Two-Dimensional (Polyacrylamide) Gel Electrophoresis Analysis of Apoptosis Induced by Harringtonine in K562 Cells

摘要

OBJECTIVE The anti-tumor drug, harringtonine (HT), has been extensively used with satisfactory results in the treatment of acute or chronic myeloid leukemia. Previous studies have shown that the anti -tumor activity of the drug is related to induced apoptosis of tumor cells, but the molecular mechanism still remains unclear. The main purpose of this research was to analyze the protein profiles formed during HT-induced apoptosis in K562 cells and to screen the apoptotic-related proteins. METHODS Annexin V and PI double staining was used in combination with flow cytometry to examine the early and the late stages of HT -induced apoptosis in K562 cells. In addition two -dimensional gel electrophoresis and computer -assisted image analysis were employed to separate and compare the HT -induced apoptotic proteins of the K562 cells and the controls. RESULTS When a concentration of 10 μg/ml HT was used to treat K562 cells, the percentage of the early-apoptotic cells (Annexin V~+/PI~-) was found to be 28.3% and 18.1% at 5 and 24 h, respectively (P < 0.01), while the rate of late-apoptotic cells (Annexin V~+/PI~+ ) was at a level of 9.1% and 20.2% , respectively (P < 0.01). Matching analysis of the proteome among the control group and the early- and late-apoptotic groups showed 1,300 ± 50 protein spots which were identified in the control K562 cells with a matching rate of 88.3 ± 2.0 % for the protein spots in the two treated groups. Ten protein spots showed overt and steady changes in both quality and quantity in the cells of the late-apoptotic group (P < 0.01), among which the level of expression for eight of the ten protein spots was up -regulated after apoptosis, one was down-regulated and one was merely expressed as in the control cells. CONCLUSION The proteins with differential expression might be important proteins involved in the process of apoptosis in K562 cells induced by HT.