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Macrophages play a pivotal role in the internalization and processing of administered nanoparticles (NPs). Furthermore, the phagocytic capacity and immunological properties of macrophages can vary depending on their microenvironment, exhibiting a spectrum of polarization states ranging from pro-inflammatory M1 to anti-inflammatory M2. However, previous research investigating this phenotype-dependent interaction with NPs has predominantly relied on semi-quantitative techniques or conventional metrics to assess intracellular NPs. Here, we focus on the interaction of human monocyte-derived macrophage phenotypes (M1-like and M2-like) with gold NPs (AuNPs) by combining population-based metrics and single-cell analysis by focused ion beam-scanning electron microscopy (FIB-SEM). The multimodal analysis revealed phenotype-dependent response and uptake behavior differences, becoming more pronounced after 48 hours. The study also highlighted phenotype-dependent cell-to-cell heterogeneity in AuNPs uptake and variability in particle number at the single-cell level, which was particularly evident in M2-like macrophages, which increases with time, indicating enhanced heteroscedasticity. Future efforts to design NPs targeting macrophages should consider the phenotypic variations and the distribution of NPs concentrations within a population, including the influence of cell-to-cell heterogeneity. This comprehensive understanding will be critical in developing safe and effective NPs to target different macrophage phenotypes., Competing Interests: The authors declare no competing interests., (This journal is © The Royal Society of Chemistry.)

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.)

The gastrointestinal tract is the main target of orally ingested nanoparticles (NPs) and at the same time is exposed to noxious substances, such as bacterial components. We investigated the interaction of 59 nm silica (SiO 2 ) NPs with differentiated Caco-2 intestinal epithelial cells in the presence of cholera toxin subunit B (CTxB) and compared the effects to J774A.1 macrophages. CTxB can affect cellular functions and modulate endocytosis via binding to the monosialoganglioside (GM1) receptor, expressed on both cell lines. After stimulating macrophages with CTxB, we observed notable changes in the membrane structure but not in Caco-2 cells, and no secretion of the pro-inflammatory cytokine tumor necrosis factor-α (TNF-α) was detected. Cells were then exposed to 59 nm SiO 2 NPs and CtxB sequentially and simultaneously, resulting in a high NP uptake in J774A.1 cells, but no uptake in Caco-2 cells was detected. Flow cytometry analysis revealed that the exposure of J774A.1 cells to CTxB resulted in a significant reduction in the uptake of SiO 2 NPs. In contrast, the uptake of NPs by highly selective Caco-2 cells remained unaffected following CTxB exposure. Based on colocalization studies, CTxB and NPs might enter cells via shared endocytic pathways, followed by their sorting into different intracellular compartments. Our findings provide new insights into CTxB's function of modulating SiO 2 NP uptake in phagocytic but not in differentiated intestine cells.

Introduction: Delivery of therapeutic nanoparticles (NPs) to cancer cells represents a promising approach for biomedical applications. A key challenge for nanotechnology translation from the bench to the bedside is the low amount of administered NPs dose that effectively enters target cells. To improve NPs delivery, several studies proposed NPs conjugation with ligands, which specifically deliver NPs to target cells via receptor binding. One such example is epidermal growth factor (EGF), a peptide involved in cell signaling pathways that control cell division by binding to epidermal growth factor receptor (EGFR). However, very few studies assessed the influence of EGF present in the cell environment, on the cellular uptake of NPs., Methods: We tested if the stimulation of EGFR-expressing lung carcinomacells A549 with EGF affects the uptake of 59 nm and 422 nm silica (SiO 2 ) NPs. Additionally, we investigated whether the uptake enhancement can be achieved with gold NPs, suitable to downregulate the expression of cancer oncogene c-MYC ., Results: Our findings show that EGF binding to its receptor results in receptor autophosphorylation and initiate signaling pathways, leading to enhanced endocytosis of 59 nm SiO 2 NPs, but not 422 nm SiO 2 NPs. Additionally, we demonstrated an enhanced gold (Au) NPs endocytosis and subsequently a higher downregulation of c-MYC ., Discussion: These findings contribute to a better understanding of NPs uptake in the presence of EGF and that is a promising approach for improved NPs delivery., Competing Interests: Conflict of interest The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest.

Commercial static cell culture substrates can usually not change their physical properties over time, resulting in a limited representation of the variation in biomechanical cues in vivo. To overcome this limitation, approaches incorporating gold nanoparticles to act as transducers to external stimuli have been employed. In this work, gold nanorods were embedded in an elastomeric matrix and used as photothermal transducers to fabricate biocompatible light-responsive substrates. The nanocomposite films analysed by lock-in thermography and nanoindentation show a homogeneous heat distribution and a greater stiffness when irradiated with NIR light. After irradiation, the initial stiffness values were recovered. In vitro experiments performed during NIR irradiation with NIH-3T3 fibroblasts demonstrated that these films were biocompatible and cells remained viable. Cells cultured on the light stiffened nanocomposite exhibited a greater proliferation rate and stronger focal adhesion clustering, indicating increased cell-surface binding strength., Competing Interests: Christoph Geers is the CEO of NanoLockin GmbH, which specialises in lock-in thermal imaging instruments for nanoparticle analysis and might benefit from potential interest in this work. The funder had no role in the design of the study; in the collection, analyses, or interpretation of data; in the writing of the manuscript; or in the decision to publish the results.

Generation of amplified stimulated emission inside mammalian cells has paved the way for a novel bioimaging and cell sensing approach. Single cells carrying gain media (e.g., fluorescent molecules) are placed inside an optical cavity, allowing the production of intracellular laser emission upon sufficient optical pumping. Here, we investigate the possibility to trigger another amplified emission phenomenon (i.e., amplified spontaneous emission or ASE) inside two different cell types, namely macrophage and epithelial cells from different species and tissues, in the presence of a poorly reflecting cavity. Furthermore, the resulting ASE properties can be enhanced by introducing plasmonic nanoparticles. The presence of gold nanoparticles (AuNPs) in rhodamine 6G-labeled A549 epithelial cells results in higher intensity and lowered ASE threshold in comparison to cells without nanoparticles, due to the effect of plasmonic field enhancement. An increase in intracellular concentration of AuNPs in rhodamine 6G-labeled macrophages is, however, responsible for the twofold increase in the ASE threshold and a reduction in the ASE intensity, dominantly due to a suppressed in and out-coupling of light at high nanoparticle concentrations., 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 © 2022 The Author(s). Published by Elsevier Inc. All rights reserved.)

Plastic particle pollution has been shown to be almost completely ubiquitous within our surrounding environment. This ubiquity in combination with a variety of unique properties (e.g. density, hydrophobicity, surface functionalization, particle shape and size, transition temperatures, and mechanical properties) and the ever-increasing levels of plastic production and use has begun to garner heightened levels of interest within the scientific community. However, as a result of these properties, plastic particles are often reported to be challenging to study in complex (i.e. real) environments. Therefore, this review aims to summarize research generated on multiple facets of the micro- and nanoplastics field; ranging from size and shape definitions to detection and characterization techniques to generating reference particles; in order to provide a more complete understanding of the current strategies for the analysis of plastic particles. This information is then used to provide generalized recommendations for researchers to consider as they attempt to study plastics in analytically complex environments; including method validation using reference particles obtained via the presented creation methods, encouraging efforts towards method standardization through the reporting of all technical details utilized in a study, and providing analytical pathway recommendations depending upon the exact knowledge desired and samples being studied., (Copyright © 2022 The Authors. Published by Elsevier Ltd.. All rights reserved.)

The approval of new nanomedicines requires a deeper understanding of the interaction between cells and nanoparticles (NPs). Silica (SiO 2 ) and gold (Au) NPs have shown great potential in biomedical applications, such as the delivery of therapeutic agents, diagnostics, and biosensors. NP-cell interaction and internalization can trigger several cellular responses, including gene expression regulation. The identification of differentially expressed genes in response to NP uptake contributes to a better understanding of the cellular processes involved, including potential side effects. We investigated gene regulation in human macrophages and lung epithelial cells after acute exposure to spherical 60 nm SiO 2 NPs. SiO 2 NPs uptake did not considerably affect gene expression in epithelial cells, whereas five genes were up-regulated in macrophages. These genes are principally related to inflammation, chemotaxis, and cell adhesion. Nuclear receptor NR4A1, an important modulator of inflammation in macrophages, was found to be up-regulated. The expression of this gene was also increased upon 1 h of macrophage exposure to spherical 50 nm AuNPs and 200 nm spherical SiO 2 NPs. NR4A1 can thus be an important immediate regulator of inflammation provoked by NP uptake in macrophages.

Control over the synthesis of anisotropic nanoparticles is crucial as slight differences in their size, shape, sharpness, or the number of tips in the case of gold nanostars, has an inordinate influence on their properties and functionality for future applications. Herein, we show that the supplier and purity of polyvinylpyrrolidone (PVP) can significantly alter the synthesis of gold nanostars, demonstrating that impurities, not PVP itself, are the main factor responsible for star-like shape formation. We demonstrate that in the presence of pure PVP and N , N -dimethylformamide, the use of hydrazine leads to the formation of branched nanoparticles. This synthetic approach opens the door to solving issues associated with the use of commercial PVP during the synthesis of gold nanostars., Competing Interests: There are no conflicts to declare., (This journal is © The Royal Society of Chemistry.)

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.

Small plastic particles such as micro- (<5 mm), sub-micro- (1 µm-100 nm) and nanoplastics (<100 nm) are known to be ubiquitous within our surrounding environment. However, to date relatively few methods exist for the reliable detection of nanoplastic particles in relevant sample matrices such as foods or environmental samples. This lack of relevant data is likely a result of key limitations (e.g., resolution and/or scattering efficiency) for common analytical techniques such as Fourier transform infrared or Raman spectroscopy. This study aims to address this knowledge gap in the field through the creation of surface-enhanced Raman scattering spectroscopy substrates utilizing spherical gold nanoparticles with 14 nm and 46 nm diameters to improve the scattering signal obtained during Raman spectroscopy measurements. The substrates are then used to analyze polystyrene particles with sizes of 161 nm or 33 nm and poly(ethylene terephthalate) particles with an average size of 62 nm. Through this technique, plastic particles could be detected at concentrations as low as 10 µg/mL, and analytical enhancement factors of up to 446 were achieved.

Upon dissolution of silver nanoparticles, silver ions are released into the environment, which are known to induce adverse effects. However, since dissolution studies are predominantly performed in water and/or at room temperature, the effects of biological media and physiologically relevant temperature on the dissolution rate are not considered. Here, we investigate silver nanoparticle dissolution trends based on their plasmonic properties under biologically relevant conditions, i.e. in biological media at 37 °C over a period of 24 h. The studied nanoparticles, surface-functionalized with polyvinylpyrrolidone, beta-cyclodextrin/polyvinylpyrrolidone, and starch/polyvinylpyrrolidone, were analysed by UV-Vis spectroscopy, lock-in thermography and depolarized dynamic light scattering to evaluate the influence of these coatings on silver nanoparticle dissolution. Transmission electron microscopy was employed to visualize the reduction of the nanoparticle core diameters. Consequently, the advantages and limitations of these analytical techniques are discussed. To assess the effects of temperature on the degree of dissolution, the results of experiments performed at biological temperature were compared to those obtained at room temperature. Dissolution is often enhanced at elevated temperatures, but has to be determined individually for every specific condition. Furthermore, we evaluated potential nanoparticle aggregation. Our results highlight that additional surface coatings do not necessarily hinder the dissolution or aggregation of silver nanoparticles., Competing Interests: Christoph Geers and Mathias Bonmarin both have equity in the company NanoLockin GmbH, which specializes in lock-in thermal imaging instruments for NP analysis and might benefit from potential interest in this work., (This journal is © The Royal Society of Chemistry.)

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 overt hazard of carbon nanotubes (CNTs) is often assessed using in vitro methods, but determining a dose-response relationship is still a challenge due to the analytical difficulty of quantifying the dose delivered to cells. An approach to accurately quantify CNT doses for submerged in vitro adherent cell culture systems using UV-VIS-near-infrared (NIR) spectroscopy is provided here. Two types of multi-walled CNTs (MWCNTs), Mitsui-7 and Nanocyl, which are dispersed in protein rich cell culture media, are studied as tested materials. Post 48 h of CNT incubation, the cellular fractions are subjected to microwave-assisted acid digestion/oxidation treatment, which eliminates biological matrix interference and improves CNT colloidal stability. The retrieved oxidized CNTs are analyzed and quantified using UV-VIS-NIR spectroscopy. In vitro imaging and quantification data in the presence of human lung epithelial cells (A549) confirm that up to 85% of Mitsui-7 and 48% for Nanocyl sediment interact (either through internalization or adherence) with cells during the 48 h of incubation. This finding is further confirmed using a sedimentation approach to estimate the delivered dose by measuring the depletion profile of the CNTs., Competing Interests: The authors declare no conflict of interest.

Poly(ethylene oxide)-poly(propylene oxide)-poly(ethylene oxide) (PEO-PPO-PEO) (F127) hydrogels have been used to deliver nitric oxide (NO) topically in biomedical applications. Here, the effect of F127 microenvironments on the photochemical NO release from S-nitrosoglutathione (GSNO) was investigated in F127 solutions 7.6 wt% 15 wt% and 22.5 wt% at 15 °C and 37 °C. Small-angle X-ray Scattering (SAXS) and Differential Scanning Calorimetry (DSC) measurements, along with proton Nuclear Magnetic Resonance ( 1 H NMR) spectral shifts and T 2 relaxation data at six different concentration-temperature conditions, allowed identifying F127 microphases characterized by: a sol phase of unimers; micelles in non-defined periodic order, and a gel phase of cubic packed micelles. Kinetic measurements showed that GSNO photodecompositon proceeds faster in micellized F127 where GSNO is segregated to the intermicellar microenvironment. Real time kinetic monitoring of NO release and T 2 relaxation profiles showed that NO is preferentially partitioned into the hydrophobic PPO cores of the F127 micelles, with the consequent decrease in its rate of release to the gas phase. These results show that F127 microphases affect both the kinetics of GSNO photodecomposition and the rate of NO escape and can be used to modulate the photochemical NO delivery from F127/GSNO solutions., (Copyright © 2019 Elsevier Inc. All rights reserved.)

Nitric oxide (NO)-mediated vasodilation plays a key role in gastric mucosal defense, and NO-donor drugs may protect against diseases associated with gastric mucosal blood flow (GMBF) deficiencies. In this study, we used the ex vivo gastric chamber method and Laser Doppler Flowmetry to characterize the effects of luminal aqueous NO-donor drug S-nitroso-N-acetylcysteine (SNAC) solution administration compared to aqueous NaNO2 and NaNO3 solutions (pH 7.4) on GMBF in Sprague-Dawley rats. SNAC solutions (600 μM and 12 mM) led to a rapid threefold increase in GMBF, which was maintained during the incubation of the solutions with the gastric mucosa, while NaNO2 or NaNO3 solutions (12 mM) did not affect GMBF. SNAC solutions (600 μM and 12 mM) spontaneously released NO at 37 °C at a constant rate of 0.3 or 14 nmol·mL-1·min-1, respectively, while NaNO2 (12 mM) released NO at a rate of 0.06 nmol·mL-1·min-1 and NaNO3 (12 mM) did not release NO. These results suggest that the SNAC-induced GMBF increase is due to their higher rates of spontaneous NO release compared to equimolar NaNO2 solutions. Taken together, our data indicate that oral SNAC administration is a potential approach for gastric acid-peptic disorder prevention and treatment.

We report herein the design, preparation and characterisation of a new, water soluble, nanoplatform for the light-triggered release of nitric oxide (NO). This nanoconstruct has been achieved by exploiting a layer-by-layer approach to assembly a polyelectrolyte modified with a NO photochemical precursor, around citrate stabilized gold colloidal template. Combined spectroscopic and transmission electron microscopy analysis show that the layered hybrid nanoparticles remain stable under physiological conditions without any significant clustering along the different coating processes. The photochemical properties of the NO photocage are well preserved upon its covalent grafting in the polymeric skeleton, before and after the of the metal colloids. The direct and in real time monitoring of NO using an ultrasensitive electrode unambiguously shows the light-stimulated NO release from modified gold nanoparticles and demonstrates that the approach used permits the accurate regulation of the NO reservoir on the nanoparticles surface., (Copyright © 2013 Elsevier Inc. All rights reserved.)