HHT-Induced Cell Apoptosis Is Associated with Reduction of Survivin mRNA Expression.

The inhibitor of apoptosis (IAP) gene family, which was discovered less than a decade ago, encodes a group of structurally related proteins that, in addition to their ability to suppress apoptotic cell death, are involved in an increasing number of seemingly unrelated cellular functions. The IAP families have evolved as highly conserved regulators of cell death. Homoharringtonine(HHT) is a plant alkaloid with antileukemia activity which is currently tested for treatment of acute and chronic leukemia. Our previous study suggested that HHT could induced apoptosis of MUTZ-1 in vitro, but little is know about the possible molecular mechanisms of HHT induced cell apoptosis. The study on the mechanisms of HHT in regulation of apoptosis will give us insight into further understanding of the role HHT. The data generated from this study will also provide theoretical ground for making use of HHT in clinical treatment. In this study, five cell lines including MUTZ-1, K562, Jurkat, RMPI and HL60 were used in the experiments to detect the effects of HHT on the induction of apoptosis and to study the possible mechanisms of HHT in regulation of apoptosis of the cells. Cell apoptosis were observed by flow cytometry (FCM). Cell apoptotic morphology was observed by transmission electron microscope. Semi-quantitative RT-PCR was used to evaluate the mRNA expression of survivin, XIAP, Bcl-2 and Bax in these cells. Our results demonstrated that HHT was capable of inducing apoptosis in all the five cell lines, with an increasing apoptotic rate in the order of K562, MUTZ-1/RMPI, and Jurkat/HL60, at a HHT concentration of 0.1 μg/ml for 12 hours (p<0.05). Survivin mRNA was not detected without HHT treatment but was detected in all the five cell lines after induction with HHT. Interestingly, the level of survivin mRNA expression was detected in the same but reversed order as with apoptotic rate: cell lines showed lowest apoptotic rate had highest survivin mRNA level, indicating a negative correlation between the two (p=0.003). The expression of Bcl-2 and Bax mRNA showed no significant correlation to HHT-induced apoptotic rate. There was no significant difference in XIAP expression in the 5 cell lines. We conclude that HHT might act via reduction of survivin expression and the level of survivin mRNA may serve as a predictor for chemotherapy sensitivity to HHT.