Search

The targeting of natural tolerogenic liver sinusoidal endothelial cells (LSEC) by nanoparticles (NPs), decorated with a stabilin receptor ligand, is capable of generating regulatory T-cells (Tregs), which can suppress antigen-specific immune responses, including to ovalbumin (OVA), a possible food allergen. In this regard, we have previously demonstrated that OVA-encapsulating poly(lactic-co-glycolic acid) (PLGA) nanoparticles eliminate allergic airway inflammation in OVA-sensitized mice, prophylactically and therapeutically. A competing approach is a nanocarrier platform that incorporates pharmaceutical agents interfering in mTOR (rapamycin) or NF-κB (curcumin) pathways, with the ability to induce a tolerogenic state in nontargeted antigen-presenting cells system-wide. First, we compared OVA-encapsulating, LSEC-targeting tolerogenic nanoparticles (TNPs) with nontargeted NPs incorporating curcumin and rapamycin (Rapa) in a murine eosinophilic airway inflammation model, which is Treg-sensitive. This demonstrated roughly similar tolerogenic effects on allergic airway inflammation by stabilin-targeting NPOVA versus nontargeted NPs delivering OVA plus Rapa. Reduction in eosinophilic inflammation and TH2-mediated immune responses in the lung was accompanied by increased Foxp3+ Treg recruitment and TGF-β production in both platforms. As OVA incorporates IgE-binding as well as non-IgE-binding epitopes, the next experiment explored the possibility of obtaining immune tolerance by non-anaphylactic T-cell epitopes. This was accomplished by incorporating OVA323-339 and OVA257-264 epitopes in liver-targeting NPs to assess the prophylactic and therapeutic impact on allergic inflammation in transgenic OT-II mice. Importantly, we demonstrated that the major histocompatibility complex (MHC)-II binding (former) but not the MHC-I binding (latter) epitope interfered in allergic airway inflammation, improving TNPOVA efficacy. The epitope-specific effect was transduced by TGF-β-producing Tregs. In the final phase of experimentation, we used an OVA-induced anaphylaxis model to demonstrate that targeted delivery of OVA and its MHC-II epitope could significantly suppress the anaphylaxis symptom score, mast cell release, and the late-phase inflammatory response. In summary, these results demonstrate comparable efficacy of LSEC-targeting versus pharmaceutical PLGA nanoparticles, as well as the ability of T-cell epitopes to achieve response outcomes similar to those of the intact allergens.

Nanotechnology is enjoying an impressive growth and the global nanotechnology industry is expected to exceed US$ 125 billion by 2024. Based on these successes, there are notions that enough is known and efforts on engineered nanomaterial environmental health and safety (nano-EHS) research should be put on the back burner. However, there are recent events showing that it is not the case. The US Food and Drug Administration found ferumoxytol (carbohydrate-coated superparamagnetic iron oxide nanoparticle) for anemia treatment could induce lethal anaphylactic reactions. The European Union will categorize TiO2 as a category 2 carcinogen due to its inhalation hazard and France banned use of TiO2 (E171) in food from January 1, 2020 because of its carcinogenic potential. Although nanoindustry is seemingly in a healthy state, growth could be hindered for the lack of certainty and more nano-EHS research is needed for the sustainable growth of nanoindustry. Herein, the current knowledge gaps and the way forward are elaborated.

The mononuclear phagocyte system in the liver is a frequent target for nanoparticles (NPs). A toxicological profiling of metal-based NPs is performed in Kupffer cell (KC) and hepatocyte cell lines. Sixteen NPs are provided by the Nanomaterial Health Implications Research Consortium of the National Institute of Environmental Health Sciences to study the toxicological effects in KUP5 (KC) and Hepa 1-6 cells. Five NPs (Ag, CuO, ZnO, SiO2 , and V2 O5 ) exhibit cytotoxicity in both cell types, while SiO2 and V2 O5 induce IL-1β production in KC. Ag, CuO, and ZnO induced caspase 3 generated apoptosis in both cell types is accompanied by ion shedding and generation of mitochondrial reactive oxygen species (ROS) in both cell types. However, the cell death response to SiO2 in KC differs by inducing pyroptosis as a result of potassium efflux, caspase 1 activation, NLRP3 inflammasome assembly, IL-1β release, and cleavage of gasdermin-D. This releases pore-performing peptide fragments responsible for pyroptotic cell swelling. Interestingly, although V2 O5 induces IL-1β release and delays caspase 1 activation by vanadium ion interference in membrane Na+ /K+ adenosine triphosphate (ATP)ase activity, the major cell death mechanism in KC (and Hepa 1-6) is caspase 3 mediated apoptosis. These findings improve the understanding of the mechanisms of metal-based engineered nanomaterial (ENM) toxicity in liver cells toward comprehensive safety evaluation.

The widespread use of metal oxide nanoparticles (MOx NPs) poses a risk of exposure that may lead to adverse health effects on humans. Even though a number of toxicological methodologies are available for assessing nanotoxicity, the effect of MOx NPs on cell metabolism in vitro and in vivo remains largely unknown, especially under the exposure to low-dose or supposedly low-toxicity MOx NPs. In this study, liquid chromatography-mass spectrometry (LC-MS) based metabolomics was used to reveal significantly altered metabolites and metabolic pathways in human bronchial epithelial cells exposed to four different types of MOx NPs (ZnO, SiO2, TiO2, and CeO2) at both high (25 μg/mL) and low (12.5 μg/mL) doses. We demonstrated that high-dose ZnO NPs caused severe cytotoxicity with altered metabolism of amino acids, nucleotides, nucleosides, tricarboxylic acid cycle, lipids, inflammation/redox, and fatty acid oxidation, as well as the elevation of toxic and DNA damage related metabolites. Fewer metabolomic alterations were induced by low-dose ZnO NPs. However, most metabolites significantly altered by high-dose ZnO NPs were also slightly changed by low-dose ZnO NPs. On the other hand, the cells exposed to SiO2, TiO2, and CeO2 NPs at either high or low dose displayed low cytotoxicity with similar metabolomic alterations, although each type of NPs induced distinct changes of certain metabolites. These three NPs significantly affected the metabolic pathways of sphingosine-1-phosphate, fatty acid oxidation, folate cycle, inflammation/redox, and lipid metabolism. In addition, dose-dependent effects were observed for a number of metabolites significantly altered by respective MOx NPs. Representative metabolites of the significantly altered metabolic pathways were successfully validated in vitro using enzymatic assays. More importantly, these representative metabolites were further validated in a mouse model after lung exposure to respective NPs, indicating that in vitro metabolomic findings may be used to effectively predict the toxicological effects in vivo. Despite functional assay results demonstrating that the changes in cellular functions were largely reflected by the metabolomic alterations, LC-MS-based metabolomics was sensitive enough to detect the subtle metabolomic changes when functional cellular assays showed no significant difference. Collectively, our studies have unveiled potential metabolic mechanisms of MOx NP-induced nanotoxicity in lung epithelial cells and demonstrated the sensitivity and feasibility of using metabolomic signatures to understand and predict nanotoxicity in vivo.

Molecular imaging can provide functional and molecular information at the cellular or subcellular level in vivo in a noninvasive manner. Activatable nanoprobes that can react to the surrounding physiological environment or biomarkers are appealing agents to improve the efficacy, specificity, and sensitivity of molecular imaging. The physiological parameters, including redox status, pH, presence of enzymes, and hypoxia, can be designed as the stimuli of the activatable probes. However, the success rate of imaging nanoprobes for clinical translation is low. Herein, the recent advances in nanoparticle-based activatable imaging probes are critically reviewed. In addition, the challenges for clinical translation of these nanoprobes are also discussed in this review.

Therapeutic strategies to promote nerve cell growth and improve their functions or stimulate nerve fiber reconnection and ameliorate the loss of neuronal functions are in high demand. A disadvantage of current conventional methods, which includes injection of nerve growth factors (NGF) either systemically or in the affected area, is rapid clearance or degradation of NGF, thereby reducing the effective concentration of NGFs that can reach the damaged nerves to stimulate the healing process. To overcome this obstacle, a nanoparticle platform based on mesoporous silica nanoparticles (MSNs) was developed to not only prevent clearance and degradation of NGFs, but also deliver the NGF directly to nerve cells to promote nerve cell proliferation and neurite growth. We synthesized (NGF)-loaded MSN (MSN-NGF) with a diameter of 65 nm. MSN-NGF significantly promoted the differentiation of neuron-like PC12 cells and growth of neurites compared to NGF alone, as confirmed by MTS cell proliferation assay and optical microscopy analysis. This study shows that MSN-NGF could be an effective therapy to speed up nerve cell growth or recovery of function.

Nanocellulose-derived carbon is a promising material in energy storage because of its sustainability, low environmental impact, and large specific surface area. Herein, the skin secretion of Andrias davidianus (SSAD) is applied for the first time as the bio-nitrogen source to dope carbon aerogels from nanocellulose. Cellulose nanocrystal (CNC) is discovered to be very effective to address the dispersion problem of SSAD in water. After being homogeneously mixed with cellulose nanofiber (CNF), honeycomb-structured nanofibrous carbon aerogels are obtained via unidirectional freeze-drying of the SSAD/CNC/CNF mixture followed by high-temperature carbonization. Impressively, unlike those fragile carbon aerogels in many early works, the present ones exhibit outstanding elasticity in repeated compression and release tests. Moreover, a symmetric binder-free supercapacitor is assembled from the carbon aerogels, which exhibits improved electrochemical capacitive properties and cycling stability. And even after 500 compression and release cycles, the supercapacitor can still maintain high capacitive performance, indicating its superiorities in durability and electrochemical stability.

The surge of applications for engineered nanomaterials (ENMs) across multiple industries raises safety concerns regarding human health and environmental impacts. ENMs can be hazardous through various mechanisms, including, particle dissolution and shedding of toxic metal ions, surface reactivity and perturbation of cellular membranes, lysosomal membrane damage, activation of inflammation pathways (e.g., NLRP3 inflammasome), etc. The aim of this review is therefore to discuss practical approaches for the safer design of ENMs through modification of their physicochemical properties that can lead to acute and/or chronic toxicity. This is premised on our understanding of how different ENMs induce toxicity within various biological systems. We will summarize studies that have investigated nanomaterial toxicity both in vitro and in vivo to understand the underlying mechanisms by which nanoparticles can cause inflammation, fibrosis, and cell death. With this knowledge, researchers have identified several design strategies to counter these mechanisms of toxicity. In particular, we will discuss how metal doping, surface coating and covalent functionalization, and adjustment of surface oxidation state and aspect ratio of ENMs could reduce their potential adverse effects. While these strategies might be effective under certain experimental and exposure scenarios, more research is required to fully apply this knowledge in real life applications of nanomaterials.

© 2015 American Chemical Society. The production of pyrogenic (fumed) silica is increasing worldwide at a 7% annual growth rate, including expanded use in food, pharmaceuticals, and other industrial products. Synthetic amorphous silica, including fumed silica, has been generally recognized as safe for use in food products by the Food and Drug Administration. However, emerging evidence from experimental studies now suggests that fumed silica could be hazardous due to its siloxane ring structure, high silanol density, and string-of-pearl-like aggregate structure, which could combine to cause membrane disruption, generation of reactive oxygen species, pro-inflammatory effects, and liver fibrosis. Based on this structure-activity analysis (SAA), we investigated whether calcination and rehydration of fumed silica changes its hazard potential in the lung due to an effect on silanol density display. This analysis demonstrated that the accompanying change in surface reactivity could indeed impact cytokine production in macrophages and acute inflammation in the lung, in a manner that is dependent on siloxane ring reconstruction. Confirmation of this SAA in vivo, prompted us to consider safer design of fumed silica properties by titanium and aluminum doping (0-7%), using flame spray pyrolysis. Detailed characterization revealed that increased Ti and Al doping could reduce surface silanol density and expression of three-membered siloxane rings, leading to dose-dependent reduction in hydroxyl radical generation, membrane perturbation, potassium efflux, NLRP3 inflammasome activation, and cytotoxicity in THP-1 cells. The reduction of NLRP3 inflammasome activation was also confirmed in bone-marrow-derived macrophages. Ti doping, and to a lesser extent Al doping, also ameliorated acute pulmonary inflammation, demonstrating the possibility of a safer design approach for fumed silica, should that be required for specific use circumstances.

The safe implementation of nanotechnology requires nanomaterial hazard assessment in accordance with the material physicochemical properties that trigger the injury response at the nano/bio interface. Since CuO nanoparticles (NPs) are widely used industrially and their dissolution properties play a major role in hazard potential, we hypothesized that tighter bonding of Cu to Fe by particle doping could constitute a safer-by-design approach through decreased dissolution. Accordingly, we designed a combinatorial library in which CuO was doped with 1–10% Fe in a flame spray pyrolysis (FSP) reactor. The morphology and structural properties were determined by XRD, BET, Raman spectroscopy, HRTEM, EFTEM and EELS, which demonstrated a significant reduction in the apical Cu-O bond length while simultaneously increasing the planar bond length (Jahn-Teller distortion). Hazard screening was performed in tissue culture cell lines and zebrafish embryos to discern the change in the hazardous effects of doped vs. non-doped particles. This demonstrated that with increased levels of doping, there was a progressive decrease in cytotoxicity in BEAS-2B and THP-1 cells, as well as an incremental decrease in the rate of hatching interference in zebrafish embryos. The dissolution profiles were determined and the surface reactions taking place in Holtfreter’s solution were validated using cyclic voltammetry (CV) measurements to demonstrate the Cu+/Cu2+ and Fe2+/Fe3+ redox species playing a major role in the dissolution process of pure and Fe doped CuO. Altogether, a safe-by-design strategy was implemented for the toxic CuO particles via Fe doping and has been demonstrated for their safe use in the environment.

Nanocellulose is increasingly considered for applications; however, the fibrillar nature, crystalline phase, and surface reactivity of these high aspect ratio nanomaterials need to be considered for safe biomedical use. Here a comprehensive analysis of the impact of cellulose nanofibrils (CNF) and nanocrystals (CNC) is performed using materials provided by the Nanomaterial Health Implications Research Consortium of the National Institute of Environmental Health Sciences. An intermediary length of nanocrystals is also derived by acid hydrolysis. While all CNFs and CNCs are devoid of cytotoxicity, 210 and 280 nm fluorescein isothiocyanate (FITC)-labeled CNCs show higher cellular uptake than longer and shorter CNCs or CNFs. Moreover, CNCs in the 200-300 nm length scale are more likely to induce lysosomal damage, NLRP3 inflammasome activation, and IL-1β production than CNFs. The pro-inflammatory effects of CNCs are correlated with higher crystallinity index, surface hydroxyl density, and reactive oxygen species generation. In addition, CNFs and CNCs can induce maturation of bone marrow-derived dendritic cells and CNCs (and to a lesser extent CNFs) are found to exert adjuvant effects in ovalbumin (OVA)-injected mice, particularly for 210 and 280 nm CNCs. All considered, the data demonstrate the importance of length scale, crystallinity, and surface reactivity in shaping the innate immune response to nanocellulose.

The lack of control over microvasculature formation remains a key roadblock to the therapeutic vascularization and regeneration of functional tissues. In the current study, the integration of plasmid DNA (pDNA) condensation and electrospraying technologies was proposed to promote the regeneration of mature blood vessels through injectable or infusible administration of microparticles. Calcium phosphate (CP) nanoparticles with encapsulated plasmids encoding vascular endothelial growth factors (pVEGF) and basic fibroblast growth factor (pbFGF) were synthesized using reverse microemulsions. Electrosprayed microparticles with the loading of CP-pDNA nanoparticles were evaluated on both endothelial cells and smooth muscle cells and after subcutaneous infusion into animals. CP-pDNA nanoparticles was obtained with an average size of around 110 nm and electrosprayed into microparticles, resulting in high loading efficiency and extended protection on pDNA from external DNase environment. The inoculation of poly(ethylene glycol) into microparticle matrices realized a gradual release for 4 weeks of CP-pDNA nanoparticles, leading to an incremental transfection efficiency and strong secretion of extracellular matrices. After subcutaneous infusion of microparticles with encapsulated both CP-pVEGF and CP-pbFGF nanoparticles, significantly higher densities of blood vessels were achieved than those containing individual nanoparticles, and induced a rapid generation of mature blood vessels with few cytotoxicity and inflammation reactions. Electrosprayed microparticle with CP-pDNA nanoparticles encapsulated promoted the formation of vascular networks, providing clinical relevance for therapeutic vascularization and regeneration of functional tissues after injection to ischemic sites or entrapment into tissue engineering scaffolds.

Background: Differences in interlaboratory research protocols contribute to the conflicting data in the literature regarding engineered nanomaterial (ENM) bioactivity. Objectives: Grantees of a National Institute of Health Sciences (NIEHS)-funded consortium program performed two phases of in vitro testing with selected ENMs in an effort to identify and minimize sources of variability. Methods: Consortium program participants (CPPs) conducted ENM bioactivity evaluations on zinc oxide (ZnO), three forms of titanium dioxide (TiO2), and three forms of multiwalled carbon nanotubes (MWCNTs). In addition, CPPs performed bioassays using three mammalian cell lines (BEAS-2B, RLE-6TN, and THP-1) selected in order to cover two different species (rat and human), two different lung epithelial cells (alveolar type II and bronchial epithelial cells), and two different cell types (epithelial cells and macrophages). CPPs also measured cytotoxicity in all cell types while measuring inflammasome activation [interleukin-1β (IL-1β) release] using only THP-1 cells. Results: The overall in vitro toxicity profiles of ENM were as follows: ZnO was cytotoxic to all cell types at ≥ 50 μg/mL, but did not induce IL-1β. TiO2 was not cytotoxic except for the nanobelt form, which was cytotoxic and induced significant IL-1β production in THP-1 cells. MWCNTs did not produce cytotoxicity, but stimulated lower levels of IL-1β production in THP-1 cells, with the original MWCNT producing the most IL-1β. Conclusions: The results provide justification for the inclusion of mechanism-linked bioactivity assays along with traditional cytotoxicity assays for in vitro screening. In addition, the results suggest that conducting studies with multiple relevant cell types to avoid false-negative outcomes is critical for accurate evaluation of ENM bioactivity.

We used a multi-functional mesoporous silica nanoparticle (MSNP) carrier to overcome doxorubicin (Dox) resistance in a multidrug resistant (MDR) human breast cancer xenograft by co-delivering Dox and siRNA that targets the P-glycoprotein (Pgp) drug exporter. The Pgp siRNA selection from among a series of drug resistance targets was achieved by performing high throughput screening in a MDR breast cancer cell line, MCF-7/MDR. Following the establishment of a MCF-7/MDR xenograft model in nude mice, we demonstrated that a 50 nm MSNP, functionalized by a polyethyleneimine-polyethylene glycol (PEI-PEG) copolymer, provides protected delivery of stably bound Dox and Pgp siRNA to the tumor site. The effective biodistribution and reduced reticuloendothelial uptake as a result of our nanocarrier design, allowed us to achieve an 8% enhanced permeability and retention effect at the tumor site. Compared to free Dox or the carrier loaded with either drug or siRNA alone, the dual delivery system resulted in synergistic inhibition of tumor growth in vivo. Analysis of multiple xenograft biopsies demonstrated significant Pgp knockdown at heterogeneous tumor sites, which correspond to the regions where Dox was released intracellularly and induced apoptosis. We emphasize that the heterogeneity originates in the tumor microenvironment, which influence the vascular access, rather than heterogeneous Pgp expression in the MDR cells. Taken together, these data provide proof-of-principle testing of the use of a dual drug/siRNA nanocarrier to overcome Dox resistance in a xenograft. The study also provides the first detailed analysis of the impact of heterogeneity in the tumor microenvironment on the efficacy of siRNA delivery in vivo.

Background Although classified as metal oxides, cobalt monoxide (CoO) and lanthanum oxide (La2O3) nanoparticles, as representative transition and rare earth oxides, exhibit distinct material properties that may result in different hazardous potential in the lung. The current study was undertaken to compare the pulmonary effects of aerosolized whole body inhalation of these nanoparticles in mice. Results Mice were exposed to filtered air (control) and 10 or 30 mg/m3 of each particle type for 4 days and then examined at 1 h, 1, 7 and 56 days post-exposure. The whole lung burden 1 h after the 4 day inhalation of CoO nanoparticles was 25 % of that for La2O3 nanoparticles. At 56 days post exposure, < 1 % of CoO nanoparticles remained in the lungs; however, 22–50 % of the La2O3 nanoparticles lung burden 1 h post exposure was retained at 56 days post exposure for low and high exposures. Significant accumulation of La2O3 nanoparticles in the tracheobronchial lymph nodes was noted at 56 days post exposure. When exposed to phagolysosomal simulated fluid, La nanoparticles formed urchin-shaped LaPO4 structures, suggesting that retention of this rare earth oxide nanoparticle may be due to complexation of cellular phosphates within lysosomes. CoO nanoparticles caused greater lactate dehydrogenase release in the bronchoalveolar fluid (BALF) compared to La2O3 nanoparticles at 1 day post exposure, while BAL cell differentials indicate that La2O3 nanoparticles generated more inflammatory cell infiltration at all doses and exposure points. Histopathological analysis showed acute inflammatory changes at 1 day after inhalation of either CoO or La2O3 nanoparticles. Only the 30 mg/m3 La2O3 nanoparticles exposure caused chronic inflammatory changes and minimal fibrosis at day 56 post exposure. This is in agreement with activation of the NRLP3 inflammasome after in vitro exposure of differentiated THP-1 macrophages to La2O3 but not after CoO nanoparticles exposure. Conclusion Taken together, the inhalation studies confirmed the trend of our previous sub-acute aspiration study, which reported that CoO nanoparticles induced more acute pulmonary toxicity, while La2O3 nanoparticles caused chronic inflammatory changes and minimal fibrosis. Electronic supplementary material The online version of this article (doi:10.1186/s12989-016-0155-3) contains supplementary material, which is available to authorized users.

In this study, we have generated and characterized Electronic Cigarette (EC) aerosols using a combination of advanced technologies. In the gas phase, the particle number concentration (PNC) of EC aerosols was found to be positively correlated with puff duration whereas the PNC and size distribution may vary with different flavors and nicotine strength. In the liquid phase (water or cell culture media), the size of EC nanoparticles appeared to be significantly larger than those in the gas phase, which might be due to aggregation of nanoparticles in the liquid phase. By using in vitro high-throughput cytotoxicity assays, we have demonstrated that EC aerosols significantly decrease intracellular levels of glutathione in NHOKs in a dose-dependent fashion resulting in cytotoxicity. These findings suggest that EC aerosols cause cytotoxicity to oral epithelial cells in vitro, and the underlying molecular mechanisms may be or at least partially due to oxidative stress induced by toxic substances (e.g., nanoparticles and chemicals) present in EC aerosols.

We have developed structure/toxicity relationships for amorphous silica nanoparticles (NPs) synthesized through low-temperature colloidal (e.g., Stöber silica) or high-temperature pyrolysis (e.g., fumed silica) routes. Through combined spectroscopic and physical analyses, we have determined the state of aggregation, hydroxyl concentration, relative proportion of strained and unstrained siloxane rings, and potential to generate hydroxyl radicals for Stöber and fumed silica NPs with comparable primary particle sizes (16 nm in diameter). On the basis of erythrocyte hemolytic assays and assessment of the viability and ATP levels in epithelial and macrophage cells, we discovered for fumed silica an important toxicity relationship to postsynthesis thermal annealing or environmental exposure, whereas colloidal silicas were essentially nontoxic under identical treatment conditions. Specifically, we find for fumed silica a positive correlation of toxicity with hydroxyl concentration and its potential to generate reactive oxygen species (ROS) and cause red blood cell hemolysis. We propose fumed silica toxicity stems from its intrinsic population of strained three-membered rings (3MRs) along with its chainlike aggregation and hydroxyl content. Hydrogen-bonding and electrostatic interactions of the silanol surfaces of fumed silica aggregates with the extracellular plasma membrane cause membrane perturbations sensed by the Nalp3 inflammasome, whose subsequent activation leads to secretion of the cytokine IL-1β. Hydroxyl radicals generated by the strained 3MRs in fumed silica, but largely absent in colloidal silicas, may contribute to the inflammasome activation. Formation of colloidal silica into aggregates mimicking those of fumed silica had no effect on cell viability or hemolysis. This study emphasizes that not all amorphous silicas are created equal and that the unusual toxicity of fumed silica compared to that of colloidal silica derives from its framework and surface chemistry along with its fused chainlike morphology established by high-temperature synthesis (1300 °C) and rapid thermal quenching.

While it has been shown that high aspect ratio nanomaterials like carbon nanotubes and TiO(2) nanowires can induce toxicity by acting as fiber-like substances that damage the lysosome, it is not clear what the critical lengths and aspect ratios are that induce this type of toxicity. To answer this question, we synthesized a series of cerium oxide (CeO(2)) nanorods and nanowires with precisely controlled lengths and aspect ratios. Both phosphate and chloride ions were shown to play critical roles in obtaining these high aspect ratio nanostructures. High-resolution TEM analysis shows that single-crystalline CeO(2) nanorods/nanowires were formed along the [211] direction by an "oriented attachment" mechanism, followed by Ostwald ripening. The successful creation of a comprehensive CeO(2) nanorod/nanowire combinatorial library allows, for the first time, the systematic study of the effect of aspect ratio on lysosomal damage, cytotoxicity, and IL-1β production by the human myeloid cell line (THP-1). This in vitro toxicity study demonstrated that, at lengths ≥200 nm and aspect ratios ≥22, CeO(2) nanorods induced progressive pro-inflammatory effects and cytotoxicity. The relatively low "critical" length and aspect ratio were associated with small nanorod/nanowire diameters (6-10 nm), which facilitates the formation of stacking bundles due to strong van der Waals and dipole-dipole attractions. Our results suggest that both length and diameter components of aspect ratio should be considered when addressing the cytotoxic effects of high aspect ratio materials.

Although the aspect ratio (AR) of engineered nanomaterials (ENMs) is one of the key physicochemical parameters that could determine biological outcome, not much is understood about how AR contributes to shaping biological outcome. By using a mesoporous silica nanoparticle (MSNP) library that has been constructed to cover a range of different lengths, we could demonstrate that the AR of rod-shaped particles determines the rate and abundance of MSNP uptake by a macropinocytosis process in HeLa and A549 cancer cell lines. MSNPs with an AR of 2.1-2.5 were taken up in larger quantities compared to shorter or longer length rods by a process that is sensitive to amiloride, cytochalasin D, azide, and 4 °C inhibition. The rods with intermediary AR also induced the maximal number of filopodia, actin polymerization, and activation of small GTP-binding proteins (e.g., Rac1, CDC42) that involve assembly of the actin cytoskeleton and filopodia formation. When assessing the role of AR in the delivery of paclitaxel or camptothecin, the rods with AR 2.1-2.5 were clearly more efficient for drug delivery and generation of cytotoxic killing in HeLa cells. All considered, our data suggest an active sensoring mechanism by which HeLa and A549 cells are capable of detecting AR differences in MSNP to the extent that accelerated macropinocytosis can be used to achieve more efficient drug delivery.

Most in vitro toxicity studies on engineered nanomaterials (ENMs) use transformed rather than primary cells for logistical reasons. However, primary cells may provide a more appropriate connection to in vivo toxicity because these cells maintain their phenotypic fidelity and are also capable of differentiating into lineages that may be differently affected by potentially hazardous ENM. Few studies to date have focused on the role of cellular differentiation in determining ENM toxicity. We compared the response of undifferentiated and differentiated primary human bronchial epithelial cells (NHBE) to cationic mesoporous silica nanoparticles (MSNP) that are coated with polyethyleneimine (PEI) since this polymer is known to exert differential cytotoxicity depending on its molecular weight and cationic density. The attachment of cationic PEI polymers to the MSNP surface was used to assess these materials' toxicological potential in undifferentiated and differentiated human bronchial epithelial cells (NHBE), using of a multi-parametric assay that screens for an integrated set of sub-lethal and lethal response outcomes. MSNP coated with high molecular weight (10 and 25 kD) polymers were more toxic in differentiated cells than particles coated with shorter length polymers. The increased susceptibility of the differentiated cells is in agreement with more abundant expression of a proteoglycan, syndecan-1, which contains copious heparin sulfate side chains. Pre-treatment with heparinase to remove the negatively charged sulfates decreased MSNP-PEI binding to the cell surface and lowered the cytotoxic potential of the cationic particles. These data demonstrate the importance of studying cellular differentiation as an important variable in the response of primary cells to toxic ENM properties.

Overexpression of drug efflux transporters such as P-glycoprotein (P-gp) protein is one of the major mechanisms for multiple drug resistance (MDR) in cancer cells. A new approach to overcome MDR is to use a co-delivery strategy that utilizes a siRNA to silence the expression of efflux transporter together with an appropriate anti-cancer drug for drug resistant cells. In this paper, we report that mesoporous silica nanoparticles (MSNP) can be functionalized to effectively deliver a chemotherapeutic agent doxorubicin (Dox) as well as Pgp siRNA to a drug-resistant cancer cell line (KB-V1 cells) to accomplish cell killing in an additive or synergistic fashion. The functionalization of the particle surface with a phosphonate group allows electrostatic binding of Dox to the porous interior, from where the drug could be released by acidification of the medium under abiotic and biotic conditions. In addition, phosphonate modification also allows exterior coating with the cationic polymer, polyethylenimine (PEI), which endows the MSNP contemporaneously deliver Pgp siRNA. The dual delivery of Dox and siRNA in KB-V1 cells was capable of increasing the intracellular as well as intranuclear drug concentration to levels exceeding that of free Dox or the drug being delivered by MSNP in the absence of siRNA co-delivery. These results demonstrate that it is possible to use the MSNP platform to effectively deliver a siRNA that knocks down gene expression of a drug exporter that can be used to improve drug sensitivity to a chemotherapeutic agent.

Although mankind stands to obtain great benefit from nanotechnology, it is important to consider the potential health impacts of nanomaterials (NMs). This consideration has launched the field of nanotoxicology, which is charged with assessing toxicological potential as well as promoting safe design and use of NMs. Although no human ailments have been ascribed to NMs thus far, early experimental studies indicate that NMs could initiate adverse biological responses that can lead to toxicological outcomes. One of the principal mechanisms is the generation of reactive oxygen species and oxidant injury. Because oxidant injury is also a major mechanism by which ambient ultrafine particles can induce adverse health effects, it is useful to consider the lessons learned from studying ambient particles. This review discusses the toxicological potential of NMs by comparing the possible injury mechanisms and adverse health effects of engineered and ambient ultrafine particles.

Ambient particulate matter (PM) is an environmental factor that has been associated with increased respiratory morbidity and mortality. The major effect of ambient PM on the pulmonary system is the exacerbation of inflammation, especially in susceptible people. One of the mechanisms by which ambient PM exerts its proinflammatory effects is the generation of oxidative stress by its chemical compounds and metals. Cellular responses to PM-induced oxidative stress include activation of antioxidant defense, inflammation, and toxicity. The proinflammatory effect of PM in the lung is characterized by increased cytokine/chemokine production and adhesion molecule expression. Moreover, there is evidence that ambient PM can act as an adjuvant for allergic sensitization, which raises the possibility that long-term PM exposure may lead to increased prevalence of asthma. In addition to ambient PM, rapid expansion of nanotechnology has introduced the potential that engineered nanoparticles (NP) may also become airborne and may contribute to pulmonary diseases by novel mechanisms that could include oxidant injury. Currently, little is known about the potential adverse health effects of these particles. In this communication, the mechanisms by which particulate pollutants, including ambient PM and engineered NP, exert their adverse effects through the generation of oxidative stress and the impacts of oxidant injury in the respiratory tract will be reviewed. The importance of cellular antioxidant and detoxification pathways in protecting against particle-induced lung damage will also be discussed.

Ultrafine particles are airborne particulates of less than 100 nm in aerodynamic diameter. Examples of ultrafine particles are diesel exhaust particles, products of cooking, heating and wood burning in indoor environments, and more recently, products generated through the use of nanotechnology. Studies have shown that ambient ultrafine particles have detrimental effects on both the cardiovascular and respiratory systems, including a higher incidence of atherosclerosis and the exacerbation rate of asthma. Ultrafine particles have been found to alter in vitro and in vivo responses of the immune system to allergens and may also play a role in allergen sensitization. The inflammatory properties of ultrafine particles may be mediated by a number of different mechanisms, including the ability to produce reactive oxygen species, leading to the generation of pro-inflammatory cytokines and airway inflammation. In addition, because of their small size, ultrafine particles also have unique distribution characteristics in the respiratory tree and circulation and may be able to alter cellular function in ways that circumvent normal signaling pathways. Additionally, ultrafine particles can penetrate intracellularly and potentially cause DNA damage. The recent advances in nanotechnology, while opening up new opportunities for the advancement of technology and medicine, could also lead to unforeseen adverse health effects in exposed humans. Further research is needed to clarify the safety of nanoscale particles, as well as the elucidation of the possible beneficial use of these particulates to treat disease.

By enabling nanoparticle-based drug delivery system to actively target cancer cells with high selectivity, active targeted molecules have attracted great attention in the application of nanoparticles for anticancer drug delivery. However, the clinical application of most active targeted molecules in breast cancer therapy is limited, due to the low expression of their receptors in breast tumors or coexpression in the normal and tumor breast tissues. Here, a neuropeptide Y Y1 receptors ligand PNBL-NPY, as a novel targeted molecule, is conjugated with anticancer drug doxorubicin encapsulating albumin nanoparticles to investigate the effect of Y1 receptors on the delivery of drug-loaded nanoparticles to breast cancer cells and its potential for breast cancer therapy. The PNBL-NPY can actively recognize and bind to the Y1 receptors that are significantly overexpressed on the surface of the breast cancer cells, and the drug-loaded nanoparticles are delivered directly into the cancer cells through internalization. This system is highly selective and able to distinguish the breast cancer cells from the normal cells, due to normal breast cells that express Y2 receptors only. It is anticipated that this study may provide a guidance in the development of Y1 receptor-based nanoparticulate drug delivery system for a safer and more efficient breast cancer therapy.

© 2015 American Chemical Society. Upconversion nanoparticles (UCNPs), which are generated by doping with rare earth (RE) metals, are increasingly used for bioimaging because of the advantages they hold over conventional fluorophores. However, because pristine RE nanoparticles (NPs) are unstable in acidic physiological fluids (e.g., lysosomes), leading to intracellular phosphate complexation with the possibility of lysosomal injury, it is important to ensure that UCNPs are safely designed. In this study, we used commercially available NaYF4:Er/Yb UCNPs to study their stability in lysosomes and simulated lysosomal fluid. We demonstrate that phosphate complexation leads to REPO4deposition on the particle surfaces and morphological transformation. This leads to a decline in upconversion fluorescence efficiency as well as inducing pro-inflammatory effects at the cellular level and in the intact lung. In order to preserve the imaging properties of the UCNPs as well as improve their safety, we experimented with a series of phosphonate chemical moieties to passivate particle surfaces through the strong coordination of the organophosphates with RE atoms. Particle screening and physicochemical characterization revealed that ethylenediamine tetra(methylenephosphonic acid) (EDTMP) surface coating provides the most stable UCNPs, which maintain their imaging intensity and do not induce pro-inflammatory effects in vitro and in vivo. In summary, phosphonate coating presents a safer design method that preserves and improves the bioimaging properties of UCNPs, thereby enhancing their biological use.

We demonstrate through PdO doping that creation of heterojunctions on Co3O4 nanoparticles can quantitatively adjust band-gap and Fermi energy levels to study the impact of metal oxide nanoparticle semiconductor properties on cellular redox homeostasis and hazard potential. Flame spray pyrolysis (FSP) was used to synthesize a nanoparticle library in which the gradual increase in the PdO content (0-8.9%) allowed electron transfer from Co3O4 to PdO to align Fermi energy levels across the heterojunctions. This alignment was accompanied by free hole accumulation at the Co3O4 interface and production of hydroxyl radicals. Interestingly, there was no concomitant superoxide generation, which could reflect the hole dominance of a p-type semiconductor. Although the electron flux across the heterojunctions induced upward band bending, the E(c) levels of the doped particles showed energy overlap with the biological redox potential (BRP). This allows electron capture from the redox couples that maintain the BRP from -4.12 to -4.84 eV, causing disruption of cellular redox homeostasis and induction of oxidative stress. PdO/Co3O4 nanoparticles showed significant increases in cytotoxicity at 25, 50, 100, and 200 μg/mL, which was enhanced incrementally by PdO doping in BEAS-2B and RAW 264.7 cells. Oxidative stress presented as a tiered cellular response involving superoxide generation, glutathione depletion, cytokine production, and cytotoxicity in epithelial and macrophage cell lines. A progressive series of acute pro-inflammatory effects could also be seen in the lungs of animals exposed to incremental PdO-doped particles. All considered, generation of a combinatorial PdO/Co3O4 nanoparticle library with incremental heterojunction density allowed us to demonstrate the integrated role of E(v), E(c), and E(f) levels in the generation of oxidant injury and inflammation by the p-type semiconductor, Co3O4.

Growing international exploitation of rare earth oxides (REOs) for commercial and biological use has increased the possibility of human exposure and adverse health effects. Occupational exposure to rare earth materials in miners and polishers leads to a severe form of pneumoconiosis, while gadolinium-containing MRI contrast agents cause nephrogenic systemic fibrosis in patients with renal impairment. The mechanisms for inducing these adverse pro-fibrogenic effects are of considerable importance for the safety assessment of REO particles as well as presenting opportunities for safer design. In this study, using a well-prepared REO library, we obtained a mechanistic understanding of how REOs induce cellular and pulmonary damage by a compartmentalized intracellular biotransformation process in lysosomes that results in pro-fibrogenic growth factor production and lung fibrosis. We demonstrate that rare earth oxide ion shedding in acidifying macrophage lysosomes leads to biotic phosphate complexation that results in organelle damage due to stripping of phosphates from the surrounding lipid bilayer. This results in nanoparticle biotransformation into urchin shaped structures and setting in motion a series of events that trigger NLRP3 inflammasome activation, IL-1β release, TGF-β1 and PDGF-AA production. However, pretreatment of REO nanoparticles with phosphate in a neutral pH environment prevents biological transformation and pro-fibrogenic effects. This can be used as a safer design principle for producing rare earth nanoparticles for biological use. © 2014 American Chemical Society.

Graphene and graphene-based nanomaterials display novel and beneficial chemical, electrical, mechanical, and optical characteristics, which endow these nanomaterials with promising applications in a wide spectrum of areas such as electronics and biomedicine. However, its toxicity on health remains unknown and is of great concern. In the present study, we demonstrated that graphene oxide (GO) induced necrotic cell death to macrophages. This toxicity is mediated by activation of toll-like receptor 4 (TLR4) signaling and subsequently in part via autocrine TNF-α production. Inhibition of TLR4 signaling with a selective inhibitor prevented cell death nearly completely. Furthermore, TLR4-deficient bone marrow-derived macrophages were resistant to GO-triggered necrosis. Similarly, GO did not induce necrosis of HEK293T/TLR4-null cells. Macrophagic cell death upon GO treatment was partially attributed to RIP1-RIP3 complex-mediated programmed necrosis downstream of TNF-α induction. Additionally, upon uptake into macrophages, GO accumulated primarily in cytoplasm causing dramatic morphologic alterations and a significant reduction of the macrophagic ability in phagocytosis. However, macrophagic uptake of GO may not be required for induction of necrosis. GO exposure also caused a large increase of intracellular reactive oxygen species (ROS), which contributed to the cause of cell death. The combined data reveal that interaction of GO with TLR4 is the predominant molecular mechanism underlying GO-induced macrophagic necrosis; also, cytoskeletal damage and oxidative stress contribute to decreased viability and function of macrophages upon GO treatment.

Quantum dots (QDs) are nanometer-sized luminescent semiconductor nanocrystals. Their unique optical properties, such as high brightness, long-term stability, simultaneous detection of multiple signals and tunable emission spectra, make them appealing as potential diagnostic and therapeutic systems in oncology. Preparing the functional QDs by modifying bio-molecules such as antibody will have potential value for cancer diagnosis and treatment. This paper summarized the recent progress of promising application of QDs in cancer diagnosis and therapy, from identifying molecular targets, to drug delivery and therapy; from limitations of toxicity issues related to QDs in living organisms to multifunctional design and development. Finally, the promising applications of QDs are also discussed.

Vascularization is one of the capital challenges in the establishment of tissue engineering constructs and recovery of ischemic and wounded tissues. The aim of this study was to assess electrospun fibers with loadings of multiple pDNA to allow a localized delivery for an efficient regeneration of mature blood vessels. To induce sufficient protein expression, a reverse microemulsion process was adopted to load pDNA into calcium phosphate nanoparticles (CP-pDNA), which were electrospun into fibers to achieve a sustained release for 4 weeks. Compared with pDNA-infiltrated fibers, the localized and gradual release of pDNA facilitated cell proliferation, gene transfection, and extracellular matrix secretion and enhanced the generation of blood vessels after subcutaneous implantation. Compared with commonly used pDNA polyplexes with poly(ethyleneimine), CP-pDNA nanoparticles induced significantly lower cytotoxicity and less inflammation reaction after implantation into animals. Fibers with encapsulated nanoparticles containing plasmids encoding vascular endothelial growth factor (pVEGF) and basic fibroblast growth factors (pbFGF) led to significantly higher density of mature blood vessels than those containing individual plasmid. It is suggested that the integration of CP-pDNA nanoparticles with loadings of multiple plasmids into fibrous scaffolds should provide clinical relevance for therapeutic vascularization, getting fully vascularized in engineered tissues and regeneration of blood vessel substitutes.

A particular challenge for nanotoxicology is the evaluation of the biological fate and toxicity of nanomaterials that dissolve in aqueous fluids. Zinc oxide nanomaterials are of particular concern because dissolution leads to release of the toxic divalent zinc ion. Although dissolved zinc ions have been implicated in ZnO cytotoxicity, direct identification of the chemical form of zinc taken up by cells exposed to ZnO nanoparticles, and its intracellular fate, has not yet been achieved. We combined high resolution X-ray spectromicroscopy and high elemental sensitivity X-ray microprobe analyses to determine the fate of ZnO and less soluble iron-doped ZnO nanoparticles following exposure to cultures of human bronchial epithelial cells, BEAS-2B. We complemented two-dimensional X-ray imaging methods with atomic force microscopy of cell surfaces to distinguish between nanoparticles that were transported inside the cells from those that adhered to the cell exterior. The data suggest cellular uptake of ZnO nanoparticles is a mechanism of zinc accumulation in cells. Following uptake, ZnO nanoparticles dissolved completely generating intracellular Zn2+ complexed by molecular ligands. These results corroborate a model for ZnO nanoparticle toxicity that is based on nanoparticle uptake followed by intracellular dissolution.

UV-light-induced electron-hole (e(-)/h(+)) pair generation with free radical production in TiO(2)-based nanoparticles is a major conceptual paradigm for biological injury. However, to date, this hypothesis has been difficult to experimentally verify due to the high energy of UV light that is intrinsically highly toxic to biological systems. Here, a versatile flame spray pyrolysis (FSP) synthetic process has been exploited to synthesize a library of iron-doped (0-10 wt%) TiO(2) nanoparticles. These particles have been tested for photoactivation-mediated cytotoxicity using near-visible light exposure. The reduction in TiO(2) band gap energy with incremental levels of Fe loading maintained the nanoparticle crystalline structure in spite of homogeneous Fe distribution (demonstrated by XRD, HRTEM, SAED, EFTEM, and EELS). Photochemical studies showed that band gap energy was reciprocally tuned proportional to the Fe content. The photo-oxidation capability of Fe-doped TiO(2) was found to increase during near-visible light exposure. Use of a macrophage cell line to evaluate cytotoxic and ROS production showed increased oxidant injury and cell death in parallel with a decrease in band gap energy. These findings demonstrate the importance of band gap energy in the phototoxic response of the cell to TiO(2) nanoparticles and reflect the potential of this material to generate adverse effects in humans and the environment during high-intensity light exposure.

We have recently shown that the dissolution of ZnO nanoparticles and Zn(2+) shedding leads to a series of sublethal and lethal toxicological responses at the cellular level that can be alleviated by iron doping. Iron doping changes the particle matrix and slows the rate of particle dissolution. To determine whether iron doping of ZnO also leads to lesser toxic effects in vivo, toxicity studies were performed in rodent and zebrafish models. First, we synthesized a fresh batch of ZnO nanoparticles doped with 1-10 wt % of Fe. These particles were extensively characterized to confirm their doping status, reduced rate of dissolution in an exposure medium, and reduced toxicity in a cellular screen. Subsequent studies compared the effects of undoped to doped particles in the rat lung, mouse lung, and the zebrafish embryo. The zebrafish studies looked at embryo hatching and mortality rates as well as the generation of morphological defects, while the endpoints in the rodent lung included an assessment of inflammatory cell infiltrates, LDH release, and cytokine levels in the bronchoalveolar lavage fluid. Iron doping, similar to the effect of the metal chelator, DTPA, interfered in the inhibitory effects of Zn(2+) on zebrafish hatching. In the oropharyngeal aspiration model in the mouse, iron doping was associated with decreased polymorphonuclear cell counts and IL-6 mRNA production. Doped particles also elicited decreased heme oxygenase 1 expression in the murine lung. In the intratracheal instillation studies in the rat, Fe doping was associated with decreased polymorphonuclear cell counts, LDH, and albumin levels. All considered, the above data show that Fe doping is a possible safe design strategy for preventing ZnO toxicity in animals and the environment.

Accurate evaluation of engineered nanomaterial toxicity requires not only comprehensive physical-chemical characterization of nanomaterials as produced, but also thorough understanding of nanomaterial properties and behavior under conditions similar to those used for in vitro and in vivo toxicity studies. In this investigation, TiO(2) nanoparticles were selected as a model nanoparticle and bovine serum albumin (BSA) was selected as a model protein for studying the effect of protein-nanoparticle interaction on TiO(2) nanoparticle dispersion in six different mammalian, bacteria, and yeast cell culture media. Great improvement in TiO(2) dispersion was observed upon the addition of BSA, even though the degree of dispersion varied from medium to medium and phosphate concentration in the cell culture media was one of the key factors governing nanoparticle dispersion. Fetal bovine serum (FBS) was an effective dispersing agent for TiO(2) nanoparticles in all six media due to synergistic effects of its multiple protein components, successfully reproduced using a simple "FBS mimic" protein cocktail containing similar concentrations of BSA, γ-globulin, and apo-transferrin.

The establishment of verifiably safe nanotechnology requires the development of assessment tools to identify hazardous nanomaterial properties that could be modified to improve nanomaterial safety. While there is a lot of debate of what constitutes appropriate safety screening methods, one approach is to use the assessment of cellular injury pathways to collect knowledge about hazardous material properties that could lead to harm to humans and the environment. We demonstrate the use of a multiparameter cytotoxicity assay that evaluates toxic oxidative stress to compare the effects of titanium dioxide (TiO(2)), cerium oxide (CeO(2)), and zinc oxide (ZnO) nanoparticles in bronchial epithelial and macrophage cell lines. The nanoparticles were chosen on the basis of their volume of production and likelihood of spread to the environment. Among the materials, dissolution of ZnO nanoparticles and Zn(2+) release were capable of ROS generation and activation of an integrated cytotoxic pathway that includes intracellular calcium flux, mitochondrial depolarization, and plasma membrane leakage. These responses were chosen on the basis of the compatibility of the fluorescent dyes that contemporaneously assess their response characteristics by a semiautomated epifluorescence procedure. Purposeful reduction of ZnO cytotoxicity was achieved by iron doping, which changed the material matrix to slow Zn(2+) release. In summary, we demonstrate the utility of a rapid throughput, integrated biological oxidative stress response pathway to perform hazard ranking of a small batch of metal oxide nanoparticles, in addition to showing how this assay can be used to improve nanosafety by decreasing ZnO dissolution through Fe doping.

Rapid growth in nanotechnology is increasing the likelihood of engineered nanomaterials coming into contact with humans and the environment. Nanoparticles interacting with proteins, membranes, cells, DNA and organelles establish a series of nanoparticle/biological interfaces that depend on colloidal forces as well as dynamic biophysicochemical interactions. These interactions lead to the formation of protein coronas, particle wrapping, intracellular uptake and biocatalytic processes that could have biocompatible or bioadverse outcomes. For their part, the biomolecules may induce phase transformations, free energy releases, restructuring and dissolution at the nanomaterial surface. Probing these various interfaces allows the development of predictive relationships between structure and activity that are determined by nanomaterial properties such as size, shape, surface chemistry, roughness and surface coatings. This knowledge is important from the perspective of safe use of nanomaterials.

Nanomaterials (NM) exhibit novel physicochemical properties that determine their interaction with biological substrates and processes. Three metal oxide nanoparticles that are currently being produced in high tonnage, TiO(2), ZnO, and CeO(2), were synthesized by flame spray pyrolysis process and compared in a mechanistic study to elucidate the physicochemical characteristics that determine cellular uptake, subcellular localization, and toxic effects based on a test paradigm that was originally developed for oxidative stress and cytotoxicity in RAW 264.7 and BEAS-2B cell lines. ZnO induced toxicity in both cells, leading to the generation of reactive oxygen species (ROS), oxidant injury, excitation of inflammation, and cell death. Using ICP-MS and fluorescent-labeled ZnO, it is found that ZnO dissolution could happen in culture medium and endosomes. Nondissolved ZnO nanoparticles enter caveolae in BEAS-2B but enter lysosomes in RAW 264.7 cells in which smaller particle remnants dissolve. In contrast, fluorescent-labeled CeO(2) nanoparticles were taken up intact into caveolin-1 and LAMP-1 positive endosomal compartments, respectively, in BEAS-2B and RAW 264.7 cells, without inflammation or cytotoxicity. Instead, CeO(2) suppressed ROS production and induced cellular resistance to an exogenous source of oxidative stress. Fluorescent-labeled TiO(2) was processed by the same uptake pathways as CeO(2) but did not elicit any adverse or protective effects. These results demonstrate that metal oxide nanoparticles induce a range of biological responses that vary from cytotoxic to cytoprotective and can only be properly understood by using a tiered test strategy such as we developed for oxidative stress and adapted to study other aspects of nanoparticle toxicity.

International audience; C 60 fullerene is a promising material because of its unique physiochemical properties. However, previous studies have reported that colloidal aggregates of C 60 (nC 60) produce toxicity in fish and human cell cultures. The preparation method of nC 60 raises questions as to whether the observed effects stem from fullerenes or from the organic solvents used during the preparation of the suspensions. In this paper, we set out to elucidate the mechanism by which tetrahydrofuran (THF) treatment to enhance the preparation of nC 60 leads to cytotoxicity in a mouse macrophage cell line. Our results demonstrate that THF/nC 60 but not fullerol or aqueous nC 60 generates cellular toxicity through a pathway that involves increased intracellular flux and mitochondrial perturbation in RAW 264.7 cells. Interestingly, the supernatant of the THF/nC 60 suspension rather than the colloidal fullerene aggregates mimics the cytotoxic effects due to the presence of γ-butyrolactone and formic acid. Thus, the role of nC 60 in the cellular responses is likely not due to the direct effect of the nC 60 material surface on the cells but is related to the conversion of THF into a toxic byproduct during preparation of the suspension.