OBJECTIVE: To screen and identify apoptosis related proteins and explore the mechanism of harringtonine (HT)-induced K562 cells apoptosis. METHODS: Flow cytometry was used to distinguish K562 cells in the earlier stage of apoptosis from those in the later stage of apoptosis by annexin V and PI staining. Two-dimensional polyacrylamide gel electrophoresis (2-D PAGE) coupled with computer image analysis was used to detect the changes in protein expression in the two stages of apoptosis. Proteins were identified by peptide mass fingerprint in combination with database searching. RESULTS: K562 cells treated with HT for 5 and 24 hours were in the early and later stages of apoptosis respectively. Statistical analysis showed 3 spots disappeared, 7 spots with decreased intensity and 10 spots with increased intensity in the 24 h HT induced apoptotic cells as compared with that in 5 h HT induced ones. Ten spots were selected on the basis of the intensity and the significant changes in abundance. Among them, 5 apoptosis related proteins were successfully identified by MALDI-TOF: keratin 9, BTF3, TrpRS, RS and prohibitin. CONCLUSIONS: Up-regulation of TrpRS, RS, prohibitin and down-regulation of BTF3 were involved in inhibition of transcription and protein synthesis in the apoptotic K562 cells induced by HT, whereas up-regulation of keratin 9 was related to apoptosis resistance.
OBJECTIVE: To screen and identify apoptosis related proteins and explore the mechanism of harringtonine (HT)-induced K562 cells apoptosis. METHODS: Flow cytometry was used to distinguish K562 cells in the earlier stage of apoptosis from those in the later stage of apoptosis by annexin V and PI staining. Two-dimensional polyacrylamide gel electrophoresis (2-D PAGE) coupled with computer image analysis was used to detect the changes in protein expression in the two stages of apoptosis. Proteins were identified by peptide mass fingerprint in combination with database searching. RESULTS: K562 cells treated with HT for 5 and 24 hours were in the early and later stages of apoptosis respectively. Statistical analysis showed 3 spots disappeared, 7 spots with decreased intensity and 10 spots with increased intensity in the 24 h HT induced apoptotic cells as compared with that in 5 h HT induced ones. Ten spots were selected on the basis of the intensity and the significant changes in abundance. Among them, 5 apoptosis related proteins were successfully identified by MALDI-TOF: keratin 9, BTF3, TrpRS, RS and prohibitin. CONCLUSIONS: Up-regulation of TrpRS, RS, prohibitin and down-regulation of BTF3 were involved in inhibition of transcription and protein synthesis in the apoptotic K562 cells induced by HT, whereas up-regulation of keratin 9 was related to apoptosis resistance.