纳米银体外抗腺病毒作用及机制研究

Inhibitory effects of silver nanoparticles and their possible mechanism on adenovirus in vitro

关键词:  纳米银  腺病毒3型  抗病毒

Key words:  silver nanoparticles  adenovirus type 3  inhibiting mechanism

摘要:

［目的］探讨纳米银（silver nanoparticles，silver-nps）在体外抗腺病毒3型（adenovirus type 3，ADV3）的作用机制。［方法］采用细胞培养方法和免疫荧光方法，观察纳米银对ADV3子代病毒体生成的抑制作用、对ADV3侵入细胞的阻断作用及直接灭活作用，同时用透射电镜、PCR方法观察体外纳米银对ADV3病毒体及其DNA的破坏作用。［结果］纳米银在Hela细胞上最大无毒浓度（TC0）为52.48 μg/mL；ADV3在Hela细胞上的半数感染浓度（TCID50）为10-2.74/100 μL；最大无毒剂量范围内，50 μg/mL的纳米银和等体积100TCID50ADV3分别以杀病毒、抗病毒、直接灭活病毒三种不同方式作用Hela细胞，细胞存活率分别为（93.58±1.79）%，（90.50±1.99）%和（95.38±2.98）%，而100TCID50ADV3感染Hela细胞后，测得细胞存活率为（31.92±2.86）%，两者相比差异有显著性意义（P<0.01）。免疫荧光显示，与腺病毒对照组形成的强特异性荧光相比，三种途径作用于Hela细胞后各组的特异性荧光颗粒均减少；电镜显示，纳米银可直接与病毒粒子相互作用，破坏ADV3粒子呈时间依赖性。PCR显示，与病毒对照组、溶剂对照组相比，纳米银组扩增目的条带亮度都相对较弱。［结论］纳米银在体外对ADV3具有明显的抑制作用，其作用机制可能与纳米银破坏病毒粒子及其表面衣壳蛋白及腺病毒DNA有关。

Abstract:

［Objective］To explore the inhibitory effects and possible mechanism of silver nanoparticles (silver-nps) on adenovirus type 3 (ADV3).［Methods］Cell culture and immuno fluorescence assay were applied to evaluate the anti-adenovirus activities of silver-nps in inhibiting the formation of ADV3 progeny virions,blocking and inactivating ADV3.The anti-adenovirus activities of silver-nps on Hela cells were observed by transmission electron microscopy.The PCR method was also used to analyze the destructive effect of silver-nps on ADV3 DNA.［Results］TC0 of silver-nps was 52.48 μg/mL.The median tissue culture infective dose(TCID50 ) of ADV3 on Hela cell was 10-2.74/100 μL.In the range of non-toxic concentration,the 50 μg/mL of silver-nps and 100TCID50 of ADV3 were analyzed in virucidal,antiviral and inactivating ways,the survival rates of Hela cells were (93.58±1.7 9)%,(90.50±1.99)% and (95.38±2.98)% respectively,when compared with the viability of control group,the differences were significant(P<0.05). Hela cells infected with ADV3 had much specific immunofluorescence when compared to the cells treated with silver -nps in virucidal,antiviral and inactivating ways.Compared to the intact shape of ADV3 virions,the silver -nps could directly interact with viral particles and damage ADV3 virions in a time dependent manner under transmission electron microscopy.The densities of DNA bands of virions treated with silver-nps were decreased in comparison to that of control group following PCR amplification.［Conclusion］Silver-nps have inhibitory effects on ADV3 in vitro,its possible mechanisms are associated with damaging viral particles,capsid protein and DNA.