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Classical Coulombic interaction, characterized by electrostatic interactions mediated through surface charges, is often regarded as the primary determinant in nanoparticles' (NPs) cellular association and internalization. However, the intricate physicochemical properties of particle surfaces, biomolecular coronas, and cell surfaces defy this oversimplified perspective. Moreover, the nanometrological techniques employed to characterize NPs in complex physiological fluids often exhibit limited accuracy and reproducibility. A more comprehensive understanding of nanoparticle-cell membrane interactions, extending beyond attractive forces between oppositely charged surfaces, necessitates the establishment of databases through rigorous physical, chemical, and biological characterization supported by nanoscale analytics. Additionally, computational approaches, such as in silico modeling and machine learning, play a crucial role in unraveling the complexities of these interactions., Competing Interests: Declaration of Competing Interest The authors declare that they have no known competing financial interests or personal relationships that could have appeared to influence the work reported in this paper., (Copyright © 2024 Elsevier Ltd. All rights reserved.)

Localized delivery of small interfering RNA (siRNA) is a promising approach for spatial control of cell responses at biomaterial interfaces. Layer-by-layer (LbL) assembly of siRNA with cationic polyelectrolytes has been used in film and nanoparticle vectors for transfection. Herein, we combine the ability of particles to efficiently deliver siRNA with the ability of film polyelectrolyte multilayers to act locally. LbL particles were prepared with alternating layers of poly(l-arginine) and siRNA and capped with hyaluronic acid. Negatively charged LbL particles were subsequently assembled on the poly(l-lysine)-functionalized substrate to form a LbL particle-decorated surface. Cells grown in contact with the particle-decorated surface were able to survive, internalize particles, and undergo gene silencing. This work shows that particle-decorated surfaces can be engineered by using electrostatic interactions and used to deliver therapeutic payloads for cell-instructive biointerfaces.

Background: In the field of nanoscience there is an increasing interest to follow dynamics of nanoparticles (NP) in cells with an emphasis on endo-lysosomal pathways and long-term NP fate. During our research on this topic, we encountered several pitfalls, which can bias the experimental outcome. We address some of these pitfalls and suggest possible solutions. The accuracy of fluorescence microscopy methods has an important role in obtaining insights into NP interactions with lysosomes at the single cell level including quantification of NP uptake in a specific cell type., Methods: Here we use J774A.1 cells as a model for professional phagocytes. We expose them to fluorescently-labelled amorphous silica NP with different sizes and quantify the colocalization of fluorescently-labelled NP with lysosomes over time. We focus on confocal laser scanning microscopy (CLSM) to obtain 3D spatial information and follow live cell imaging to study NP colocalization with lysosomes., Results: We evaluate different experimental parameters that can bias the colocalization coefficients (i.e., Pearson's and Manders'), such as the interference of phenol red in the cell culture medium with the fluorescence intensity and image post-processing (effect of spatial resolution, optical slice thickness, pixel saturation and bit depth). Additionally, we determine the correlation coefficients for NP entering the lysosomes under four different experimental set-ups. First, we found out that not only Pearson's, but also Manders' correlation coefficient should be considered in lysosome-NP colocalization studies; second, there is a difference in NP colocalization when using NP of different sizes and fluorescence dyes and last, the correlation coefficients might change depending on live-cell and fixed-cell imaging set-up., Conclusions: The results summarize detailed steps and recommendations for the experimental design, staining, sample preparation and imaging to improve the reproducibility of colocalization studies between the NP and lysosomes., (© 2022. The Author(s).)

The study of plastic particles, particularly those in the micro-, sub-micro-, and nano-size ranges, within food and beverages has gained increasing interest within recent years. However, many analytical techniques have limits of detection which hinder their use for the study of these particles in these sample matrices. In addition, remaining contaminants from the matrices can interfere with the signals from plastic particles. Thus, great care must be given to sample preparation and data interpretation to ensure accurate results. This study proposes the use of sample purification through chemical digestion protocols to facilitate the study of plastic particles present in tea samples, and serves to highlight technical limitations which must be overcome in future studies.

Nanoparticles (NPs) have attracted considerable attention in various fields, such as cosmetics, the food industry, material design, and nanomedicine. In particular, the fast-moving field of nanomedicine takes advantage of features of NPs for the detection and treatment of different types of cancer, fibrosis, inflammation, arthritis as well as neurodegenerative and gastrointestinal diseases. To this end, a detailed understanding of the NP uptake mechanisms by cells and intracellular localization is essential for safe and efficient therapeutic applications. In the first part of this review, we describe the several endocytic pathways involved in the internalization of NPs and we discuss the impact of the physicochemical properties of NPs on this process. In addition, the potential challenges of using various inhibitors, endocytic markers and genetic approaches to study endocytosis are addressed along with the principal (semi) quantification methods of NP uptake. The second part focuses on synthetic and bio-inspired substances, which can stimulate or decrease the cellular uptake of NPs. This approach could be interesting in nanomedicine where a high accumulation of drugs in the target cells is desirable and clearance by immune cells is to be avoided. This review contributes to an improved understanding of NP endocytic pathways and reveals potential substances, which can be used in nanomedicine to improve NP delivery.

Silica nanoparticles (NPs) are widely used in various industrial and biomedical applications. Little is known about the cellular uptake of co-exposed silica particles, as can be expected in our daily life. In addition, an inflamed microenvironment might affect a NP's uptake and a cell's physiological response. Herein, prestimulated mouse J774A.1 macrophages with bacterial lipopolysaccharide were post-exposed to micron- and nanosized silica particles, either alone or together, i.e., simultaneously or sequentially, for different time points. The results indicated a morphological change and increased expression of tumor necrosis factor alpha in lipopolysaccharide prestimulated cells, suggesting a M1-polarization phenotype. Confocal laser scanning microscopy revealed the intracellular accumulation and uptake of both particle types for all exposure conditions. A flow cytometry analysis showed an increased particle uptake in lipopolysaccharide prestimulated macrophages. However, no differences were observed in particle uptakes between single- and co-exposure conditions. We did not observe any colocalization between the two silica (SiO 2 ) particles. However, there was a positive colocalization between lysosomes and nanosized silica but only a few colocalized events with micro-sized silica particles. This suggests differential intracellular localizations of silica particles in macrophages and a possible activation of distinct endocytic pathways. The results demonstrate that the cellular uptake of NPs is modulated in inflamed macrophages but not in the presence of micron-sized particles.

The long-term fate of biomedically relevant nanoparticles (NPs) at the single cell level after uptake is not fully understood yet. We report that lysosomal exocytosis of NPs is not a mechanism to reduce the particle load. Biopersistent NPs such as nonporous silica and gold remain in cells for a prolonged time. The only reduction of the intracellular NP number is observed via cell division, e.g. , mitosis. Additionally, NP distribution after cell division is observed to be asymmetrical, likely due to the inhomogeneous location and distribution of the NP-loaded intracellular vesicles in the mother cells. These findings are important for biomedical and hazard studies as the NP load per cell can vary significantly. Furthermore, we highlight the possibility of biopersistent NP accumulation over time within the mononuclear phagocyte system.