4种典型纳米材料对小鼠胚胎成纤维细胞毒性的初步研究

为探讨不同种类纳米材料对原代培养小鼠胚胎成纤维细胞(Mouse embryo fibroblasts,MEF)的毒性效应及作用机制,选择4种典型的纳米材料(纳米碳、单壁碳纳米管、纳米氧化锌、纳米二氧化硅)制备颗粒悬液,设立5个剂量组(5、10、20、50、100μg·mL-1)对BALB/c小鼠MEF细胞进行24、48、72h染毒培养,利用细胞形态学观察和噻唑蓝实验(MTT比色法)检测上述4种纳米材料对MEF细胞活性的影响,同时,测定染毒24h后细胞培养液上清中乳酸脱氢酶(LDH)活性以探讨纳米颗粒对细胞膜完整性的影响.结果显示:1)4种纳米材料均能明显影响MEF细胞的生长形态.染毒24h后,MEF细胞发生不同程度的回缩变形,细胞间隙增大,排列稀疏,胞内颗粒物增多,细胞透明度下降.2)纳米碳、纳米氧化锌、纳米二氧化硅对MEF细胞增殖的抑制作用和对细胞膜完整性的损伤作用均随染毒剂量的升高而增强,具有明显的剂量-效应关系,其半数致死浓度(24h-IC50)分别为21.85、21.94、461.10μg·mL-1;碳纳米管组的剂量-效应之间不呈对数线性关系,未能得出其24h-IC50.3)在不同染毒剂量水平上,4种纳米材料的毒性对比差异显著:低剂量水平上纳米碳与碳纳米管的毒性强于纳米氧化锌和纳米二氧化硅,随着剂量的升高纳米氧化锌的细胞毒性升高最为显著.结果提示,纳米材料能够对MEF细胞造成毒性损伤,破坏细胞膜的完整性可能只是作用途径之一;纳米材料的毒性可能受粒径、形状、化学组成等许多因素的影响.

Study on Cytotoxicity of Four Typical Nanomaterials in Mouse Embryo Fibroblasts

This study was initiated to explore the toxic effects of different kinds of nanomaterials in vitro. Mouse embryo fibroblasts(MEF) were exposed to four nanomaterials(carbon nanoparticles, single wall carbon nanotubes, silica dioxide nanoparticles and zinc oxide nanoparticles)at five different concentrations(5, 10, 20, 50, 100μg·mL-1). Cellular morphology and MTT assay were carried out to evaluate the viability of particle-treated cells after 24h, 48h and 72h exposure. As a measurement of membrane damage, the leakage of Lactate dehydrogenase(LDH) into extracellular fluid was detected after 24h exposure. Results showed that, 1) MEF cells exposed for 24 hours retracted in different degrees, which resulted in increased cell spaces and sparse arrangement, as well as increased intracellular particles and reduced cellular transparency. 2) The effects of carbon, zinc oxide and silica dioxide nanoparticles on viability inhibition and membrane impairment were significantly enhanced in a dose-dependent manner. Median lethal concentrations(24h-IC50) of the three types of nanoparticles were 21.85μg·mL-1, 21.94μg·mL-1 and 461.10μg·mL-1, respectively. No logarithmic dose-effect relationship was observed in carbon nanotubes groups. The toxic effects of four nanomaterials were significantly different at varying concentrations:at lower dosage levels, the cytotoxicities of carbon nanopartices and carbon nanotubes were greater than that of zinc oxide and silica oxide nanoparticles;with the increase of dosage level, the toxic effect of zinc oxide nanoparticles was increased most significantly. In conclusion, our results indicated that nanomaterials could induce cytotoxicity in MEF cells, and destroying the membrane integrity might be merely one of the possible routs. The toxicity of nanomaterials may be attributed to several parameters including particle size, shape and chemical composition.