伊马替尼体外诱导Kasumi-1细胞增殖、分化与凋亡作用

目的探讨酪氨酸激酶抑制剂伊马替尼诱导携带c-k it突变的细胞增殖、分化与凋亡的作用。方法应用MTT比色法观察伊马替尼对Kasum i-1细胞的生长抑制作用,流式细胞术分析细胞周期、c-k it抗原和髓系分化抗原CD11b、CD13和CD15的表达,AnnexinⅤ标记和DNA凝胶电泳分析细胞凋亡,应用W estern b lot方法分析Kasum i-1细胞c-k it蛋白酪氨酸磷酸化水平。结果伊马替尼呈时间和剂量依赖性抑制Kasum i-1细胞生长,72 h半数抑制浓度(IC50)为4.45μmol/L;伊马替尼5.00μmol/L使Kasum i-1细胞阻滞于G0/G1期,S期细胞减少;髓系分化抗原CD11b、CD13和CD15表达增加;作用24 h,早期凋亡细胞比例由9.04%升至86.84%(P<0.05),培养第5天出现明显DNA梯形条带;作用72 h,Kasum i-1细胞中c-k it蛋白酪氨酸磷酸化水平下降。结论酪氨酸激酶抑制剂伊马替尼能够抑制携带c-k it突变的细胞的生长,诱导细胞分化和凋亡。

Effect of tyrosine kinase inhibitor Imatinib mesylate on proliferation,differentiation and apoptosis of Kasumi-1 leukemia cell line

OBJECTIVE: To explore the effect of Imatinib mesylate on proliferation, differentiation and apoptosis of leukemic Kasumi-1 cells bearing c-kit mutation. METHODS: Kasumi-1 cells were treated with Imatinib at different concentrations in culture. Cell proliferation was assayed by MTT assay, expressions of c-kit antigen, surface myeloid antigen and cell cycle by flow cytometry, cell apoptosis by annexin V staining and agarose gel electrophoresis. Western blot was used to analyze the level of c-kit protein tyrosine phosphorylation. RESULTS: Imatinib treatment caused a time- and dose-dependent inhibition of the cell proliferation, with a 72 h IC50 of 4.45 micromol/L. Imatinib treatment induced a decrease in the mean fluorescence value of c-kit antigen, a progressive decline in S-phase cell fraction and an increase in G0/G1 cells. Treatment with 5.00 micromol/L of imatinib for 72 h induced an increase in expression of myeloid surface protein CD11, CD13 and CD15, and for 24 h induced an increase in early apoptosis cells [from 9.04% to 86.84% (P < 0.05)]. The apoptosis ladder was observed on agarose gel electrophoresis on 5-day treatment. Tyrosine phosphorylation level of c-kit protein was decreased by Imatinib treatment. CONCLUSION: Tyrosine kinase inhibitor Imatinib mesylate treatment could inhibit proliferation of Kasumi-1 cells which bear a c-kit mutation, induce differentiation, apoptosis and G0/G1 cells accumulation.