纳米二氧化硅影响人肺癌细胞的定量糖蛋白质组学研究

摘要

Engineered nanopartilces (NPs) are beginning to the safety concerns on engineered NPs have been increased be used in many areas as their uniqpe properties. Recent years, as the absence of the systemic understanding of biological systerns exposed to NPs. Although amorphous silica NPs are gradually used in cosmetics, foods and medicinal products, the potential biological effects of them exposed to biological systems are still not comprehensively elaborated. Herein, we studied the biological effects of silica NPs to human lung cancer cells by quantitative glycoproteomic strategy. A549 cells exposed to silica NPs were incubated with sodium periodate oxidation buffer to oxidize glycoproteins and then the cells were collected and cell lysis was carried out with the aid of ultrasonication in ice bath. The protein concentration was determined with Commasie brilliant blue stain. For control experiment, normal A549 cells were disposed as above mentioned method. Then the same amount of proteins extracted from A549 cells exposed to silica NPs or not were incubated with hydrazide beads to capture glycosylated proteins, respectively. The captured glycoproteins were digested with trypsin and PNGase F sequentially and both the non-glycopeptides and glycopeptides were collected, respectively. The obtained glycopeptides were used for the identification of the captured glycoproteins in both groups. And the collected non-glycopeptides were used for quantification of the captured glycoproteins through stable isotope dimethyl labeling. Therefore, the identification and quan- tification of glycoproteins was more reliable and sensitive for collecting both the information from non-glycopeptides and glycopeptides over only collecting the glycopeptides because of the quantification of glycoproteins with a large number of non-glycoproteins. The quantification results showed that the expression of several critical proteins relating to cellular car- bohydrate metabolism, cell cycle, cell-cell signaling and interaction, cellular movement etc. have been changed significantly. Some of the differently expressed proteins are membrane proteins, which may relate with the cellular uptake of silica NPs. The proteomic platform is showing a promising power to quantitatively investigate the biological response of a biological system to NPs exposure.%纳米二氧化硅（纳米SiO2）是一种正在规模化生产的纳米材料，无定型纳米SiO2因其吸入和口服对生命体不会造成直接的危害被认为是生物安全的纳米材料，已被广泛用于疾病诊断、生物分析和成像、药物载体等的研究中，导致其进入人体的方式日益增多，因此它对人体健康影响的研究对于其作为生物材料真正实现广泛应用尤为重要．本文采用肼化学方法为基础的定量蛋白质组学对无定型纳米SiO2进入人肺癌细胞后产生的影响进行了分析，结果表明其进入细胞后，导致细胞内的平衡状态发生变化，从而影响了细胞内许多重要的蛋白质的表达水平．部分跨膜蛋白质表达的变化对纳米SiO2进入细胞的途径的阐明有一定的指导意义．