Both excess brain Fe and air pollution (AP) exposures are associated with increased risk for multiple neurodegenerative disorders. Fe is a redox-active metal that is abundant in AP and even further elevated in U.S. subway systems. Exposures to AP and associated contaminants, such as Fe, are lifelong and could therefore contribute to elevated brain Fe observed in neurodegenerative diseases, particularly via nasal olfactory uptake of ultrafine particle AP. These studies tested the hypotheses that exogenously generated Fe oxide nanoparticles could reach the brain following inhalational exposures and produce neurotoxic effects consistent with neurodegenerative diseases and disorders in adult C57/Bl6J mice exposed by inhalation to Fe nanoparticles at a concentration similar to those found in underground subway systems ( 150 µg/m3) for 20 days. Olfactory bulb sections and exposure chamber TEM grids were analyzed for Fe speciation. Measures included brain volumetric and diffusivity changes; levels of striatal and cerebellar neurotransmitters and trans-sulfuration markers; quantification of frontal cortical and hippocampal Aβ42, total tau, and phosphorylated tau; and behavioral alterations in locomotor activity and memory. Particle speciation confirmed similarity of Fe oxides (mostly magnetite) found on chamber TEM grids and in olfactory bulb. Alzheimer’s disease (AD) like characteristics were seen in Fe-exposed females including increased olfactory bulb diffusivity, impaired memory, and increased accumulation of total and phosphorylated tau, with total hippocampal tau levels significantly correlated with increased errors in the radial arm maze. Fe-exposed males showed increased volume of the substantia nigra pars compacta, a region critical to the motor impairments seen in Parkinson’s disease (PD), in conjunction with reduced volume of the trigeminal nerve and optic tract and chiasm. Inhaled Fe oxide nanoparticles appeared to lead to olfactory bulb uptake. Further, these exposures reproduced characteristic features of neurodegenerative diseases in a sex-dependent manner, with females evidencing features similar to those seen in AD and effects in regions in males associated with PD. As such, prolonged inhaled Fe exposure via AP should be considered as a source of elevated brain Fe with aging, and as a risk factor for neurodegenerative diseases. The bases for dichotomous sex effects of inhaled Fe nanoparticles is as of yet unclear. Also as of yet unknown is how duration of such Fe exposures affect outcome, and/or whether exposures to inhaled Fe during early brain development enhances vulnerability to subsequent Fe exposures. Collectively, these findings suggest that regulation of air Fe levels, particularly in enclosed areas like subway stations, may have broad public health protective effects.

Background Air pollution has been associated with neurodevelopmental disorders in epidemiological studies. In our studies in mice, developmental exposures to ambient ultrafine particulate (UFP) matter either postnatally or gestationally results in neurotoxic consequences that include brain metal dyshomeostasis, including significant increases in brain Fe. Since Fe is redox active and neurotoxic to brain in excess, this study examined the extent to which postnatal Fe inhalation exposure, might contribute to the observed neurotoxicity of UFPs. Mice were exposed to 1 µg/m 3 Fe oxide nanoparticles alone, or in conjunction with sulfur dioxide (Fe (1 µg/m 3 ) + SO 2 (SO 2 at 1.31 mg/m 3 , 500 ppb) from postnatal days 4–7 and 10–13 for 4 h/day. Results Overarching results included the observations that Fe + SO 2 produced greater neurotoxicity than did Fe alone, that females appeared to show greater vulnerability to these exposures than did males, and that profiles of effects differed by sex. Consistent with metal dyshomeostasis, both Fe only and Fe + SO 2 exposures altered correlations of Fe and of sulfur (S) with other metals in a sex and tissue-specific manner. Specifically, altered metal levels in lung, but particularly in frontal cortex were found, with reductions produced by Fe in females, but increases produced by Fe + SO 2 in males. At PND14, marked changes in brain frontal cortex and striatal neurotransmitter systems were observed, particularly in response to combined Fe + SO2 as compared to Fe only, in glutamatergic and dopaminergic functions that were of opposite directions by sex. Changes in markers of trans-sulfuration in frontal cortex likewise differed in females as compared to males. Residual neurotransmitter changes were limited at PND60. Increases in serum glutathione and Il-1a were female-specific effects of combined Fe + SO2. Conclusions Collectively, these findings suggest a role for the Fe contamination in air pollution in the observed neurotoxicity of ambient UFPs and that such involvement may be different by chemical mixture. Translation of such results to humans requires verification, and, if found, would suggest a need for regulation of Fe in air for public health protection.

An amendment to this paper has been published and can be accessed via a link at the top of the paper.

Background: Exposure to air pollution has been identified as a possible environmental contributor to Alzheimer's Disease (AD) risk. As the number of people with AD worldwide continues to rise, it becomes vital to understand the nature of this potential gene-environment interaction. This study assessed the effects of short-term exposures to concentrated ambient ultrafine particulates (UFP, <100 nm) on measurements of amyloid-beta, tau, and microglial morphology. Methods: Two cohorts of aged (12.5-14 months) 3xTgAD and NTg mice were exposed to concentrated ambient UFP or filtered air for 2 weeks (4-h/day, 4 days/week). Bronchoalveolar lavage fluid and brain tissue were collected twenty-four hours following the last exposure to evaluate lung inflammation, tau pathology, amyloid-beta pathology, and glial cell morphology. Results: No exposure- or genotype-related changes were found with any of the measures of lung inflammation or in the hippocampal staining density of astrocyte marker glial fibrillary acidic protein. The microglia marker, ionized calcium binding adaptor molecule 1, and amyloid-beta marker, 6E10, exhibited significant genotype by exposure interactions such that levels were lower in the UFP-exposed as compared to filtered air-exposed 3xTgAD mice. When microglia morphology was assessed by Sholl analysis, microglia from both NTg mouse groups were ramified. The 3xTgAD air-exposed mice had the most ameboid microglia, while the 3xTgAD UFPexposed mice had microglia that were comparatively more ramified. The 3xTgAD air-exposed mice had more plaques per region of interest as measured by Congo red staining as well as more plaque-associated microglia than the 3xTgAD UFP-exposed mice. The number of non-plaque-associated microglia was not affected by genotype or exposure. Levels of soluble and insoluble human amyloid-beta 42 protein were measured in both 3xTgAD groups and no exposure effect was found. In contrast, UFP-exposure led to significant elevations in phosphorylated tau in 3xTgAD mice as compared to those that were exposed to air, as measured by pT205 staining. Conclusions: Exposure to environmentally relevant levels of ultrafine particulates led to changes in tau phosphorylation and microglial morphology in the absence of overt lung inflammation. Such changes highlight the need to develop greater mechanistic understanding of the link between air pollution exposure and Alzheimer's disease.

Barium sulfate (BaSO4) was considered to be poorly-soluble and of low toxicity, but BaSO4 NM-220 showed a surprisingly short retention after intratracheal instillation in rat lungs, and incorporation of Ba within the bones. Here we show that static abiotic dissolution cannot rationalize this result, whereas two dynamic abiotic dissolution systems (one flow-through and one flow-by) indicated 50% dissolution after 5 to 6 days at non-saturating conditions regardless of flow orientation, which is close to the in vivo half-time of 9.6 days. Non-equilibrium conditions were thus essential to simulate in vivo biodissolution. Instead of shrinking from 32 nm to 23 nm (to match the mass loss to ions), TEM scans of particles retrieved from flow-cells showed an increase to 40 nm. Such transformation suggested either material transport through interfacial contact or Ostwald ripening at super-saturating conditions and was also observed in vivo inside macrophages by high-resolution TEM following 12 months inhalation exposure. The abiotic flow cells thus adequately predicted the overall pulmonary biopersistence of the particles that was mediated by non-equilibrium dissolution and recrystallization. The present methodology for dissolution and transformation fills a high priority gap in nanomaterial hazard assessment and is proposed for the implementation of grouping and read-across by dissolution rates.

Infants born prematurely often require supplemental oxygen, which contributes to aberrant lung development and increased pulmonary morbidity following a respiratory viral infection. We have been using a mouse model to understand how early-life hyperoxia affects the adult lung response to influenza A virus (IAV) infection. Prior studies showed how neonatal hyperoxia (100% oxygen) increased sensitivity of adult mice to infection with IAV [IAV (A/Hong Kong/X31) H3N2] as defined by persistent inflammation, pulmonary fibrosis, and mortality. Since neonatal hyperoxia alters lung structure, we used a novel fluorescence-expressing reporter strain of H1N1 IAV [A/Puerto Rico/8/34 mCherry (PR8-mCherry)] to evaluate whether it also altered early infection of the respiratory epithelium. Like Hong Kong/X31, neonatal hyperoxia increased morbidity and mortality of adult mice infected with PR8-mCherry. Whole lung imaging and histology suggested a modest increase in mCherry expression in adult mice exposed to neonatal hyperoxia compared with room air-exposed animals. However, this did not reflect an increase in airway or alveolar epithelial infection when mCherry-positive cells were identified and quantified by flow cytometry. Instead, a modest increase in the number of CD45-positive macrophages expressing mCherry was detected. While neonatal hyperoxia does not alter early epithelial infection with IAV, it may increase the activity of macrophages toward infected cells, thereby enhancing early epithelial injury.

Previous work has demonstrated size, surface charge and skin barrier dependent penetration of nanoparticles into the viable layers of mouse skin. The goal of this work was to characterize the tissue distribution and mechanism of transport of nanoparticles beyond skin, with and without Ultraviolet Radiation (UVR) induced skin barrier disruption. Atomic absorption spectroscopy (AAS), flow cytometry and confocal microscopy were used to examine the effect of UVR dose (180 and 360 mJ/cm2 UVB) on the skin penetration and systemic distribution of quantum dot (QD) nanoparticles topically applied at different time-points post UVR using a hairless C57BL/6 mouse model.

An aberrant oxygen environment at birth increases the severity of respiratory viral infections later in life through poorly understood mechanisms. Here, we show that alveolar epithelial cell (AEC) 2 cells (AEC2s), progenitors for AEC1 cells, are depleted in adultmice exposed to neonatal hypoxia or hyperoxia. Airway cells expressing surfactant protein (SP)-C and ATP binding cassette subfamily A member 3, alveolar pod cells expressing keratin (KRT) 5, and pulmonary fibrosis were observed when these mice were infected with a sublethal dose of HKx31, H3N2 influenza A virus. This was not seen in infected siblings birthed into room air. Genetic lineage tracing studies in mice exposed to neonatal hypoxia or hyperoxia revealed pre-existing secretoglobin 1a11 cells produced airway cells expressing SP-C and ATP binding cassette subfamily A member 3. Pre-existing Kr51 progenitor cells produced squamous alveolar cells expressing receptor for advanced glycation endproducts, aquaporin 5, and T1a in alveoli devoid of AEC2s. They were not the source of KRT51 alveolar pod cells. These oxygendependent changes in epithelial cell regeneration and fibrosis could be recapitulated by conditionally depletingAEC2s in mice using diphtheriaA toxin and then infecting with influenza A virus. Likewise, airway cells expressing SP-C and alveolar cells expressing KRT5 were observed in human idiopathic pulmonary fibrosis. These findings suggest that alternative progenitor lineages are mobilized to regenerate the alveolar epithelium when AEC2s are severely injured or depleted by previous insults, such as an adverse oxygen environment at birth. Because these lineages regenerate AECs in spatially distinct compartments of a lung undergoing fibrosis, they may not be sufficient to prevent disease.

Alveolar epithelial type II cells (AEC2) maintain pulmonary homeostasis by producing surfactant, expressing innate immune molecules, and functioning as adult progenitor cells for themselves and alveolar epithelial type I cells (AEC1). How the proper number of alveolar epithelial cells is determined in the adult lung is not well understood. Here, BrdU labeling, genetic lineage tracing, and targeted expression of the anti-oxidant extracellular superoxide dismutase in AEC2s are used to show how the oxygen environment at birth influences postnatal expansion of AEC2s and AEC1s in mice. Birth into low (12%) or high (≥60%) oxygen stimulated expansion of AEC2s through self-renewal and differentiation of the airway Scgb1a1 + lineage. This non-linear or hormesis response to oxygen was specific for the alveolar epithelium because low oxygen stimulated and high oxygen inhibited angiogenesis as defined by changes in V-cadherin and PECAM (CD31). Although genetic lineage tracing studies confirmed adult AEC2s are stem cells for AEC1s, we found no evidence that postnatal growth of AEC1s were derived from self-renewing Sftpc + or the Scbg1a1 + lineage of AEC2s. Taken together, our results show how a non-linear response to oxygen at birth promotes expansion of AEC2s through two distinct lineages. Since neither lineage contributes to the postnatal expansion of AEC1s, the ability of AEC2s to function as stem cells for AEC1s appears to be restricted to the adult lung. Stem Cells 2016;34:1396-1406.

Animal dung is a biomass fuel burned by vulnerable populations who cannot afford cleaner sources of energy, such as wood and gas, for cooking and heating their homes. Exposure to biomass smoke is the leading environmental risk for mortality, with over 4,000,000 deaths each year worldwide attributed to indoor air pollution from biomass smoke. Biomass smoke inhalation is epidemiologically associated with pulmonary diseases, including chronic obstructive pulmonary disease (COPD), lung cancer, and respiratory infections, especially in low and middle-income countries. Yet, few studies have examined the mechanisms of dung biomass smoke-induced inflammatory responses in human lung cells. Here, we tested the hypothesis that dung biomass smoke causes inflammatory responses in human lung cells through signaling pathways involved in acute and chronic lung inflammation. Primary human small airway epithelial cells (SAECs) were exposed to dung smoke at the air-liquid interface using a newly developed, automated, and reproducible dung biomass smoke generation system. The examination of inflammatory signaling showed that dung biomass smoke increased the production of several proinflammatory cytokines and enzymes in SAECs through activation of the activator protein (AP)-1 and arylhydrocarbon receptor (AhR) but not nuclear factor-κB (NF-κB) pathways. We propose that the inflammatory responses of lung cells exposed to dung biomass smoke contribute to the development of respiratory diseases.

The increased production of nanomaterials has caused a corresponding increase in concern about human exposures in consumer and occupational settings. Studies in rodents have evaluated dose–response relationships following respiratory tract (RT) delivery of nanoparticles (NPs) in order to identify potential hazards. However, these studies often use bolus methods that deliver NPs at high dose rates that do not reflect real world exposures and do not measure the actual deposited dose of NPs. We hypothesize that the delivered dose rate is a key determinant of the inflammatory response in the RT when the deposited dose is constant.

Significance The causes of the sporadic form of Alzheimer’s disease (AD) are unknown. In this study we show that copper (Cu) critically regulates low-density lipoprotein receptor-related protein 1–mediated Aβ clearance across the blood–brain barrier (BBB) in normal mice. Faulty Aβ clearance across the BBB due to increased Cu levels in the aging brain vessels may lead to accumulation of neurotoxic Aβ in brains. In a mouse model of AD low levels of Cu also influences Aβ production and neuroinflammation. Our study suggests that Cu may also increase the severity of AD.

Iron is critical in multiple aspects of CNS development, but its role in neurodevelopment-the ability of iron deficiency to alter normal development-is difficult to dissociate from the effects of anemia. We developed a novel dietary restriction model in the rat that allows us to study the effects of iron deficiency in the absence of severe anemia. Using a combination of auditory brainstem response analyses (ABR) and electron microscopy, we identified an unexpected impact of nonanemic iron deficiency on axonal diameter and neurofilament regulation in the auditory nerve. These changes are associated with altered ABR latency during development. In contrast to models of severe iron deficiency with anemia, we did not find consistent or prolonged defects in myelination. Our data demonstrate that iron deficiency in the absence of anemia disrupts normal development of the auditory nerve and results in altered conduction velocity.

Ultraviolet radiation (UVR) skin exposure is a common exogenous insult that can alter skin barrier and immune functions. With the growing presence of nanoparticles (NPs) in consumer goods and technological applications the potential for NPs to contact UVR-exposed skin is increasing. Therefore it is important to understand the effect of UVR on NP skin penetration and the potential for systemic translocation. Previous studies qualitatively showed that UVR skin exposure can increase the penetration of NPs below the stratum corneum. In this work, an in vivo mouse model was used to quantitatively examine the skin penetration of carboxylated (CdSe/ZnS, core/shell) quantum dots (QDs) through intact and UVR barrier-disrupted murine skin by organ Cd mass analysis. Transepidermal water loss was used to measure the magnitude of the skin barrier defect as a function of UVR dose and time post-UVR exposure. QDs were applied to mice 3-4 days post-UVR exposure at the peak of the skin barrier disruption. Our results reveal unexpected trends that suggest these negative-charged QDs can penetrate barrier intact skin and that penetration and systemic transport depends on the QD application time post-UVR exposure. The effect of UVR on skin-resident dendritic cells and their role in the systemic translocation of these QDs are described. Our results suggest that NP skin penetration and translocation may depend on the specific barrier insult and the inflammatory status of the skin.

There is an urgent need for in vitro screening assays to evaluate nanoparticle (NP) toxicity. However, the relevance of in vitro assays is still disputable. We administered doses of TiO2 NPs of different sizes to alveolar epithelial cells in vitro and the same NPs by intratracheal instillation in rats in vivo to examine the correlation between in vitro and in vivo responses. The correlations were based on toxicity rankings of NPs after adopting NP surface area as dose metric, and response per unit surface area as response metric. Sizes of the anatase TiO2 NPs ranged from 3 to 100 nm. A cell-free assay for measuring reactive oxygen species (ROS) was used, and lactate dehydrogenase (LDH) release, and protein oxidation induction were the in vitro cellular assays using a rat lung Type I epithelial cell line (R3/1) following 24 h incubation. The in vivo endpoint was number of PMNs in bronchoalveolar lavage fluid (BALF) after exposure of rats to the NPs via intratracheal instillation. Slope analyses of the dose response curves shows that the in vivo and in vitro responses were well correlated. We conclude that using the approach of steepest slope analysis offers a superior method to correlate in vitro with in vivo results of NP toxicity and for ranking their toxic potency.