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目的研究纳米氧化锌(Zn O-NP)对小鼠腹腔源巨噬细胞(Ana-1)的毒性作用及机制。方法Zn O-NP 2.5~160 mg·L~(-1)与Ana-1细胞作用24 h后,MTT法检测Ana-1细胞的存活率;吖啶橙-溴化乙锭(AO-EB)染色及流式细胞术观察Ana-1细胞凋亡;流式细胞术检测细胞对Zn O-NP的摄取;锌离子荧光探针测定细胞内的锌离子;乙二胺四乙酸(EDTA)螯合锌离子,检测细胞存活率。结果 Zn O-NP在2.5,5,10和20 mg·L~(-1)浓度范围内能够浓度依赖性地抑制Ana-1细胞存活(r=0.905,P<0.05);Zn O-NP20 mg·L~(-1)组细胞存活率为细胞对照组的27.9%,细胞出现明显的凋亡样改变;Zn O-NP 40,80和160 mg·L~(-1)组细胞摄取的Zn O-NP比细胞对照组显著增加(P<0.05);EDTA 2.5 mmol·L~(-1)能够明显降低细胞内的自由锌离子,改善Zn O-NP 10 mg·L~(-1)引起的细胞存活率下降(P<0.05)。螯合锌离子后,Zn O-NP对Ana-1细胞的毒性明显降低(P<0.05)。结论 Zn O-NP可浓度依赖性增加Ana-1细胞的毒性,引起凋亡样改变,其毒性可能与其进入细胞并释放锌离子有关。
Objective To study the toxic effects of zinc oxide (Zn O-NP) on mouse peritoneal macrophages (Ana-1) and its mechanism. Methods The survival rate of Ana-1 cells was assayed by MTT assay after treated with 2.5-160 mg · L -1 of Zn O-NP and Ana-1 cells for 24 h. Acridine orange-ethidium bromide (AO-EB) The apoptosis of Ana-1 cells was observed by flow cytometry, the uptake of Zn O-NP by flow cytometry, the zinc ion in the cells by zinc-ion fluorescence probe, the sequestration of ethylenediaminetetraacetic acid (EDTA) Zinc ions were tested for cell viability. Results Zn O-NP inhibited the survival of Ana-1 cells in a concentration-dependent manner (2.5, 10, 10 and 20 mg · L -1) (r = 0.905, P < · The cell viability in L-1 group was 27.9% of the cell-control group, and the apoptosis-like cells changed obviously. The Zn uptake by Zn O-NP 40, 80 and 160 mg · L -1 groups O-NP significantly increased (P <0.05). Compared with the control group, 2.5 mmol·L -1 EDTA could significantly reduce the free zinc ions in the cells and improve the concentration of Zn O-NP 10 mg · L -1 The cell survival rate decreased (P <0.05). After chelating zinc ions, the toxicity of Zn O-NP to Ana-1 cells was significantly decreased (P <0.05). Conclusion Zn O-NP can increase the toxicity of Ana-1 cells in a concentration-dependent manner, resulting in apoptosis-like changes. The toxicity may be related to its entry into cells and the release of zinc ions.