Brucella species-induced brucellosis: Antimicrobial effects, potential resistance and toxicity of silver and gold nanosized particles.

Brucellosis is an endemic zoonotic disease caused by Brucella species, which are intramacrophage pathogens that make treating this disease challenging. The negative effects of the treatment regime have prompted the development of new antimicrobials against brucellosis. A new treatment modality for antibiotic-resistant microorganisms is the use of nanoparticles (NPs). In this study, we examined the antibacterial activities of silver and gold NPs (SNPs and GNPs, respectively), the resistance developed by Brucella melitensis (B. melitensis) and Brucella abortus (B. abortus) strains and the toxicity of both of these NPs in experimental rats. To test the bactericidal effects of the SNPs and GNPs, we used 22 multidrug-resistant Brucella isolates (10 B. melitensis and 12 B. abortus). The minimal inhibitory concentrations (MICs) of both types of NPs were determined utilizing the microdilution technique. To test the stability of resistance, 7 B. melitensis and 6 B. abortus isolates were passaged ten times in culture with subinhibitory concentrations of NPs and another ten times without NPs. Histopathological analysis was completed after rats were given 0.25, 0.5, 1, and 2 mg/kg NPs orally for 28 consecutive days. The MIC values (µg/ml) of the 10-nm SNPs and 20-nm GNPs against B. melitensis were 22.43 ± 2.32 and 13.56 ± 1.22, while these values were 18.77 ± 1.33 and 12.45 ± 1.59 for B. abortus, respectively. After extensive in vitro exposure, most strains showed no resistance to the 10-nm SNPs or 20-nm GNPs. The NPs and antibiotics did not cross-react in any of the evolved Brucella strains. SNPs and GNPs at doses below 2 mg/kg were not harmful to rat tissue according to organ histopathological examinations. However, a greater dose of NPs (2 mg/kg) harmed all of the tissues studied. The bactericidal properties of NPs are demonstrated in this work. Brucella strains develop similar resistance to SNPs and GNPs, and at low dosages, neither SNPs nor GNPs were hazardous to rats.

Gold and silver nanoparticles effects to the earthworm Eisenia fetida - the importance of tissue over soil concentrations.

To address and to compare the respective impact of gold and silver nanoparticles (Au and Ag NPs) in soil invertebrate, the earthworm Eisenia fetida was exposed to soil containing 2, 10, and 50 mg/kg of Au and Ag in both nanoparticulate and ionic forms for 10 days. Both metal NPs were 2-15 times less bioavailable than their ionic forms, and displayed similar transfer coefficients from soil to earthworm tissues. Both metal NPs triggered the onset of an oxidative stress as illustrated by increased glutathione S-transferase levels, decreased catalase levels, and increased malondialdehyde concentrations. Protein carbonylation distinguished the nanoparticular from the ionic forms as its increase was observed only after exposure to the highest concentration of both metal NPs. Au and Ag NPs triggered DNA modifications even at the lowest concentration, and both repressed the expression of genes involved in the general defense and stress response at high concentrations as did their ionic counterparts. Despite the fact that both metal NPs were less bioavailable than their ionic forms, at equivalent concentrations accumulated within earthworms tissues they exerted equal or higher toxic potential than their ionic counterparts.Capsule: At equivalent concentrations accumulated within earthworm tissues Au and Ag NPs exert equal or higher toxic potential than their ionic forms.

Beyond gold nanoparticles cytotoxicity: Potential to impair metastasis hallmarks.

Gold nanoparticle (AuNP)-based systems have been extensively investigated as diagnostic and therapeutic agents due to their tunable properties and easy surface functionalization. Upon cell uptake, AuNPs present an inherent cell impairment potential based on organelle and macromolecules damage, leading to cell death. Such cytotoxicity is concentration-dependent and completely undesirable, especially if unspecific. However, under non-cytotoxic concentrations, internalized AuNPs could potentially weaken cells and act as antitumor agents. Therefore, this study aimed to investigate the antitumor effect of ultrasmall AuNPs (~3 nm) stabilized by the anionic polysaccharide gum arabic (GA-AuNPs). Other than intrinsic cytotoxicity, the focus was downregulation of cancer hallmarks of aggressive tumors, using a highly metastatic model of melanoma. We first demonstrated that GA-AuNPs showed excellent stability under biological environment. Non-cytotoxic concentrations to seven different cell lines, including tumorigenic and non-tumorigenic cells, were determined by standard 2D in vitro assays. Gold concentrations ≤ 2.4 mg L-1 (16.5 nM AuNPs) were non-cytotoxic and therefore chosen for further analyses. Cells exposed to GA-AuNPs were uptaken by melanoma cells through endocytic processes. Next we described remarkable biological properties using non-cytotoxic concentrations of this nanomaterial. Invasion through an extracellular matrix barrier as well as 3D growth capacity (anchorage-independent colony formation and spheroids growth) were negatively affected by 2.4 mg L-1 GA-AuNPs. Additionally, exposed spheroids showed morphological changes, suggesting that GA-AuNPs could penetrate into the preformed tumor and affect its integrity. All together these results demonstrate that side effects, such as cytotoxicity, can be avoided by choosing the right concentration, nevertheless, preserving desirable effects such as modulation of key tumor cell malignancy features.

Evaluation of the skin-sensitizing potential of gold nanoparticles and the impact of established dermal sensitivity on the pulmonary immune response to various forms of gold.

Gold nanoparticles (AuNP) are largely biocompatible; however, many studies have demonstrated their potential to modulate various immune cell functions. The potential allergenicity of AuNP remains unclear despite the recognition of gold as a common contact allergen. In these studies, AuNP (29 nm) dermal sensitization potential was assessed via Local Lymph Node Assay (LLNA). Soluble gold (III) chloride (AuCl3) caused lymph node (LN) expansion (SI 10.9), whereas bulk particles (Au, 942 nm) and AuNP did not. Next, the pulmonary immune effects of AuNP (10, 30, 90 µg) were assessed 1, 4, and 8 days post-aspiration. All markers of lung injury and inflammation remained unaltered, but a dose-responsive increase in LN size was observed. Finally, mice were dermally-sensitized to AuCl3 then aspirated once, twice, or three times with Au or AuNP in doses normalized for mass or surface area (SA) to assess the impact of existing contact sensitivity to gold on lung immune responses. Sensitized animals exhibited enhanced responsivity to the metal, wherein subsequent immune alterations were largely conserved with respect to dose SA. The greatest increase in bronchoalveolar lavage (BAL) lymphocyte number was observed in the high dose group - simultaneous to preferential expansion of BAL/LN CD8+ T-cells. Comparatively, the lower SA-based doses of Au/AuNP caused more modest elevations in BAL lymphocyte influx (predominantly CD4+ phenotype), exposure-dependent increases in serum IgE, and selective expansion/activation of LN CD4+ T-cells and B-cells. Overall, these findings suggest that AuNP are unlikely to cause sensitization; however, established contact sensitivity to gold may increase immune responsivity following pulmonary AuNP exposure.

Assessment of oxidative stress induced by gold nanorods following intra-tracheal instillation in rats.

Gold nanorods (GNRs) are used for their wide variety of applications in various industries. There is a little availability of data related to toxicity and ecological implications of these GNRs. The study evaluated the oxidative stress induction following intra-tracheal instillation of 1 and 5 mg/kg b.w. doses of 10 and 25 nm GNRs by estimating various oxidative stress markers including lipid peroxidation (malondialdehyde; MDA), glutathione (GSH), superoxide dismutase (SOD), catalase and total antioxidant capacity (TAC) after 1 day, 1 week, 1 month, and 3 months post exposure periods. The results have shown increased MDA levels and decreased GSH levels following 1 day and 1 week post exposure periods, indicating induction of oxidative stress. Also, the SOD, catalase and TAC levels were significantly decreased following exposure of both 10 and 25 nm GNRs after 1 day and 1 week after exposures, indicating the inhibition of antioxidant defense mechanisms. Moreover, the 10 nm GNRs at 5 mg/kg dose displayed greater changes in all the estimated parameters, representing dose and size based induction of oxidative stress by GNRs. In contrast, a little change was observed during 1 month and 3 months post exposure periods, may be due to recovery. Finally, the GNRs induced dose-size-dependent oxidative stress induction by various oxidative stress markers following intra-tracheal instillation in rats.

Synergistic Toxicity of Copper and Gold Compounds in Cupriavidus metallidurans.

The bacterium Cupriavidus metallidurans can reduce toxic gold(I/III) complexes and biomineralize them into metallic gold (Au) nanoparticles, thereby mediating the (trans)formation of Au nuggets. In Au-rich soils, most transition metals do not interfere with the resistance of this bacterium to toxic mobile Au complexes and can be removed from the cell by plasmid-encoded metal efflux systems. Copper is a noticeable exception: the presence of Au complexes and Cu ions results in synergistic toxicity, which is accompanied by an increased cytoplasmic Cu content and formation of Au nanoparticles in the periplasm. The periplasmic Cu-oxidase CopA was not essential for formation of the periplasmic Au nanoparticles. As shown with the purified and reconstituted Cu efflux system CupA, Au complexes block Cu-dependent release of phosphate from ATP by CupA, indicating inhibition of Cu transport. Moreover, Cu resistance of Au-inhibited cells was similar to that of mutants carrying deletions in the genes for the Cu-exporting PIB1-type ATPases. Consequently, Au complexes inhibit export of cytoplasmic Cu ions, leading to an increased cellular Cu content and decreased Cu and Au resistance. Uncovering the biochemical mechanisms of synergistic Au and Cu toxicity in C. metallidurans explains the issues this bacterium has to face in auriferous environments, where it is an important contributor to the environmental Au cycle.IMPORTANCEC. metallidurans lives in metal-rich environments, including auriferous soils that contain a mixture of toxic transition metal cations. We demonstrate here that copper ions and gold complexes exert synergistic toxicity because gold ions inhibit the copper-exporting P-type ATPase CupA, which is central to copper resistance in this bacterium. Such a situation should occur in soils overlying Au deposits, in which Cu/Au ratios usually are ≫1. Appreciating how C. metallidurans solves the problem of living in environments that contain both Au and Cu is a prerequisite to understand the molecular mechanisms underlying gold cycling in the environment, and the significance and opportunities of microbiota for specific targeting to Au in mineral exploration and ore processing.

Biosynthesis of gold nanoparticles by two bacterial and fungal strains, Bacillus cereus and Fusarium oxysporum, and assessment and comparison of their nanotoxicity in vitro by direct and indirect assays

Background: Although nanoparticles (NPs) have many advantages, it has been proved that they may be absorbed by and have toxic effects on the human body. Recent research has tried to evaluate and compare the nanotoxicity of gold nanoparticles (AuNPs) produced by two types of microorganisms in vitro by two different methods. AuNPs were produced by Bacillus cereus and Fusarium oxysporum, and their production was confirmed by visible spectral, transmission electron microscope, and X-ray diffraction (XRD) analyses. The human fibroblast cell line CIRC-HLF was treated with AuNPs, and the induced nanotoxicity was measured using direct microscopic and 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazolium bromide (MTT) assays. Results: The results showed that the produced AuNPs had a maximum absorbance peak around 510­530 nanometer (nm), with spherical, hexagonal, and octagonal shapes and average sizes around 20­50 nm. The XRD results confirmed the presence of GNPs in the microbial culture supernatants. An MTT assay showed that GNPs had dose-dependent toxic effects, and microscopic analysis showed that GNPs induced cell abnormalities in doses lower than the determined half-maximal inhibitory concentrations (IC50s). Conclusions: In conclusion, the biologically produced AuNPs had toxic effects in the cell culture, and direct techniques such as microscopic evaluation instead of indirect methods such as MTT assay were more useful for assessing the nanotoxicity of the biologically produced AuNPs. Thus, the use of only MTT assay for nanotoxicity evaluation of AuNPs is not desirable.

Dinuclear Gold(I) Pyrrolidinedithiocarbamato Complex: Cytotoxic and Antimigratory Activities on Cancer Cells and the Use of Metal-Organic Framework.

A dinuclear gold(I) pyrrolidinedithiocarbamato complex (1) with a bidentate carbene ligand has been constructed and shows potent in vitro cytotoxic activities towards cisplatin-resistant ovarian cancer cells A2780cis. Its rigid scaffold enables a zinc(II)-based metal-organic framework (Zn-MOF) to be used as a carrier in facilitating the uptake and release of 1 in solutions. Instead of using a conventional dialysis approach for the drug-release testing, in this study, a set of transwell assay-based experiments have been designed and employed to examine the cytotoxic and antimigratory activities of 1@Zn-MOF towards A2780cis.

Theranostic Gold Nanomicelles made from Biocompatible Comb-like Polymers for Thermochemotherapy and Multifunctional Imaging with Rapid Clearance.

A new generation of photothermal theranostic agents based on assembling 6 nm gold nanoparticles (AuNPs) is developed by using a novel comb-like amphipathic polymer as the template. The small AuNPs are assembled into DOX@gold nanomicelles, which show strong absorbance in the near-infrared region, for multimodal bioimaging and highly effective in vivo chemotherapy and photothermal therapy.

Evaluation of gold nanoparticles toxicity towards human endothelial cells under static and flow conditions.

A new in vitro model system, adding advection and shear stress associated with a flowing medium, is proposed for the investigation of nanoparticles uptake and toxicity towards endothelial cells, since these processes are normally present when nanoparticles formulations are intravenously administered. In this model system, mechanical forces normally present in vivo, such as advection and shear stress were applied and carefully controlled by growing human umbilical vein endothelial cells inside a microfluidic device and continuously infusing gold nanoparticle (Au NPs) solution in the device. The tests performed in the microfluidic device were also run in multiwells, where no flow is present, so as to compare the two model systems and evaluate if gold nanoparticles toxicity differs under static and flow culture conditions. Full characterization of Au NPs in water and in culture medium was accomplished by standard methods. Two-photon fluorescence correlation spectroscopy was also employed to map the flow speed of Au NPs in the microfluidic device and characterize Au NPs before and after interactions with the cells. Au NPs uptake in both in vitro systems was investigated through electron and fluorescence microscopy and ICP-AES, and NPs toxicity measured through standard bio-analytical tests. Comparison between experiments run in multiwells and in microfluidic device plays a pivotal role for the investigation of nanoparticle-cell interaction and toxicity assessment: our work showed that administration of equal concentrations of Au NPs under flow conditions resulted in a reduced sedimentation of nanoparticle aggregates onto the cells and lower cytotoxicity with respect to experiments run in ordinary static conditions (multiwells).

Size-dependent clearance of gold nanoparticles from lungs of Sprague-Dawley rats after short-term inhalation exposure.

Gold nanoparticles are known to be distributed to many tissues following their oral, inhalation, or intravenous exposure. Information on the biodistribution and clearance of gold nanoparticles from these tissues is, therefore, important to understand their behavior in vivo. To study the effect of size on the biodistribution of gold nanoparticles, Sprague-Dawley rats were exposed by inhalation to small gold nanoparticles (13 nm in diameter on average) at an exposure concentration of 12.8 ± 2.42 µg/m(3), and to large gold nanoparticles (105 nm in diameter on average) at an exposure concentration of 13.7 ± 1.32 µg/m(3). The experimental animals were exposed to the gold nanoparticles and the control animals to fresh air for 5 days (6 h/day), followed by a recovery period of 1, 3, and 28 days in fresh air. None of the exposed animals exhibited any toxic response to the gold nanoparticles. Despite the difference in size, both small and large gold nanoparticles deposited mainly in rat lungs. Their biodistribution from the lungs to secondary target organs was significantly higher with the small compared to the large gold nanoparticles. While the large gold nanoparticles were only found in the blood, the small gold nanoparticles were detected in the liver, spleen, brain, testes, and blood. In addition, the elimination half-life of the small gold nanoparticles from the lungs was significantly shorter than that of the large gold nanoparticles. The present data may, therefore, suggest that the smaller gold nanoparticles are able to translocate from the lungs, the primary exposure organ to extrapulmonary organs at a faster rate than the larger gold nanoparticles and thus confirming previous observations reported in the literature.

Toxicological aspects of injectable gold-hyaluronan combination as a treatment for neuroinflammation.

Secondary inflammatory reactions to stroke or trauma contribute to irreplaceable loss of brain tissue of the affected patients. Likewise, neuroinflammatory processes are the main pathophysiological feature in Multiple Sclerosis (MS), a common neurodegenerative disease among young adults. In the search for safe and efficient ways to reduce inflammation within nervous tissue older immunosuppressive remedies have been re-investigated. The anti-inflammatory properties of gold salts are well known but result in uncontrollable systemic spread of gold ions, generating side effects such as nephrotoxicity, limiting their use. Recent studies have circumvented this obstacle by introducing metallic gold implants as a localized source of immune-modulating gold ions and suspension in hyaluronic acid (HA) enables injection of small amounts of gold in the natural spaces of the brain. By injecting >25 µm gold beads in HA intracerebrally we recently showed a slowing of disease progression in a rodent model of MS. The toxicological aspects were, however, not assessed. The present study investigates the viability of neuronal and macrophage cell cultures exposed to the gold/HA combination and the possible risk associated with unilateral gold/HA injection in young Balb/CA mice in the first 7 to 21 days of gold-exposure. Tracing by autometallography of gold accumulations throughout the brain exhibited sparse gold uptake in glia and neurons of hippocampus and cortex, and striatum and cerebellum were void of staining. No systemic spread of gold was seen in liver or kidney, nor were there signs of obstruction of the ventricular system. Both cell cultures of J774 macrophages and CCL neurons accumulated gold from gold/HA-exposure with no signs of reduced viability. In conclusion, our findings indicate that gold/HA is not overtly neuro- or cytotoxic, nor does intraventricular exposure result in widespread gold accumulation or tissue damage, warranting further studies into the pharmacological properties of this novel form of gold treatment.

Evaluation of the toxicity of intravenous delivery of auroshell particles (gold-silica nanoshells).

Gold nanoshells (155 nm in diameter with a coating of polyethylene glycol 5000) were evaluated for preclinical biocompatibility, toxicity, and biodistribution as part of a program to develop an injectable device for use in the photothermal ablation of tumors. The evaluation started with a complete good laboratory practice (GLP) compliant International Organization for Standardization (ISO)-10993 biocompatibility program, including cytotoxicity, pyrogenicity (US Pharmacopeia [USP] method in the rabbit), genotoxicity (bacterial mutagenicity, chromosomal aberration assay in Chinese hamster ovary cells, and in vivo mouse micronucleus), in vitro hemolysis, intracutaneous reactivity in the rabbit, sensitization (in the guinea pig maximization assay), and USP/ISO acute systemic toxicity in the mouse. There was no indication of toxicity in any of the studies. Subsequently, nanoshells were evaluated in vivo by intravenous (iv) infusion using a trehalose/water solution in a series of studies in mice, Sprague-Dawley rats, and Beagle dogs to assess toxicity for time durations of up to 404 days. Over the course of 14 GLP studies, the gold nanoshells were well tolerated and, when injected iv, no toxicities or bioincompatibilities were identified.

Susceptibility to gold nanoparticle-induced hepatotoxicity is enhanced in a mouse model of nonalcoholic steatohepatitis.

Although the safety of gold nanoparticle (AuNP) use is of growing concern, most toxicity studies of AuNPs had focused on their chemical characteristics, including their physical dimensions, surface chemistry, and shape. The present study examined the susceptibility of rodents with healthy or damaged livers to AuNP-induced hepatotoxicity. To induce a model of liver injury, mice were fed a methionine- and choline-deficient (MCD) diet for 4 weeks. Sizes and biodistribution of 15-nm PEGylated AuNPs were analyzed by transmission electron microscopy. Levels of alanine aminotransferase (ALT) and aspartate aminotransferase (AST) were estimated with an automatic chemical analyzer, and liver sections were subjected to pathological examination. Activities of antioxidant enzymes were determined by biochemical assay. Lateral tail vein injection of MCD diet-fed mice with 5 mg kg(-1) AuNPs significantly elevated the serum ALT and AST levels compared to MCD diet-fed mice injected with mPEG (methylpolyethylene glycol). Similarly, severe hepatic cell damage, acute inflammation, and increased apoptosis and reactive oxygen species (ROS) production were observed in the livers of AuNP-injected mice on the MCD diet; these liver injuries were attenuated in mice fed a normal chow diet. The results suggest that AuNPs display toxicity in a stressed liver environment by stimulating the inflammatory response and accelerating stress-induced apoptosis. These conclusions may point to the importance of considering health conditions, including liver damage, in medical applications of AuNPs.

Toxic effects of gold nanoparticles on Salmonella typhimurium bacteria.

Nanometer-sized gold, due to its beautiful and bountiful color and unique optical properties, is a versatile material for many industrial and societal applications. We have studied the effect of gold nanoparticles on Salmonella typhimurium strain TA 102. The gold nanoparticles in solution prepared using the citrate reduction method is found not to be toxic or mutagenic but photomutagenic to the bacteria; however, careful control experiments indicate that the photomutagenicity is due to the co-existing citrate and Au³âº ions, not due to the gold nanoparticle itself. Au³âº is also found to be photomutagenic to the bacteria at concentrations lower than 1 µM, but toxic at higher concentrations. The toxicity of Au³âº is enhanced by light irradiation. The photomutagenicity of both citrate and Au³âº is likely due to the formation of free radicals, as a result of light-induced citrate decarboxylation or Au³âº oxidation of co-existing molecules. Both processes can generate free radicals that may cause DNA damage and mutation. Studies of the interaction of gold nanoparticles with the bacteria indicate that gold nanoparticles can be absorbed onto the bacteria surface but not able to penetrate the bacteria wall to enter the bacteria.

Metal patch test results from 1990-2009.

Although metals are common contact allergens, clinical findings of metal contact dermatitis have varied. Such patients have subsequently become rare in Japan as gold dermatitis caused by ear piercing or baboon syndrome by broken thermometers. To evaluate such clinical findings and to determine the frequency of metal allergy, we analyzed the results of patch testing with 18 metals from 1990-2009. Nine hundred and thirty-one patients (189 men and 742 women, mean age 39.0years [standard deviation±17.8]) were tested. Metals were applied on the back for 2days, and the results read with the International Contact Dermatitis Research Group (ICDRG) scoring system 3days after application. Reactions of + to +++ were regarded as positive. Differences of positive rates between men and women, and patients from 1990-1999 and those from 2000-2009 were analyzed with the χ(2) -test. Differences were considered significant at P<0.05. The metal to which the most patients reacted was 5% nickel sulfate (27.2%), irrespective of sex and phase. Significantly more women reacted to nickel sulfate (P<0.01), mercuric chloride (P<0.05) and gold chloride (P<0.01) than men. Significantly more patients in the 1990s reacted to palladium chloride, mercuric chloride and gold chloride (all P<0.01) than from 2000-2009. Nickel has been the most common metal allergen and mercury-sensitivity has decreased over 19years in Japan.

Biocompatible gellan gum-reduced gold nanoparticles: cellular uptake and subacute oral toxicity studies.

Currently gold nanoparticles are being explored for drug delivery and other biomedical applications; therefore it is necessary to study the fate of such nanoparticles inside the body. The objective of the present study was to investigate the cellular uptake and toxicity of the gold nanoparticles synthesized using a microbial polysaccharide, gellan gum, as a capping and reducing agent. The cellular uptake of gold nanoparticles was studied on mouse embryonic fibroblast cells, NIH3T3 and human glioma cell line, LN-229. The cellular uptake study indicated that the gellan gum-reduced gold nanoparticles were located in cancer cells (LN-229) while no uptake was observed in normal mouse embryonic fibroblast cells (NIH3T3). The toxicity of the gold nanoparticles was evaluated by carrying out subacute 28 day oral toxicity studies in rats. Subacute administration of gum-reduced gold nanoparticles to the rats did not show any hematological or biochemical abnormalities. The weight and normal architecture of various organs did not change compared with control. The current findings, while establishing the specific uptake of nanoparticles into cancerous cells, also demonstrates that the gellan gum-reduced gold nanoparticles are devoid of toxicity in animals following oral administration.

Impact of agglomeration state of nano- and submicron sized gold particles on pulmonary inflammation.

BACKGROUND: Nanoparticle (NP) toxicity testing comes with many challenges. Characterization of the test substance is of crucial importance and in the case of NPs, agglomeration/aggregation state in physiological media needs to be considered. In this study, we have addressed the effect of agglomerated versus single particle suspensions of nano- and submicron sized gold on the inflammatory response in the lung. Rats were exposed to a single dose of 1.6 mg/kg body weight (bw) of spherical gold particles with geometric diameters of 50 nm or 250 nm diluted either by ultrapure water or by adding phosphate buffered saline (PBS). A single dose of 1.6 mg/kg bw DQ12 quartz was used as a positive control for pulmonary inflammation. Extensive characterization of the particle suspensions has been performed by determining the zetapotential, pH, gold concentration and particle size distribution. Primary particle size and particle purity has been verified using transmission electron microscopy (TEM) techniques. Pulmonary inflammation (total cell number, differential cell count and pro-inflammatory cytokines), cell damage (total protein and albumin) and cytotoxicity (alkaline phosphatase and lactate dehydrogenase) were determined in bronchoalveolar lavage fluid (BALF) and acute systemic effects in blood (total cell number, differential cell counts, fibrinogen and C-reactive protein) 3 and 24 hours post exposure. Uptake of gold particles in alveolar macrophages has been determined by TEM. RESULTS: Particles diluted in ultrapure water are well dispersed, while agglomerates are formed when diluting in PBS. The particle size of the 50 nm particles was confirmed, while the 250 nm particles appear to be 200 nm using tracking analysis and 210 nm using TEM. No major differences in pulmonary and systemic toxicity markers were observed after instillation of agglomerated versus single gold particles of different sizes. Both agglomerated as well as single nanoparticles were taken up by macrophages. CONCLUSION: Primary particle size, gold concentration and particle purity are important features to check, since these characteristics may deviate from the manufacturer's description. Suspensions of well dispersed 50 nm and 250 nm particles as well as their agglomerates produced very mild pulmonary inflammation at the same mass based dose. We conclude that single 50 nm gold particles do not pose a greater acute hazard than their agglomerates or slightly larger gold particles when using pulmonary inflammation as a marker for toxicity.

Acute toxicity and pharmacokinetics of 13 nm-sized PEG-coated gold nanoparticles.

In general, gold nanoparticles are recognized as being as nontoxic. Still, there have been some reports on their toxicity, which has been shown to depend on the physical dimension, surface chemistry, and shape of the nanoparticles. In this study, we carry out an in vivo toxicity study using 13 nm-sized gold nanoparticles coated with PEG (MW 5000). In our findings the 13 nm sized PEG-coated gold nanoparticles were seen to induce acute inflammation and apoptosis in the liver. These nanoparticles were found to accumulate in the liver and spleen for up to 7 days after injection and to have long blood circulation times. In addition, transmission electron microscopy showed that numerous cytoplasmic vesicles and lysosomes of liver Kupffer cells and spleen macrophages contained the PEG-coated gold nanoparticles. These findings of toxicity and kinetics of PEG-coated gold nanoparticles may have important clinical implications regarding the safety issue as PEG-coated gold nanoparticles are widely used in biomedical applications.

Sublethal concentrations of diverse gold compounds inhibit mammalian cytosolic thioredoxin reductase (TrxR1).

UNLABELLED: Thioredoxin reductase (TrxR) reduces thioredoxin (Trx), thereby contributing to cellular redox balance, facilitating the synthesis of deoxy-ribose sugars for DNA synthesis, and regulating redox-sensitive gene expression. Auranofin is a gold compound that potently inhibits TrxR. This inhibition is one suspected mechanism of auranofin's therapeutic benefit in the treatment of rheumatoid arthritis. The use of other gold compounds to treat cancer or inflammatory disease may rely on their ability to inhibit TrxR. In the current study, we tested the hypothesis that a variety of gold compounds may inhibit TrxR. METHODS: We exposed rat-TrxR1 to auranofin, gold sodium thiomalate, sodium aurothiosulfate, triphenyl phosphine gold chloride, or gold acetate, and measured TrxR activity ex vivo. We then compared TrxR1 inhibitory levels of gold compounds to those that inhibited mitochondrial activity of THP1 monocytes and OSC2 epithelial cells, estimated by succinate dehydrogenase activity. RESULTS: All gold compounds inhibited TrxR1 at concentrations ranging from 5 to 4000 nM (50% inhibitory concentration). The oxidation state of gold did not correlate with inhibitory potency, but ligand configuration was important. Au(I)-phosphine compounds (triphenyl phosphine gold chloride and auranofin) were the most potent inhibitors of TrxR. All TrxR1 inhibitory concentrations were sublethal to mitochondrial activity in both THP1 and OSC2 cells. CONCLUSIONS: Diverse types of gold compounds may be effective inhibitors of TrxR1 at concentrations that do not suppress cellular mitochondrial function. Inhibition may be optimized to some degree by altering the ligand configuration of the compounds. These results support future study of a variety of Au compounds for therapeutic development as inhibitors of TrxR1.