纳米银对果蝇化蛹、羽化、寿命及细胞凋亡和蛋白表达的影响

纳米银因具有强有效的抗菌作用而被广泛应用于医药领域,然而它存在潜在的生物毒性.本研究选取经典的模式生物黑腹果蝇,探究纳米银对生物体的毒性作用机理.测定不同浓度纳米银处理下的Oregon R、w1118、MTF突变体等品系的化蛹率、羽化时间、羽化率及寿命;用rpr-lacZ品系的lacZ活性测定不同纳米银浓度处理下翅成虫盘的细胞凋亡情况; 通过SDS-PAGE研究不同浓度纳米银浓度处理下MTF突变体中肠蛋白的表达差异,并利用质谱分析差异蛋白的氨基酸序列.结果显示: 纳米银处理浓度升高至200 μg·mL^-1及以上时,MTF突变体化蛹率和羽化率显著降低; 纳米银处理浓度升高至800 μg·mL^-1时,Oregon R和w1118化蛹率和羽化率显著降低,而化蛹时间和羽化时间与纳米银处理浓度无显著关系.不同浓度纳米银处理对Oregon R和w1118的寿命无影响,而200 μg·mL^-1纳米银显著降低MTF突变体平均寿命.随纳米银处理浓度升高,细胞凋亡也逐渐增多.SDS-PAGE及质谱分析表明,随纳米银浓度上升,质子泵ATP激酶、热激蛋白、糖代谢相关酶系等蛋白的表达量上升.本研究表明,高浓度的纳米银会促进细胞凋亡致使部分蛋白表达量发生改变,从而降低果蝇存活率,为后续进一步了解其毒性作用机理提供理论依据.

Effects of silver nanoparticles on pupation,eclosion, life span,apoptosis and protein expression in Drosophila melanogaster

Silver nanoparticle is widely used in the field of medicine because of its strong and effective antibacterial action. However, it has potential biological toxicity. In this study, the classical model organism, Drosophila melanogaster, was used to explore underlying mechanism for the toxic effects of silver nanoparticle. The pupation rate, eclosion time, eclosion rate and lifespan of Oregon R, w1118, and MTF mutants under different concentrations of silver nanoparticle were measured. The lacZ activity of rpr-lacZ strain was used to determine apoptosis of imaginal disc after treated with different concentrations of silver nanoparticle. The difference of intestinal protein expression in MTF mutants treated with different concentrations of silver nanoparticle was studied by SDS-PAGE. The amino acid sequence of differential proteins was further analyzed by mass spectrometry. The results showed that pupation rate and eclosion rate of MTF mutants significantly decreased when the concentration of silver nanoparticle increased to 200 μg·mL^-1 and above. When the concentration of silver nanoparticle increased to 800 μg·mL^-1, the rate of pupation and eclosion was significantly reduced, with the time of pupation and eclosion being not correlated to the concentration of silver nanoparticle. The concentrations of silver nanoparticle had no effect on the lifespan of Oregon R and w1118, while 200 μg·mL^-1 silver nanoparticle significantly reduced the average lifespan of MTF mutant. Apoptosis increased with increasing concentration of silver nanoparticle. Results from SDS-PAGE and mass spectrometry analysis showed that the expression levels of proteins such as ATP kinase, heat shock protein and glucose metabolism related enzymes increased with increasing concentration of silver nanoparticle. Our results showed that high concentration of silver nanoparticle would reduce the survival rate of Drosophila, promote apoptosis and the expression of some proteins, which provided a theoretical basis for further understanding of the toxic mechanism of silver nanoparticle.