Fibrinogen enhances the inflammatory response of alveolar macrophages to TiO₂, SiO₂ and carbon nanomaterials

<p>Many studies have shown that the composition of the protein corona dramatically affects the response of cells to nanomaterials (NMs). However, the role of each single protein is still largely unknown. Fibrinogen (FG), one of the most abundant plasma proteins, is believed to mediate foreign-body reactions. Since this protein is absent in cell media used in <i>in vitro</i> toxicological tests the possible FG-mediated effects have not yet been assessed. Here, the effect of FG on the toxicity of three different kinds of inorganic NMs (carbon, SiO₂ and TiO₂) on alveolar macrophages has been investigated. A set of integrated techniques (UV–vis spectroscopy, dynamic light scattering and sodium dodecyl sulphate-polyacrylamide gel electrophoresis) have been used to study the strength and the kinetics of interaction of FG with the NMs. The inflammatory response of alveolar macrophages (MH-S) exposed to the three NMs associated with FG has also been investigated. We found that FG significantly enhances the cytotoxicity (lactate dehydrogenase leakage) and the inflammatory response (increase in nitric oxide (NO) concentration and NO synthase activation) induced by SiO₂, carbon and TiO₂ NMs on alveolar macrophages. This effect appears related to the amount of FG interacting with the NMs. In the case of carbon NMs, the activation of fibrinolysis, likely related to the exposure of cryptic sites of FG, was also observed after 24 h. These findings underline the critical role played by FG in the toxic response to NMs.</p

Crystalline Phase Modulates the Potency of Nanometric TiO₂ to Adhere to and Perturb the Stratum Corneum of Porcine Skin under Indoor Light

Nanometric TiO₂ is largely employed in cosmetics, but in vitro toxic effects have been reported when nano-TiO₂ is exposed to UV light. The photoreactivity of TiO₂ largely depends on its crystal phase, namely, anatase and rutile. Surface acidity, which is also dependent on crystal structure, may impart a positive or negative charge to the nanomaterial surface and ultimately modulate particle adhesion to tissues. Three nanometric TiO₂ powders with a different crystal lattice and surface charge (anatase, rutile, and anatase/rutile) have been employed here to investigate their interaction with the skin and to examine the molecular mechanisms of the TiO₂-induced oxidative damage. The strength of the interaction of nano-TiO₂ with skin has been revealed by chemiometric mapping (μ-XRF and SEM–EDS) after tissue washing. Positively charged anatase and anatase/rutile, but not negatively charged rutile, were strongly held on the skin surface and were able to promote a structural rearrangement of the lipid bilayer in the stratum corneum (DSC and Raman) under controlled indoor illumination (UVA < 1 mW/m²). Under the same conditions, cell-free reactivity tests (ROS-mediated free-radical release and lipoperoxidation) indicated that anatase and anatase/rutile are more reactive than rutile, suggesting a ROS-mediated oxidative mechanism that may alter the structure of the stratum corneum. Both the higher oxidative potential and the stronger adhesion to skin of anatase and anatase/rutile TiO₂ may explain the stronger disorganization induced by these two samples and suggests the use of rutile to produce safer TiO₂-based cosmetic and pharmaceutical products

Identification of physicochemical properties that modulate nanoparticle aggregation in blood

Inorganic materials are receiving significant interest in medicine given their usefulness for therapeutic applications such as targeted drug delivery, active pharmaceutical carriers and medical imaging. However, poor knowledge of the side effects related to their use is an obstacle to clinical translation. For the development of molecular drugs, the concept of safe-by-design has become an efficient pharmaceutical strategy with the aim of reducing costs, which can also accelerate the translation into the market. In the case of materials, the application these approaches is hampered by poor knowledge of how the physical and chemical properties of the material trigger the biological response. Hemocompatibility is a crucial aspect to take into consideration for those materials that are intended for medical applications. The formation of nanoparticle agglomerates can cause severe side effects that may induce occlusion of blood vessels and thrombotic events. Additionally, nanoparticles can interfere with the coagulation cascade causing both pro-and anti-coagulant properties. There is contrasting evidence on how the physicochemical properties of the material modulate these effects. In this work, we developed two sets of tailored carbon and silica nanoparticles with three different diameters in the 100-500 nm range with the purpose of investigating the role of surface curvature and chemistry on platelet aggregation, activation and adhesion. Substantial differences were found in the composition of the protein corona depending on the chemical nature of the nanoparticles, while the surface curvature was found to play a minor role. On the other hand, large carbon nanoparticles (but not small carbon nanoparticles or silica nanoparticles) have a clear tendency to form aggregates both in plasma and blood. This effect was observed both in the presence or absence of platelets and was independent of platelet activation. Overall, the results presented herein suggest the existence of independent modes of action that are differently affected by the physicochemical properties of the materials, potentially leading to vessel occlusion and/or formation of thrombi in vivo

Identification of the physical-chemical properties that modulate the nanoparticles aggregation in blood

Inorganic materials are receiving significant interest in medicine given their usefulness for therapeutic applications such as targeted drug delivery, carriers of active pharmaceutical and medical imaging. However, the poor knowledge of the side effects related to their use is an obstacle to their clinical translation. For the molecular drug development, safe-by-design has become as a novel pharmaceutical strategy that allows a reduction of the costs and an acceleration of the translation of research to market. In the case of materials, the application of such approaches is hampered by a poor knowledge of how the physical and chemical properties of the material trigger biological response. Hemocompatibility is a crucial factor for those materials that are intended for medical applications. In particular, the formation of agglomerates is a serious side effect that may induce occlusion of blood vessels and thrombotic events. Additionally, nanoparticles can interfere with the coagulation cascades where they have been reported to induce both pro- and anti-coagulant properties where their properties like size, shape and surface charge have been see to be critical parameters.   Here, we developed two sets of tailored carbon and silica nano/submicron-particles with three different sizes (100-500 nm) with the purpose of investigating the role of surface curvature and chemistry on platelet aggregation, activation and adhesion. We show that that large carbon nanoparticles, but not small carbon nanoparticles or silica nanoparticles, have a strong tendency to form aggregates both in plasma and blood, as a consequence of the formation of a protein corona and not of platelets activation. Substantial differences were found in the composition of the protein corona depending upon the chemical nature of the nanoparticles, while the surface curvature plays a minor role. On the other hand, coagulation proteins were abundant in the corona of both silica and carbon nanoparticles.  The results presented herein suggest that vessel occlusion and formation of thrombi in vivo may occur through independent mode of action (MoA), differently affected by the physico-chemical properties of the materials

Distinctive Toxicity of TiO₂ Rutile/Anatase Mixed Phase Nanoparticles on Caco-2 Cells

Titanium dioxide has a long-standing use as a food additive. Micrometric powders are, e.g., applied as whiteners in confectionary or dairy products. Possible hazards of ingested nanometric TiO₂ particles for humans and the potential influence of varying specific surface area (SSA) are currently under discussion. Five TiO₂-samples were analyzed for purity, crystallinity, primary particle size, SSA, ζ potential, and aggregation/agglomeration. Their potential to induce cytotoxicity, oxidative stress, and DNA damage was evaluated in human intestinal Caco-2 cells. Only anatase-rutile containing samples, in contrast to the pure anatase samples, induced significant LDH leakage or mild DNA damage (Fpg-comet assay). Evaluation of the metabolic competence of the cells (WST-1 assay) revealed a highly significant correlation between the SSA of the anatase samples and cytotoxicity. The anatase/rutile samples showed higher toxicity per unit surface area than the pure anatase powders. However, none of the samples affected cellular markers of oxidative stress. Our findings suggest that both SSA and crystallinity are critical determinants of TiO₂-toxicity toward intestinal cells

Additional file 5: Figure S5. of Multi-walled carbon nanotubes directly induce epithelial-mesenchymal transition in human bronchial epithelial cells via the TGF-β-mediated Akt/GSK-3β/SNAIL-1 signalling pathway

Representative microscope images of BEAS-2B cells (A-B) and relative expression of epithelial and mesenchymal markers (C). A-B) BEAS-2B cells were incubated for 96 h in either the absence (0 μg/ml, CTRL) or presence of [Fe3+] (released by FeCl3) similar to the one potentially released at the highest concentration of MWCNT used, Fe-depleted MWCNTg (44 μg/ml) and MWCNTg (44 μg/ml). After the incubation, the cells were rinsed with PBS and observed by OM (A) or fixed with paraformaldehyde and fluorescently labelled as described in Methods (B). A) Representative images are shown (10×; scale bar = 50 μm). B) Actin filaments were visualized in red, nuclei in blue. Representative images are shown (63×; scale bar = 10 μm). C). Relative expression of epithelial and mesenchymal markers were checked by WB. Relative expression of E-cadherin, β-catenin, α-SMA and vimentin proteins in BEAS-2B cells incubated in either the absence (0 μg/ml, CTRL, lane 1) or presence of [Fe3+] (released by FeCl3, lane 2), Fe-depleted MWCNTg (44 μg/ml, lane 3) and MWCNTg (44 μg/ml, lane 4). GAPDH was used as loading control for cytosolic extracts. Each figure is representative of three experiments giving similar results. (TIF 1558 kb

Efficacy, biocompatibility and degradability of carbon nanoparticles for photothermal therapy of lung cancer

Aim: To investigate near infrared-induced phototoxicity toward lung cancer cells, and the biodegradability and effect on immune cells of glucose-derived carbon nanoparticles (CNPs).  Methods: The human A549 lung adenocarcinoma cell line was used as a model to study the phototoxicity of CNPs. The biodegradability and the effect on immune cells was demonstrated in primary human neutrophils and macrophages.  Results: Near infrared-Activated CNPs elicited rapid cell death, characterized by the elevation of heat shock proteins and the induction of DNA damage. CNPs were found to be noncytotoxic toward primary human macrophages and were susceptible to biodegradation when cocultured with human neutrophils.  Conclusions: Our results identify CNPs as promising platforms for photothermal therapy of lung cancer.</p