Effects of heat shock protein 90 inhibitor 17-DMAG on proliferation and apoptosis of colon cancer cell line HT-29

Abstract

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. Key words: Heat shock protein 90 ; 17-DMAG ; Cell proliferation ; Apoptosis