奥佐米星对急性髓细胞性白血病的细胞毒性及其与CD33表达水平的关系研究

Study on cytotoxicity of Gemtuzumab Ozogamicin in acute myeloid leukemia and its relationship with expression level of CD33

目的探索奥佐米星(GO)对急性髓细胞性白血病(AML)细胞毒性与CD33表达水平的关系及作用机制。方法选取人髓系OCI-AML3和KG-1a细胞,采用含CD33质粒的慢病毒对其进行转染,然后根据CD33表达量将细胞分为CD33高表达组和CD33低表达组。两组均采用15μg/L的GO进行处理,然后通过MTT实验测定两组细胞的存活率。采用不同浓度(0、5、10、15μg/L)的GO处理OCI-AML3细胞,然后采用蛋白质印迹法检测细胞的凋亡相关蛋白Bcl-2、cleaved-caspase 3、caspase 3的表达水平。结果转染后,OCI-AML3和KG-1a细胞表面CD33表达水平高于转染前,差异有统计学意义(P<0.05)。相同浓度的GO处理后,CD33高表达组的OCI-AML3和KG-1a细胞存活率均低于CD33低表达组,差异有统计学意义(P<0.05)。GO浓度为5、10、15μg/L时,OCI-AML3细胞Bcl-2水平低于GO浓度为0时,cleaved-caspase 3、caspase 3水平均高于GO浓度为0时;GO浓度为10、15μg/L时OCI-AML3细胞Bcl-2水平低于GO浓度为5μg/L时,cleaved-caspase 3、caspase 3水平均高于GO浓度为5μg/L时;GO浓度为15μg/L时,OCI-AML3细胞Bcl-2水平低于GO浓度为10μg/L时,cleaved-caspase 3、caspase 3水平均高于GO浓度为10μg/L时,差异有统计学意义(P<0.05)。结论GO对CD33高表达的人髓系细胞增殖的抑制作用增强,可通过调控凋亡蛋白的表达发挥促进AML细胞凋亡的作用。

Objective To explore the relationship between the cytotoxicity of Gemtuzumab Ozogamicin(GO)on acute myeloid leukemia(AML)cells and the expression level of CD33 and the mechanism of action.Methods Human myeloid OCI-AML3 and KG-1a cells were transfected with lentivirus containing CD33 plasmid,then the cells were divided into CD33 high expression group and CD33 low expression group according to CD33 expression level.Both groups were treated with 15μg/L GO,and then the survival rate of cells in the two groups was determined by MTT assay.OCI-AML3 cells were treated with different concentrations of GO(0,5,10,15μg/L),and then the expression levels of apoptosis-related proteins Bcl-2,cleaved caspase 3,and caspase 3 were detected by Western blot.Results After transfection,the expression levels of CD33 on OCI-AML3 and KG-1a cells were higher than that before transfection,and the differences were statistically significant(P<0.05).After the same concentration of GO treatment,the survival rates of OCI-AML3 and KG-1a cells in CD33 high expression group were lower than those in CD33 low expression group,the differences were statistically significant(P<0.05).When GO concentration was 5,10,15μg/L,the Bcl-2 level of OCI-AML3 cells was lower than those when GO concentration was 0,and cleaved-caspase 3 and caspase 3 levels were higher than those when GO concentration was 0.When GO concentration was 10,15μg/L,the Bcl-2 level of OCI-AML3 cells was lower than that when GO concentration was 5μg/L,and cleaved-caspase 3 and caspase 3 levels were higher than those when GO concentration was 5μg/L.When GO concentration was 15μg/L,the Bcl-2 level of OCI-AML3 cells was lower than those when GO concentration was 10μg/L,and cleaved-caspase 3 and caspase 3 levels were higher than those when GO concentration was 10μg/L,the differences were statistically significant(P<0.05).Conclusion GO has enhanced inhibitory effect on proliferation of human myeloid cells with high CD33 expression,and can promote apoptosis of AML cells by regulating the expression of apoptotic proteins.