The biodistribution of polystyrene nanoparticles administered intravenously in the chicken embryo.

Nanoplastics can cause severe malformations in chicken embryos. To improve our understanding of the toxicity of nanoplastics to embryos, we have studied their biodistribution in living chicken embryos. We injected the embryos in the vitelline vein at stages 18-19. We injected polystyrene nanoparticles (PS-NPs) tagged with europium- or fluorescence. Their biodistribution was tracked using inductively-coupled plasma mass spectrometry on tissue lysates, paraffin histology, and vibratome sections analysed by machine learning algorithms. PS-NPs were found at high levels in the heart, liver and kidneys. Furthermore, PS-NPs crossed the endocardium of the heart at sites of epithelial-mesenchymal transformation; they also crossed the liver endothelium. Finally, we detected PS-NPs in the allantoic fluid, consistent with their being excreted by the kidneys. Our study shows the power of the chicken embryo model for analysing the biodistribution of nanoplastics in embryos. Such experiments are difficult or impossible in mammalian embryos. These findings are a major advance in our understanding of the biodistribution and tissue-specific accumulation of PS-NPs in developing animals.

Harnessing PET to track micro- and nanoplastics in vivo.

The proliferation of plastics in the environment continues at an alarming rate. Plastic particles have been found to be persistent and ubiquitous pollutants in a variety of environments, including sea water, fresh water, soil, and air. In light of this phenomenon, the scientific and medical communities have become increasingly wary of the dangers posed to human health by chronic exposure to microplastics (< 5 mm diameter) and nanoplastics (< 100 nm diameter). A critical component of the study of the health effects of these pollutants is the accurate determination of their pharmacokinetic behavior in vivo. Herein, we report the first use of molecular imaging to track polystyrene (PS) micro- and nanoplastic particles in mammals. To this end, we have modified PS particles of several sizes-diameters of 20 nm, 220 nm, 1 µm, and 6 µm-with the chelator desferrioxamine (DFO) and radiolabeled these DFO-bearing particles with the positron-emitting radiometal zirconium-89 (89Zr; t1/2 ~ 3.3 d). Subsequently, positron emission tomography (PET) was used to visualize the biodistribution of these radioplastics in C57BL/6J mice at 6, 12, 24, and 48 h after ingestion. The imaging data reveal that the majority of the radioplastics remain in the gastrointestinal tract and are eliminated through the feces by 48 h post-ingestion, a result reinforced by acute biodistribution studies. Ultimately, this work suggests that nuclear imaging-and PET in particular-can be a sensitive and effective tool in the urgent and rapidly growing effort to study the in vivo behavior and potential toxicity of micro- and nanoplastics.

Inhalation toxicity of polystyrene micro(nano)plastics using modified OECD TG 412.

Recently, there have been reports that many microplastics are found in the air, which has raised concerns about their toxicity. To date, however, only limited research has investigated the effects of micro(nano)plastics on human health, and even less the potential for inhalation toxicity. To fill this research gap, we investigated the potential inhalation toxicity of micro(nano)plastics using a modified OECD Guideline for Testing of Chemicals No. 412 '28-Day (subacute) inhalation toxicity study' using a whole-body inhalation system. Sprague-Dawley rats were exposed to three different exposure concentrations of polystyrene micro(nano)plastics (PSMPs), as well as control, for 14 days of inhalation exposure. After 14 days, alterations were observed on sevral endpoints in physiological, serum biochemical, hematological, and respiratory function markers measured on the samples exposed to PSMPs. However, no concentration-response relationships were observed, suggesting that these effects may not be definitively linked to exposure of PSMPs. On the other hand, the expression of inflammatory proteins (TGF-ß and TNF-α) increased in the lung tissue in an exposure concentration-dependent manner. The overall results indicate that 14-day inhalation exposure of PSMPs to rats has a more pronounced effect at the molecular level than at the organismal one. These results suggest that if the exposure sustained, alterations at the molecular level may lead to subsequent alterations at the higher levels, and consequently, the health risks of inhalation exposed micro(nano)plastics should not be neglected.

Pulmonary and systemic toxicity in rats following inhalation exposure of 3-D printer emissions from acrylonitrile butadiene styrene (ABS) filament.

BACKGROUND: Fused filament fabrication 3-D printing with acrylonitrile butadiene styrene (ABS) filament emits ultrafine particulates (UFPs) and volatile organic compounds (VOCs). However, the toxicological implications of the emissions generated during 3-D printing have not been fully elucidated. AIM AND METHODS: The goal of this study was to investigate the in vivo toxicity of ABS-emissions from a commercial desktop 3-D printer. Male Sprague Dawley rats were exposed to a single concentration of ABS-emissions or air for 4 hours/day, 4 days/week for five exposure durations (1, 4, 8, 15, and 30 days). At 24 hours after the last exposure, rats were assessed for pulmonary injury, inflammation, and oxidative stress as well as systemic toxicity. RESULTS AND DISCUSSION: 3-D printing generated particulate with average particle mass concentration of 240 ± 90 µg/m³, with an average geometric mean particle mobility diameter of 85 nm (geometric standard deviation = 1.6). The number of macrophages increased significantly at day 15. In bronchoalveolar lavage, IFN-γ and IL-10 were significantly higher at days 1 and 4, with IL-10 levels reaching a peak at day 15 in ABS-exposed rats. Neither pulmonary oxidative stress responses nor histopathological changes of the lungs and nasal passages were found among the treatments. There was an increase in platelets and monocytes in the circulation at day 15. Several serum biomarkers of hepatic and kidney functions were significantly higher at day 1. CONCLUSIONS: At the current experimental conditions applied, it was concluded that the emissions from ABS filament caused minimal transient pulmonary and systemic toxicity.

Rapid ingestion and egestion of spherical microplastics by bacteria-feeding nematodes.

Microplastics, anthropogenically released into freshwaters, settle in sediments, where they are directly ingested by benthic organisms. However, to the best of our knowledge, fine-scale studies of microplastic ingestion and egestion by nematodes, one of the most abundant meiofaunal taxa, are lacking. We therefore conducted a time series of the ingestion and egestion by adult Caenorhabditis elegans and Pristionchus pacificus of 0.5- and 1.0-µm fluorescent polystyrene (PS) beads along with bacteria. The nematodes were exposed to 107 beads ml-1 in aqueous medium for 5 min-24 h and pumping rates of C. elegans were determined. In the egestion study, PS bead egestion was monitored in nematodes with high microplastic body burdens for 5 min-24 h in microplastic-free medium. Ingested beads were detected already within 5 min and up to 203 ± 15 PS beads (1.0 µm; C. elegans) were found after 30 min. Overall, significantly more 1.0-µm than 0.5-µm PS beads were taken up. The distinct feeding behaviors of the two species influenced their PS bead body burdens. Ingested PS beads were almost completely egested within the first 20-40 min in the presence of sufficient food. In C. elegans, 1.0-µm beads were egested less rapidly than 0.5-µm PS beads. Given the rapid ingestion and egestion of the beads, our study demonstrates that the actual amount of ingested and egested microplastics by nematodes in the environment may be several times higher than the microplastic body burdens may imply. However, spherical PS beads did not bioconcentrate in nematodes.

Differentially charged nanoplastics demonstrate distinct accumulation in Arabidopsis thaliana.

Although the fates of microplastics (0.1-5 mm in size) and nanoplastics (<100 nm) in marine environments are being increasingly well studied1,2, little is known about the behaviour of nanoplastics in terrestrial environments3-6, especially agricultural soils7. Previous studies have evaluated the consequences of nanoplastic accumulation in aquatic plants, but there is no direct evidence for the internalization of nanoplastics in terrestrial plants. Here, we show that both positively and negatively charged nanoplastics can accumulate in Arabidopsis thaliana. The aggregation promoted by the growth medium and root exudates limited the uptake of amino-modified polystyrene nanoplastics with positive surface charges. Thus, positively charged nanoplastics accumulated at relatively low levels in the root tips, but these nanoplastics induced a higher accumulation of reactive oxygen species and inhibited plant growth and seedling development more strongly than negatively charged sulfonic-acid-modified nanoplastics. By contrast, the negatively charged nanoplastics were observed frequently in the apoplast and xylem. Our findings provide direct evidence that nanoplastics can accumulate in plants, depending on their surface charge. Plant accumulation of nanoplastics can have both direct ecological effects and implications for agricultural sustainability and food safety.

Exposure of bay scallop Argopecten irradians to micro-polystyrene: Bioaccumulation and toxicity.

Marine microplastic pollution poses a threat to aquatic organisms, including bivalves. In this study, we investigated the accumulation of microplastics and their elicited antioxidant stress response in the bay scallop Argopecten irradians. Scallops were exposed to 1 µm diameter micro-polystyrene (MP) beads at 10, 100, and 1000 beads/mL concentrations for a 7 day period. Bead presence in the digestive diverticula and defense responses in the digestive diverticula and hemolymph were measured at 1, 3, 5, and 7 days. The activity and expression of the antioxidant enzymes superoxide dismutase (SOD) and catalase (CAT) and H2O2 in the digestive diverticula and/or hemolymph of scallops increased with microplastic concentration and exposure duration. These results suggest that microplastics can accumulate in the digestive diverticula of A. irradians, and that exposure to microplastics induces oxidative stress in bivalves. It is likely that exposure to high concentrations of micro- or nano-sized plastic particles could potentially have adverse effects in bivalves.

Intraductal Drug Delivery to the Breast: Effect of Particle Size and Formulation on Breast Duct and Lymph Node Retention.

Drug delivery by direct intraductal administration can achieve high local drug concentration in the breast and minimize systemic levels. However, the clinical application of this approach for breast cancer treatment is limited by the rapid clearance of the drug from the ducts. With the goal of developing strategies to prolong drug retention in the breast, this study was focused on understanding the influence of particle size and formulation on breast duct and lymph node retention. Fluorescent-labeled polystyrene (PS) particles ranging in size from 100 to 1000 nm were used to study the influence of particle size. Polylactic acid-co-glycolic acid (PLGA) was used to develop and test formulations for intraductal delivery. Cy 5.5, a near-IR dye, was encapsulated in PLGA microparticles, nanoparticles, and the in situ gel to study the biodistribution in rats using an in vivo imager. PS microparticles (1 µm) showed longer retention in the duct compared to 100 and 500 nm nanoparticles. The ductal retention half-life was 5-fold higher for PS microparticles compared to the nanoparticles. On the other hand, the free dye was cleared from the breast within 6 h. PLGA nanoparticles sustained the release of Cy 5.5 for >4 days. Microparticles and gel showed a much slower release than nanoparticles. PLGA in situ gel and microparticles were retained in the breast for up to 4 days, while the nanoparticles were retained in the breast for 2 days. PLGA nanoparticles and microparticles drained to the axillary lymph node and were retained for up to 24 and 48 h, respectively, while the in situ gel and the free dye did not show any detectable fluorescence in the lymph nodes. Taken together, the results demonstrate the feasibility of prolonged retention in the breast duct and lymph node by optimal formulation design. The findings can serve as a framework to design formulations for localized treatment of breast cancer.

Size-dependent elimination of ingested microplastics in the Mediterranean mussel Mytilus galloprovincialis.

Filter feeding organisms have been reported to ingest microplastics (MP) in marine environments. However, information regarding how long the ingested MPs are retained in their digestive tracts remains limited. Here, we report the gut retention time (GRT90) and the long-term egestion time of three different sized polystyrene microspheres (1, 10, and 90 µm) in the Mediterranean mussel Mytilus galloprovincialis. We found significant differences in GRT90 with respect to MP size. With respect to the long-term egestion of MPs, most of the smaller MPs were excreted immediately, although some were detected intermittently until day 40. In comparison, larger MPs were slowly excreted in bulk, after which they were not detected. The results indicate that different sized MPs are retained differently in the digestive tract of mussels. The size-dependent effects of MPs should thus be considered when evaluating the effects of MPs in mussels.

The uptake and elimination of polystyrene microplastics by the brine shrimp, Artemia parthenogenetica, and its impact on its feeding behavior and intestinal histology.

Microplastics are a ubiquitous contaminant of marine ecosystems that have received considerable global attention. The effects of microplastic ingestion on some marine biota have been evaluated, but the uptake, elimination, and histopathological impacts of microplastics remain under-investigated especially for zooplankton larvae. Here, we show that 10 µm polystyrene microspheres can be ingested and egested by Artemia parthenogenetica larvae, which impact their health. The results indicate that A. parthenogenetica larvae have a varying capacity to consume 10 µm polystyrene microspheres that is dependent on microplastic exposure concentrations, exposure times, and the availability of food. The lowest level of microplastics that was ingested by A. parthenogenetica was 0.15 particles/individual when exposed to 10 particles/mL and 0.05 particles/individual when exposed to 1 particle/mL over 24 h and 14 d, respectively. A. parthenogenetica larvae were able to egest feces with microplastics within 3 h of ingestion. However, ingested microplastics persisted in individuals for up to 14 days. Furthermore, microalgal feeding was significantly reduced by 27.2% in the presence of 102 particles/mL microplastics over 24 h. Histological analyses indicated that a greater abundance of lipid droplets was present among epithelia after 24 h of exposure at a concentration of 10 particles/mL. Moreover, intestinal epithelia were deformed and disorderedly arranged after 14 d of exposure. Overall, these results indicate that marine microplastic pollution could pose a threat to A. parthenogenetica health, especially that of larvae. Consequently, further research is required to evaluate the potential physiological and histopathological effects of microplastics for other marine invertebrate species.

Uptake and effects of orally ingested polystyrene microplastic particles in vitro and in vivo.

Evidence exists that humans are exposed to plastic microparticles via diet. Data on intestinal particle uptake and health-related effects resulting from microplastic exposure are scarce. Aim of the study was to analyze the uptake and effects of microplastic particles in human in vitro systems and in rodents in vivo. The gastrointestinal uptake of microplastics was studied in vitro using the human intestinal epithelial cell line Caco-2 and thereof-derived co-cultures mimicking intestinal M-cells and goblet cells. Different sizes of spherical fluorescent polystyrene (PS) particles (1, 4 and 10 µm) were used to study particle uptake and transport. A 28-days in vivo feeding study was conducted to analyze transport at the intestinal epithelium and oxidative stress response as a potential consequence of microplastic exposure. Male reporter gene mice were treated three times per week by oral gavage with a mixture of 1 µm (4.55 × 107 particles), 4 µm (4.55 × 107 particles) and 10 µm (1.49 × 106 particles) microplastics at a volume of 10 mL/kg/bw. Effects of particles on macrophage polarization were investigated using the human cell line THP-1 to detect a possible impact on intestinal immune cells. Altogether, the results of the study demonstrate the cellular uptake of a minor fraction of particles. In vivo data show the absence of histologically detectable lesions and inflammatory responses. The particles did not interfere with the differentiation and activation of the human macrophage model. The present results suggest that oral exposure to PS microplastic particles under the chosen experimental conditions does not pose relevant acute health risks to mammals.

Combining vascular targeting and the local first pass provides 100-fold higher uptake of ICAM-1-targeted vs untargeted nanocarriers in the inflamed brain.

New advances in intra-arterial (IA) catheters offer clinically proven local interventions in the brain. Here we tested the effect of combining local IA delivery and vascular immunotargeting. Microinjection of tumor necrosis factor alpha (TNFα) in the brain parenchyma causes cerebral overexpression of Inter-Cellular Adhesion Molecule-1 (ICAM-1) in mice. Systemic intravenous injection of ICAM-1 antibody (anti-ICAM-1) and anti-ICAM-1/liposomes provided nearly an order of magnitude higher uptake in the inflamed vs normal brain (from ~0.1 to 0.8%ID/g for liposomes). Local injection of anti-ICAM-1 and anti-ICAM-1/liposomes via carotid artery catheter provided an additional respective 2-fold and 5-fold elevation of uptake in the inflamed brain vs levels attained by IV injection. The uptake in the inflamed brain of respective untargeted IgG counterparts was markedly lower (e.g., uptake of anti-ICAM-1/liposomes was 100-fold higher vs IgG/liposomes). These data affirm the specificity of the combined effect of the first pass and immunotargeting. Intravital real-time microscopy via cranial window revealed that anti-ICAM-1/liposomes, but not IgG/liposomes bind to the lumen of blood vessels in the inflamed brain within minutes after injection. This straightforward framework provides the basis for translational efforts towards local vascular drug targeting to the brain.

Role of the Basement Membrane as an Intestinal Barrier to Absorption of Macromolecules and Nanoparticles.

Full understanding of the barrier property of mucosal tissues is imperative for development of successful mucosal drug delivery strategies, particularly for biologics and nanomedicines. The contribution of the mucosal basement membrane (BM) to this barrier is currently not fully appreciated. This work examined the role of the BM as a barrier to intestinal absorption of model macromolecules (5 and 10 kDa dextrans) and 100 nm polystyrene nanoparticles. Dextrans and nanoparticles were applied either directly to BM-coated inserts or to an intestinal model, namely, differentiated intestinal epithelial monolayers (Caco-2) cultured on BM-modified inserts. The work shows that the BM per se does not impact the diffusion of dextran macromolecules but severely hinders the movement of nanoparticles. However, importantly, Caco-2 monolayers cultured on BM-coated inserts, which show a remarkably different morphology, display a significantly larger barrier to the translocation of one dextran, as well as nanoparticle systems compared to cells cultured on unmodified inserts. Therefore, this work shows that, in addition to presenting a direct physical barrier to the movement of nanoparticles, the BM also exerts an indirect barrier effect, likely due to its influence on epithelial cell physiology. This work is important as it highlights the currently unmet need to consider and further study the barrier properties of the BM in mucosal delivery of biologics and nanomedicines.

Indium-111 labeling of high-density lipoprotein-mimicking phospholipid-styrene maleic acid copolymer complexes and its biodistribution in mice.

Discoidal lipid nanoparticles mimicking native high-density lipoproteins (HDL) are promising delivery vehicles of drugs and/or imaging agents. However, little is known about the in vivo biodistribution of such discoidal lipid nanoparticles compared to liposomes, clinically available spherical lipid nanoparticles. Recently, it has been reported that synthetic polymers instead of apolipoproteins can be complexed with phospholipid to form discoidal nanoparticles. In the present study, with the aim of developing phospholipid-synthetic polymer complexes for future clinical applications, the biodistribution of such particles in normal mice was investigated. Lipid nanoparticles comprising 1-palmitoyl-2-oleoyl-glycero-3-phosphocholine (POPC) and styrene maleic acid copolymer (SMA), having sizes similar to native HDL, were prepared using the freeze-sonication method. POPC-SMA complexes remained stable at 37°C for at least 3 days in buffer. By devising ways to avoid detrimental effects accompanied by pH reduction and nonspecific binding of 111 In to SMA, POPC-SMA complexes were successfully labeled with 111 In without affecting particle integrity. The biodistribution of POPC-SMA complexes in normal mice was similar to that of discoidal lipid nanoparticles composed of POPC and apolipoprotein A-I, the major protein constituent of native HDL. Unlike liposomes, the accumulation of POPC-SMA complexes in the spleen was low, suggesting that these complexes are not recognized as foreign substances. To the best of our knowledge, this is the first in vivo study of HDL-mimicking phospholipid-synthetic polymer complexes.

Alveolar epithelial cell processing of nanoparticles activates autophagy and lysosomal exocytosis.

Using confocal microscopy, we quantitatively assessed uptake, processing, and egress of near-infrared (NIR)-labeled carboxylated polystyrene nanoparticles (PNP) in live alveolar epithelial cells (AEC) during interactions with primary rat AEC monolayers (RAECM). PNP fluorescence intensity (content) and colocalization with intracellular vesicles in a cell were determined over the entire cell volume via z stacking. Isotropic cuvette-based microfluorimetry was used to determine PNP concentration ([PNP]) from anisotropic measurements of PNP content assessed by confocal microscopy. Results showed that PNP uptake kinetics and steady-state intracellular content decreased as diameter increased from 20 to 200 nm. For 20-nm PNP, uptake rate and steady-state intracellular content increased with increased apical [PNP] but were unaffected by inhibition of endocytic pathways. Intracellular PNP increasingly colocalized with autophagosomes and/or lysosomes over time. PNP egress exhibited fast Ca2+ concentration-dependent release and a slower diffusion-like process. Inhibition of microtubule polymerization curtailed rapid PNP egress, resulting in elevated vesicular and intracellular PNP content. Interference with autophagosome formation led to slower PNP uptake and markedly decreased steady-state intracellular content. At steady state, cytosolic [PNP] was higher than apical [PNP], and vesicular [PNP] (~80% of intracellular PNP content) exceeded both cytosolic and intracellular [PNP]. These data are consistent with the following hypotheses: 1) autophagic processing of nanoparticles is essential for maintenance of AEC integrity; 2) altered autophagy and/or lysosomal exocytosis may lead to AEC injury; and 3) intracellular [PNP] in AEC can be regulated, suggesting strategies for enhancement of nanoparticle-driven AEC gene/drug delivery and/or amelioration of AEC nanoparticle-related cellular toxicity.

Protein machineries defining pathways of nanocarrier exocytosis and transcytosis.

The transport of nanocarriers through barriers like the gut in a living organism involves the transcytosis of these nanocarriers through the cell layer dividing two compartments. Understanding how this process works is not only essential to further developing strategies for a more effective nanocarrier transport system but also for providing fundamental insights into the barrier function as a means of protection against micro- and nanoplastics in the food chain. We therefore set out to investigate the different uptake mechanisms, intracellular trafficking and the routes for exocytosis for small polystyrene nanoparticles (PS-NPs ca. 100 nm) as mimicking nanocarriers in a Caco-2 cell model for gut-blood transition. We used label-free, quantitative mass spectrometry (MS) for determining the proteome that adhered to transversed nanoparticles. From this rich proteomics dataset, as well as previous studies, we generated stable-transfected Caco-2 cell lines carrying the green fluorescent protein (GFP) coupled to proteins of interest for uptake, early, late and exocytotic endosomes. We detected the spatial and temporal overlap of such marked endosomes with the nanocarrier signal in confocal laser scanning and super-resolution microscopy. There was a clear distinction in the time course of nanoparticle trafficking between groups of proteins for endocytosis, intracellular storage and putatively transcytosis and we identified several key transcytotic markers like Rab3 and Copine1. Moreover, we postulate the necessity of a certain protein composition on endosomes for successful transcytosis of nanocarriers. Finally, we define the two-sided impasse of the lysosome as a dead end for nano-plastic and the limit of nanocarriers in the 100 nm range. STATEMENT OF SIGNIFICANCE: Here we focus on mechanisms of transcytosis and how we can follow these with methods not used before. First, we use mass spectrometry of transcytosed nanoparticles to pick proteins of the transcytosis machinery describing key proteins involved. We can detect the complex mixtures of proteins. As this is a dynamic process involving whole families of proteins interacting with each other and as this is an orchestrated process we coined the term protein machineries for this active interplay. By genetically modifying the proteins attaching GFP we are able to follow the transcytosis pathway. We evaluate the process in a quantitative manner over time. This reveals that the most obvious obstacle to transcytosis is a routing of the nanocarriers to the lysosomes.

Polystyrene nanoplastics inhibit reproduction and induce abnormal embryonic development in the freshwater crustacean Daphnia galeata.

We assayed the toxicity of polystyrene nanoparticles (PS-NP, 52 nm) to Daphnia galeata. Survival and reproduction were significantly decreased in individuals exposed to 5 mg/L of PS-NP for 5 days, and embryos showed abnormal development, including a low hatching rate. Using fluorescence confocal microscopy, we recorded the transfer of PS-NP from the external surface of the body to the internal organs, including the thoracic appendices, ovaries, caudal appendices, and brood chamber, as well as PS-NP storage in lipid droplets. Although embryos were exposed to PS-NP in the brood chamber, they did not internalize PS-NP. Exposed D. galeata adults that were not pregnant stored significantly fewer lipid droplets than did the control group, and the lipid droplets that they did store were smaller; meanwhile, there were no significant changes in lipid storage in exposed pregnant individuals. Some embryos showed a high level of lipid storage, a response that occurs when embryos experience an abnormal state, and these embryos showed a very low hatching rate. However, the offspring of exposed adults showed normal survival and lipid storage. This study provides visual evidence that confirms the transfer and effects of PS-NP in Daphnia species, and suggests a relationship between toxicity and lipid storage.

Exploring uptake and biodistribution of polystyrene (nano)particles in zebrafish embryos at different developmental stages.

In ecotoxicology, it is continuously questioned whether (nano)particle exposure results in particle uptake and subsequent biodistribution or if particles adsorb to the epithelial layer only. To contribute to answering this question, we investigated different uptake routes in zebrafish embryos and how they affect particle uptake into organs and within whole organisms. This is addressed by exposing three different life stages of the zebrafish embryo in order to cover the following exposure routes: via chorion and dermal exposure; dermal exposure; oral and dermal exposure. How different nanoparticle sizes affect uptake routes was assessed by using polystyrene particles of 25, 50, 250 and 700nm. In our experimental study, we showed that particle uptake in biota is restricted to oral exposure, whereas the dermal route resulted in adsorption to the epidermis and gills only. Ingestion followed by biodistribution was observed for the tested particles of 25 and 50nm. The particles spread through the body and eventually accumulated in specific organs and tissues such as the eyes. Particles larger than 50nm were predominantly adsorbed onto the intestinal tract and outer epidermis of zebrafish embryos. Embryos exposed to particles via both epidermis and intestine showed highest uptake and eventually accumulated particles in the eye, whereas uptake of particles via the chorion and epidermis resulted in marginal uptake. Organ uptake and internal distribution should be monitored more closely to provide more in depth information of the toxicity of particles.

Pathway analysis of systemic transcriptome responses to injected polystyrene particles in zebrafish larvae.

Microplastics are a contaminant of emergent concern in the environment, however, to date there is a limited understanding on their movement within organisms and the response of organisms. In the current study zebrafish embryos at different development stages were exposed to 700nm fluorescent polystyrene (PS) particles and the response pathway after exposure was investigated using imaging and transcriptomics. Our results show limited spreading of particles within the larvae after injection during the blastula stage. This is in contrast to injection of PS particles in the yolk of 2-day old embryos, which resulted in redistribution of the PS particles throughout the bloodstream, and accumulation in the heart region. Although injection was local, the transcriptome profiling showed strong responses of zebrafish embryos exposed to PS particle, indicating a systemic response. We found several biological pathways activated which are related to an immune response in the PS exposed zebrafish larvae. Most notably the complement system was enriched as indicated by upregulation of genes in the alternative complement pathway (e.g. cfhl3, cfhl4, cfb and c9). The fact that complement pathway is activated indicates that plastic microparticles are integrated in immunological recognition processes. This was supported by fluorescence microscopy results, in which we observed co-localisation of neutrophils and macrophages around the PS particles. Identifying these key events can be a first building block to the development of an adverse outcome pathway (AOP). These data subsequently can be used within ecological and human risk assessment.

Preferential accumulation of gold nanorods into human skin hair follicles: Effect of nanoparticle surface chemistry.

HYPOTHESIS: Gold nanoparticles (GNP) are considered an ideal model to help understanding the nano-skin interface. The surface functionality of gold nanorods (GNR) is expected to influence the uptake of nanoparticles into specific targets of skin such as hair follicles or dermis. Hence, it should be possible to modify the surface chemistry of GNP to achieve more targeted and safe skin therapy. EXPERIMENTS: GNR functionalized with various surface ligands (neutral, anionic, cationic, and hydrophobic) were evaluated for their accumulation into hair follicles of human skin sheets using ex-vivo setup. The extent of GNR accumulation into hair follicles and other skin compartments was quantified by inductively coupled plasma-optical emission spectroscopy (ICP-OES), and their spatial distribution through skin layers was investigated by laser ablation-inductively coupled plasma-mass spectroscopy (LA-ICP-MS). RESULTS: The lipophilic properties of sebum-rich hair follicles enhanced the accumulation of hydrophobic polystyrene (PS)-GNR into hair follicles (∼13% of the total applied dose), while neutral polyethylene glycol (PEG)-GNR were distributed into all skin compartments, especially the dermis (∼11.5% of the total applied dose), which exhibits hydrophilic characteristics. Charged GNR showed a negligible percentage of penetration into any of the skin compartments. GNR could be a promising approach for targeted skin disease treatment and transdermal administration of drugs and therapy.