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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 investigate the effects of bortezomib combined with natural killer (NK) cell killing and apoptosis induction in multiple myeloma cell line KM-3. Methods: The killing effect of NK cells on KM-3 cells was observed by WST-1 method. Annexin-Ⅴ, PI and CD45 triple immunofluorescence staining were used to detect the Annexin-Ⅴ + / PI- Changes of transmembrane potential of apoptotic cells and mitochondria. Results: The killing rates of KM-3 cells (P = 0.003) were significantly higher than that of NK cells treated with 5: 1, 10:1 and 20:1 NK cells in combination with 5nmol / L bortezomib at 12,24 and 48h (P = 0.002), which was significantly higher than that of NK cells alone (P <0.01). (P = 0.003), and the ratio of Annexin-V + / PI-cells was increased in a time-dependent manner (P = 0.002), and higher than NK cells alone, P <0.01. The mitochondrial transmembrane potential of KM-3 cells was significantly decreased (P = 0.025) at 6, 12 and 24 h after treatment with 5: 1, 10: 1 and 20: 1 NK cells in combination with 5 nmol / L bortezomib Time-dependent (P = 0.022), and lower than NK cells alone, P <0.05. CONCLUSION: Bortezomib combined with NK cells has a more significant killing effect and induces the apoptosis of KM-3 cells, suggesting a synergistic effect of bortezomib and NK cells.