Microplastics: first extended proteomic analysis on kidney tubular cells

Abstract Background and Aims Several studies have shown the ubiquitous presence of microplastics (plastic fragments between 5 mm and 1 μm in diameter) and nanoplastics (<1 μm) in the environment and their toxicity. Furthermore, microplastics (MPs) absorb a lot of environmental pollutants, such as bisphenol A (BPA), and release them into tissues increasing their toxicity; this is the so called “Trojan Horse” effect. Recent studies have also proved their presence in human blood and human tissues of healthy people, such as placenta and lung, and in cirrhotic liver. Most frequent biological effects of MPs are inflammation, oxidative stress and alteration of metabolic pathways. In this study proteomic analysis was performed to evaluate the toxicity of polyethylene (PE) and bisphenol-A (BPA) MPs on renal tubular cells (HK2) in vitro. This is the first extended proteomic study on human cells. Method HK-2 cultures were exposed for 5-24-48 hours to BPA, PE Microspheres (PE-MP) and MP combined with BPA. Then it was performed a proteomic analysis by mass spectrometry (MS). MS data were obtain by Orbitrap Fusion Tribrid mass spectrometer (ThermoScientific). Analysis of data were performed using unsupervised hierarchical clustering using multidimensional scaling, non-linear support vector machine (SVM) learning, and partial least squares discriminant analysis. In SVM learning, a fourfold cross-validation approach was applied to estimate the prediction and classification accuracy. Results The proteomic analysis showed a clear differentiation of the HK2 proteome based on conditioning and identified a “core” of proteins, significant at ANOVA and above the 95th percentile for “fold increase” and significant at T-test compared with controls, highly discriminatory between groups. Finally, among these, a final set of 6 proteins was selected to be validated for distinguishing features: Nephronectin, GDF15, Vasorin, IGFBP7, Midkine, Tissue factor-F3. Nephronectin is a structural membrane's protein involved in cellular adhesion. GDF15, tissue factor F3 and midkine are markers of stress conditions, including inflammation and oxidative stress. IGFBP7 is a biomarker of acute kidney damage. Vasorin is a transmembrane glycoprotein that protects against apoptosis and fibrosis. Conclusion MP and BPA significantly modify the protein expression in renal tubular cells. These findings highlight the urgent need for additional research into the toxic effects of plastic debris on human kidneys and the eventual link to kidney diseases.