Considerations for the Uptake Characteristic of Inorganic Nanoparticles into Mammalian Cells—Insights Gained by TEM Investigations

Abstract

between individual uptake mechanisms for respective nanoparticle systems. Cells internalizing nanoparticles respond to the nanoparticles and different intracellular processes will be initiated, possibly resulting in the impairment of the cellular key functions. [4] Even though these investigations provide data with statistical relevance, they can merely provide an overall picture of the uptake mechanism, as issues of different uptake mechanisms in anisotropic nanoparticle suspensions are barely assessable. Simultaneously, the influence of the biological environment, e.g., the presence of serum proteins, salts, or altered pH conditions, may have a severe impact on the particle properties, which can entail various biological implications. As such, the role of high resolution imaging techniques, which provide a clear evidence for the nanoparticles' fate in the cellular environment, [5] their localization and targeting to different cellular organelles, as well as evidence for uptake and release mechanisms, have gained increasing importance. In particular, transmission electron microscopy (TEM) investigations provide attractive possibilities to study the interaction of particles with biological barriers, i.e., the extracellular membrane or the nuclear membrane, their accumulation into cellular organelles or their spatial distribution on the scale of individual nanoparticles [6] (e.g., uptake as individual nanoparticles or in form of large clusters, etc.). In addition TEM can simultaneously image also the cellular membranes and organelles to determine respective morphological changes [7] upon nanoparticle interaction. [8][9][10][11] Thus, attractive possibilities to acquire additional information on the particle fate inside the cell emerge from TEM investigations.It can be stated here, that the utilization of electron microscopy has been conducted during the last decades [5] and, in view of some extent electron microscopy investigations of particle uptake, can be regarded in this sense as a routinely applied technique. However, frequently TEM is only utilized to characterize the nanoparticle system or to prove particle internalization. Only a fraction of reported studies utilize TEM analysis to reveal details of nanoparticle uptake and internalization or even deduce uptake pathways from these studies. This might be related to the fact that sometimes the interpretation of TEM data is not straightforward but a number of experimental limitations have to be taken into account for such studies. In a recent review, the advantages and disadvantages of TEM investigations in the context of the The internalization of nanoparticles into mammalian cells relies on a complex interplay of several parameters, which enable and dictate uptake and fate of nanoparticles in the cellular system. Due to the complexity of the involved processes, a careful experimental design has to be developed to elucidate peculiarities of the uptake processes of new nanoparticle systems into cells. Individual parameters can be hardly considered alone but h...

Understanding nanoparticle endocytosis to improve targeting strategies in nanomedicine

“… 243 Inorganic NPs such as gold and heavy metal NPs are easily identified inside or outside the cell due to their high electron density that enhances imaging contrast. 240 Dobay et al investigated the subcellular distribution of AuNPs in lung epithelial cells by TEM. 245 They showed that AuNP uptake by cells occurs through different mechanisms including caveolae-dependent endocytosis.…”

“…EM is used to access the ultrastructural information of biological and non-biological specimens, providing a resolution in the nanometer range. 240 In particular, transmission electron microscopy (TEM) provides valuable insight into the localization of NPs in the cellular compartments. 241 Several drawbacks are associated with TEM, including the laborious sample preparation techniques, which includes fixation, dehydration, embedding, sectioning (o100 nm) and staining, [240][241][242] thus does not allow one to visualize dynamic events.…”

“…240 In particular, transmission electron microscopy (TEM) provides valuable insight into the localization of NPs in the cellular compartments. 241 Several drawbacks are associated with TEM, including the laborious sample preparation techniques, which includes fixation, dehydration, embedding, sectioning (o100 nm) and staining, [240][241][242] thus does not allow one to visualize dynamic events. TEM is regularly used as a characterization method for NPs and it is also applied to visualize the localization and distribution of NPs within tissues, cells and subcellular structures.…”

“…clathrin vesicles, and early and late endosomes) can be challenging. 240 To overcome these limitations, immunogold labelling of subcellular compartments can be performed. 244 It is important to consider that organic NPs, such as polymer NPs, are composed by elements with a lower atomic number that are commonly present in the cell (e.g., carbon, hydrogen, nitrogen, and oxygen), which remarkably increases the difficulty to analyze them by EM techniques.…”

Cellular Uptake and Magneto-Hyperthermia Induced Cytotoxicity using Photoluminescent Fe3O4 Nanoparticle/Si Quantum Dot Hybrids

“…90,91 Entry into the cell nucleus is typically limited to small NPs (<5 nm) and can be extended to 20 nm by employing an NLS peptide to assist with passage through the nuclear envelope. 92 Iron oxide NPs, in particular, tend to remain in the perinuclear region without further modification. 93,94 Our magnetofluorescent nano-hybrids cross both the cell and nucleus membranes without a need for an NLS tag, thus overcoming typical limitations of nanomaterials for nuclear targeting.…”