Silke Kiefer

Background: Quantum dots (QDs) have great potential as fluorescent labels but cytotoxicity relating to extra- and intracellular degradation in biological systems has to be addressed prior to biomedical applications. In this study, human intestinal cells (Caco-2) grown on transwell membranes were used to study penetration depth, intracellular localization, translocation and cytotoxicity of CdSe/ZnS QDs with amino and carboxyl surface modifications. The focus of this study was to compare the penetration depth of QDs in differentiated vs undifferentiated cells using confocal microscopy and image processing.
Results: Caco-2 cells were exposed to QDs with amino (NH2) and carboxyl (COOH) surface groups for 3 days using a concentration of 45 µg cadmium ml−1. Image analysis of confocal/multiphoton microscopy z-stacks revealed no penetration of QDs into the cell lumen of differentiated Caco-2 cells. Interestingly, translocation of cadmium ions onto the basolateral side of differentiated monolayers was observed using high resolution inductively coupled plasma mass spectrometry (ICP-MS). Membrane damage was neither detected after short nor long term incubation in Caco-2 cells. On the other hand, intracellular localization of QDs after exposure to undifferentiated cells was observed and QDs were partially located within lysosomes.Conclusions: In differentiated Caco-2 monolayers, representing a model for small intestinal enterocytes, no penetration of amino and carboxyl functionalized CdSe/ZnS QDs into the cell lumen was detected using microscopy analysis and image processing. In contrast, translocation of cadmium ions onto the basolateral side could be detected using ICP-MS. However, even after long term incubation, the integrity of the cell monolayer was not impaired and no cytotoxic effects could be detected. In undifferentiated Caco-2 cells, both QD modifications could be found in the cell lumen. Only to some extend, QDs were localized in endosomes or lysosomes in these cells. The results indicate that the differentiation status of Caco-2 cells is an important factor in internalization and localization studies using Caco-2 cells. Furthermore, a combination of microscopy analysis and sensitive detection techniques like ICP-MS are necessary for studying the interaction of cadmium containing QDs with cells.

For a detailed analysis of the biological effects of silver nanoparticles, discrimination between effects related to the nano-scale size of the particles and effects of released silver ions is required. Silver ions are either present in the initial particle dispersion or released by the nanoparticles over time. The aim of this study is to monitor the free silver ion activity {Ag+} in the presence of silver nanoparticles using a silver ion selective electrode. Therefore, silver in the form of silver nanoparticles, 4.2 ± 1.4 nm and 2-30 nm in size, or silver nitrate was added to cell culture media in the absence or presence of A549 cells as a model for human type II alveolar epithelial cells. The free silver ion activity measured after the addition of silver nanoparticles was determined by the initial ionic silver content. The p {Ag+} values indicated that the cell culture media decrease the free silver ion activity due to binding of silver ions by constituents of the media. In the presence of A549 cells, the free silver ion activity was further reduced. The morphology of A549 cells, cultivated in DME medium containing 9.1% (v/v) FBS, was affected by adding AgNO3 at concentrations of ≥30 μM after 24 h. In comparison, silver nanoparticles up to a concentration of 200 μM Ag did not affect cellular morphology. Our experiments indicate that the effect of silver nanoparticles is mainly mediated by silver ions. An effect of silver on cellular morphology was observed at p {Ag+} ≤ 9.2.