p21(WAF1) transfection decreases sensitivity of K562 cells to VP-16

The objective of this study was to explore the effect of p21(WAF1) on the sensitivity to chemotherapeutic agent VP-16, etoposide, in leukemia cell line K562. A p21(WAF1) retroviral expression vector was constructed, and mediated by FuGENE(TM)6, it was transfected into K562 cells, which without p21(WAF1) expression. The ecotropic expression of p21(WAF1) in K562 cells was identified by RT-PCR and Western blot, and named K562-p21(WAF1) cell. The K562-p21(WAF1) cells were exposed to VP-16 at different concentrations and different times, then, the sensitivity to VP-16 was examined by cell viability and MTT assay, the apoptosis induced by VP-16 was examined by DNA fragments electrophoresis and flow cytometric Annexin V-PI dual labeling technique. The results showed that the ecotropic expression of p21(WAF1) decreased the sensitivity of K562 cells to VP-16. After treatment of VP-16, the DNA ladder was examined in control K562-neo cells at 20 microgram/ml, but in K562-p21(WAF1) cells at 80 microgram/ml. With FCM, the number of apoptotic K562-neo cells was 14.9% and 25.4% respectively after treated with 20 microgram/ml VP-16 for 12 and 24 hours, and it was 6.94% and 10.96% in K562-p21(WAF1) cells. Our results suggest that the expression of p21(WAF1) could reduce the sensitivity of K562 cells to VP-16 and inhibit the apoptosis of K562 cells