Effect of small interfering RNA-mediated down-regulation expression of X-linked inhibitor of apoptosis protein in leukemic cell-lines on sensitivity to chemotherapy

Abstract

Objective To investigate the effect of small interfering RNA (siRNA)-mediated downregulated expression of X-linked inhibitor of apoptosis protein (XIAP) in leukemic cell-lines MOLT-4, HL-60 and K562 on the sensitivity to chemotherapy. Methods Real-time fluorescent quantitative PCR and Western blotting were used to detect the expression of XIAP mRNA and protein in leukemic cell-lines MOLT-4, HL-60,K562 and healthy peripheral blood mononuclear cells (PBMCs). The K562 cells with high expression of XIAP were transfected with XIAP siRNA (experimental group) or non-homologous siRNA (negative control group) by using NucleofectorTM nucleic acid transfection device, and those not transfected with any siRNA were assigned to blank control group. The expression levels of XIAP mRNA and protein in three groups were then detected.Meanwhile, the K562 cells of all groups were exposed to different concentrations of etoposide (vp-16: 0.01,0.1,1, 10, 100 mg/L)and Ara-C (0.1, 1, 10, 100, 1000, 10 000 mg/L)for determination of the cell growth inhibition rate with CC K-8 assay. Results The expression levels of XIAP mRNA and protein were elevated in MOLT-4,HL-60 and K562 cell lines compared with those in healthy PBMCs (all P＜0.05). Notably, K562 had the highest expression of XIAP mRNA and protein, which was statistically different from MOLT-4 and HL-60 (all P＜0.05 ).Compared with negative and blank control groups, the experimental group had a significant decline in the expressions of XIAP mRNA and protein after transfection with XIAP siRNA for 48 hours(mRNA: 0.37±0.10 vs 1.41 ±0.13 vs 1.00±0.12, protein: 0.37±0.03 vs 0.99±0.08 vs 0.98±0.07, all P＜0.05 ). The cell growth inhibition rate in the experimental group was remarkably different from those in negative and blank control groups when treated with 1 mg/L vp-16, 10 mg/L and 100 mg/L Ara-c (all P＜0.05 ), whereas no statistical difference was found among three groups when treated with other doses of vp-16 and Ara-c (all P＞0.05 ). Conclusion XIAP siRNA may specifically down-regulate the expressions of XIAP mRNA and protein in K562 cells and thereby enhance the sensitivity of K562 cells to vp-16 and Ara-C of chemotherapy. Key words: RNA, small interfering; Leukemic cell lines; X-linked inhibitor of apoptosis protein; Antineoplastic agents; Drug sensitivity