三氧化二砷联合硼替佐米对骨髓瘤细胞株增殖、凋亡及β-连环蛋白水平的影响

Effect of Arsenic Trioxide Combined with Bortezomib on Proli-feration,Apoptosis and β-Catenin Level in Myeloma Cell Lines

本研究探讨三氧化二砷(arsenic trioxide,AS2O3)联合硼替佐米对骨髓瘤细胞株增殖、凋亡及β-连环蛋白(β-catenin)水平的影响。硼替佐米、As2O3单用及联合处理RPMI8226、CZ-l、NCI-H929骨髓瘤细胞株48小时后,采用台盼蓝拒染法检测活细胞比例,用改良的四甲基偶氮唑蓝(MTT)法检测细胞增殖,AnnexinV/PI双染色流式细胞术检测凋亡细胞比例,免疫印迹法比较用前药后β-连环蛋白的水平。结果表明:硼替佐米对RPMI8226、CZ-l、NCI-H929的半数抑制浓度(IC50)为46.9、20.7、6.8nmol/L;As2O3(1μmol/L)联合硼替佐米(5nmol/L)作用后,活细胞比例分别由88.99%、72.23%、51.06%降至54.01%、39.59%、25.00%(p<0.05),凋亡细胞比例由(11.1±0.1)%、(26.8±1.7)%、(36.8±5.5)%增加为(36.1±2.2)%、(60.4±3.8)%、(76±5.6)%,两组Q值介于增强与明显增强之间(1.198-3.75)。联合用药组RPMI8226、CZ-l、NCI-H929胞浆β-连环蛋白水平与单药组相比,分别下降24.15%,31.85%,33.72%。结论:As2O3能增强硼替佐米对骨髓瘤细胞的增殖抑制和凋亡诱导,降低β-连环蛋白表达水平,提高骨髓瘤细胞对硼替佐米的敏感性。

The aim of this study was to investigate the effect of arsenic trioxide （ AS2O3 ） combined with bortezomib on the proliferation, apoptosis and β-catenin level in myeloma cell lines. Myeloma cell lines RPMI8226, CZ-1 and NCl- H929 were treated with As2O3 and bortezomib alone or in combination for 48 hours. Trypan blue dye exclusion and modified MIT were used to assess the cell viability. Flow cytometry with Annexin V-FITC and PI staining was used to detect the apoptosis rate. The β-catenin level was analyzed by Western blot. The results showed that IC50 of bortezomib to RPMI8226 .CZ-1 and NCI-H929 were 46.9,20.7 and 6.8 nmol/L, respectively. After the combination treatment with bortezomib （5 nmol/L） and As203 （1 μmol/L）, the cell viability of RPMI8226, CZ-1 and NCI-H929 decreased from 88. 99%, 72.23%, 51.06% to 54.01%, 39.59%, 25.00% （p 〈 0.05 ）, the apoptosis rate increased from 11. 1 ± 0.1%, 26.8 ± 1.7 %, 46.8 ± 5.5 % to 36.1 ± 2.2%, 60.4 ± 3.8%, 76 ± 5.6 % （p 〈 0.01 ） respectively. The Q value of two groups lies between enhancement and significant enhancement （ 1. 198 - 3.75 ）. Besides, β-catenin levels in tested cell lines were decreased to 24.15% ,31.85% ,33.72% of their basic constitutions respectively （p 〈0.05 ）. It is concluded that combination treatment of As2O3 and bortezomib can enhance the proliferation inhibition and apoptosis induction of bortezomib to myeloma ceil lines, reduce β-catenin level, and increase the sensitivity of myeloma cell lines to bortezomib.