Engineered nanopartilces (NPs) are beginning to be used in many areas as their uniqμe properties. Recent years, the safety concerns on engineered NPs have been increased as the absence of the systemic understanding of biological systems 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 quantification 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 carbohydrate 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.

Key words: silica NPs, hydrazide chemistry, quantitative glycoproteomics, cell membrane proteins, human lung cancer cells