The potential effects of in vitro digestion on the physicochemical and biological characteristics of polystyrene nanoplastics.

The presence of plastic waste in our environment has continued growing and become an important environmental concern. Because of its degradation into micro- and nanoplastics (MNPLs), MNPLs are becoming environmental pollutants of special environmental/health concern. Since ingestion is one of the main exposure routes to MNPLs, the potential effects of digestion on the physicochemical/biological characteristics of polystyrene nanoplastics (PSNPLs) were determined. The results indicated a high tendency of digested PSNPLs to agglomerate and a differential presence of proteins on their surface. Interestingly, digested PSNPLs showed greater cell uptake than undigested PSNPLs in all three tested cell lines (TK6, Raji-B, and THP-1). Despite these differences in cell uptake, no differences in toxicity were observed except for high and assumed unrealistic exposures. When oxidative stress and genotoxicity induction were determined, the low effects observed after exposure to undigested PDNPLs were not observed in the digested ones. This indicated that the greater ability of digested PSNPLs to internalize was not accompanied by a greater hazard. This type of analysis should be performed with other MNPLs of varying sizes and chemical compositions.

Nanometers-Thick Ferromagnetic Surface Produced by Laser Cutting of Diamond.

In this work, we demonstrate that cutting diamond crystals with a laser (532 nm wavelength, 0.5 mJ energy, 200 ns pulse duration at 15 kHz) produced a ≲20 nm thick surface layer with magnetic order at room temperature. We measured the magnetic moment of five natural and six CVD diamond crystals of different sizes, nitrogen contents and surface orientations with a SQUID magnetometer. A robust ferromagnetic response at 300 K was observed only for crystals that were cut with the laser along the (100) surface orientation. The magnetic signals were much weaker for the (110) and negligible for the (111) orientations. We attribute the magnetic order to the disordered graphite layer produced by the laser at the diamond surface. The ferromagnetic signal vanished after chemical etching or after moderate temperature annealing. The obtained results indicate that laser treatment of diamond may pave the way to create ferromagnetic spots at its surface.

Characterization of Total RNA, CD44, FASN, and PTEN mRNAs from Extracellular Vesicles as Biomarkers in Gastric Cancer Patients.

In-depth characterization has introduced new molecular subtypes of gastric cancer (GC). To identify these, new approaches and techniques are required. Liquid biopsies are trendsetting and provide an easy and feasible method to identify and to monitor GC patients. In a prospective cohort of 87 GC patients, extracellular vesicles (EVs) were isolated from 250 µL of plasma. The total RNA was isolated with TRIZOL. The total RNA amount and the relative mRNA levels of CD44, PTEN, and FASN were measured by qRT-PCR. The isolation of EVs and their contained mRNA was possible in all 87 samples investigated. The relative mRNA levels of PTEN were higher in patients already treated by chemotherapy than in chemo-naïve patients. In patients who had undergone neoadjuvant chemotherapy followed by gastrectomy, a decrease in the total RNA amount was observed after neoadjuvant chemotherapy and gastrectomy, while FASN and CD44 mRNA levels decreased only after gastrectomy. The amount of RNA and the relative mRNA levels of FASN and CD44 in EVs were affected more significantly by chemotherapy and gastrectomy than by chemotherapy alone. Therefore, they are a potential biomarker for monitoring treatment response. Future analyses are needed to identify GC-specific key RNAs in EVs, which could be used for the diagnosis of gastric cancer patients in order to determine their molecular subtype and to accompany the therapeutic response.

Raman spectroscopy on blood serum samples of patients with end-stage liver disease.

Raman spectroscopy has shown to be a promising method for the examination of biomedical samples. However, until now, its efficacy has not been established in clinical diagnostics. In this study, Raman spectroscopy's potential application in medical laboratories is evaluated for a large variety (38) of biomarkers. Given 234 serum samples from a cohort of patients with different stages of liver disease, we performed Raman spectroscopy at 780nm excitation wavelength. The Raman spectra were analyzed in combination with the results of routine diagnostics using specifically developed complex mathematical algorithms, including fluorescence filtering, frequency subset selection and several overfitting circumventing strategies, such as independent validation. With the results of this cohort, which were validated in 328 independent samples, a significant proof-of-concept study was completed. This study highlights the need to prevent overfitting and to use independent data for validation. The results reveal that Raman spectroscopy has high potential for use in medical laboratory diagnostics to simultaneously quantify multiple biomarkers.

Enzymatic degradation of polyethylene terephthalate nanoplastics analyzed in real time by isothermal titration calorimetry.

Plastics are globally used for a variety of benefits. As a consequence of poor recycling or reuse, improperly disposed plastic waste accumulates in terrestrial and aquatic ecosystems to a considerable extent. Large plastic waste items become fragmented to small particles through mechanical and (photo)chemical processes. Particles with sizes ranging from millimeter (microplastics, <5 mm) to nanometer (nanoplastics, NP, <100 nm) are apparently persistent and have adverse effects on ecosystems and human health. Current research therefore focuses on whether and to what extent microorganisms or enzymes can degrade these NP. In this study, we addressed the question of what information isothermal titration calorimetry, which tracks the heat of reaction of the chain scission of a polyester, can provide about the kinetics and completeness of the degradation process. The majority of the heat represents the cleavage energy of the ester bonds in polymer backbones providing real-time kinetic information. Calorimetry operates even in complex matrices. Using the example of the cutinase-catalyzed degradation of polyethylene terephthalate (PET) nanoparticles, we found that calorimetry (isothermal titration calorimetry-ITC) in combination with thermokinetic models is excellently suited for an in-depth analysis of the degradation processes of NP. For instance, we can separately quantify i) the enthalpy of surface adsorption ∆AdsH = 129 ± 2 kJ mol-1, ii) the enthalpy of the cleavage of the ester bonds ∆EBH = -58 ± 1.9 kJ mol-1 and the apparent equilibrium constant of the enzyme substrate complex K = 0.046 ± 0.015 g L-1. It could be determined that the heat production of PET NP degradation depends to 95% on the reaction heat and only to 5% on the adsorption heat. The fact that the percentage of cleaved ester bonds (η = 12.9 ± 2.4%) is quantifiable with the new method is of particular practical importance. The new method promises a quantification of enzymatic and microbial adsorption to NP and their degradation in mimicked real-world aquatic conditions.

Calcium-sensing receptor-mediated NLRP3 inflammasome response to calciprotein particles drives inflammation in rheumatoid arthritis.

Increased extracellular Ca2+ concentrations ([Ca2+]ex) trigger activation of the NLRP3 inflammasome in monocytes through calcium-sensing receptor (CaSR). To prevent extraosseous calcification in vivo, the serum protein fetuin-A stabilizes calcium and phosphate into 70-100 nm-sized colloidal calciprotein particles (CPPs). Here we show that monocytes engulf CPPs via macropinocytosis, and this process is strictly dependent on CaSR signaling triggered by increases in [Ca2+]ex. Enhanced macropinocytosis of CPPs results in increased lysosomal activity, NLRP3 inflammasome activation, and IL-1ß release. Monocytes in the context of rheumatoid arthritis (RA) exhibit increased CPP uptake and IL-1ß release in response to CaSR signaling. CaSR expression in these monocytes and local [Ca2+] in afflicted joints are increased, probably contributing to this enhanced response. We propose that CaSR-mediated NLRP3 inflammasome activation contributes to inflammatory arthritis and systemic inflammation not only in RA, but possibly also in other inflammatory conditions. Inhibition of CaSR-mediated CPP uptake might be a therapeutic approach to treating RA.

Picomolar glyphosate sensitivity of an optical particle-based sensor utilizing biomimetic interaction principles.

The continually growing use of glyphosate and its critically discussed health and biodiversity risks ask for fast, low cost, on-site sensing technologies for food and water. To address this problem, we designed a highly sensitive sensor built on the remarkably specific recognition of glyphosate by its physiological target enzyme 5-enolpyruvyl-shikimate-3-phosphate synthase (EPSPs). This principle is implemented in an interferometric sensor by using the recently established soft colloidal probe (SCP) technique. EPSPs was site-specifically immobilized on a transparent surface utilizing the self-assembling properties of circadian clock gene 2 hydrophobin chimera and homogeneity of the layer was evidenced by atomic force microscopy. Exposure of the enzyme decorated biochip to glyphosate containing samples causes formation of enzyme-analyte complexes and a competitive loss of available binding sites for glyphosate-functionalized poly(ethylene glycol) SCPs. Functionalization of the SCPs with different types of linker molecules and glyphosate was assessed employing confocal laser scanning microscopy as well as confocal Raman microspectroscopy. Overall, reflection interference contrast microscopy analysis of SCP-biochip interactions revealed a strong influence of linker length and glyphosate coupling position on the sensitivity of the sensor. In employing a combination of pentaglycine linker and tethering glyphosate via its secondary amino group, concentrations in aqueous solutions down to 100 pM could be measured by the differential adhesion between SCP and biochip surface, supported by automated image analysis algorithms. This sensing concept could even prove its exceptional pM sensitivity in combination with a superior discrimination against structurally related compounds.

Docetaxel-loaded solid lipid nanoparticles prevent tumor growth and lung metastasis of 4T1 murine mammary carcinoma cells.

BACKGROUND: Metastasis causes the most breast cancer-related deaths in women. Here, we investigated the antitumor effect of solid lipid nanoparticles (SLN-DTX) when used in the treatment of metastatic breast tumors using 4T1-bearing BALB/c mice. RESULTS: Solid lipid nanoparticles (SLNs) were produced using the high-energy method. Compritol 888 ATO was selected as the lipid matrix, and Pluronic F127 and Span 80 as the surfactants to stabilize nanoparticle dispersion. The particles had high stability for at least 120 days. The SLNs' dispersion size was 128 nm, their polydispersity index (PDI) was 0.2, and they showed a negative zeta potential. SLNs had high docetaxel (DTX) entrapment efficiency (86%), 2% of drug loading and showed a controlled drug-release profile. The half-maximal inhibitory concentration (IC50) of SLN-DTX against 4T1 cells was more than 100 times lower than that of free DTX after 24 h treatment. In the cellular uptake test, SLN-DTX was taken into the cells significantly more than free DTX. The accumulation in the G2-M phase was significantly higher in cells treated with SLN-DTX (73.7%) than in cells treated with free DTX (23.0%), which induced subsequent apoptosis. TEM analysis revealed that SLN-DTX internalization is mediated by endocytosis, and fluorescence microscopy showed DTX induced microtubule damage. In vivo studies showed that SLN-DTX compared to free docetaxel exhibited higher antitumor efficacy by reducing tumor volume (p < 0.0001) and also prevented spontaneous lung metastasis in 4T1 tumor-bearing mice. Histological studies of lungs confirmed that treatment with SLN-DTX was able to prevent tumor. IL-6 serum levels, ki-67 and BCL-2 expression were analyzed and showed a remarkably strong reduction when used in a combined treatment. CONCLUSIONS: These results indicate that DTX-loaded SLNs may be a promising carrier to treat breast cancer and in metastasis prevention.

Organ burden of inhaled nanoceria in a 2-year low-dose exposure study: dump or depot?

No detailed information on in vivo biokinetics of CeO2 nanoparticles (NPs) following chronic low-dose inhalation is available. The CeO2 burden for lung, lung-associated lymph nodes, and major non-pulmonary organs, blood, and feces, was determined in a chronic whole-body inhalation study in female Wistar rats undertaken according to OECD TG453 (6 h per day for 5 days per week for a 104 weeks with the following concentrations: 0, 0.1, 0.3, 1.0, and 3.0 mg/m3, animals were sacrificed after 3, 12, 24 months). Different spectroscopy methods (ICP-MS, ion-beam-microscopy) were used for the quantification of organ burden and for visualization of NP distribution patterns in tissues. After 24 months of exposure, the highest CeO2 lung burden (4.41 mg per lung) was associated with the highest aerosol concentration and was proportionally lower for the other groups in a dose-dependent manner. Imaging techniques confirmed the presence of CeO2 agglomerates of different size categories within lung tissue with a non-homogenous distribution. For the highest exposure group, after 24 months in total 1.2% of the dose retained in the lung was found in the organs and tissues analyzed in this study, excluding lymph nodes and skeleton. The CeO2 burden per tissue decreased from lungs > lymph nodes > hard bone > liver > bone marrow. For two dosage groups, the liver organ burden showed a low accumulation rate. Here, the liver can be regarded as depot, whereas kidneys, the skeleton, and bone marrow seem to be dumps due to steadily increasing NP burden over time.

Photodynamic therapy mediated by aluminium-phthalocyanine nanoemulsion eliminates primary tumors and pulmonary metastases in a murine 4T1 breast adenocarcinoma model.

Photodynamic therapy (PDT) is effective in the treatment of different types of cancer, such as basal cell carcinoma and other superficial cancers. However, improvements in photosensitizer delivery are still needed, and the use of PDT against more deeply located tumors has been the subject of many studies. Thus, the goal of this study was to evaluate the efficacy of a nanoemulsion containing aluminium-phthalocyanine (AlPc-NE) as a mediator of photodynamic therapy (PDT-AlPc-NE) against grafted 4T1 breast adenocarcinoma tumors in mice (BALB/c). Short after the appearance of the tumor, the animals were divided into groups (n = 5) as follows: untreated; only AlPc-NE and treated with PDT-AlPc-NE. The tumor volume was measured with a digital calliper at specific times. The presence of metastasis in the lungs was evaluated by microtomography and histopathological analyses. The results show that the application of PDT-AlPc-NE eradicated the transplanted tumors in all the treated animals, while the animals from control groups presented a robust increase in the tumor volume. Still more significantly, microtomography showed the animals submitted the PDT-AlPc-NE to be free of detectable metastasis in the lungs. The histological analysis of the lungs further confirmed the results verified by the microtomography. Therefore, this study suggests that PDT-AlPc-NE is effective in the elimination of experimentally grafted breast tumors in mice and also in preventing the formation of metastasis in the lungs.

Gold Nanoparticles as Boron Carriers for Boron Neutron Capture Therapy: Synthesis, Radiolabelling and In vivo Evaluation.

Background: Boron Neutron Capture Therapy (BNCT) is a binary approach to cancer therapy that requires accumulation of boron atoms preferentially in tumour cells. This can be achieved by using nanoparticles as boron carriers and taking advantage of the enhanced permeability and retention (EPR) effect. Here, we present the preparation and characterization of size and shape-tuned gold NPs (AuNPs) stabilised with polyethylene glycol (PEG) and functionalized with the boron-rich anion cobalt bis(dicarbollide), commonly known as COSAN. The resulting NPs were radiolabelled with 124I both at the core and the shell, and were evaluated in vivo in a mouse model of human fibrosarcoma (HT1080 cells) using positron emission tomography (PET). Methods: The thiolated COSAN derivatives for subsequent attachment to the gold surface were synthesized by reaction of COSAN with tetrahydropyran (THP) followed by ring opening using potassium thioacetate (KSAc). Iodination on one of the boron atoms of the cluster was also carried out to enable subsequent radiolabelling of the boron cage. AuNPs grafted with mPEG-SH (5 Kda) and thiolated COSAN were prepared by ligand displacement. Radiolabelling was carried out both at the shell (isotopic exchange) and at the core (anionic absorption) of the NPs using 124I to enable PET imaging. Results: Stable gold nanoparticles simultaneously functionalised with PEG and COSAN (PEG-AuNPs@[4]-) with hydrodynamic diameter of 37.8 ± 0.5 nm, core diameter of 19.2 ± 1.4 nm and ξ-potential of -18.0 ± 0.7 mV were obtained. The presence of the COSAN on the surface of the NPs was confirmed by Raman Spectroscopy and UV-Vis spectrophotometry. PEG-AuNPs@[4]- could be efficiently labelled with 124I both at the core and the shell. Biodistribution studies in a xenograft mouse model of human fibrosarcoma showed major accumulation in liver, lungs and spleen, and poor accumulation in the tumour. The dual labelling approach confirmed the in vivo stability of the PEG-AuNPs@[4]-. Conclusions: PEG stabilized, COSAN-functionalised AuNPs could be synthesized, radiolabelled and evaluated in vivo using PET. The low tumour accumulation in the animal model assayed points to the need of tuning the size and geometry of the gold core for future studies.

Oily core/amphiphilic polymer shell nanocapsules change the intracellular fate of doxorubicin in breast cancer cells.

The aim of this work was to develop and test the in vitro biological activity of nanocapsules loaded with a doxorubicin (DOX) free base dissolved in a core of castor oil shelled by poly(methyl vinyl ether-co-maleic anhydride) conjugated to n-octadecylamine residues. This system was stable and monodisperse, with a hydrodynamic diameter of about 300 nm. These nanocapsules changed the intracellular distribution of DOX, from the nuclei to the cytoplasm, and exhibited higher toxicity towards cancer cells - 4T1 and MCF-7 - and significantly lower toxicity towards normal cells - NIH-3T3 and MCF-10A - in vitro. In conclusion, these nanocapsules are suitable DOX carriers, which remain to be studied in in vivo tumor models.

Single cell study of adipose tissue mediated lipid droplet formation and biochemical alterations in breast cancer cells.

Obesity is a known risk factor for breast cancer and a negative prognostic factor for cancer recurrence and survival. Several studies demonstrated that aggressive breast tumor cells contain higher numbers of intracellular lipid droplets (LDs). Here we applied simultaneous visualization, identification and quantification of the lipid accumulation in lipid droplets (LDs) of aggressive, human triple-negative MDA-MB-231 breast cancer cells treated with adipose tissue-conditioned medium (ACM) derived from overweight and obese patients. In addition to Oil Red O and AdipoRed fluorescent staining, label-free confocal Raman microspectroscopy (CRM) has been applied. CRM enables imaging of cell compartments as well as quantification and monitoring of specific biomolecules and metabolic processes on a single cell level. Interestingly, breast cancer cells incubated with ACM showed a significantly higher number of intracellular LDs. Cultivation of breast tumor cells with ACM of obese patients induced the formation of LDs with a 20-fold higher lipid concentration than cultivation with basal medium. This is in line with the significantly higher levels of NEFAs (non-esterified fatty acids) detected in the ACM obtained from obese patient compared to ACM obtained from overweight patients or basal medium. Further, by principal component analysis, we identified a significant increase in unsaturation, esterification and lipid to protein ratio in LDs in breast cancer cells incubated with ACM. CRM analyses might function as a valuable diagnostic tool to identify metabolic alterations in biological samples which in turn could provide more detailed insights in the pathogenesis of breast cancer in association with obesity.

Uptake and molecular impact of aluminum-containing nanomaterials on human intestinal caco-2 cells.

Aluminum (Al) is one of the most common elements in the earth crust and increasingly used in food, consumer products and packaging. Its hazard potential for humans is still not completely understood. Besides the metallic form, Al also exists as mineral, including the insoluble oxide, and in soluble ionic forms. Representatives of these three species, namely a metallic and an oxidic species of Al-containing nanoparticles and soluble aluminum chloride, were applied to human intestinal cell lines as models for the intestinal barrier. We characterized physicochemical particle parameters, protein corona composition, ion release and cellular uptake. Different in vitro assays were performed to determine potential effects and molecular modes of action related to the individual chemical species. For a deeper insight into signaling processes, microarray transcriptome analyses followed by bioinformatic data analysis were employed. The particulate Al species showed different solubility in biological media. Metallic Al nanoparticles released more ions than Al2O3 nanoparticles, while AlCl3 showed a mixture of dissolved and agglomerated particulate entities in biological media. The protein corona composition differed between both nanoparticle species. Cellular uptake, investigated in transwell experiments, occurred predominantly in particulate form, whereas ionic Al was not taken up by intestinal cell lines. Transcellular transport was not observed. None of the Al species showed cytotoxic effects up to 200 µg Al/mL. The transcriptome analysis indicated mainly effects on oxidative stress pathways, xenobiotic metabolism and metal homeostasis. We have shown for the first time that intestinal cellular uptake of Al occurs preferably in the particle form, while toxicological effects appear to be ion-related.

Humic acid attenuation of silver nanoparticle toxicity by ion complexation and the formation of a Ag3+ coating.

The use of silver nanoparticles (AgNPs) result in an inevitable contact with aquatic environments. Here we study the behavior of AgNPs and the developmental toxicity in zebrafish embryos exposed to these nanoparticles (0-10 mg/L) with and without the presence of HA (20 mg/L), using zebrafish facility water (ZFW) and zebrafish growing media (ZGM). The presence of cations and HA gave rise to a decrease in Ag ion release and ζ-potential, an increase in the hydrodynamic diameter and oxidation of the AgNP surface. The results show that the presence of HA and cations in the media, as well as the silver speciation, i.e., the unusual presence of Ag3+, decreases the toxicity of AgNPs (LC50AgNPs: 1.19 mg/L; LC50AgNPs + HA: 3.56 mg/L), as well as silver bioavailability and toxicity in zebrafish embryos. Developmental alterations and the LC50 (1.19 mg/L) of AgNPs in ZFW were more relevant (p ≤ 0.05) than for AgNPs in ZGM (LC50 ˃ 10 mg/L). It was demonstrated that the bioaccumulation and toxicity of AgNPs depends on several factors including AgNPs concentration, nanoparticle aggregation, dissolved silver ions, speciation of silver ions, the amount of salt in the environment, the presence of humic substances and others, and different combinations of all of these factors.

Detection of ZrO2 Nanoparticles in Lung Tissue Sections by Time-of-Flight Secondary Ion Mass Spectrometry and Ion Beam Microscopy.

The increasing use of nanoparticles (NP) in commercial products requires elaborated techniques to detect NP in the tissue of exposed organisms. However, due to the low amount of material, the detection and exact localization of NP within tissue sections is demanding. In this respect, Time-of-Flight Secondary Ion Mass Spectrometry (ToF-SIMS) and Ion Beam Microscopy (IBM) are promising techniques, because they both offer sub-micron lateral resolutions along with high sensitivities. Here, we compare the performance of the non-material-consumptive IBM and material-consumptive ToF-SIMS for the detection of ZrO2 NP (primary size 9-10 nm) in rat lung tissue. Unfixed or methanol-fixed air-dried cryo-sections were subjected to IBM using proton beam scanning or to three-dimensional ToF-SIMS (3D ToF-SIMS) using either oxygen or argon gas cluster ion beams for complete sample sputtering. Some sample sites were analyzed first by IBM and subsequently by 3D ToF-SIMS, to compare results from exactly the same site. Both techniques revealed that ZrO2 NP particles occurred mostly agglomerated in phagocytic cells with only small quantities being associated to the lung epithelium, with Zr, S, and P colocalized within the same biological structures. However, while IBM provided quantitative information on element distribution, 3D ToF-SIMS delivered a higher lateral resolution and a lower limit of detection under these conditions. We, therefore, conclude that 3D ToF-SIMS, although not yet a quantitative technique, is a highly valuable tool for the detection of NP in biological tissue.

Nanocapsules for the co-delivery of selol and doxorubicin to breast adenocarcinoma 4T1 cells in vitro.

Nanocapsules (NCS-DOX) with an oily core of selol and a shell of poly(methyl vinyl ether-co-maleic anhydride) covalently conjugated to doxorubicin were developed. These nanocapsules are spherical, with an average hydrodynamic diameter of about 170 nm, and with negative zeta potential. NCS-DOX effectively co-delivered the selol and the doxorubicin into 4T1 cells and changed the intracellular distribution of DOX from the nuclei to the mitochondria. Moreover, a significantly increased cytotoxicity against 4T1 cells was observed, which is suggestive of additive or synergic effect of selol and doxorubicin. In conclusion, PVM/MA nanocapsules are suitable platforms to co-deliver drugs into cancer cells.

Biokinetics of Nanomaterials: the Role of Biopersistence.

Nanotechnology risk management strategies and environmental regulations continue to rely on hazard and exposure assessment protocols developed for bulk materials, including larger size particles, while commercial application of nanomaterials (NMs) increases. In order to support and corroborate risk assessment of NMs for workers, consumers, and the environment it is crucial to establish the impact of biopersistence of NMs at realistic doses. In the future, such data will allow a more refined future categorization of NMs. Despite many experiments on NM characterization and numerous in vitro and in vivo studies, several questions remain unanswered including the influence of biopersistence on the toxicity of NMs. It is unclear which criteria to apply to characterize a NM as biopersistent. Detection and quantification of NMs, especially determination of their state, i.e., dissolution, aggregation, and agglomeration within biological matrices and other environments are still challenging tasks; moreover mechanisms of nanoparticle (NP) translocation and persistence remain critical gaps. This review summarizes the current understanding of NM biokinetics focusing on determinants of biopersistence. Thorough particle characterization in different exposure scenarios and biological matrices requires use of suitable analytical methods and is a prerequisite to understand biopersistence and for the development of appropriate dosimetry. Analytical tools that potentially can facilitate elucidation of key NM characteristics, such as ion beam microscopy (IBM) and time-of-flight secondary ion mass spectrometry (ToF-SIMS), are discussed in relation to their potential to advance the understanding of biopersistent NM kinetics. We conclude that a major requirement for future nanosafety research is the development and application of analytical tools to characterize NPs in different exposure scenarios and biological matrices.

Dosimetric Quantification of Coating-Related Uptake of Silver Nanoparticles.

The elucidation of mechanisms underlying the cellular uptake of nanoparticles (NPs) is an important topic in nanotoxicological research. Most studies dealing with silver NP uptake provide only qualitative data about internalization efficiency and do not consider NP-specific dosimetry. Therefore, we performed a comprehensive comparison of the cellular uptake of differently coated silver NPs of comparable size in different human intestinal Caco-2 cell-derived models to cover also the influence of the intestinal mucus barrier and uptake-specialized M-cells. We used a combination of the Transwell system, transmission electron microscopy, atomic absorption spectroscopy, and ion beam microscopy techniques. The computational in vitro sedimentation, diffusion, and dosimetry (ISDD) model was used to determine the effective dose of the particles in vitro based on their individual physicochemical characteristics. Data indicate that silver NPs with a similar size and shape show coating-dependent differences in their uptake into Caco-2 cells. The internalization of silver NPs was enhanced in uptake-specialized M-cells while the mucus did not provide a substantial barrier for NP internalization. ISDD modeling revealed a fivefold underestimation of dose-response relationships of NPs in in vitro assays. In summary, the present study provides dosimetry-adjusted quantitative data about the influence of NP coating materials in cellular uptake into human intestinal cells. Underestimation of particle effects in vitro might be prevented by using dosimetry models and by considering cell models with greater proximity to the in vivo situation, such as the M-cell model.