Iron Speciation in Respirable Particulate Matter and Implications for Human Health.

Particulate matter (PM) air pollution poses a major global health risk, but the role of iron (Fe) is not clearly defined because chemistry at the particle-cell interface is often not considered. Detailed spectromicroscopy characterizations of PM2.5 samples from the San Joaquin Valley, CA identified major Fe-bearing components and estimated their relative proportions. Iron in ambient PM2.5 was present in spatially and temporally variable mixtures, mostly as Fe(III) oxides and phyllosilicates, but with significant fractions of metallic iron (Fe(0)), Fe(II,III) oxide, and Fe(III) bonded to organic carbon. Fe(0) was present as aggregated, nm-sized particles that comprised up to ∼30% of the Fe spectral fraction. Mixtures reflect anthropogenic and geogenic particles subjected to environmental weathering, but reduced Fe in PM originates from anthropogenic sources, likely as abrasion products. Possible mechanistic pathways involving Fe(0) particles and mixtures of Fe(II) and Fe(III) surface species may generate hydrogen peroxide and oxygen-centered radical species (hydroxyl, hydroperoxyl, or superoxide) in Fenton-type reactions. From a health perspective, PM mixtures with reduced and oxidized Fe will have a disproportionate effect in cellular response after inhalation because of their tendency to shuttle electrons and produce oxidants and electrophiles that induce inflammation and oxidative stress.

Iron Speciation in Particulate Matter (PM2.5) from Urban Los Angeles Using Spectro-microscopy Methods.

The speciation, oxidation states, and relative abundance of iron (Fe) phases in PM2.5 samples from two locations in urban Los Angeles were investigated using a combination of bulk and spatially resolved, element-specific spectroscopy and microscopy methods. Synchrotron X-ray absorption spectroscopy (XAS) of bulk samples in situ (i.e., without extraction or digestion) was used to quantify the relative fractions of major Fe phases, which were corroborated by spatially resolved spectro-microscopy measurements. Ferrihydrite (amorphous Fe(III)-hydroxide) comprised the largest Fe fraction (34-52%), with hematite (α-Fe2O3; 13-23%) and magnetite (Fe3O4; 10-24%) identified as major crystalline oxide components. An Fe-bearing phyllosilicate fraction (16-23%) was fit best with a reference spectrum of a natural illite/smectite mineral, and metallic Fe(0) was a relatively small (2-6%) but easily identified component. Sizes, morphologies, oxidation state, and trace element compositions of Fe-bearing PM from electron microscopy, electron energy loss spectroscopy (EELS), and scanning transmission X-ray microscopy (STXM) revealed variable and heterogeneous mixtures of Fe species and phases, often associated with carbonaceous material with evidence of surface oxidation. Ferrihydrite (or related Fe(III) hydroxide phases) was ubiquitous in PM samples. It forms as an oxidation or surface alteration product of crystalline Fe phases, and also occurs as coatings or nanoparticles dispersed with other phases as a result of environmental dissolution and re-precipitation reactions. The prevalence of ferrihydrite (and adsorbed Fe(III) has likely been underestimated in studies of ambient PM because it is non-crystalline, non-magnetic, more soluble than crystalline phases, and found in complex mixtures. Review of potential sources of different particle types suggests that the majority of Fe-bearing PM from these urban sites originates from anthropogenic activities, primarily abrasion products from vehicle braking systems and engine emissions from combustion and/or wear. These variable mixtures have a high probability for electron transfer reactions between Fe, redox-active metals such as copper, and reactive carbon species such as quinones. Our findings suggest the need to assess biological responses of specific Fe-bearing phases both individually and in combination to unravel mechanisms of adverse health effects of particulate Fe.

Colloidal structure and proton conductivity of the gel within the electrosensory organs of cartilaginous fishes.

Cartilaginous fishes possess gel-filled tubular sensory organs called Ampullae of Lorenzini (AoL) that are used to detect electric fields. Although recent studies have identified various components of AoL gel, it has remained unclear how the molecules are structurally arranged and how their structure influences the function of the organs. Here we describe the structure of AoL gel by microscopy and small-angle X-ray scattering and infer that the material is colloidal in nature. To assess the relative function of the gel's protein constituents, we compared the microscopic structure, X-ray scattering, and proton conductivity properties of the gel before and after enzymatic digestion with a protease. We discovered that while proteins were largely responsible for conferring the viscous nature of the gel, their removal did not diminish proton conductivity. The findings lay the groundwork for more detailed studies into the specific interactions of molecules inside AoL gel at the nanoscale.

Evidence of chitin in the ampullae of Lorenzini of chondrichthyan fishes.

We previously reported that the polysaccharide chitin, a key component of arthropod exoskeletons and fungal cell walls, is endogenously produced by fishes and amphibians in spite of the widely held view that it was not synthesized by vertebrates [1]. Genes encoding chitin synthase enzymes were found in the genomes of a number of fishes and amphibians and shown to be correspondingly expressed at the sites where chitin was localized [1,2]. In this report, we present evidence suggesting that chitin is prevalent within the specialized electrosensory organs of cartilaginous fishes (Chondrichthyes). These organs, the Ampullae of Lorenzini (AoL), are widely distributed and comprise a series of gel-filled canals emanating from pores in the skin (Figure 1A). The canals extend into bulbous structures called alveoli that contain sensory cells capable of detecting subtle changes in electric fields (Figure 1B) [3,4]. The findings described here extend the number of vertebrate taxa where endogenous chitin production has been detected and raise questions regarding chitin's potential function in chondrichthyan fishes and other aquatic vertebrates.

Surface characterization and chemical speciation of adsorbed iron(iii) on oxidized carbon nanoparticles.

Carbonaceous nanomaterials represent a significant portion of ultra-fine airborne particulate matter, and iron is the most abundant transition metal in air particles. Owing to their high surface area and atmospheric oxidation, carbon nanoparticles (CNP) are enriched with surface carbonyl functional groups and act as a host for metals and small molecules. Using a synthetic model, concentration-dependent changes in the chemical speciation of iron adsorbed on oxidized carbon surfaces were investigated by a combination of X-ray and electron microscopic and spectroscopic methods. Carbon K-edge absorption spectra demonstrated that the CNP surface was enriched with carboxylic acid groups after chemical oxidation but that microporosity was unchanged. Oxidized CNP showed a high affinity for sorption of Fe(iii) from solution (75-95% uptake) and spectroscopic measurements confirmed a 3+ oxidation state of Fe on CNP irrespective of surface loading. The bonding of adsorbed Fe(iii) at variable loadings was determined by iron K-edge X-ray absorption spectroscopy. At low loadings (3 and 10 µmol Fe m-2 CNP), mononuclear Fe was octahedrally coordinated to oxygen atoms of carboxylate groups. As Fe surface coverage increased (21 and 31 µmol Fe m-2 CNP), Fe-Fe backscatters were observed at interatomic distances indicating iron (oxy)hydroxide particle formation on CNP. Electron-donating surface carboxylate groups on CNP coordinated and stabilized mononuclear Fe(iii). Saturation of high-affinity sites may have promoted hydroxide particle nucleation at higher loading, demonstrating that the chemical form of reactive metal ions may change with surface concentration and degree of CNP surface oxidation. Model systems such as those discussed here, with controlled surface properties and known chemical speciation of adsorbed metals, are needed to establish structure-activity models for toxicity assessments of environmentally relevant nanoparticles.

The bismuth oxyhalide family: thin film synthesis and periodic properties.

Bismuth oxyhalides (BiOX, where X = F, Cl, Br, I) are interesting materials due to their layered structure, which can be useful for different applications. In this work, we present the synthesis of the complete BiOX family in the thin film form. The tetragonal phase Bi2O3 film deposited onto a glass substrate was transformed into BiOF, BiOCl or BiOBr by a simple immersion at ambient temperature in a halide (X = F, Cl, Br) containing solution. For these films, a residual phase from the oxide was present and for BiOF another phase (tentatively identified as Bi7O5F11) was present too. For the BiOI film synthesis, an iodine and bismuth containing solution was sprayed onto the glass substrate heated at 275 °C and a pure phase was obtained. Microstructural and morphological characterization was performed by X-ray diffraction and scanning electron microscopy, while the chemical environment was studied by X-ray photoelectron spectroscopy. Optical and photocatalytic properties were also obtained. The physical and chemical characteristics of the BiOX films follow a correlation with the atomic radius of the halogen atom incorporated into the corresponding lattice. All the BiOX films showed a photocatalytic response for the photodiscoloration of indigo carmine dye under simulated sunlight irradiation in an alkaline medium. The photocatalytic reactions occurred via 2 proton-electron transfer from the oxide or oxyhalide to the adsorbed IC dye, favoring its reduction to the corresponding leuco IC form.

Low dose inflammatory potential of silica particles in human-derived THP-1 macrophage cell culture studies - Mechanism and effects of particle size and iron.

Silica and iron are major constituents in ambient particulate matter, and iron is a common impurity in many engineered nanomaterials. The purpose of this work was to determine the pro-inflammatory and other biological effects and mechanism of particle size and iron presence under low dose, non-cytotoxic conditions that are likely to approximate actual exposure levels, in contrast with higher dose studies in which cytotoxicity occurs. Specifically, human-derived THP-1 macrophages were exposed to 1 µg/ml of pristine and iron-coated 50 nm and 2 µm engineered silica nanoparticles. Particles were first characterized for size, size distribution, surface area, iron concentration, phase and aggregation in cell culture media. Then, biological assays were conducted to determine a non-lethal dose used in subsequent experiments. Superoxide production, lipid peroxidation, and increased pro-inflammatory cytokine (TNF-α and IL-1ß) mRNA expression were measured as a function of particle size and iron presence. Smaller particle size and the presence of iron increased superoxide production, lipid peroxidation, and the induction of pro-inflammatory cytokine mRNA expression. Separate addition of an iron-chelator, a scavenger of superoxide and hydrogen peroxide, and an inhibitor of phosphatidylcholine specific phospholipase C (PC-PLC), suppressed the increase in cytokine mRNA expression. Furthermore, free iron itself showed none of the aforementioned effects. The results highlight the importance of particle size and iron in lung inflammation for both natural and engineered nanomaterials, under low dose, non-toxic conditions, and support the role of an oxidant, lipid peroxidation and PC-PLC dependent inflammatory mechanism.

Delayed Nrf2-regulated antioxidant gene induction in response to silica nanoparticles.

Silica nanoparticles with iron on their surface cause the production of oxidants and stimulate an inflammatory response in macrophages. Nuclear factor erythroid-derived 2 - like factor 2 (Nrf2) signaling and its regulated antioxidant genes play critical roles in maintaining redox homeostasis. In this study we investigated the regulation of four representative Nrf2-regulated antioxidant genes; i.e., glutamate cysteine ligase (GCL) catalytic subunit (GCLC), GCL modifier subunit (GCLM), heme oxygenase 1 (HO-1), and NAD(P)H:quinone oxidoreductase-1 (NQO-1), by iron-coated silica nanoparticles (SiO2-Fe) in human THP-1 macrophages. We found that the expression of these four antioxidant genes was modified by SiO2-Fe in a time-dependent manner. At 6h, their expression was unchanged except for GCLC, which was reduced compared with controls. At 18h, the expression of these antioxidant genes was significantly increased compared with controls. In contrast, the Nrf2 activator sulforaphane induced all antioxidant genes at as early as 3h. The nuclear translocation of Nrf2 occurred later than that for NF-κB p65 protein and the induction of proinflammatory cytokines (TNFα and IL-1ß). NF-κB inhibitor SN50 prevented the reduction of GCLC at 6h and abolished the induction of antioxidant genes at 18h by SiO2-Fe, but did not affect the basal and sulforaphane-induced expression of antioxidant genes, suggesting that NF-κB signaling plays a key role in the induction of Nrf2-mediated genes in response to SiO2-Fe. Consistently, SN50 inhibited the nuclear translocation of Nrf2 caused by SiO2-Fe. In addition, Nrf2 silencing decreased the basal and SiO2-induced expression of the four reprehensive antioxidant genes. Taken together, these data indicated that SiO2-Fe induced a delayed response of Nrf2-regulated antioxidant genes, likely through NF-κB-Nrf2 interactions.

Endothelial cells derived from embryonic stem cells respond to cues from topographical surface patterns.

The generation of micro- and nano-topography similar to those found in the extra cellular matrix of three-dimensional tissues is one technique used to recapitulate the cell-tissue physiology found in the native tissues. Despite the fact that ample studies have been conducted on the physiological significance of endothelial cells alignment parallel to shear stress, as this is the normal physiologic arrangement for healthy arterial EC, very few studies have examined the use of topographical signals to initiate endothelial cell alignment. Here, we have examined the ability for our mouse embryonic stem cell-derived endothelial cells (ESC-EC) to align on various microchip topographical systems. Briefly, we generated metal molds with 'wrinkled' topography using 1) 15 nm and 2) 30 nm of gold coating on the pre-strained polystryene (PS) sheets. After thermal-induced shrinkage of the PS sheets, polydimethylsiloxane (PDMS) microchips were then generated from the wrinkled molds. Using similar Shrink™-based technology, 3) larger selectively crazed acetone-etched lines in the PS sheets, and 4) fully crazed acetone-treated PS sheets of stochastic topographical morphology were also generated. The 15 nm and 30 nm gold coating generated 'wrinkles' of uniaxial anisotropic channels at nano-scaled widths while the crazing generated micron-sized channels. The ESC-EC were able to respond and align on the 320 nm, 510 nm, and the acetone-etched 10.5 µm channels, but not on the fully 'crazed' topographies. Moreover, the ESC-EC aligned most robustly on the wrinkles, and preferentially to ridge edges on the 10.5 µm-sized channels. The ability to robustly align EC on topographical surfaces enables a variety of controlled physiological studies of EC-EC and EC-ECM contact guidance, as well as having potential applications for the rapid endothelialization of stents and vascular grafts.

Iron-mediated lipid peroxidation and lipid raft disruption in low-dose silica-induced macrophage cytokine production.

Silica inhalation can induce respiratory disease. Iron is suspected of playing an important role in silica-mediated respiratory toxicity, but unambiguously determining its role has been hampered by incomplete characterization, use of high particle doses, and lack of understanding of proinflammatory mechanisms. In this study, we investigated a novel hypothesis for the mechanism of silica particle-induced increase in cytokine production. We studied the role of iron in lipid peroxidation-dependent transcription of cytokines in macrophages by ground natural silica particles at low sublethal doses. Particle size, size distribution, surface area, and structure were determined using electron microscopy, nitrogen adsorption, and X-ray diffraction. Iron impurity concentrations before and after acid treatment were determined by energy-dispersive X-ray and inductively coupled plasma mass spectroscopy. At a low noncytotoxic dose (1 µg/ml) of 2-µm silica, the presence of iron significantly increased superoxide (O(2)(•-)), lipid peroxidation, lipid raft disruption, and cytokine production in macrophages. The iron chelators deferoxamine mesylate and diethylenetriaminepentaacetic acid were found to abrogate O(2)(•-) production and inhibit lipid peroxidation, raft disruption, and cytokine induction. Tricyclodecan-9-yl xanthate, a competitive inhibitor of phosphatidylcholine-specific phospholipase C (PC-PLC), which is an upstream participant in NF-κB activation, and manganese(III) tetrakis(N-ethylpyridinium-2-yl) porphyrin, a superoxide dismutase and catalase mimic, blocked silica-stimulated cytokine production. We propose a pathway of iron-induced lipid peroxidation disrupting lipid rafts and signaling for the production of cytokines through PC-PLC in silica-exposed macrophages.

Spatially resolved energy electron loss spectroscopy studies of iron oxide nanoparticles.

The oxidation state of iron oxide nanoparticles co-generated with soot during a combustion process was studied using electron energy-loss spectroscopy (EELS). Spatially resolved EELS spectra in the scanning transmission electron microscopy mode were collected to detect changes in the oxidation state between the cores and surfaces of the particles. Quantification of the intensity ratio of the white lines of the iron L-ionization edge was used to measure the iron oxidation state. Quantitative results obtained from Pearson's method, which can be directly compared with the literature data, indicated that the L3 /L2-intensity ratio for these particles changes from 5.5 +/- 0.3 in the particles' cores to 4.4 +/- 0.3 at their surfaces. This change can be directly related to the reduction of the iron oxidation state at the surface of the particles. Experimental results indicate that the cores of the particles are composed of gamma-Fe2O3, which seems to be reduced to FeO at their surfaces. These results were also supported by the fine structure of the oxygen K-edge and by the significant chemical shift of the iron L-edge.

Oxidative stress and NFkappaB activation in the lungs of rats: a synergistic interaction between soot and iron particles.

Particulate matter (PM) has been associated with a variety of adverse health effects primarily involving the cardiopulmonary system. However, the precise biological mechanisms to explain how exposure to PM exacerbates or directly causes adverse effects are unknown. Particles of varying composition may play a critical role in these effects. To study such a phenomenon, a simple, laminar diffusion flame was used to generate aerosols of soot and iron particles in the ultrafine size range. Exposures of healthy adult rats were for 6 h/day for 3 days. Conditions used included exposure to soot only, iron only, or a combination of soot and iron. We found animals exposed to soot particles at 250 microg/m3 had no adverse respiratory effects. Exposure to iron alone at a concentration of 57 microg/m3 also had no respiratory effects. However, the addition of 45 microg/m3 of iron to soot with a combined total mass concentration of 250 microg/m3 demonstrated significant pulmonary ferritin induction, oxidative stress, elevation of IL-1beta, and cytochrome P450s, as well as activation of NFkappaB. These findings suggest that a synergistic interaction between soot and iron particles account for biological responses not found with exposure to iron alone or to soot alone.

Reduced lung cell proliferation following short-term exposure to ultrafine soot and iron particles in neonatal rats: key to impaired lung growth?

Particulate matter (PM) has been associated with a variety of negative health outcomes in children involving the respiratory system and early development. However, the precise mechanisms to explain how exposure to airborne particles may cause adverse effects in children are unknown. To study their influence on early postnatal development, a simple, laminar diffusion flame was used to generate an aerosol of soot and iron particles in the size range of 10 to 50 nm. Exposure of 10-day-old rat pups to soot and iron particles was for 6 h/day for 3 days. The lungs were examined following a single injection of bromodeoxyuridine (BrdU) 2 h prior to necropsy. Neonatal rats exposed to these particles demonstrated no effect on the rate of cell proliferation within terminal bronchioles or the general lung parenchyma. In contrast, within those regions arising immediately beyond the terminal bronchioles (defined as the proximal alveolar region), the rate of cell proliferation was significantly reduced compared with filtered air controls. These findings strongly suggest exposure to airborne particles during early neonatal life has significant direct effects on lung growth by altering cell division within critical sites of the respiratory tract during periods of rapid postnatal development. Such effects may result in altered growth in the respiratory system that may be associated with lifelong consequences.