.57)％,0.25μmol/L、0.5μmol/L、1.0μmol/L和2.5μmol/L 17-DMAG干预24 h后细胞总凋亡率分别为(5.38±0.46)％、(6.88±0.52)％、(10.44±0.32)％与(17.87±4.66)％,不同浓度组的细胞凋亡率比较差异有统计学意义(P＜0.05).17-DMAG药物作用HT-29细胞24h,随着药物剂量的增加,cleaved Caspase-3的表达有增加趋势.结论 17-DMAG体外呈时间-剂量依赖性抑制HT-29细胞增生,诱导其通过Caspase-3途径凋亡.
目的 觀察17-二甲胺乙胺基-17-去甲氧基格爾德黴素(17-dimethylaminoethylamino-17-demethoxygeldanamyein,17-DMAG)對人結腸癌HT-29細胞增生抑製及誘導其凋亡的作用.方法 用CCK-8檢測17-DMAG對結腸癌HT-29細胞增生的影響；DAPI染色在倒置熒光顯微鏡下觀察17-DMAG作用于HT-29細胞24 h後細胞覈的形態；AnnexinV-FITC/PI雙染法流式細胞檢測細胞凋亡率；免疫印跡實驗分析Caspase-3蛋白錶達水平變化.結果 17-DMAG呈時間-劑量-依賴性抑製HT-29細胞增生.0.1μmol/L、0.25μmol/L 、0.5μmol/L、1.0μmol/L、2.5μmol/L和0.5μmol/L作用于HT-29細胞24 h後,細胞增生抑製率分彆為(14.36±0.95)％、(22.17±1.15)％、(28.45±1.16)％、(35.04±1.58)％、(46.85±2.44)％和(57.19±2.06)％；作用48h後,細胞增生抑製率分彆為(20.80±1.17)％、(27.55±0.65)％、(33.33±1.23)％、(46.20±4.76)％、(55.45±4.47)％和(61.75±2.72)％；作用72h,細胞增生抑製率分彆為(29.62±2.27)％、(39.19±1.74)％、(44.29±2.00)％、(50.66±2.17)％、(58.84±3.18)％和(70.74±2.65)％.DAPI染色倒置熒光顯微鏡觀察0.25μmol/L、0.5μmol/L、1.0μmol/L與2.5μmol/L的17-DMAG作用HT-29細胞24h,均可見到細胞凋亡的改變.AnnexinV-FITC雙染法檢測結果顯示,正常對照組HT-29細胞24h的自然總凋亡率(早期+晚期)為(2.72±0.57)％,0.25μmol/L、0.5μmol/L、1.0μmol/L和2.5μmol/L 17-DMAG榦預24 h後細胞總凋亡率分彆為(5.38±0.46)％、(6.88±0.52)％、(10.44±0.32)％與(17.87±4.66)％,不同濃度組的細胞凋亡率比較差異有統計學意義(P＜0.05).17-DMAG藥物作用HT-29細胞24h,隨著藥物劑量的增加,cleaved Caspase-3的錶達有增加趨勢.結論 17-DMAG體外呈時間-劑量依賴性抑製HT-29細胞增生,誘導其通過Caspase-3途徑凋亡.
목적 관찰17-이갑알을알기-17-거갑양기격이덕매소(17-dimethylaminoethylamino-17-demethoxygeldanamyein,17-DMAG)대인결장암HT-29세포증생억제급유도기조망적작용.방법 용CCK-8검측17-DMAG대결장암HT-29세포증생적영향；DAPI염색재도치형광현미경하관찰17-DMAG작용우HT-29세포24 h후세포핵적형태；AnnexinV-FITC/PI쌍염법류식세포검측세포조망솔；면역인적실험분석Caspase-3단백표체수평변화.결과 17-DMAG정시간-제량-의뢰성억제HT-29세포증생.0.1μmol/L、0.25μmol/L 、0.5μmol/L、1.0μmol/L、2.5μmol/L화0.5μmol/L작용우HT-29세포24 h후,세포증생억제솔분별위(14.36±0.95)％、(22.17±1.15)％、(28.45±1.16)％、(35.04±1.58)％、(46.85±2.44)％화(57.19±2.06)％；작용48h후,세포증생억제솔분별위(20.80±1.17)％、(27.55±0.65)％、(33.33±1.23)％、(46.20±4.76)％、(55.45±4.47)％화(61.75±2.72)％；작용72h,세포증생억제솔분별위(29.62±2.27)％、(39.19±1.74)％、(44.29±2.00)％、(50.66±2.17)％、(58.84±3.18)％화(70.74±2.65)％.DAPI염색도치형광현미경관찰0.25μmol/L、0.5μmol/L、1.0μmol/L여2.5μmol/L적17-DMAG작용HT-29세포24h,균가견도세포조망적개변.AnnexinV-FITC쌍염법검측결과현시,정상대조조HT-29세포24h적자연총조망솔(조기+만기)위(2.72±0.57)％,0.25μmol/L、0.5μmol/L、1.0μmol/L화2.5μmol/L 17-DMAG간예24 h후세포총조망솔분별위(5.38±0.46)％、(6.88±0.52)％、(10.44±0.32)％여(17.87±4.66)％,불동농도조적세포조망솔비교차이유통계학의의(P＜0.05).17-DMAG약물작용HT-29세포24h,수착약물제량적증가,cleaved Caspase-3적표체유증가추세.결론 17-DMAG체외정시간-제량의뢰성억제HT-29세포증생,유도기통과Caspase-3도경조망.
Objective To investigate the effects of Hsp90 inhibitor 17-DMAG on proliferation and apoptosis of human colon cancer cell line HT-29.Methods HT-29 cells were treated with 17-DMAG.The cell proliferation inhibition rate was evaluated by CCK-8 assay.Apoptosis of HT-29 cells by 17-DMAG was delineated by DAPI staining assay and Annexin V PI double labeling FCM was used to determine cell apoptotic rate.Furthermore,Westen blotting analysis was used to determine caspase-3 and cleaved caspase-3 protein expression.Results 17-DMAG time-dose-dependently inhibited the proliferation of HT-29 cells.After 0.1μmol/L,0.25μmol/L,0.5μmol/L,1.0μmol/L,2.5μmol/L and 5μmol/L 17-DMAG exposured for 24 hours,the cell proliferation inhibition rate was (14.36±0.95)％,(22.17± 1.15)％,(28.45±1.16)％,(35.04±1.58)％,(46.85 ±2.44)％,(57.19 ± 2.06)％ respectively,after exposured for 48 hours,the cell proliferation inhibition rate was increased to (20.80±1.17)％,(27.55 ±0.65)％,(33.33 ±1.23)％,(46.20±4.76)％,(55.45 ±4.47)％,(61.75 ±2.72) ％ respectively,after exposure for 72 hours,the cell proliferation inhibition rate was to (29.62 ± 2.27) ％,(39.19 ± 1.74)％,(44.29 ±2.00)％,(50.66 ±2.17)％,(58.84 ±3.18)％,(70.74 ±2.65)％.DAPI staining showed that HT-29 cells treated with 17-DMAG displayed chromatin condensation and nuclear fragmentation which are typical changes of apoptosis.Annexin V PI double labeling FCM showed that when HT-29 cells were exposed to 0,0.25,0.5,1.0 and 2.5(μmoL/L) 17-DMAG for 24 hours,the total apoptotic rate for 24 hours was (2.72 ±0.57)％,(5.38 ±0.46)％,(6.88 ±0.52)％,(10.44 ±0.32)％ and (17.87 ±4.66)％ respectively.(P ＜0.05).In addition,the expression of procaspase-3 decreased,while cleaved caspase-3 increased in the presence of 17-DMAG at different concentrations for 24 hours.Conclusion 17-DMAG can time-dose-dependently inhibit proliferation and promote apoptosis of HT-29 cells in vitro.
