目的 观察载小干扰RNA(siRNAs)纳米微粒沉默B细胞淋巴瘤/白血病-2(bcl-2)基因联合纳米砷剂协同抗急性髓性白血病效应.方法 凝胶阻滞电泳、荧光显微镜、流式细胞仪及噻唑蓝(MTT)法分别用于评估正聚乙二醇(PEG)-多聚赖氨酸(PLL)负载siRNAs能力、细胞转染效率及对细胞活力影响,Western blot法观察siRNAs沉默靶基因,MTT法评估siRNAs联合纳米砷剂对U937协同抑制效应.结果 N/P-10时,PEG-PLL和siRNAs基本复合,U937细胞活力为(85.06±5.80)％,转染效率为(83.70±0.37)％,转染后bcl-2蛋白表达下降为(44.1 l±4.39)％.空载体、纯砷单药、纳米As单药及联合用药组细胞活力分别为(99.44±1.45)％、(87.76±2.21)％、(92.98±4.34)％、(67.93±8.16)％(P＜0.05)(砷1.25 μmol/L);(99.56±2.53)％、(54.08±4.46)％、(57.85±2.11)％、(38.60±6.24)％(P＜0.05)(砷2.50 μmol/L); (98.88±1.59)％、(37.62±1.38)％、(51.31±3.14)％、(28.92±4.97)％(p＜0.05)(砷5.00 μmol/L);(97.72±2.55)％、(29.44±4.14)％、(40.18±3.72)％、(20.56±4.97)％(P＜0.05)(砷10.00 μmol/L); (96.65±2.03)％、(21.49±1.60)％、(26.34±0.97)％、(15.90±1.70)％(P＜0.05)(砷20.00 μmol/L).结论 PEG-PLL对U937细胞毒性较低、转染效率较高,PEG-PLL/siRNAs沉默bel-2联合纳米As获得协同抗白血病效应.%Objective To observed the synergetic inhibitory effects on human acute myelogenous leukemia (AML) by nanoparticle-mediated small interfering RNA (siRNAs) and arsenic therapy in vitro.Methods Gel retardation assay,fluorescence microscopy,flow cytometry assay and methyl thiazol tetrazolium (MTT) assay was performed to evaluate siRNAs complexation capacity,transfection efficiency and in-fluence on cell viability of polyethylene glycol (PEG)-poly (L-lysine) (PLL).The down regulation of B cell lymphoma/leukemia-2 (bcl-2) gene expression was comfirmed by Western blotting assay.The MTT assay was further performed to evaluate the synergetic inhibitory effects on U937 cells by siRNAs and arse-nic nanoparticles.Results At N/P ratio of 10,PEG-PLL and siRNAs complexed completely,the cell viability was (85.06 ± 5.80) ％,the transfection efficiency was (83.70 ± 0.37) ％,and the siRNAs knocked down bcl-2 protein expression up to (44.11 ± 4.39) ％.When bcl-2 targeted siRNAs was com-bined with As for the therapy,the enhanced cytotoxicity on U937 cells was observed.The cell viabilities of bPDLLA-,As-sol-,As-NPs-and si + As-NPs-treated group were (99.44 ± 1.45) ％,(87.76 ± 2.21) ％,(92.98 ± 4.34) ％,(67.93 ± 8.16) ％ (As 1.25 μmol/L,P ＜ 0.05).(99.56 ± 2.53) ％,(54.08 ± 4.46)％,(57.85±2.ll)％,(38.60±6.24)％ (As 2.50 μmol/L,P＜0.05).(98.88 ±1.59)％,(37.62±1.38)％,(51.31 ±3.14)％,(28.92 ±4.97)％ (As 5.00 μmol/L,P＜0.05).(97.72±2.55)％,(29.44±4.14)％,(40.18±3.72)％,(20.56±4.97)％ (As 10.00 μmol/L,P＜0.05) and (96.65 ±2.03)％,(21.49± 1.60)％,(26.34 ±0.97)％,(15.90±1.70)％ (As 20.00 μmol/L,P ＜ 0.05).Conclusion PEG-PLL was charaterized potent siRNAs complexation capacity,low cytotoxicity and high transfection efficiency.Combination of bcl-2 gene silencing and As delivery using nanoparticles exerted enhanced cytotoxicity on U937 cells.
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