Effects of synchrotron-based X-rays and gold nanoparticles on normal and cancer cell morphology and migration.

It has been shown lately that gold nanoparticles (AuNPs) and ionizing radiation (IR) have inhibitory effects on cancer cell migration while having promoting effects on normal cells' motility. Also, IR increases cancer cell adhesion with no significant effects on normal cells. In this study, synchrotron-based microbeam radiation therapy, as a novel pre-clinical radiotherapy protocol, is employed to investigate the effects of AuNPs on cell migration. Experiments were conducted utilizing synchrotron X-rays to investigate cancer and normal cell morphology and migration behaviour when they are exposed to synchrotron broad beams (SBB) and synchrotron microbeams (SMB). This in vitro study was conducted in two phases. In phase I two cancer cell lines - human prostate (DU145) and human lung (A549) - were exposed to various doses of SBB and SMB. Based on the phase I results, in phase II two normal cell lines were studied: human epidermal melanocytes (HEM) and human primary colon epithelial (CCD841), along with their respective cancerous counterparts, human primary melanoma (MM418-C1) and human colorectal adenocarcinoma (SW48). The results show that radiation-induced damage in cells' morphology becomes visible with SBB at doses greater than 50 Gy, and incorporating AuNPs increases this effect. Interestly, under the same conditions, no visible morphological changes were observed in the normal cell lines post-irradiation (HEM and CCD841). This can be attributed to the differences in cell metabolic and reactive oxygen species levels between normal and cancer cells. The outcome of this study highlights future applications of synchrotron-based radiotherapy, where it is possible to deliver extremely high doses to cancer tissues whilst preserving surrounding normal tissues from radiation-induced damage.

Ultrasound-Stimulated Microbubbles Enhance Radiation-Induced Cell Killing.

Recent in vivo studies using ultrasound-stimulated microbubbles as a localized radiosensitizer have had impressive results. While in vitro studies have also obtained similar results using human umbilical vein endothelial cells (HUVEC), studies using other cell lines have had varying results. This study was aimed at investigating any increases in radiation-induced cell killing in vitro using two carcinoma lines not previously investigated before (metastatic follicular thyroid carcinoma cells [FTC-238] and non-small cell lung carcinoma cells [NCI-H727]), in addition to HUVEC. Cells were treated using a combination of 1.6% (v/v) microbubbles, ∼90 s of 2-MHz ultrasound (mechanical index = 0.8) and 0-6 Gy of kilovolt or MV X-rays. Cell viability assays obtained 72 h post-treatment were normalized to untreated controls, and analysis of variance was used to determine statistical significance. All cells treated with combined ultrasound-stimulated microbubbles and radiation exhibited decreased normalized survival, with statistically significant effects observed for the NCI-H727 cells. No statistically significant differences in effects were observed using kV compared with MV radiation. Further studies using increased microbubble concentrations may be required to achieve statistically significant results for the FTC-238 and HUVEC lines.

Oxidative Damage to Mitochondria Enhanced by Ionising Radiation and Gold Nanoparticles in Cancer Cells.

Gold nanoparticles (AuNP) can increase the efficacy of radiation therapy by sensitising tumor cells to radiation damage. When used in combination with radiation, AuNPs enhance the rate of cell killing; hence, they may be of great value in radiotherapy. This study assessed the effects of radiation and AuNPs on mitochondrial reactive oxygen species (ROS) generation in cancer cells as an adjunct therapeutic target in addition to the DNA of the cell. Mitochondria are considered one of the primary sources of cellular ROS. High levels of ROS can result in an intracellular state of oxidative stress, leading to permanent cell damage. In this study, human melanoma and prostate cancer cell lines, with and without AuNPs, were irradiated with 6-Megavolt X-rays at doses of 0-8 Gy. Indicators of mitochondrial stress were quantified using two techniques, and were found to be significantly increased by the inclusion of AuNPs in both cell lines. Radiobiological damage to mitochondria was quantified via increased ROS activity. The ROS production by mitochondria in cells was enhanced by the inclusion of AuNPs, peaking at ~4 Gy and then decreasing at higher doses. This increased mitochondrial stress may lead to more effectively kill of AuNP-treated cells, further enhancing the applicability of functionally-guided nanoparticles.

Marine Bile Natural Products as Agonists of the TGR5 Receptor.

Agonism of the G protein-coupled bile acid receptor "Takeda G-protein receptor 5" (TGR5) aids in attenuating cholesterol accumulation due to atherosclerotic progression. Although mammalian bile compounds can activate TGR5, they are generally weak agonists, and more effective compounds need to be identified. In this study, two marine bile compounds (5ß-scymnol and its sulfate) were compared with mammalian bile compounds deoxycholic acid (DCA) and ursodeoxycholic acid (UDCA) using an in vitro model of TGR5 agonism. The response profiles of human embryonic kidney 293 cells (HEK293) transfected to overexpress TGR5 (HEK293-TGR5) and incubated with subcytotoxic concentrations of test compounds were compared to nontransfected HEK293 control cells using the specific calcium-binding fluorophore Fura-2AM to measure intracellular calcium [Ca2+]i release. Scymnol and scymnol sulfate caused a sustained increase in [Ca2+]i within TGR5 cells only, which was abolished by a specific inhibitor for Gαq protein (UBO-QIC). Sustained increases in [Ca2+]i were seen in both cell types with DCA exposure; this was unaffected by UBO-QIC, indicating that TGR5 activation was not involved. Exposure to UDCA did not alter [Ca2+]i, suggesting a lack of TGR5 bioactivity. These findings demonstrated that both scymnol and scymnol sulfate are novel agonists of TGR5 receptors, showing therapeutic potential for treating atherosclerosis.

An advanced method for quantitative measurements of cholesterol crystallization.

BACKGROUND: Cholesterol crystallization within an atherosclerotic plaque significantly contributes to the acceleration of plaque rupture - a problematic event due to the current lack of specific treatments to prevent such formations. Modelling this pathogenic process is also difficult due to the lack of suitable experimental models that enable quantitative analysis of crystal formation and bioactivity screening of potential therapeutic compounds. AIM: To develop an in vitro human cell model of cholesterol crystallization combined with an imaging system that incorporates both quantitative analysis and real-time continuous imaging of cholesterol crystal formation. METHODS AND RESULTS: An enhanced in vitro model of cholesterol crystallization was developed through the use of acetylated low-density lipoprotein (AcLDL) and 7-ketocholesterol as agents of foam cell induction within a human THP-1 monocytic cell line. Advanced confocal and polarizing microscopies were incorporated into the model so as to allow for quantitation of cholesterol crystallization, with the lipid-loaded group producing significantly greater numbers of cholesterol crystals than the untreated group. The utility of this system was also demonstrated by investigating the effects of the cholesterol-lowering drug lovastatin and therapeutic bile compound ursodeoxycholic acid (UDCA), showing that these drugs influence different aspects of cholesterol crystal formation. CONCLUSIONS: The in vitro human THP-1 monocyte model of cholesterol crystallization provides an effective and efficient means of quantitating cholesterol crystallization in the pre-clinical stage of research. The model also allows for the screening of potentially therapeutic compounds that may be used in attenuating or preventing cholesterol crystallization.

Samarium doped titanium dioxide nanoparticles as theranostic agents in radiation therapy.

PURPOSE: Titanium dioxide nanoparticles (TiO2 NPs) have been investigated for their role as radiosensitisers for radiation therapy. The study aims to increase the efficiency of these NPs by synthesising them with samarium. METHODS: Samarium-doped TiO2 NPs (Ti(Sm)O2 NPs) were synthesised using a solvothermal method. Transmission electron microscopy (TEM), X-ray diffraction (XRD), and energy-dispersive X-ray spectroscopy (EDS) were performed for characterising of the Ti(Sm)O2 NPs. The intracellular uptake and cytotoxicity were assessed in vitro using A549 and DU145 cancer cell lines. Furthermore, the effect of dose enhancement and generation of reactive oxygen species (ROS) in response to 6 MV X-rays was evaluated. Additionally, the image contrast properties were investigated using computed tomography (CT) images. RESULTS: The synthesised Ti(Sm)O2 NPs were about 13 nm in diameter as determined by TEM. The XRD pattern of Ti(Sm)O2 NPs was consistent with that of anatase-type TiO2. EDS confirmed the presence of samarium in the nanoparticles. At 200 µg/ml concentration, no differences in cellular uptake and cytotoxicity were observed between TiO2 NPs and Ti(Sm)O2 NPs in both A549 and DU145 cells. However, the combination of Ti(Sm)O2 NPs and X-rays elicited higher cytotoxic effect and ROS generation in the cells than that with TiO2 NPs and X-rays. The CT numbers of Ti(Sm)O2 NPs were systematically higher than that of TiO2 NPs. CONCLUSIONS: The Ti(Sm)O2 NPs increased the dose enhancement of MV X-ray beams than that elicited by TiO2 NPs. Samarium improved the efficiency of TiO2 NPs as potential radiosensitising agent.

The Effects of a Weight-Loss Herbal Formula RCM-107 and Its Eight Individual Ingredients on Glucagon-Like Peptide-1 Secretion-An In Vitro and In Silico Study.

Obesity is a multifactorial disease that can lead to other health issues. Glucagon-like peptide-1(GLP-1), as one of the satiety signal, has been linked with appetite suppression and weight loss. Due to the limitations of GLP-1 and its analogues, alternative treatments such as herbal therapies have become popular. The herbal formula RCM-107 has demonstrated its inhibitory effects on lipid and carbohydrate absorption in our previous work. However, no published data described its effects on GLP-1 secretion. Therefore, this study aimed to determine the effects of RCM-107 and its individual ingredients on GLP-1 secretion via enzyme-linked immunosorbent assay (ELISA). Furthermore, molecular docking was performed to predict the key chemical compounds that are likely to be GLP-1 receptor agonists. Gardeniae fructus, one of the ingredients in RCM-107, demonstrated significantly greater effects on inducing GLP-1 secretion than the positive control epigallocatechin gallate (EGCG). Two Gardeniae fructus ligands, 3-epioleanolic acid and crocin were predicted to bind to the active form of GLP-1 receptor at the binding pocket with residues known for the receptor activation, suggesting that they could potentially serve as receptor agonists. Overall, this study reported the effects of researched herbs on GLP-1 secretion and proposed two compounds that may be responsible for antiobesity via GLP-1 receptor activation.

Combined Effects of Gold Nanoparticles and Ionizing Radiation on Human Prostate and Lung Cancer Cell Migration.

The effect of 15 nm-sized gold nanoparticles (AuNPs) and/or ionizing radiation (IR) on the migration and adhesion of human prostate (DU145) and lung (A549) cancer cell lines was investigated. Cell migration was measured by observing the closing of a gap created by a pipette tip on cell monolayers grown in 6-well plates. The ratio of the gap areas at 0 h and 24 h were used to calculate the relative migration. The relative migration of cells irradiated with 5 Gy was found to be 89% and 86% for DU145 and A549 cells respectively. When the cells were treated with 1 mM AuNPs this fell to ~75% for both cell lines. However, when the cells were treated with both AuNPs and IR an additive effect was seen, as the relative migration rate fell to ~60%. Of interest was that when the cells were exposed to either 2 or 5 Gy IR, their ability to adhere to the surface of a polystyrene culture plate was significantly enhanced, unlike that seen for AuNPs. The delays in gap filling (cell migration) in cells treated with IR and/or AuNPs can be attributed to cellular changes which also may have altered cell motility. In addition, changes in the cytoskeleton of the cancer cells may have also affected adhesiveness and thus the cancer cell's motility response to IR.

Bisphosphonate activation of crystallized bioglass scaffolds for enhanced bone formation.

The interplay between bone formation by osteoblasts and bone resorption by osteoclasts has a critical effect on bone remodelling processes, and resultant bone quality. Bone scaffolds combined with anti-resorptive bisphosphonate drugs are a promising approach to achieving bone regeneration. Here, we have examined the synergistic effects of the bisphosphonate alendronate (ALD) coated onto calcium phosphate (CaP) modified, sintered bioactive glass 45S5 (BG) scaffolds, on osteoblast stimulation and osteoclast inhibition. After BG pre-treatment with ALD (10-8 M) for 5 days, human MG-63 osteoblasts displayed increased cellular proliferation and significantly enhanced alkaline phosphatase activity (ALP), in comparison with a non-ALD control BG. In contrast, human THP-1-derived osteoclasts cultured with 10-8 M ALD pretreated BG scaffolds showed a significant decrease in tartrate-resistant acid phosphatase (TRAcP) activity, and morphological changes indicative of functional inhibition, including reduced cell size and disruption of the osteoclast sealing zone (F-actin rings). These findings indicate that ALD-coated BG scaffolds promote osteoblast activity and inhibit osteoclast function to enhance bone formation.

Effect of pre-treatment of crystallized bioactive glass with cell culture media on structure, degradability, and biocompatibility.

The silicate glass 45S5 Bioglass® (BG) is a potential scaffold material for bone regeneration because of its excellent bioactivity, biocompatibility and ability to form a strong bond with bone tissues, via the formation of an apatite layer on its surface. The evaluation of in vitro bioactivity in physiological body fluids, whilst challenging, can offer some insights for developing the bone-bonding ability of these glasses in vivo. In this study, we investigated the influence of three different cell culture and tissue fluid-like solutions on the dissolution and calcium-phosphate (CaP) based re-precipitation behaviour at the glass-liquid interface. We also examined pre-treatment of BG with these biological solutions, and how its influence on bone-forming MG-63 osteoblastic cell proliferation, viability and adhesion. The biological solutions used in this comparative study were: commercial cell culture medium (DMEM), a DMEM solution without organic components (DML) and a simulated body fluid (SBF), incorporating TRIS-buffer. Incubation of BG in these solutions over 28 days resulted in differences in weight loss, solution pH and ion release, and the development of CaP-based surface layers. XRD and FT-IR analyses showed clear differences in the characteristics of the CaP-based coating layers formed by the different solutions. The interfacial reactivity between the glass and the solutions depended on the composition and properties of the solutions. The formation of the CaP layer occurred more rapidly in SBF due to the presence of TRIS-buffer, which also significantly accelerated glass dissolution, further reducing the BG mass in SBF. MG-63 osteoblasts proliferated and spread more rapidly across the surfaces of all pre-conditioned BG, compared to fresh BG. The experimental results of this work help clarify differences between in vitro bioactivity of BG observed in cell culture solutions and in vivo BG bioactivity.

Titanium Dioxide Nanoparticles as Radiosensitisers: An In vitro and Phantom-Based Study.

Objective: Radiosensitisation caused by titanium dioxide nanoparticles (TiO2-NPs) is investigated using phantoms (PRESAGE® dosimeters) and in vitro using two types of cell lines, cultured human keratinocyte (HaCaT) and prostate cancer (DU145) cells. Methods: Anatase TiO2-NPs were synthesised, characterised and functionalised to allow dispersion in culture-medium for in vitro studies and halocarbons (PRESAGE® chemical compositions). PRESAGE® dosimeters were scanned with spectrophotometer to determine the radiation dose enhancement. Clonogenic and cell viability assays were employed to determine cells survival curves from which the dose enhancement levels "radiosensitisation" are deduced. Results: Comparable levels of radiosensitisation were observed in both phantoms and cells at kilovoltage ranges of x-ray energies (slightly higher in vitro). Significant radiosensitisation (~67 %) of control was also noted in cells at megavoltage energies (commonly used in radiotherapy), compared to negligible levels detected by phantoms. This difference is attributed to biochemical effects, specifically the generation of reactive oxygen species (ROS) such as hydroxyl radicals (•OH), which are only manifested in aqueous environments of cells and are non-existent in case of phantoms. Conclusions: This research shows that TiO2-NPs improve the efficiency of dose delivery, which has implications for future radiotherapy treatments. Literature shows that Ti2O3-NPs can be used as imaging agents hence with these findings renders these NPs as theranostic agents.

Porous 45S5 Bioglass®-based scaffolds using stereolithography: Effect of partial pre-sintering on structural and mechanical properties of scaffolds.

Scaffolds made from 45S5 Bioglass® ceramic (BG) show clinical potential in bone regeneration due to their excellent bioactivity and ability to bond to natural bone tissue. However, porous BG scaffolds are limited by their mechanical integrity and by the substantial volume contractions occurring upon sintering. This study examines stereolithographic (SLA) methods to fabricate mechanically robust and porous Bioglass®-based ceramic scaffolds, with regular and interconnected pore networks and using various computer-aided design architectures. It was found that a diamond-like (DM) architecture gave scaffolds the most controllable results without any observable closed porosity in the fired scaffolds. When the pore dimensions of the DM scaffolds of the same porosity (~60vol%) were decreased from 700 to 400µm, the compressive strength values increased from 3.5 to 6.7MPa. In addition, smaller dimensional shrinkage could be obtained by employing partially pre-sintered bioglass, compared to standard 45S5 Bioglass®. Scaffolds derived from pre-sintered bioglass also showed marginally improved compressive strength.

ZnO nanoparticles and organic chemical UV-filters are equally well tolerated by human immune cells.

An important part of assessing the toxic potential of nanoparticles for specific applications should be the direct comparison of biological activities with those of alternative materials for the same application. Nanoparticulate inorganic ultraviolet (UV) filters, such as zinc oxide (ZnO), are commonly incorporated into transparent sunscreen and cosmetic formulations. However, concerns have been raised about potential unwanted effects, despite their negligible skin penetration and inherent advantages over organic chemical UV-filters. To provide useful application-relevant assessments of their potential hazard with/without UVA co-exposure, we directly compared cytotoxic and immune response profiles of human THP-1 monocytic cells to ZnO nanoparticles (30 nm) with bulk ZnO particulates (200 nm) and five conventional organic chemical UV-filters - butylmethoxydibenzoylmethane (avobenzone), octylmethoxycinnamate, octylsalicylate, homosalate and 4-methylbenzylidene camphor. High exposure concentrations of both organic and particulate UV-filters were required to cause cytotoxicity in monocyte and macrophage cultures after 24 h. Co-exposure with UVA (6.7 J/cm(2)) did not alter cytotoxicity profiles. Particle surface area-based dose responses showed that ZnO NPs were better tolerated than bulk ZnO. Organic and particulate UV-filters increased apoptosis at similar doses. Only particulates increased the generation of reactive oxygen species. Interleukin-8 (IL-8) release was increased by all particulates, avobenzone, homosalate and octylsalicylate. IL-1ß release was only increased in macrophages by exposure to avobenzone and homosalate. In conclusion, direct effects were caused in monocytes and macrophages at similar concentrations of both organic UV-filters and ZnO nanoparticulates - indicating that their intrinsic cytotoxicity is similar. With their lower skin penetration, ZnO nanoparticles are expected to have lower bioactivity when used in sunscreens.

Investigating the immunomodulatory nature of zinc oxide nanoparticles at sub-cytotoxic levels in vitro and after intranasal instillation in vivo.

BACKGROUND: This study evaluates the time-dependent pro-inflammatory response of the model human lung epithelial cells (A549) to industrially relevant zinc oxide nanoparticles (ZnO NPs). In terms of toxicity, ZnO-NPs are categorised into the group of high toxicity nanomaterials. However information on pro-inflammatory potential of these NPs at sub-toxic concentrations is limited. Understanding how cellular defense mechanisms function in the presence of sub-cytotoxic concentrations of these NPs is vital. Moreover, there is an urgent need for additional in vivo studies addressing pulmonary toxicity due to accidental inhalation of ZnO NPs. RESULTS: Exposure to sub-cytotoxic ZnO NP concentrations (20 µg/mL) induced significant up-regulation of mRNA for the pro-inflammatory cytokine IL-8 and redox stress marker heme oxygenase-1, along with increased release of IL-8. The highest pro-inflammatory response was recorded after 4 to 6 hr exposure to ZnO NPs over a 24 hr period. Pre-treatment of A549 cells with the sulfhydryl antioxidant N-acetyl cysteine (at 5 mM) resulted in significant reduction of the up-regulation of inflammatory markers, confirming the role of reactive oxygen species in the observed immunomodulatory effects, independent of cytotoxicity. Furthermore, we report for the first time that, intranasal instillation of a single dose (5 mg/kg) of pristine or surfactant-dispersed ZnO NPs can cause pulmonary inflammation, already after 24 hr in a murine model. This was confirmed by up-regulation of eotaxin mRNA in the lung tissue and release of pro-inflammatory cytokines in the sera of mice exposed to ZnO NPs. CONCLUSION: Our study highlights that even at sub-cytotoxic doses ZnO NPs can stimulate a strong inflammatory and antioxidant response in A549 cells. ZnO NP mediated cytotoxicity may be the outcome of failure of cellular redox machinery to contain excessive ROS formation. Moreover exposure to a single but relatively high dose of ZnO NPs via intranasal instillation may provoke acute pulmonary inflammatory reactions in vivo.

Reducing ZnO nanoparticle cytotoxicity by surface modification.

Nanoparticulate zinc oxide (ZnO) is one of the most widely used engineered nanomaterials and its toxicology has gained considerable recent attention. A key aspect for controlling biological interactions at the nanoscale is understanding the relevant nanoparticle surface chemistry. In this study, we have determined the disposition of ZnO nanoparticles within human immune cells by measurement of total Zn, as well as the proportions of extra- and intracellular dissolved Zn as a function of dose and surface coating. From this mass balance, the intracellular soluble Zn levels showed little difference in regard to dose above a certain minimal level or to different surface coatings. PEGylation of ZnO NPs reduced their cytotoxicity as a result of decreased cellular uptake arising from a minimal protein corona. We conclude that the key role of the surface properties of ZnO NPs in controlling cytotoxicity is to regulate cellular nanoparticle uptake rather than altering either intracellular or extracellular Zn dissolution.

Comparison of UVA-induced ROS and sunscreen nanoparticle-generated ROS in human immune cells.

Oxidative damage to cells and tissues from free radicals induced by ultraviolet (UV) irradiation can be attenuated by sunscreen components, such as ZnO and TiO2 nanoparticles (NPs). Although it is known that reactive oxygen species (ROS) are generated by cells upon exposure to ZnO and TiO2 NPs, it is unknown to what extent the amount generated is altered with UV co-exposure. As it is a critical component for determining the relative risk of these NPs when used in sunscreen formulations, we have investigated ROS generation by these NPs in human THP-1 monocyte immune cells following UVA co-exposure. Whilst the applied UVA dose (6.7 J cm(-2)) did not alter cell viability after 24 h, it induced significant ROS production - causing a 7-fold increase in intracellular peroxide and 3.3-fold increase in mitochondrial superoxide levels after 1 h. However, co-exposure to NPs and UVA generated the same or less ROS than with UVA exposure alone, with the exception of anatase TiO2, which showed significantly increased levels. These findings indicate that ROS generation from nanosunscreens is, in most cases, an insignificant contributor to the overall risk associated with oxidative stress from UVA exposure itself.

Quantification of ZnO nanoparticle uptake, distribution, and dissolution within individual human macrophages.

The usefulness of zinc oxide (ZnO) nanoparticles has led to their wide distribution in consumer products, despite only a limited understanding of how this nanomaterial behaves within biological systems. From a nanotoxicological viewpoint the interaction(s) of ZnO nanoparticles with cells of the immune system is of specific interest, as these nanostructures are readily phagocytosed. In this study, rapid scanning X-ray fluorescence microscopy was used to assay the number ZnO nanoparticles associated with ∼1000 individual THP-1 monocyte-derived human macrophages. These data showed that nanoparticle-treated cells endured a 400% elevation in total Zn levels, 13-fold greater than the increase observed when incubated in the presence of an equitoxic concentration of ZnCl2. Even after excluding the contribution of internalized nanoparticles, Zn levels in nanoparticle treated cells were raised ∼200% above basal levels. As dissolution of ZnO nanoparticles is critical to their cytotoxic response, we utilized a strategy combining ion beam milling, X-ray fluorescence and scanning electron microscopy to directly probe the distribution and composition of ZnO nanoparticles throughout the cellular interior. This study demonstrated that correlative photon and ion beam imaging techniques can provide both high-resolution and statistically powerful information on the biology of metal oxide nanoparticles at the single-cell level. Our approach promises ready application to broader studies of phenomena at the interface of nanotechnology and biology.

Surface coatings of ZnO nanoparticles mitigate differentially a host of transcriptional, protein and signalling responses in primary human olfactory cells.

BACKGROUND: Inhaled nanoparticles have been reported in some instances to translocate from the nostril to the olfactory bulb in exposed rats. In close proximity to the olfactory bulb is the olfactory mucosa, within which resides a niche of multipotent cells. Cells isolated from this area may provide a relevant in vitro system to investigate potential effects of workplace exposure to inhaled zinc oxide nanoparticles. METHODS: Four types of commercially-available zinc oxide (ZnO) nanoparticles, two coated and two uncoated, were examined for their effects on primary human cells cultured from the olfactory mucosa. Human olfactory neurosphere-derived (hONS) cells from healthy adult donors were analyzed for modulation of cytokine levels, activation of intracellular signalling pathways, changes in gene-expression patterns across the whole genome, and compromised cellular function over a 24 h period following exposure to the nanoparticles suspended in cell culture medium. RESULTS: ZnO nanoparticle toxicity in hONS cells was mediated through a battery of mechanisms largely related to cell stress, inflammatory response and apoptosis, but not activation of mechanisms that repair damaged DNA. Surface coatings on the ZnO nanoparticles mitigated these cellular responses to varying degrees. CONCLUSIONS: The results indicate that care should be taken in the workplace to minimize generation of, and exposure to, aerosols of uncoated ZnO nanoparticles, given the adverse responses reported here using multipotent cells derived from the olfactory mucosa.

Relating cytotoxicity, zinc ions, and reactive oxygen in ZnO nanoparticle-exposed human immune cells.

Although zinc oxide (ZnO) nanoparticles (NPs) have been widely formulated in sunscreens, the relationship between reactive oxygen species (ROS) generation induced by these particles, zinc ions, and cytotoxicity is not clearly understood. This study explores whether these factors can be accurately quantified and related. The study demonstrates a strong correlation between ZnO NP-induced cytotoxicity and free intracellular zinc concentration (R (2) = .945) in human immune cells, indicating a requirement for NP dissolution to precede cytotoxicity. In addition, although direct exposure to ZnO NPs was found to induce cytotoxicity at relatively high concentrations, indirect exposure (via dialysis) was not cytotoxic, even at extremely high concentrations, highlighting a requirement for NP-to-cell contact. Elevated levels of ROS present in NP-exposed cells also correlated to both cytotoxicity and intracellular free zinc. Although the addition of antioxidant was able to reduce ROS, cytotoxicity to ZnO NPs was unaffected, suggesting ROS may be, in part, a result of cytotoxicity rather than a causal factor. This study highlights both the requirement and role of intracellular dissolution of zinc nanomaterials to elicit a cytotoxic response. This response is only partially ROS dependent, and therefore, modification of NP uptake and their intracellular solubility are key components in modulating the bioactivity of ZnO NPs.

Formation of zinc-containing nanoparticles from Zn²âº ions in cell culture media: implications for the nanotoxicology of ZnO.

Zinc ions generate a range of poorly soluble Zn-containing nanoparticles when added to commonly used mammalian cell culture media. The formation of these nanoparticles confounds the use of soluble Zn salts as positive controls during cytotoxicity testing of other Zn-containing nanoparticles, such as ZnO. These nanoprecipitates can either be crystalline or amorphous and vary in composition depending upon the concentration of Zn(II) within the medium. The cytotoxicity and immune system response of these nanoparticles in situ are similar to those of 30 nm ZnO nanoparticles. The low residual level of truly soluble Zn species (taken as species passing through a 2 kDa membrane) in cell culture media with serum is insufficient to elicit any appreciable cytotoxicity. These observations highlight the importance of employing appropriate controls when studying ZnO nanoparticle toxicity and suggest a re-evaluation of the conclusions drawn in some previous cytotoxicity studies.