High-Content Imaging and Gene Expression Approaches To Unravel the Effect of Surface Functionality on Cellular Interactions of Silver Nanoparticles

Abstract

The toxic effects of Ag nanoparticles (NPs) remain an issue of debate, where the respective contribution of the NPs themselves and of free Ag(+) ions present in the NP stock suspensions and after intracellular NP corrosion are not fully understood. Here, we employ a recently set up methodology based on high-content (HC) imaging combined with high-content gene expression studies to examine the interaction of three types of Ag NPs with identical core sizes, but coated with either mercaptoundecanoic acid (MUA), dodecylamine-modified poly(isobutylene-alt-maleic anhydride) (PMA), or poly(ethylene glycol) (PEG)-conjugated PMA with two types of cultured cells (primary human umbilical vein endothelial cells (HUVEC) and murine C17.2 neural progenitor cells). As a control, cells were also exposed to free Ag(+) ions at the same concentration as present in the respective Ag NP stock suspensions. The data reveal clear effects of the NP surface properties on cellular interactions. PEGylation of the NPs significantly reduces their cellular uptake efficiency, whereas MUA-NPs are more prone to agglomeration in complex tissue culture media. PEG-NPs display the lowest levels of toxicity, which is in line with their reduced cell uptake. MUA-NPs display the highest levels of toxicity, caused by autophagy, cell membrane damage, mitochondrial damage, and cytoskeletal deformations. At similar intracellular NP levels, PEG-NPs induce the highest levels of reactive oxygen species (ROS), but do not affect the cell cytoskeleton, in contrast to MUA- and PMA-NPs. Gene expression studies support the findings above, defining autophagy and cell membrane damage-related necrosis as main toxicity pathways. Additionally, immunotoxicity, DNA damage responses, and hypoxia-like toxicity were observed for PMA- and especially MUA-NPs. Together, these data reveal that Ag(+) ions do contribute to Ag NP-associated toxicity, particularly upon intracellular degradation. The different surface properties of the NPs however result in distinct toxicity profiles for the three NPs, indicating clear NP-associated effects.

The role of intracellular trafficking of CdSe/ZnS QDs on their consequent toxicity profile

“…In contrast, exposure to COOH-QDs resulted in a decrease in an array of genes mainly involved in cellular hypoxia. This finding is in line with earlier findings, where the involvement of genes linked to hypoxia have been associated with cellular NP toxicity [50]. The induction of high levels of mitochondrial metabolism, as indicated by the induction of mitochondrial ROS, may result in an artificial hypoxia-like scenario.…”

Presence of an Immune System Increases Anti‐Tumor Effect of Ag Nanoparticle Treated Mice

“…Silver NPs have been frequently studied regarding their potential toxicity and antibacterial effects . Recent data have shown that despite the strong contribution of intracellularly released Ag + ions, clear NP‐associated effects contribute to their cytotoxicity profile . Here, we use 5 nm diameter Ag NPs ( Figure ) coated with citrate that have been optimized for cytotoxicity research and which have been approved by the Organisation for Economic Co‐operation and Development (OECD) as standard reference materials for toxicity testing.…”

“…Toxicity appears to be mainly associated with the generation of reactive oxygen species (ROS), induction of mitochondrial damage, and alterations in autophagy levels. These parameters are very general and have been described to play an important role in the cytotoxicity of a wide range of different NPs, including silver NPs . Typically, when silver NPs are present in the endosomal environment of the cells, they are subjected to the more acidic micro‐environment that encompasses the endosomal lumen.…”

“…Silver NPs have specifically been shown to have a high potential for biomedical applications, as their dissolution rates are far slower than those observed for ZnO and CuO NPs and the associated toxicity of Ag NPs has therefore also been found to be lower . Although the toxicological profile of Ag NPs is slightly different than the one for free Ag + ions, the intracellular release of Ag + is generally accepted to be the main cause of Ag NP toxicity . A recent preclinical in vivo study has shown that high levels of Ag NPs can cause tumor destruction and has highlighted one important benefit of NPs as the toxic agent compared to common chemotherapeutic agents.…”

Involvement of Programmed Cell Death in Neurotoxicity of Metallic Nanoparticles: Recent Advances and Future Perspectives

“…In addition, the cathepsin D protein levels were up-regulated, which suggested that autophagy in HCECs was induced by NPs. Manshian et al [100] treated murine C17.2 neural progenitor cells with silver NPs and found that the LC3 fluorescent intensity was enhanced, which indicated that autophagy in C17.2 was induced by silver NPs. Since OS can induce autophagy [101–103], which was implicated in the neurotoxicity of metallic NPs [104–106], the role of autophagy in the neurotoxicity of metallic NPs should be not ignored.…”