The Study Of Hydroxycamptothecin-Induced Apoptosis Of Cell Line MUTZ-1 And The Mechanisms In The Human Myelodysplastic Syndromes

Objective To study the inductive effect and the possible mechanisms of hydroxycamptothecine(HCPT) on MUTZ-1 apoptosis in myelodysplastic syndromes (MDS), which could possibly be the theoretical basis of treatment of MDS in clinical practice.Methods Method of methyl thiazolyl tetrazolium (MTT) was applied to study the growth inhibition of MUTZ-1 cells induced by HCPT. The cell apoptosis and its cycle changes induced by HCPT were analyzed with flow cytometry (FCM). Apoptotic morphological features were observed through microscope and transmission electronmicroscope. The mRNA expression level of Survivin, B-cell lymphoma/leukemia 2 (Bcl-2), Bcl-2 associated X protein (Bax) in cells treated with HCPT is determined by semi-quantitative reverse transcriptase polymerase chain reaction (RT-PCR).Results 1 HCPT can inhibit the growth of MUTZ-1 cells. There are significant differences in inhibition of proliferation between groups with treatment of HCPT of different concentrations (F=10.24,p<0.0001) and different periods (F=3.78,p=0.009). 2 HCPT can induce apoptosis of MUTZ-1 cells. There are significant differences in apoptosis rates between groups with treatment of HCPT of different concentrations(F=16.35,p<0.0001) and different periods (F=57.93,p<0.0001). 3 Through microscope, typical apoptotic morphological features were observed followed by treatment of HCPT, such as condensation of cells and nuclear chromatin, disintegration of nuclear chromatin. Through transmission electronmicroscope, aggregation and margination of apoptotic nuclear chromatin could be observed. Some parts of the nuclear membrane disappeared; the concentration of plasm increased and chromatin mass of irregular size could be seen in the plasm. 4 After treatment by HCPT of different concentrations for 24 h, compared with that of the control group, thepercentage of cells in the S phase increased. The majority of the cells were remained in S phage. 5 Under certain condition, there were no significant changes in the mRNA expression of Bcl-2 and Bax after treatment by HCPT of different concentrations(p>0.05) and different periods, while mRNA expression of survivin decreased(p<0.05).Conclusions HCPT can inhibit the growth of MUTZ-1 cells in dose- and time-dependent manners. HCPT can induce the apoptosis of MUTZ-1 cells in dose- and time-dependent manners. Typical apoptotic cells can be observed through microscope and transmission electronmicroscope. HCPT can change the cell cycle, the majority of the cells are detained in S phage. The apoptosis of the MUTZ-1 cells may be correlated with the down-regulation of the mRNA expression of Survivin.