Proteomics and Metabolomics Analysis Reveals the Toxicity of ZnO Quantum Dots on Human SMMC-7721 Cells

Abstract

Purpose: ZnO quantum dots (QDs) are composed of less toxic metals than other QDs but have the same interesting photochemical properties. Thus, they have received considerable attention recently. Nevertheless, their toxicity cannot be ignored.

Methods: In this study, we incubated ZnO QDs with human SMMC-7721 cells for 24 h to assess their nanotoxicity through proteomics (Fold change >1.5 and p-value <0.05) and metabolomics (Fold change ≥ 1.5; VIP ≥ 1; p-value < 0.05) analyses.

Results: Both of 174 and 219 significantly changed metabolites were identified in human SMMC-7721 cells treated with 20 and 50 µg/mL ZnO QDs, respectively. ZnO QDs significantly modified metabolic pathways, including purine metabolism, ferroptosis, morphine addiction, alcoholism, cGMP-PKG signaling, and Cushing syndrome. Moreover, we identified 105 and 8 differentially expressed proteins in cells treated with 20 and 50 µg/mL ZnO QDs, and the pathways of alcoholism and Cushing syndrome were enriched.

Conclusion: ZnO QDs did not affect cell viability in a CCK8 assay, but disturbed the level of intracellular metabolites and proteins at 20 µg/mL. The KEGG analyses of the metabolomics and proteomics data both enriched the alcoholism and Cushing syndrome pathways. These results provide an experimental basis for future research on the safe use of nanomaterials.

Keywords: ZnO quantum dots; cytotoxicity; metabolomics; proteomics.

Figure 1

Cytotoxicity of ZnO QDs against SMMC-7721 cells after 24 h exposure. Cell viability was determined by CCK-8 assay and calculated relative to negative controls. All data are presented as mean ± SEM (n = 3). *p < 0.05, and ***p < 0.001 versus control according to ANOVA followed by Dunnett’s test.

Figure 2

Discrimination plots of control and ZnO QDs-treated cells from the PCA of UPLC-MS/MS data: (A) positive mode, (B) negative mode.

Figure 3

Metabolic profiles of the different ZnO QDs doses and controls at 24 h. (A–D) score plots of the PCA model, (E–H) score plots of the PLS-DA model, (I–L) plot of the permutation test (200 times) of the PLS-DA model.

Figure 4

Scatterplot of significantly enriched KEGG pathways based on the differential metabolites induced by 20 µg/mL ZnO QDs (data obtained in positive mode UPLC-MS/MS). The size of the dots represents the number of metabolites.

Figure 5

Scatterplot of significantly enriched KEGG pathways based on differential proteins induced by 20 µg/ mL ZnO QDs. The size of the dots represents the numbers of proteins.