Park Hee-Yon, Chung Christopher, Eiken Madeline K, Baumgartner Karl V, Fahy Kira M, Leung Kaitlyn Q, Bouzos Evangelia, Asuri Prashanth, Wheeler Korin E, Riley Kathryn R
Department of Chemistry and Biochemistry, Swarthmore College, Swarthmore, PA, United States.
Department of Chemistry and Biochemistry, Santa Clara University, Santa Clara, CA, United States.
Front Toxicol. 2023 Feb 28;5:1081753. doi: 10.3389/ftox.2023.1081753. eCollection 2023.
Biomolecules bind to and transform nanoparticles, mediating their fate in biological systems. Despite over a decade of research into the protein corona, the role of protein modifications in mediating their interaction with nanomaterials remains poorly understood. In this study, we evaluated how glycation of the most abundant blood protein, human serum albumin (HSA), influences the formation of the protein corona on 40 nm silver nanoparticles (AgNPs) and the toxicity of AgNPs to the HepG2 human liver cell line. The effects of glycation on AgNP-HSA interactions were quantified using circular dichroism spectroscopy to monitor protein structural changes, dynamic light scattering to assess AgNP colloidal stability, zeta potential measurements to measure AgNP surface charge, and UV-vis spectroscopy and capillary electrophoresis (CE) to evaluate protein binding affinity and kinetics. The effect of the protein corona and HSA glycation on the toxicity of AgNPs to HepG2 cells was measured using the WST cell viability assay and AgNP dissolution was measured using linear sweep stripping voltammetry. Results from UV-vis and CE analyses suggest that glycation of HSA had little impact on the formation of the AgNP protein corona with protein-AgNP association constants of ≈2x10 M for both HSA and glycated HSA (gHSA). The formation of the protein corona itself (regardless of whether it was formed from HSA or glycated HSA) caused an approximate 2-fold decrease in cell viability compared to the no protein AgNP control. While the toxicity of AgNPs to cells is often attributed to dissolved Ag(I), dissolution studies showed that the protein coated AgNPs underwent less dissolution than the no protein control, suggesting that the protein corona facilitated a nanoparticle-specific mechanism of toxicity. Overall, this study highlights the importance of protein coronas in mediating AgNP interactions with HepG2 cells and the need for future work to discern how protein coronas and protein modifications (like glycation) may alter AgNP reactivity to cellular organisms.
生物分子与纳米颗粒结合并使其发生转化，从而在生物系统中决定其命运。尽管对蛋白质冠层已进行了十多年的研究，但蛋白质修饰在介导其与纳米材料相互作用中的作用仍知之甚少。在本研究中，我们评估了最丰富的血液蛋白——人血清白蛋白（HSA）的糖基化如何影响40纳米银纳米颗粒（AgNP）上蛋白质冠层的形成以及AgNP对HepG2人肝癌细胞系的毒性。使用圆二色光谱法监测蛋白质结构变化、动态光散射评估AgNP胶体稳定性、zeta电位测量AgNP表面电荷以及紫外可见光谱法和毛细管电泳（CE）评估蛋白质结合亲和力和动力学，对糖基化对AgNP - HSA相互作用的影响进行了量化。使用WST细胞活力测定法测量蛋白质冠层和HSA糖基化对AgNP对HepG2细胞毒性的影响，并使用线性扫描伏安法测量AgNP的溶解情况。紫外可见光谱和CE分析结果表明，HSA的糖基化对AgNP蛋白质冠层的形成影响很小，HSA和糖基化HSA（gHSA）与蛋白质 - AgNP的缔合常数约为2×10 M。与无蛋白质的AgNP对照相比，蛋白质冠层本身的形成（无论其由HSA还是糖基化HSA形成）导致细胞活力下降约2倍。虽然AgNP对细胞的毒性通常归因于溶解的Ag(I)，但溶解研究表明，蛋白质包被的AgNP比无蛋白质对照的溶解更少，这表明蛋白质冠层促进了纳米颗粒特异性的毒性机制。总体而言，本研究强调了蛋白质冠层在介导AgNP与HepG2细胞相互作用中的重要性，以及未来工作中辨别蛋白质冠层和蛋白质修饰（如糖基化）如何改变AgNP对细胞生物体反应性的必要性。
