硼替佐米联合NK细胞对骨髓瘤细胞株KM-3作用的影响

Influence of bortezomib and NK cells on multiple myeloma cell line KM-3

目的:探讨硼替佐米联合自然杀伤(NK)细胞杀伤及诱导多发性骨髓瘤细胞株KM-3凋亡的作用。方法:WST-1法观察加入硼替佐米后,NK细胞对KM-3细胞杀伤作用的变化;Annexin-Ⅴ、PI及CD45三重免疫荧光标记,流式细胞术检测Annexin-Ⅴ+/PI-凋亡细胞及线粒体跨膜电位的变化。结果:效靶比为5∶1、10∶1和20∶1的NK细胞联合5nmol/L硼替佐米处理后12、24和48h,均显著杀伤KM-3细胞(P=0.003),杀伤率的升高呈时间依赖性(P=0.002),且显著高于NK细胞单独处理,P<0.01。效靶比为5∶1、10∶1和20∶1的NK细胞联合5nmol/L硼替佐米处理,Annexin-Ⅴ+/PI-细胞比例均增加(P=0.003),呈时间依赖性(P=0.002),且高于NK细胞单独处理,P<0.01。效靶比为5∶1、10∶1和20∶1的NK细胞联合5nmol/L硼替佐米处理后6、12和24h,KM-3细胞线粒体跨膜电位明显降低(P=0.025),呈时间依赖性(P=0.022),且低于NK细胞单独处理,P<0.05。结论:硼替佐米联合NK细胞具有更显著地杀伤并诱导KM-3细胞凋亡的作用,提示硼替佐米与NK细胞具有协同作用。

OBJECTIVE：To study the killing and inducing apoptotic effect of bortezomib and NK cells on multiple myeloma cell line KM-3. METHODS：WST-1 assay was used to observe the changes of the killing effect of KM-3 cells induced by NK cells and bortezomib. The changes of Annexin-Ⅴ＋/PI-apoptotic cells and the mitochondrial transmembrane potential were examined by triplicate immunofluorescence of Annexin-Ⅴ,PI and CD45 and flowcytometry. RESULTS：NK cells （E：T ratio at 5：1,10：1,20：1） with 5 nmol/L bortezomib could significantly kill KM-3 cells after 12,24 and 48 hours （P=0.003）. The growth killing rates increased in time dependence （P=0.002）,which were higher than those of NK cells at the same time （P0.01）. The Annexin-Ⅴ＋/PI-cells increased in time dependence （P=0.002） and in dose dependence （P=0.003） when treated with NK cells （E：T ratio at 5：1,10：1,20：1） and 5 nmol/L bortezomib,which were higher than those of NK cells at the same time （P0.01）. After 6,12 and 24 hours,the mitochondrial transmembrane potential of KM-3 cells treated with NK cells （E：T ratio at 5：1,10：1,20：1） and 5 nmol/L bortezomib significantly decreased （P=0.025） in time dependence （P=0.022）,and the decreasing rates were higher than those of NK cells at the same time （P0.05）. CONCLUSION：Bortezomib with NK cells can kill KM-3 cells and induce them apoptosis more significantly,which indicates that bortezomib and NK cells are synergistic.