Sage Journals: Discover world-class research

Abstract

Epidemiological studies report associations between air pollution (AP) exposures and several neurodevelopmental disorders including autism, attention deficit disorder, and cognitive delays. Our studies in mice of postnatal (human third trimester brain equivalent) exposures to concentrated ambient ultrafine particles (CAPs) provide biological plausibility for these associations, producing numerous neuropathological and behavioral features of these disorders, including male-biased vulnerability. These findings raise questions about the specific components of AP that underlie its neurotoxicity, which our studies suggest could involve trace elements as candidate neurotoxicants. X-ray fluorescence analyses of CAP chamber filters confirm contamination of AP exposures by multiple elements, including iron (Fe) and sulfur (S). Correspondingly, laser ablation inductively coupled plasma mass spectrometry of brains of male mice indicates marked postexposure elevations of Fe and S and other elements. Elevations of brain Fe and S in particular are consistent with potential ferroptotic, oxidative stress, and altered antioxidant capacity-based mechanisms of CAPs-induced neurotoxicity, supported by observations of increased serum oxidized glutathione and increased neuronal cell death in nucleus accumbens with no corresponding significant increase in caspase-3, in male brains following postnatal CAP exposures. Understanding the role of trace element contaminants of particulate matter AP as a source of neurotoxicity is critical for public health protection.

Keywords

ultrafine particles iron sulfur glutathione ventriculomegaly corpus callosum nucleus accumbens

Introduction

Numerous epidemiological studies report associations of various metrics of air pollution (AP) exposures with characteristics of neurodevelopmental disorders and neurodegenerative diseases. These have included reports of increased risk for autism spectrum disorder (ASD), attention-deficit/hyperactivity disorder (ADHD), slower cognitive development in children, schizophrenia, and a more rapid cognitive decline with age in adults, as summarized in a series of recent reviews.^1

-10 Several studies have also reported neuropathological changes in response to AP exposures, which to date, include reductions in brain white matter (myelin) and corresponding brain disconnectivity, reductions in gray matter volume, and altered basal ganglia structure.^11

-16 Air pollution as a risk factor for neurodevelopmental disorders and neurodegenerative diseases would carry heavy social and economic burdens. Annual US costs just for ASD are US$11 to US$60 billion according to the Centers for Disease Control, while estimated costs for Alzheimer’s are US$290 billion (Alzheimer’s Association).

Biological plausibility for the neurotoxic effects of AP has also begun to emerge from studies using animal models.^6,17
-19 Based on the fact that for different particle sizes of the same chemistry, ultrafine particulate (UFP) matter is considered the most reactive component of AP per unit mass because of its greater surface area/mass ratio available for adsorption of organic and inorganic contaminants,²⁰ we have examined the impact of developmental exposures to concentrated ambient ultrafine particles (CAPs) on brain and behavior in mice since 2013 as reviewed here. Exposures were carried out from postnatal days (PND) 4 to 7 and 10 to 13 for 4 h/d. This period is considered analogous to human third trimester brain development^21,22 and is characterized by significant neuro- and gliogenesis.²³ Mass CAPs concentrations across these studies to date have ranged from 22 to 121 µg/m³; levels at the lower-mid part of this range are similar to PM_2.5 levels in global cities.^24
-26

These postnatal (PN) CAPs exposures in mice resulted in marked neuropathological effects, as well as behavioral dysfunctions, with these effects occurring predominantly in males, as previously reported.^27

-33 One dramatic neuropathological effect was a male-specific persistent ventriculomegaly (ie, enlarged lateral ventricles) occurring at 96 µg/m³ and observed even at PND 270, that is, the last time point examined. Subsequent exposure cohorts revealed that postnatal CAPs produced male-specific ventriculomegaly even at 44 μg/m³, the lowest levels yet examined with sufficient sample sizes to date. In addition, marked reductions in size and myelination of the corpus callosum (CC), the largest white matter tract of the brain and a structure critical to interhemispheric connectivity, were found. Male biased microglial activation (inflammation) was found in CC (340% of control) as late as PND 270, the last time point examined to date. A profile of male-specific elevated brain glutamate, with consequent excitatory–inhibitory imbalance was also observed. Behavioral impairments, as assessed to date, reveal increased impulsivity/cognitive inflexibility and altered social interactions, including reduced social preference for a novel mouse versus a cagemate.^28,31,31,34

Collectively, these findings following postnatal CAPs exposures suggest that AP can reproduce numerous characteristics of a number of neurodevelopmental disorders, including the enhanced vulnerability of males to such conditions. While ASD has unique features, it also shares features with several other neurodevelopmental disorders.^35

-40 Autism spectrum disorder and early childhood-onset schizophrenia^41

-44 share abnormalities of glutamatergic systems,^45
-47 inflammatory mechanisms,⁴⁸ and neuropathological features, including ventriculomegaly and disconnectivity.^49
-51 Attention-deficit/hyperactivity disorder occurs at higher rates in children with ASD.⁵² Both involve increases in impulsive behavior and are considered disorders of hemispheric disconnectivity.³⁶ Like ASD, ADHD is male-biased and its rates have increased significantly from 7.8% in 2003 to 9.5% in 2007 and 11.0% in 2011 to 2012 according to the Centers for Disease Control (CDC). Attention-deficit/hyperactivity disorder can also include ventriculomegaly⁵³ and alterations in glutamatergic function,⁵⁴ brain white matter,⁵⁵ brain connectivity,⁵⁶ and inflammation.⁵⁷ Thus, the neurotoxic consequences of AP seen in these experimental models may contribute risk not only for ASD but for other neurodevelopmental disorders as well. Differences in timing of AP exposure during pregnancy could contribute to the heterogeneity and subgroups of ASD⁵⁸ or other neurodevelopmental disorders.

A collaborative study carried out at NYU (Tuxedo, New York)^59
-61 obtained brains of mice following gestational (GE, days 0.5-16.5) fine/UFP CAPs exposures, that is, a 1st + 2nd trimester human brain equivalent time point. The GE exposures produced ventriculomegaly in both sexes (see^31,59,60) at PND 11 to 15. In contrast to postnatal CAPs, GE exposures increased rather than decreased CC size and myelination, effects that were still present at PND 57 to 61, the last time point available for examination. Increased CC size and myelination are actually seen early in the ASD brain,⁶² followed only later by reductions in size and myelination, as seen with our CAPs exposures.^63,64 The findings produced by GE CAPs exposures to date confirm that AP-induced neurotoxicity is not unique to exposures carried out in Rochester, and despite the heterogeneity of AP compositions, similarities exist in their potential to disrupt white matter development in the central nervous system (CNS). In addition, they reconfirm sex differences in outcome and also underscore the importance of timing of AP exposures during brain development.

The observations of neuropathological effects from particulate AP exposures at 2 different sites also suggested that similar AP components could be involved in this neurotoxicity. Consequently, we carried out studies to determine whether the carbon-based nanoparticle itself, that is, the base of UFPs, or perhaps a known inflammatory component of AP, for example, endotoxin,⁶⁵ could produce such effects by generating atmospheres of each and exposing mice to them alone or in combination. However, neither exposure alone nor the 2 combined were able to reproduce the developmental neurotoxicity of postnatal exposure to ambient CAPs.⁶⁵ Trace element contaminants were subsequently considered as another potential source of postnatal CAPs neurotoxicity. Subsequent analyses, elaborated here, reveal that trace element contaminants of AP may be a likely source of its neurotoxic consequences, based on the presence of known elemental neurotoxicants in the exposures and in excess in brain following such exposures. The increases in these elements, which can be taken up into brain from bloodstream but also via the olfactory and trigeminal nerves,^66
-68 moreover, suggest specific mechanisms of CAP-induced neurotoxicity via ferroptosis, as schematized in Figure 1, and their actions in brain would be likely to influence pathways consistent with reported biomarkers of neurodevelopmental disorders.^69

-75 In addition, data across different exposures were utilized to determine preliminary concentration–effect functions for various outcomes where available.

Figure 1.

Schematic diagram showing entry routes of ultrafine particles to brain via olfactory and trigeminal nerves, vagal nerve, and bloodstream. The consequent elevation of brain metals as contaminants of ultrafine particles, for example, Fe, raise the potential for ferroptotic mechanisms of brain neuropathology associated with air pollution. Both transferrin-bound and nontransferrin (labile) Fe can be released into cells with Fe3⁺ (ferric iron) reduced to Fe2⁺ (ferrous iron) followed by movement into cytosol via the divalent metal transporter 1. Potential increases in the redox active Fe2⁺ can lead to reactive oxygen species via the Fenton reaction and lipid peroxidation. Ferroptosis is negatively regulated by glutathione and glutathione peroxidase 4, derived from the trans-sulfuration pathway, which if reduced can facilitate ferroptosis, an iron-and lipid peroxidation-dependent form of cell death. DMT1 indicates divalent metal transporter 1; GPx4, glutathione peroxidase 4; ROS, reactive oxygen species; TRF1, transferrin.

Methods and Materials

Animals and Postnatal CAP Exposures

Male and female C57BL6/J mice 8 weeks of age from Jackson Laboratories (Bar Harbor, Maine) were acclimated to a housing room for 1 week and then bred monogamously for 3 days. Pregnant dams were singly housed with litters until weaning on PND 21. To preclude litter-specific effects, only single pups/sex/litter were used in all of these studies. All mice used in this study were treated humanely and with regard for alleviation of suffering and approved by the University of Rochester Institutional Animal Care and Use Committee.

Pups were placed singly in compartmentalized whole-body exposure chambers. They were exposed to filtered air (Air) or CAPs using the Harvard University Concentrated Ambient Particle System (HUCAPS) fitted with a size-selective inlet and a high-volume UFP (≤100 nm) concentrator (10-20x) that takes in outdoor air at 5000 L/min and concentrates ambient UFP, as previously described.^27

-30 Exposures lasted for 4 hours per day from 0700 to 1100 for 4 days per week from PND 4 to 7 and PND 10 to 13, with exposure timing corresponding to peak vehicular traffic outside the intake valve of the HUCAPS instrumentation. A condensation particle counter (TSI 3022A) provided particle counts. Mass concentration was calculated using idealized particle density (1.5 g/cm³). A scanning mobility particle sizer was used to determine particle size distribution and median particle diameter + geometric standard deviation. Flow of CAPs-enriched and filtered air was maintained at 35% to 40% relative humidity and 77°F to 79°F. Average exposure mass concentrations from these exposures were 22, 44, 53, 96, and 121 μg/m³.Teflon filter samples from the breathing zone inside the exposure chamber were taken for chemical analysis of the constituents. Teflon filter samples were analyzed by the Desert Research Institute (Reno, Nevada) in a temperature-controlled package at below 4° C for elemental analyses by an energy dispersive X-ray fluorescence analyzer (Epsilon 5; PANalytical Company, Almelo, the Netherlands) as previously described.⁷⁶

Brain Pathology

To preclude any effect of anesthetics on measured end points, brains were extracted following decapitation without sedation and placed into 4% paraformaldehyde for 24 hours and then cryoprotected in 30% sucrose until they sank. Brains were stored at 4°C until they were sectioned and then stored in sucrose- and ethylene glycol containing cryoprotectant and kept at −20°C.

Corpus callosum area determination

The size of the CC (approximate adult-equivalent Bregma range 0.74-0.2 mm) was determined by tracing the area of interest in at least 3 adjacent coronal sections of slide-mounted brain using Neurolucida (MBF, Williston, Vermont). Software enumerated the area of interest in square micrometers.

Myelin basic protein immunostaining and image analysis

To determine the extent of myelination, every fourth section (PND 14) or every sixth section (PND 270) was stained for myelin basic protein (MBP). Briefly, brain sections were washed of cryoprotectant and placed into primary antibody for MBP (EMD Millipore; MAB386, Burlington, MA). Tissue was then placed into biotinylated secondary antibody (BA-9401; 1:200 dilution; Vector Labs, Burlingame, California) for 1 hour and the stain was visualized using 3-3′-diaminobenzidine (DAB). Expression of MBP was analyzed using Image Pro Plus 7.0 (Media Cybernetics, Rockville, MD) using segmentation and thresholding to estimate a percentage area of the region of interest that was positive for MBP expression.

Laser Ablation Inductively Coupled Plasma Mass Spectrometry

Brain thin sections of 40 µm were prepared by cryosectioning and mounted onto a microscope slide for elemental imaging by laser ablation inductively coupled plasma mass spectrometry (LA-ICP-MS). Slides were not subbed (coated with chromium potassium sulfate to prevent metal contamination). The LA-ICP-MS system used was an ESI Lasers NWR 213 laser ablation unit coupled to an Agilent 8900 ICP-MS operated in O₂ reaction mode to enhance detection of S by mass shifting to SO at 48 m/z. The laser “spot” size was 50 µm² and the laser scan speed was 250 µm/s and the laser shot frequency was 20 Hz. The sample was analyzed in a series of line scans with a spacing of 50 µm. The ICP-MS collected data for Al, S, Ca, Cr, Mn, Fe, Ni, Cu, Zn, Se, Cd, and Pb with a total acquisition time of 0.2 seconds. The raw ICP-MS data were concatenated into an Excel sheet, and relative X and Y coordinates for each data point were added. The resulting file was then imported into SMAK elemental imaging software to generate 2D elemental images of each section.⁷⁷ The images show the relative concentration differences in each element across the thin section as no quantitative calibration was performed during this analysis.

Atomic Absorption Spectrophotometry

Atomic absorption spectrophotometry was used to measure levels of metals in olfactory bulb. After wet weights were recorded, tissue were digested with a 1:1 mixture of nitric and perchloric acids, samples brought to a constant volume, and determination of metal levels performed by graphite AAS (Hitachi 170-70, Tokyo, Japan). For each analysis, controls (blanks) and standards were carried out in 0.7% nitric acid and calibration curves determined at the beginning and end of a sample run.

Serum Oxidized Glutathione

Blood was collected during sacrifice into prechilled tubes and spun at 1500g for 20 minutes and serum was removed. Serum oxidized glutathione (GSH) was measured using a commercially available colorimetric assay kit (Arbor Assays, Ann Arbor, Michigan) according to the manufacturer’s instructions. Quality controls fell within appropriate ranges with intra-assay sample variability below 15% variance.

Immunohistochemistry

Caspase-3

Briefly, brain sections incubated in primary antibody solution (1:2000; 9662; Cell Signaling Technology, Danvers, Massachusetts) at 4°C for 24 hours. Sections were then washed and incubated with biotinylated secondary antibody (1:200, anti-rabbit, BA1000; Vector Labs) for 1 hour; the stain was visualized using DAB and quantified using ImageJ with established thresholding methods.⁷⁸

Silver staining

The method was carried out on brain sections using the FD NeuroSilver Kit II (FD NeuroTechnologies, Inc, Columbia, Maryland), according to the manufacturer’s instructions.

Statistical Analyses

Statistical analysis was carried out using JMP11 (Cary, North Carolina) with initial between-group analyses of variance that included all experimental factors (sex and treatment) in full factorial designs. Main effects and interactions were then pursued in post hoc t tests as appropriate. P values ≤.05 were considered statistically significant.

Results

Concentration Effect Functions for Postnatal UFPs

Postnatal CAPs exposure concentrations across our studies to date have ranged from mass concentrations of 22 to 121 μg/m³. In order to ascertain the impact of concentration, we plotted magnitude of effect against exposure concentrations for outcomes in males that were measured at 2 or more of the exposure concentrations. The results are depicted in Figure 2. As shown in the left panel, ventriculomegaly was not observed in preliminary assessments following a 22 μg/m³ exposure, but trending increases in size of the lateral ventricle were observed at 44 μg/m³,F_(1,17) = 3.98, P = .062, in males and significantly further increased at 96 μg/m³ as previously reported and shown in Figure 3.^28,31 No changes in myelination were evident at 22 or 44 μg/m³, while, as previously reported, marked reductions occurred as concentrations increased to 96 μg/m³.^28,31

Figure 2.

Concentration–effect functions for neuropathological (left), neurochemical (middle), and behavioral (right) consequences for concentrated ambient ultrafine particle (CAP) exposure. Data are from males only and presented as percentage of the corresponding filtered air control data and the CAP concentration (x-axis). Unpublished and previously reported data.^{27

–32,82}

Figure 3.

Coronal unstained brain sections from a male brain exposed to filtered air (left) or to 96 µg/m³ concentrated ambient ultrafine particles (CAPs; right) during the postnatal period representing the extreme of ventriculomegaly observed. The enlarged lateral ventricle (LV) below the corpus callosum (CC) of the CAPs-exposed brain on the right panel is indicated by an arrow.²⁸

The middle panel depicts alterations in dopamine and glutamate levels and function. Even at the lowest concentrations generated in our studies to date (22 μg/m³), increases in levels of glutamate have been observed, with slight increases in the magnitude of elevation at the highest concentration generated to date (121 μg/m³).^28
-30,33 Findings also suggest increases in levels of dopamine turnover in brain, again increasing with increasing CAPs exposure concentration.^29,30 With increases in dopamine turnover, levels of dopamine appear to decline with increasing CAPs exposure concentration.^29,30

The right panel depicts 2 behavioral baselines that have been measured in 2 different exposure cohorts, that is, behavior on a fixed interval (FI) schedule of food reward, as a measure of temporal learning, and novel object recognition, an index of short-term memory, as previously described.^30,17 Slight increases in rates of response on the FI schedule (delayed learning) at 44 μg/m³ were observed, while at 96 μg/m³, marked reductions in response rates (delayed learning and delayed reward) were found that persisted across sessions. Novel object recognition was already significantly impaired at 44 μg/m³, with even further impairments in performance observed at 96 μg/m³.

Assessment of Trace Elements in CAPs Exposures

X-ray fluorescence analyses were utilized to examine the elemental exposures associated with various CAPs exposure studies. As shown in the tree maps for one such exposure from Rochester, New York and another from Tuxedo, New York in Figure 4, all such analyses revealed multiple trace elements were contaminants of CAPs, regardless of the site of the exposure. In addition, as expected, the profile of elevation of trace elements in CAPs differed somewhat by exposure site, with S being the highest in Tuxedo with a relatively high Fe levels as well; S was also highest in Rochester, but Fe was not as high as in Tuxedo.

Figure 4.

Tree maps of elements from ambient ultrafine particulate exposures via X-ray fluorescence analysis of filters during postnatal (left; Rochester, New York)^28
–30 and gestational exposures (right; Tuxedo, New York). ^59,61,60 Both sulfur (S) and iron (Fe) are circled.

Elemental contaminants of the AP exposures are further characterized in Figure 5 where the top row depicts fold changes in various elements relative to the filtered air chamber exposure levels for 3 different exposures. As it shows, levels of sulfur (S) showed dramatic increases in virtually all these exposures. In addition, increases in iron (Fe) levels were also consistently evident. Potassium levels likewise showed increases. Increases in numerous other elements were also observed, but fold-changes were less systematic across such exposures.

Figure 5.

Top row shows fold changes in various elements relative to levels in filtered air control chambers for gestational (left column) and for 2 postnatal exposures (middle and right columns) following X-ray fluorescence analyses. Corresponding data in nanograms per cubic meters are depicted along the bottom row.

Total concentrations in nanograms per cubic meters are depicted in the bottom row of Figure 5 relative to filtered air controls for these same exposure cohorts. As these plots show, S levels ranged from 2000 to 4000 ng/m³ and Fe levels from approximately 500 to 1000 ng/m³. Other metals of note in these exposures included zinc (Zn: range from 2.7 to 297 ng/m³), copper (Cu: range from 0.68 to 69 ng/m³), aluminum (Al: range from 1.65 to 164 ng/m³), potassium (K: range from 0.88 to 313 ng/m³), and silica (Si: range from 0.65 to 64 ng/m³).

Assessment of Trace Elements in Brain

Subsequently, LA-ICP-MS was carried out on 40-µm sagittal sections after postnatal CAPs exposure from a randomly chosen filtered air control male and a CAPs-exposed male brain. Figure 6 contrasts the levels of Fe and S in these brains at PND 14, that is, 24 hours after completion of exposures. As it shows, postnatal CAPs markedly elevated the levels of both elements in brain relative to levels in filtered air control brain. These elevations, moreover, were seen diffusely across the brain rather than regionally, as would be consistent with the broad functions of Fe and S in brain. As would be suggested by the analyses of filters from exposure chambers, other metals were likewise elevated in CAPs brain post exposure (Figure 7), in particular levels of Cu and to some degree calcium (Ca). Notably, increases in Al were also seen that were somewhat diffuse across regions but showed a selectively high accumulation in hippocampus. In contrast, levels of 2 essential metals in brain appeared to be of a lower concentration in brain, specifically manganese (Mn) and Zn. Collectively, these findings suggest brain metal dyshomeostasis.

Figure 6.

Brain iron (Fe) and sulfur (S) levels are increased in postnatal concentrated ambient ultrafine particles-exposed males after analysis by inductively coupled plasma mass spectrometry in one randomly chosen filtered air control male and 1 male exposed to concentrated ambient ultrafine particles. These images show a relatively diffuse pattern of increases across brain regions. Brains from females not yet examined. Exposure concentrations averaged 45 µg/m³.

Figure 7.

Brain levels of aluminum (Al), calcium (Ca), copper (Cu), manganese (Mn), and zinc (Zn) from a randomly chosen male (air) and concentrated ambient ultrafine particles–exposed male following postnatal exposures. Exposure concentrations averaged 45 µg/m³.

Given the demonstrated olfactory and trigeminal nerve uptake of elemental contaminants of UFPs, and since the LA-ICP-MS analysis is not quantitative, olfactory bulb tissue from PND 14 CAPs exposed and filtered air-exposed brains of males and females were analyzed by atomic absorption spectrophotometry for levels of Al, Fe, and Zn. As can be seen from Figure 8, in support of the ICP-MS images, significantly increased levels of both Fe, 65%, F_(1,11) = 47.58, P < .0001, and Zn, 43%, F_(1,11) = 20.09, P = .0008, were found in males, with a similar trend for Al increases, F_(1,11) = 3.495, P = .086, relative to filtered air controls after exposures to 44 µg/m³. Females showed a significant increase only in levels of Al by 120%, F_(1,11) = 6.56, P = .0265, but not in Fe or Zn.

Figure 8.

Mean ± standard error levels (µg/g) of aluminum (Al), iron (Fe), and zinc (Zn) in olfactory bulb following postnatal concentrated ambient ultrafine particles exposure concentrations of 45 µg/m³ in females (left) and males (right). * indicates statistical significance; ∼ indicates marginal significance.

Potential Ferroptotic Mechanisms

Increases in Fe and S in brain suggested the potential for ferroptotic mechanisms could underlie the neuropathological changes observed.^79

-82 Ferroptosis is a nonapoptotic fatal combination of Fe toxicity, antioxidant depletion due to disruption of glutathione peroxidase-4 (GPx4), that is, trans-sulfuration, and lipid peroxidation-induced membrane damage.⁸³ Ferroptosis is negatively regulated by GSH, a product of the trans-sulfuration pathway, making S levels critical. An inverse relationship between Fe overload and GSH, hydrogen peroxide, and reactive oxygen species (ROS) are well-documented.^84
-86 Thus, high brain Fe and low GSH or GPx4 would facilitate ferroptosis.

Oxidized GSH

To determine the potential involvement of ferroptotic mechanisms in these effects, levels of serum-oxidized GSH were measured and revealed male-specific increases as shown in Figure 9, F_(1,8) = 10.9, P = .0108. These effects were seen following postnatal CAPs exposures where the exposure concentrations averaged only 22 µg/m³.

Figure 9.

Mean ± standard error levels of oxidized glutathione in serum from males (black) and females (gray) exposed postnatally to filtered air or to 45 µg/m³ concentrated ambient ultrafine particles (CAPs). Statistically significant increases in serum oxidized glutathione were seen in males after CAPs, but not in females. * indicates statistical significance; ∼ indicates marginal significance.

Nucleus Accumbens Neuronal Cell Death

In addition, measures of neuronal death were examined at PND 14 in nucleus accumbens (NAc), a key region of the mesocorticolimbic circuit of brain critical to executive functions, after exposure concentrations averaging 44 µg/m³. For this purpose, tissue was silver stained as well as examined by caspase immunohistochemistry. Males exhibited significant increases in numbers of dead cells and reductions in numbers of live cells and total numbers of cells (Figure 10), males: dead cells, F_(1,8) = 8.92, P = .0174; live cells, F_(1,8) = 7.46, P = .0258; total cells, F_(1,8) = 6.97, P = .0297, as measured via silver staining. While a similar trend toward an increase in dead cells was seen in females at PND 14, effects were not statistically significant. Figure 11 shows a photomicrograph depicting the increased silver staining in CAPs-exposed male NAc; importantly, these increases were not seen with caspase staining. When measured at PND 25 (Figure 12), effects were no longer evident in males, whereas significant increases in live cells (F_(1,6) = 8.99, P = .024) and total cells (F_(1,6) = 8.95, P = .043) were seen in females at this time point.

Figure 10.

Mean ± standard error levels of dead cells (left column), live cells (middle column), and total cells (right column) in nucleus accumbens at PND 14 as evaluated by silver staining of males (top row) and females (bottom row) exposed postnatally to filtered air (air) or to 45 µg/m³ concentrated ambient ultrafine particles. Males but not females exhibited increases in dead cells and reductions in both live cells and total cells.

Figure 11.

Male-specific degeneration of neurons in nucleus accumbens (arrow) as measured by silver staining as described in the Methods at postnatal day 14 following postnatal exposure to 45 µg/m³ concentrated ambient ultrafine particles. No comparable effects were seen in males exposed to filtered air, nor did caspase staining show increases.

Figure 12.

Mean ± standard error levels of dead cells (left column), live cells (middle column), and total cells (right column) in nucleus accumbens at postnatal day 25 as evaluated by silver staining of males (top row) and females (bottom row) exposed postnatally to filtered air (air) or to 45 µg/m³ concentrated ambient ultrafine particles. At this time point, changes were no longer evident in males, whereas females showed increases in both live cells and total cells.

Discussion

Collectively, our studies of postnatal and GE CAPs exposures provide biological plausibility supporting an impact of AP on brain development and behavior.^{17,27

-31,59,61,60} Importantly, these studies demonstrate that the impacts of UFP matter exposures are sex dependent and differ by the specific developmental window of time in which it occurs. Further, they suggest that the adverse impacts are not geographically unique to Rochester and that similar components of AP could be responsible for the observed neuropathological consequences.

Similarities of Effects of Postnatal CAPs Exposures in Mice to Features and Hypothesized Mechanisms of Autism Spectrum Disorder and Other Neurodevelopmental Disorders in Humans

The similarity of the effects of CAPs exposures to characteristics and hypothesized mechanisms of ASD is striking as depicted for postnatal CAPs exposures in Table 1. Autism spectrum disorder is a male-biased disorder diagnosed on the basis of impaired social behaviors and restricted/repetitive behaviors.^48,87,88 Neuropathology of ASD in humans is dynamic across time and can include persistent ventriculomegaly,^64,89
-91 initial increases followed by sustained loss of white matter ^63,64 and gray matter,^92

-96 loss of myelin connectivity across the hemispheres^97
-99 particularly in CC,⁶² persistent brain inflammation,^100,101 and excitatory (glutamate) inhibitory imbalance¹⁰² that correlates with IQ reductions and social and cognitive deficits. Notably, in mice, increases in white matter were seen following GE CAPs exposures, whereas postnatal exposures markedly reduced white matter.^63,64 Nucleus accumbens dysfunction in ASD has been related to social and repetitive behavior deficits and response inhibition^{95,96,103,104,96} and to altered self-control in ADHD.^105,106 While ASD etiology remains unknown, inflammation is considered key, and biomarkers include altered trans-sulfuration pathways and increased oxidative stress.^{69,107

-112}

Table 1.

Comparison of ASD and PN CAPs characteristics.

	ASD	PN CAPs
Male biased	√	√
Ventriculomegaly	√	√
Reductions in white matter	√	√
Reductions in corpus callosum size	√	√
Interhemispheric disconnectivity	√	√
Persistent inflammation	√	√
Excitatory/inhibitory imbalance	√	√
Oxidative stress	√	√
Neuronal cell death	√	√

Abbreviations: ASD, autism spectrum disorder; CAPs, concentrated ambient ultrafine particles; PN, postnatal.

It is important to recognize, however, that many of the phenotypic features and hypothesized mechanisms of ASD are also seen in other neurodevelopmental disorders.^35

-40 Autism spectrum disorder and early childhood-onset schizophrenia^41

-44 share abnormalities of glutamatergic systems,^45
-47 inflammatory mechanisms,⁴⁸ and neuropathological features, including ventriculomegaly and disconnectivity.^49
-51 Attention-deficit/hyperactivity disorder occurs at higher rates in children with ASD.⁵² Both involve increases in impulsive behavior and are considered disorders of hemispheric disconnectivity.³⁶ Like ASD, ADHD is male biased and its rates have increased significantly from 7.8% in 2003 to 9.5% in 2007 and 11.0% in 2011 to 2012 according to CDC. Attention-deficit/hyperactivity disorder can also include ventriculomegaly⁵³ and alterations in glutamatergic function,⁵⁴ brain white matter,⁵⁵ brain connectivity,⁵⁶ and inflammation.⁵⁷ Thus, the neurotoxic consequences of AP seen in our experimental models may contribute risk not only for ASD but for other neurodevelopmental disorders as well.

Olfactory Uptake of Elemental Contaminants of AP

Inductively coupled plasma mass spectrometry analyses of brains from mice after CAP exposures suggest that AP exposures could produce brain metal dyshomeostasis, with increases in several brain metals, for example, Fe and S, and suggestive depletion of others. The observed increases in olfactory bulb Fe levels in males are consistent with the assertion that some component of this uptake occurred via nasal olfactory uptake after postnatal CAPs exposures.^113,114 Similarly, increases in brain Fe were seen in cerebellum of both sexes following GE CAPs exposures.¹¹⁵ Nasal olfactory uptake of elemental AP particles has been demonstrated in both rats and concluded from studies in humans,¹¹⁶ including Fe, but also Mn, Cd, Ni, Hg, Al, Co, Zn, and Cu,^{113,116

-122} across olfactory epithelial barriers in the nasal passages followed by transport to olfactory bulb and movement to other brain regions.¹²³ In one study, intranasal instillation of either ferric or ferrous iron subsequently increased blood Fe levels in rats, with uptake relying on divalent metal transporter 1.¹¹³ Accumulation of ferric iron in olfactory bulb was found following inhalation exposures to iron soot (40 µg/m³ Fe oxide nanoparticles) in adult female mice.¹²⁴

Inhaled exposures to SO² show that virtually all of it is removed from air by the nose.¹²⁵ Early studies indicate that subsequently, its derivatives enter the blood and dissociate to form the derivatives bisulfite and sulfite,¹²⁶ both of which have been found in hippocampal slices from rat brain.¹²⁷

With olfactory nerve uptake, elements bypass the blood–brain barrier such that increased levels of trace elements in brain may not be reflected in peripheral markers. In addition, sensory nerves in the upper and lower respiratory tract can translocate particles¹²⁸ that reach for example, the trigeminal ganglion or the vagal nerve.^{66,68,129,130} As Fe regulatory proteins 1 and 2, and Fe transporters such as the transferrin receptor and divalent metal transporter-1, critical to tight brain regulation of Fe levels, are not fully expressed until PNDs 15 to 20 in rodents,^131,132 an extended window is open during which excess Fe or S can reach and directly influence brain development. Similarly, homeostatic regulation of Fe absorption appears in human infants at about 6 to 9 months of age.¹³³

Plausibility of Elemental Contaminants of AP for the Postnatal CAPs-Induced Phenotype

Notably, reported consequences of excess brain Fe or SO₂ are consistent with ASD and other neurodevelopmental disorders. Normally, Fe²⁺ oxidation to Fe³⁺ releases a free radical quenching electron, with nanoparticulate Fe³⁺ existing in a biomineralized solid or particle. However, excess Fe²⁺, that is, the labile pool (like other ionic species, eg, Ce³⁺), has greater bioactivity than Fe³⁺ and produces oxidative stress and inflammation, reductions in antioxidant capacity, increased cell death markers, and altered prefrontal cortex and hippocampal glutamate function.¹³⁴ Fe(III)Cl injections into rodent brain produce ventriculomegaly.^135,136 Developing brain is particularly susceptible to excess ionic Fe²⁺ given its immature antioxidant defense systems, low total Fe binding capacity, and low concentrations of Fe²⁺ binding proteins.¹³⁷ Indeed, increased non-protein-bound Fe in umbilical cord blood is an early predictive marker of neonatal brain damage.¹³⁸ The uptake, transport, and sequestration of extrinsic Fe _x O _y nanoparticles may be accompanied by breakdown and in vivo processing, making it ultimately critical to distinguish extrinsic Fe _x O _y from intrinsic ferritin nanoparticle formation.

In what might seem contradictory to our findings, some studies have reported reduced, rather than increased, serum Fe levels in ASD,^139
-141 although this was not confirmed by a recent meta-analysis.¹⁴² Even if such a finding was confirmed, it does not preclude a concurrent elevation of brain Fe. For example, neuroferritinopathy resulting from a mutation in the ferritin light chain gene results in elevated brain Fe and ferritin levels but reduced serum ferritin.¹⁴³ Additionally, intranasal instillation of Fe₂O₃ nanoparticles to rats over 7 days increased Fe brain levels, while significantly reducing serum Fe levels.¹⁴⁴ Such discrepancies are also seen in neurodegenerative diseases that include elevated brain Fe, for example, Parkinson disease,^145
-147 suggesting a broader metal dyshomeostasis under such conditions.

Further exacerbating the potential toxicity of elevated brain Fe, studies have demonstrated its extremely slow turnover in rodent brain (half-life of ∼9 months), with estimates on the order of decades when extrapolated to humans^148
-150; such slow turnover may relate to the potential role of Fe in multiple neurodegenerative diseases, including Alzheimer and Parkinson disease,^83,151 given that such exposures are cumulative across the life span.

Sulfur is critical for Fe-S cluster proteins, amino acid metabolism, and brain trans-sulfuration pathways critical to GSH production necessary to maintain cellular redox states and scavenge free radicals. Glutathione activity is very low in fetal brain, reaching adult levels only at PND 14 in rat, indicating incomplete regulation of this pathway during brain development.^137,152 Alterations in the trans-sulfuration pathway adversely influence brain and behavior.^153,154 Sulfite, taken up during SO₂ inhalation,¹⁵⁵ can auto-oxidize, as mediated by O₂ or Fe³⁺ (the most stable such complex), to produce sulfite radicals and ROS.^156
-158 Inhaled SO₂ increases levels of brain sulfite,¹⁵⁹ hippocampal neuronal death, protein oxidation, lipid peroxidation,¹⁶⁰ brain inflammation, and oxidative damage in brain; it also alters glutamate function and reduces GSH and GPx4 levels.¹⁶¹ These features correspond to the key role that chronic neuroinflammation is considered to play in ASD.¹⁶² Of direct relevance to our findings, recent studies report that 1 ppb increases in SO₂ in AP increase the odds ratios for ASD by 17%.¹⁶³

Elevated Brain Fe and S Suggest Potential Ferroptotic and Oxidative Stress-Based Mechanisms for Postnatal CAPs Effects

Elevated brain Fe and S also suggest ferroptosis and oxidative stress mechanisms of postnatal CAPs effects. Ferroptosis is a nonapoptotic fatal combination of Fe toxicity, antioxidant depletion due to disruption of GPx4, that is, trans-sulfuration, and lipid peroxidation-induced membrane damage.⁸³ It is negatively regulated by GSH, a product of the trans-sulfuration pathway, making S levels critical. An inverse relationship between Fe overload and GSH, hydrogen peroxide, and ROS are well-documented.^84
-86 Thus, high brain Fe, and low GSH or GPx4 would facilitate ferroptosis, as proposed in Figure 1. These mechanisms would be consistent with our observations of postnatal CAPs-induced increases in NAc neuronal cell death that were not revealed by caspase staining (Figure11), as well as the increases in levels of serum oxidized GSH (Figure 9). While ferroptosis is seen in multiple neurodegenerative diseases,⁸¹ its role in neurodevelopmental disorders has not been investigated, although it is seen in periventricular leukomalacia of prematurity, that is, brain white matter injury arising from free radical damage to oligodendrocytes.^82,164

Male Vulnerability to Postnatal CAPs Exposures and Heterogeneity of Neurodevelopmental Disorders

Although numerous different and perhaps even multiple mechanisms could account for the enhanced vulnerability of the male brain to postnatal CAPs exposures, one attractive hypothesis relates to differences in the trajectory of colonizing activated brain microglia. Specifically, males have both a greater number of, as well as more activated (amoeboid), microglia than females in early postnatal development,¹⁶⁵ which could impact early neurogenesis and oligodendrogenesis and late cell toxicity. Additionally, it is interesting to consider that differences not only in the timing but also in the components of AP exposure during development could contribute to the heterogeneity and subgroups of ASD⁵⁸ or other neurodevelopmental disorders.

But Our Air Is Getting Cleaner?

One seeming inconsistency relates to the fact that US air quality has improved, whereas rates of ASD have continued to increase. Air pollution health effects are related to particulate matter (PM) size, designated as PM₁₀ (coarse, <10 μm), PM_2.5 (fine, <2.5 μm), or PM_0.1 (ultrafine:UFPs; <100 nm). Ultrafine particulates, being the subject of our studies, and the same size as engineered nanoparticles, are considered the most reactive component of AP,²⁰ with greater surface area/mass for adsorption. Levels of PM₁₀ and PM_2.5,but not UFPs, are regulated by the U.S. Environmental Protection Agency (EPA). While it might be anticipated that UFP levels would decline with PM_2.5 regulation, this has not necessarily been the case, with several studies reporting no correlations between levels of PM_2.5 and UFPs.^166

-170 Ultrafine particulates are part of PM_2.5 and PM₁₀, due to the fact that UFPs agglomerate rapidly into the PM_2.5 mode¹⁷¹ and PM₁₀ mode: as larger agglomerated particles, they have a high deposition efficiency on nasal mucosa, whereas de-agglomeration occurs on epithelial lining fluid so that neuronal uptake and CNS translocation takes place (olfactory and trigeminal pathways). Further understanding of how UFP and its most toxic components, alter sex-differentiated brain development in children, and alter risk for neurobehavioral disorders, is critical for public health protection.

Limitations of Animal Models for Human Risk and Research Needs

Of course rodent models have limitations in regard to characterizing human risk. Among these is the fact that rodents are altricial and thus the third trimester human equivalent of brain development occurs postnatally. Therefore, rodents are nose-breathing rather than being exposed in utero during this period. There are also differences in the anatomy of the nasal cavity between rodents and humans that can influence extent of deposition and thus transfer of particulates to brain via olfactory routes.^172
-174

To date, our studies have only begun to examine the neurotoxicity of particulate AP on brain development, but they have raised innumerable research questions that will be critical to ultimately understanding the impacts and consequences of AP. Our studies have focused primarily on white matter to date, leaving many open questions about developmental AP exposures and changes in gray matter as well as susceptibility of various brain regions. In addition, more complete characterization of effects during different windows of development will be required and would be assisted by a more complete assessment of the trajectory of effects across time; such information will also be critical to the ultimate understanding of the mechanisms by which neuropathological consequences occur. For example, with GE CAPs, we see a correlation between increased brain Fe and hypermyelination in females and a premature maturationals shift in oligodendrocytes and the myelinating cell of the CNS and suggest an impact on oligodendrocyte precursor cells.⁶⁰

Certainly, interventions would be desirable. In that regard, the elevation of brain metals always raises the suggestion by some to employ chelating agents that would bind metals and enhance their excretion. As noted by others,¹⁷⁵ such approaches are problematic for multiple reasons, including the complexity of brain metal homeostaisis and its differences within different cellular components and the lack of specificity of most chelating agents. Other approaches may be devised following a more complete understanding of the dynamics of particle chemistry and tissue interactions in vivo, that is, their in vivo bioprocessing. Aside from primary prevention, an understanding of the specific components of AP that contribute to risk could be used to alter regulatory policies to minimize exposures. For example, while levels of Pb in air are currently regulated by EPA, should this be extended to other metals?

Footnotes

Authors’ Note

Data will be made available to any qualified individuals within the scientific community to the extent that resources are possible to accommodate the nature of the request

Declaration of Conflicting Interests

The author(s) declared no potential conflicts of interest with respect to the research, authorship, and/or publication of this article.

Funding

The author(s) disclosed receipt of the following financial support for the research, authorship, and/or publication of this article: Supported in part by NIH Grants P30 ES001247 and R01 ES025541.

ORCID iD

Deborah A. Cory-Slechta

References

Sunyer

Dadvand

. Pre-natal brain development as a target for urban air pollution. Basic Clin Pharmacol Toxicol. 2019;125(suppl 3):81–88.

De Prado Bert

Mercader

EMH

Pujol

Sunyer

Mortamais

. The effects of air pollution on the brain: a review of studies interfacing environmental epidemiology and neuroimaging. Curr Environ Health Rep. 2018;5(3):351–364.

Sram

Veleminsky

Jr Veleminsky

Sr Stejskalova

. The impact of air pollution to central nervous system in children and adults. Neuro Endocrinol Lett. 2017;38(6):389–396.

Palacios

. Air pollution and parkinson’s disease—evidence and future directions. Rev Environ Health. 2017;32(4):303–313.

Jayaraj

Rodriguez

Wang

Block

. Outdoor ambient air pollution and neurodegenerative diseases: the neuroinflammation hypothesis. Curr Environ Health Rep. 2017;4(2):166–179.

Costa

Chang

Cole

. Developmental neurotoxicity of traffic-related air pollution: focus on autism. Curr Environ Health Rep. 2017;4(2):156–165.

Basnet

. A review of epidemiological research on adverse neurological effects of exposure to ambient air pollution. Front Public Health. 2016;4:157.

Power

Adar

Yanosky

Weuve

. Exposure to air pollution as a potential contributor to cognitive function, cognitive decline, brain imaging, and dementia: a systematic review of epidemiologic research. Neurotoxicol. 2016;56:235–253.

Clifford

Lang

Chen

Anstey

Seaton

. Exposure to air pollution and cognitive functioning across the life course—a systematic literature review. Environ Res. 2016;147:383–398.

10.

Suades-Gonzalez

Gascon

Guxens

Sunyer

. Air Pollution and neuropsychological development: areview of the latest evidence. Endocrinol. 2015;156(10):3473–3482.

11.

Wilker

Preis

Beiser

, et al. Long-term exposure to fine particulate matter, residential proximity to major roads and measures of brain structure. Stroke. 2015;46(5):1161–1166.

12.

Pujol

Martinez-Vilavella

Macia

, et al. Traffic pollution exposure is associated with altered brain connectivity in school children. NeuroImage. 2016;129:175–184.

13.

Pujol

Fenoll

Macia

, et al. Airborne copper exposure in school environments associated with poorer motor performance and altered basal ganglia. Brain Behav. 2016;6(6):e00467.

14.

Power

Lamichhane

Liao

, et al. The association of long-term exposure to particulate matter air pollution with brain MRI findings: the ARIC study. Environ Health Perspect. 2018;126(2):027009.

15.

Chen

Wang

Wellenius

, et al. Ambient air pollution and neurotoxicity on brain structure: evidence from women’s health initiative memory study. Ann Neurol. 2015;78(3):466–476.

16.

Alemany

Vilor-Tejedor

Garcia-Esteban

, et al. Traffic-related air pollution, APOEepsilon4 Status, and neurodevelopmental outcomes among school children enrolled in the BREATHE project (catalonia, spain). Environ Health Perspect. 2018;126(8):087001.

17.

Cory-Slechta

Allen

Conrad

Marvin

Sobolewski

. Developmental exposure to low level ambient ultrafine particle air pollution and cognitive dysfunction. Neurotoxicol. 2018;69:217–231.doi:10.1016/j.neuro.2018.08.009.

18.

Costa

Cole

Coburn

Chang

Dao

Roque

. Neurotoxicants are in the air: convergence of human, animal, and in vitro studies on the effects of air pollution on the brain. Biomed Res Int. 2014;2014:736385.

19.

Heusinkveld

Wahle

Campbell

, et al. Neurodegenerative and neurological disorders by small inhaled particles. Neurotoxicol. 2016;56:94–106.

20.

Lin

Chen

Huang

Hwang

Chang-Chien

Lin

. Characteristics of metals in nano/ultrafine/fine/coarse particles collected beside a heavily trafficked road. Environ Sci Technol. 2005;39(21):8113–8122.

21.

Clancy

Finlay

Darlington

Anand

. Extrapolating brain development from experimental species to humans. Neurotoxicol. 2007;28(5):931–937.

22.

Clancy

Kersh

Hyde

Darlington

Anand

Finlay

. Web-based method for translating neurodevelopment from laboratory species to humans. Neuroinformatic. 2007;5(1):79–94.

23.

Bandeira

Lent

Herculano-Houzel

. Changing numbers of neuronal and non-neuronal cells underlie postnatal brain growth in the rat. Proc Natl Acad Sci U S A. 2009;106(33):14108–14113.

24.

Kaur

Nieuwenhuijsen

Colvile

Personal exposure of street canyon intersection users to PM2.5, ultrafine particle counts and carbon monoxide in Central London, UK. Atmospheric Environ. 2005;39(20):3629–3641.

25.

Cattaneo

Garramone

Taronna

Peruzzo

Cavallo

. Personal exposure to airborne ultrafine particles in the urban area of milan. J Phys. 2009;151(1):012039.

26.

Zhang

Appendix

Intensive Characterization of Air Pollution in Oxford Street and Hyde Park. Boston, MA: Health Effects Institute; 2009.

27.

Allen

Conrad

Oberdorster

Johnston

Sleezer

Cory-Slechta

. Developmental exposure to concentrated ambient particles and preference for immediate reward in mice. Environ Health Perspec. 2013;121(1):32–38.

28.

Allen

Liu

Pelkowski

, et al. Early postnatal exposure to ultrafine particulate matter air pollution: persistent ventriculomegaly, neurochemical disruption, and glial activation preferentially in male mice. Environ Health Perspec. 2014;122(9):939–945.

29.

Allen

Liu

Weston

Conrad

Oberdorster

Cory-Slechta

. Consequences of developmental exposure to concentrated ambient ultrafine particle air pollution combined with the adult paraquat and maneb model of the parkinson’s disease phenotype in male mice. Neurotoxicol. 2014;41:80–88.

30.

Allen

Liu

Weston

, et al. Developmental exposure to concentrated ambient ultrafine particulate matter air pollution in mice results in persistent and sex-dependent behavioral neurotoxicity and glial activation. Toxicol Sci. 2014;140(1):160–178.

31.

Allen

Oberdorster

Morris-Schaffer

, et al. Developmental neurotoxicity of inhaled ambient ultrafine particle air pollution: parallels with neuropathological and behavioral features of autism and other neurodevelopmental disorders. Neurotoxicology. 2017;59:140–154. doi:10.1016/j.neuro.2015.12.014.

32.

Morris-Schaffer

Sobolewski

Allen

, et al. Effect of neonatal hyperoxia followed by concentrated ambient ultrafine particle exposure on cumulative learning in C57Bl/6 J mice. Neurotoxicol. 2018;67:234–244.

33.

Morris-Schaffer

Sobolewski

Welle

, et al. Cognitive flexibility deficits in male mice exposed to neonatal hyperoxia followed by concentrated ambient ultrafine particles. Neurotoxicol Teratol. 2018;70:51–59.

34.

Sobolewski

Anderson

Conrad

, et al. Developmental exposures to ultrafine particle air pollution reduces early testosterone levels and adult male social novelty preference: risk for children’s sex-biased neurobehavioral disorders. Neurotoxicol. 2018;68:203–211.

35.

Hamshere

Stergiakouli

Langley

, et al. Shared polygenic contribution between childhood attention-deficit hyperactivity disorder and adult schizophrenia. Br J Psychiatry. 2013;203(2):107–111.

36.

Kern

Geier

King

Sykes

Mehta

Geier

. Shared brain connectivity issues, symptoms, and comorbidities in autism spectrum disorder, attention deficit/hyperactivity disorder, and tourette syndrome. Brain Connect. 2015;5(6):321–335.

37.

Mitchell

Goldstein

. Inflammation in children and adolescents with neuropsychiatric disorders: a systematic review. J Am Acad Child Adolesc Psychiatry. 2014;53(3):274–296.

38.

Paul

. Developmental malformation of the corpus callosum: a review of typical callosal development and examples of developmental disorders with callosal involvement. J Neurodev Disord. 2011;3(1):3–27.

39.

Rommelse

Geurts

Franke

Buitelaar

Hartman

. A review on cognitive and brain endophenotypes that may be common in autism spectrum disorder and attention-deficit/hyperactivity disorder and facilitate the search for pleiotropic genes. Neurosci Biobehav Rev. 2011;35(6):1363–1396.

40.

Takahashi

Sakai

Tomita

. Linking activation of microglia and peripheral monocytic cells to the pathophysiology of psychiatric disorders. Front Cell Neurosci. 2016;10:144.

41.

Cheung

Fung

, et al. Autistic disorders and schizophrenia: related or remote? an anatomical likelihood estimation. PloS One. 2010;5(8):e12233.

42.

King

Lord

. Is schizophrenia on the autism spectrum? Brain Res. 2011;1380:34–41.

43.

Meyer

Feldon

Dammann

. Schizophrenia and autism: both shared and disorder-specific pathogenesis via perinatal inflammation? Pediatr Res. 2011;69(5 pt 2):26R–33R.

44.

Rapoport

Chavez

Greenstein

Addington

Gogtay

. Autism spectrum disorders and childhood-onset schizophrenia: clinical and biological contributions to a relation revisited. J Am Acad Child Adolesc Psychiatry. 2009;48(1):10–18.

45.

Brown

Singel

Hepburn

Rojas

. Increased glutamate concentration in the auditory cortex of persons with autism and first-degree relatives: a (1)H-MRS study. Autism Res. 2013;6(1):1–10.

46.

Shimmura

Suzuki

Iwata

, et al. Enzymes in the glutamate-glutamine cycle in the anterior cingulate cortex in postmortem brain of subjects with autism. Mole Aut. 2013;4(1):6.

47.

Sodhi

Sanders-Bush

. Serotonin and brain development. Int Rev Neurobiol. 2004;59:111–174.

48.

van Rees

Lago

Cox

, et al. Evidence of microglial activation following exposure to serum from first-onset drug-naive schizophrenia patients. Brain Behav Immun. 2018;67:364–373.

49.

Palmen

Hulshoff Pol

Kemner

, et al. Increased gray-matter volume in medication-naive high-functioning children with autism spectrum disorder. Psychologic Med. 2005;35(4):561–570.

50.

Sanfilipo

Lafargue

Arena

, et al. Fine volumetric analysis of the cerebral ventricular system in schizophrenia: further evidence for multifocal mild to moderate enlargement. Schizophr Bull. 2000;26(1):201–216.

51.

Shen

Nordahl

Young

, et al. Early brain enlargement and elevated extra-axial fluid in infants who develop autism spectrum disorder. Brain. 2013;136(Pt 9):2825–2835.

52.

Taurines

Schwenck

Westerwald

Sachse

Siniatchkin

Freitag

. ADHD and autism: differential diagnosis or overlapping traits? A selective review. Atten Defic Hyperact Disord. 2012;4(3):115–139.

53.

Ball

Abuhamad

Mason

Burket

Katz

Deutsch

. Clinical outcomes of mild isolated cerebral ventriculomegaly in the presence of other neurodevelopmental risk factors. J Ultrasound Med. 2013;32(11):1933–1938.

54.

Bauer

Werner

Kohl

, et al. Hyperactivity and impulsivity in adult attention-deficit/hyperactivity disorder is related to glutamatergic dysfunction in the anterior cingulate cortex. World J Biol Psychiatry. 2016;19(7):538–546.

55.

Aoki

Yoncheva

Chen

, et al. Association of white matter structure with autism spectrum disorder and attention-deficit/hyperactivity disorder. JAMA Psychiatry. 2017;74(11):1120–1128.

56.

Bos

Oranje

Achterberg

, et al. Structural and functional connectivity in children and adolescents with and without attention deficit/hyperactivity disorder. J Child Psychol Psychiatry. 2017;58(7):810–818.

57.

Donfrancesco

Nativio

Borrelli

, et al. Serum cytokines in paediatric neuropsychiatric syndromes: focus on attention deficit hyperactivity disorder. Minerva Pediatr. 2016, December 22, Epub ahead of print.

58.

Cholemkery

Medda

Lempp

Freitag

. Classifying autism spectrum disorders by ADI-R: subtypes or severity gradient? J Autism Dev Disord. 2016;46(7):2327–2339.

59.

Klocke

Allen

Sobolewski

, et al. Neuropathological consequences of gestational exposure to concentrated ambient fine and ultrafine particles in the mouse. Toxicol Sci. 2017;156(2):492–508.

60.

Klocke

Allen

Sobolewski

Blum

Zelikoff

Cory-Slechta

. Exposure to fine and ultrafine particulate matter during gestation alters postnatal oligodendrocyte maturation, proliferation capacity, and myelination. Neurotoxicology. 2018;65:196–206.

61.

Klocke

Sherina

Graham

, et al. Enhanced cerebellar myelination with concomitant iron elevation and ultrastructural irregularities following prenatal exposure to ambient particulate matter in the mouse. Inhal Toxicol. 2018;30(9-10):381–396.

62.

Levman

Vasung

MacDonald

, et al. Regional volumetric abnormalities in pediatric autism revealed by structural magnetic resonance imaging. Int J Dev Neurosci. 2018;71:34–45.

63.

Ouyang

Cheng

Mishra

, et al. Atypical age-dependent effects of autism on white matter microstructure in children of 2-7 years. Hum Brain Mapp. 2016;37(2):819–832.

64.

Lange

Travers

Bigler

, et al. Longitudinal volumetric brain changes in autism spectrum disorder ages 6-35 years. Autism Res. 2015;8(1):82–93.

65.

Morris-Schaffer

Merrill

Jew

, et al. Effects of neonatal inhalation exposure to ultrafine carbon particles on pathology and behavioral outcomes in C57BL/6 J mice. Part Fibre Toxicol. 2019;16(1):10.

66.

Wang

Zong

, et al. The trigeminal pathway dominates the nose-to-brain transportation of intact polymeric nanoparticles: evidence from aggregation-caused quenching probes. J Biomed Nanotechnol. 2019;15(4):686–702.

67.

Lochhead

Wolak

Pizzo

Thorne

. Rapid transport within cerebral perivascular spaces underlies widespread tracer distribution in the brain after intranasal administration. J Cereb Blood Flow Metab. 2015;35(3):371–381.

68.

Kozlovskaya

Abou-Kaoud

Stepensky

. Quantitative analysis of drug delivery to the brain via nasal route. J Control Release. 2014;189:133–140.

69.

Bjorklund

Meguid

El-Ansary

, et al. Diagnostic and severity-tracking biomarkers for autism spectrum disorder. J Mol Neurosci. 2018;66(4):492–511.

70.

Leffa

Torres

ILS

Rohde

. A Review on the role of inflammation in attention-deficit/hyperactivity disorder. Neuroimmunomodulation. 2018;25(5-6):328–333.

71.

Fuentes-Albero

Cauli

. Homocysteine levels in autism spectrum disorder: aclinical update. Endocr Metab Immune Disord Drug Targets. 2018;18(4):289–296.

72.

Prata

Santos

Almeida

Coelho

Barbosa

. Bridging autism spectrum disorders and schizophrenia through inflammation and biomarkers—pre-clinical and clinical investigations. J Neuroinflammation. 2017;14(1):179.

73.

Al-Yafee

Al-Ayadhi

Haq

El-Ansary

. Novel metabolic biomarkers related to sulfur-dependent detoxification pathways in autistic patients of Saudi Arabia. BMC Neurol. 2011;11:139.

74.

El-Ansary

Al-Ayadhi

. Neuroinflammation in autism spectrum disorders. J Neuroinflammation. 2012;9:265.

75.

El-Ansary

Al-Ayadhi

. GABAergic/glutamatergic imbalance relative to excessive neuroinflammation in autism spectrum disorders. J Neuroinflammation. 2014;11:189.

76.

Chen

X-C

Chow

Ward

, et al. Estimation of personal exposure to fine particles (PM2.5) of ambient origin for healthy adults in hong kong. Sci Total Environ. 2019;654:514–524.

77.

Webb

SM.

The microanalysis toolkit:X-ray fluorescence image processing software. AIP Conf Proc. 2011;1365(1):196–199.

78.

Montgomery

Narrow

Mastrangelo

Olschowka

O’Banion

Bowers

. Chronic neuron- and age-selective down-regulation of TNF receptor expression in triple-transgenic Alzheimer disease mice leads to significant modulation of amyloid- and tau-related pathologies. Am J Pathol. 2013;182(6):2285–2297.

79.

Bertrand

. Iron accumulation, glutathione depletion, and lipid peroxidation must occur simultaneously during ferroptosis and are mutually amplifying events. Med Hypotheses. 2017;101:69–74.

80.

Latunde-Dada

. Ferroptosis: role of lipid peroxidation, iron and ferritinophagy. Biochim Biophys Acta. 2017;1861(8):1893–1900.

81.

Masaldan

Bush

Devos

Rolland

Moreau

Striking while the iron is hot:Iron metabolism and ferroptosis in neurodegeneration. Free Radic Biol Med. 2019;133:221–233.

82.

Weiland

Wang

, et al. Ferroptosis and its role in diverse brain diseases. Mol Neurobiol. 2019;56(7):4880–4893.

83.

Morris

Berk

Carvalho

Maes

Walker

Puri

. Why should neuroscientists worry about iron? the emerging role of ferroptosis in the pathophysiology of neuroprogressive diseases. Behav Brain Res. 2018;341:154–175.

84.

Radu

Munteanu

Petrache

, et al. Depletion of intracellular glutathione and increased lipid peroxidation mediate cytotoxicity of hematite nanoparticles in MRC-5 cells. Acta Biochim Pol. 2010;57(3):355–360.

85.

Hohnholt

Dringen

. Iron-dependent formation of reactive oxygen species and glutathione depletion after accumulation of magnetic iron oxide nanoparticles by oligodendroglial cells. J Nanopart Res. 2011;13(12):6761–6774.

86.

Viktorinova

Ursinyova

Trebaticka

Uhnakova

Durackova

Masanova

. Changed plasma levels of zinc and copper to zinc ratio and their possible associations with parent- and teacher-rated symptoms in children with attention-deficit hyperactivity disorder. Biol Trace Elem Res. 2016;169(1):1–7.

87.

Mintz

. Evolution in the understanding of autism spectrum disorder: historical perspective. Indian J Pediatr. 2017;84(1):44–52.

88.

Goldson

. Advances in autism-2016. Adv Pediatr. 2016;63(1):333–355.

89.

Turner

Greenspan

van Erp

TGM

. Pallidum and lateral ventricle volume enlargement in autism spectrum disorder. Psychiatry Res. 2016;252:40–45.

90.

Damasio

Maurer

Damasio

Chui

. Computerized tomographic scan findings in patients with autistic behavior. Arch Neurol. 1980;37(8):504–510.

91.

Lyall

Woolson

Wolfe

, et al. Prenatal isolated mild ventriculomegaly is associated with persistent ventricle enlargement at ages 1 and 2. Early Hum Dev. 2012;88(8):691–698.

92.

Riva

Bulgheroni

Aquino

Di Salle

Savoiardo

Erbetta

. Basal forebrain involvement in low-functioning autistic children: a voxel-based morphometry study. AJNR Am J Neuroradiol. 2011;32(8):1430.

93.

Wegiel

Flory

Kuchna

, et al. Brain-region–specific alterations of the trajectories of neuronal volume growth throughout the lifespan in autism. Acta Neuropathol Commun. 2014;2(1):28.

94.

Wegiel

Flory

Kuchna

, et al. Stereological study of the neuronal number and volume of 38 brain subdivisions of subjects diagnosed with autism reveals significant alterations restricted to the striatum, amygdala and cerebellum. Acta Neuropathol Commun. 2014;2(1):141.

95.

D’Cruz

Mosconi

Ragozzino

Cook

Sweeney

. Alterations in the functional neural circuitry supporting flexible choice behavior in autism spectrum disorders. Transl Psychiatry. 2016;6(10):e916.

96.

Shafritz

Bregman

Ikuta

Szeszko

. Neural systems mediating decision-making and response inhibition for social and nonsocial stimuli in autism. Prog Neuropsychopharmacol Biol Psychiatry. 2015;60:112–120.

97.

Geschwind

Levitt

. Autism spectrum disorders: developmental disconnection syndromes. Curr Opin Neurobiol. 2007;17(1):103–111.

98.

Frith

. Is autism a disconnection disorder? Lancet Neurol. 2004;3(10):577.

99.

Blackmon

Ben-Avi

Wang

, et al. Periventricular white matter abnormalities and restricted repetitive behavior in autism spectrum disorder. Neuroimage Clin. 2016;10:36–45.

100.

Pardo

Vargas

Zimmerman

. Immunity, neuroglia and neuroinflammation in autism. Int Rev Psychiatry. 2005;17(6):485–495.

101.

Zimmerman

Jyonouchi

Comi

, et al. Cerebrospinal fluid and serum markers of inflammation in autism. Pediatr Neurol. 2005;33(3):195–201.

102.

Rojas

. The role of glutamate and its receptors in autism and the use of glutamate receptor antagonists in treatment. J Neural Transm (Vienna). 2014;121(8):891–905.

103.

Akkermans

SEA

Rheinheimer

Bruchhage

MMK

, et al. Frontostriatal functional connectivity correlates with repetitive behaviour across autism spectrum disorder and obsessive-compulsive disorder. Psychol Med. 2018:1–9.doi:10.1017/S0033291718003136.

104.

Supekar

Kochalka

Schaer

, et al. Deficits in mesolimbic reward pathway underlie social interaction impairments in children with autism. Brain. 2018;141(9):2795–2805.

105.

von Rhein

Cools

Zwiers

, et al. Increased neural responses to reward in adolescents and young adults with attention-deficit/hyperactivity disorder and their unaffected siblings. J Am Acad Child Adolesc Psychiatry. 2015;54(5):394–402.

106.

Costa Dias

Wilson

Bathula

, et al. Reward circuit connectivity relates to delay discounting in children with attention-deficit/hyperactivity disorder. Eur Neuropsychopharmacol. 2013;23(1):33–45.

107.

Frye

Melnyk

Fuchs

, et al. Effectiveness of methylcobalamin and folinic acid treatment on adaptive behavior in children with autistic disorder is related to glutathione redox status. Autism Res Treat. 2013;2013:609705.

108.

Chauhan

Audhya

Chauhan

. Brain region-specific glutathione redox imbalance in autism. Neurochem Res. 2012;37(8):1681–1689.

109.

Frye

James

. Metabolic pathology of autism in relation to redox metabolism. Biomark Med. 2014;8(3):321–330.

110.

Puig-Alcaraz

Fuentes-Albero

Calderon

Garrote

Cauli

. Increased homocysteine levels correlate with the communication deficit in children with autism spectrum disorder. Psychiatry Res. 2015;229(3):1031–1037.

111.

Nagiah

Phulukdaree

Naidoo

, et al. Oxidative stress and air pollution exposure during pregnancy: a molecular assessment. Hum Exp Toxicol. 2015;34(8):838–847.

112.

Nie

Beyea

, et al. Exposure to traffic emissions: associations with biomarkers of antioxidant status and oxidative damage. Environ Res. 2013;121:31–38.

113.

Ruvin Kumara

Wessling-Resnick

. Olfactory ferric and ferrous iron absorption in iron-deficient rats. Am J Physiol Lung Cell Mol Physiol. 2012;302(12):L1280–L1286.

114.

Wang

Feng

Wang

, et al. Transport of intranasally instilled fine Fe2O3 particles into the brain: micro-distribution, chemical states, and histopathological observation. Biol Trace Elem Res. 2007;118(3):233–243.

115.

Klocke

Sherina

Graham

116.

Tjalve

Henriksson

. Uptake of metals in the brain via olfactory pathways. Neurotoxicol. 1999;20(2-3):181–195.

117.

Maher

Ahmed

Karloukovski

, et al. Magnetite pollution nanoparticles in the human brain. Proc Natl Acad Sci U S A. 2016;113(39):10797–10801.

118.

Gonzalez-Maciel

Reynoso-Robles

Torres-Jardon

Mukherjee

Calderon-Garciduenas

. Combustion-derived nanoparticles in key brain target cells and organelles in young urbanites: culprit hidden in plain sight in Alzheimer’s disease development. J Alzheimers Dis. 2017;59(1):189–208.

119.

Calderón-Garcidueñas

Gónzalez-Maciel

Reynoso-Robles

, et al. Hallmarks of Alzheimer disease are evolving relentlessly in metropolitan mexico city infants, children and young adults. APOE4 carriers have higher suicide risk and higher odds of reaching NFT stage V at ≤40 years of age. Environ Res. 2018;164:475–487.

120.

Persson

Henriksson

Tjalve

. Uptake of cobalt from the nasal mucosa into the brain via olfactory pathways in rats. Toxicol Lett. 2003;145(1):19–27.

121.

Ibanez

Suhard

Tessier

, et al. Intranasal exposure to uranium results in direct transfer to the brain along olfactory nerve bundles. Neuropathol Appl Neurobiol. 2014;40(4):477–488.

122.

Sunderman

JR . Nasal toxicity, carcinogenicity, and olfactory uptake of metals. Ann Clin Lab Sci. 2001;31(1):3–24.

123.

Oberdörster

Sharp

Atudorei

, et al. Translocation of inhaled ultrafine particles to the brain. Inhalat Toxicol. 2004;16(6-7):437–445.

124.

Hopkins

Laing

Peake

, et al. Repeated iron-soot exposure and nose-to-brain transport of inhaled ultrafine particles. Toxicol Pathol. 2018;46(1):75–84.

125.

Speizer

Frank

. The uptake and release of SO2 by the human nose. Arch Environ Health. 1966;12(6):725–728.

126.

Meng

. Modulation of sodium currents in rat dorsal root ganglion neurons by sulfur dioxide derivatives. Brain Res. 2004;1010(1-2):127–133.

127.

Chen

Wang

, et al. Direct imaging of biological sulfur dioxide derivatives in vivo using a two-photon phosphorescent probe. Biomaterials. 2015;63:128–136.

128.

Hunter

Dey

. Identification and neuropeptide content of trigeminal neurons innervating the rat nasal epithelium. Neuroscience. 1998;83(2):591–599.

129.

Oberdorster

Sharp

Atudorei

, et al. Translocation of inhaled ultrafine particles to the brain. Inhal Toxicol. 2004;16(6-7):437–445.

130.

Lochhead

Thorne

. Intranasal delivery of biologics to the central nervous system. Adv Drug Deliv Rev. 2012;64(7):614–628.

131.

Moos

Morgan

. A morphological study of the developmentally regulated transport of iron into the brain. Dev Neurosci. 2002;24(2-3):99–105.

132.

Siddappa

Rao

Wobken

Leibold

Connor

Georgieff

. Developmental changes in the expression of iron regulatory proteins and iron transport proteins in the perinatal rat brain. J Neurosci Res. 2002;68(6):761–775.

133.

Lonnerdal

Georgieff

Hernell

. Developmental physiology of iron absorption, homeostasis, and metabolism in the healthy term infant. J Pediatr. 2015;167(suppl 4):S8–S14.

134.

Han

Kim

. Effect of dietary iron loading on recognition memory in growing rats. PLoS One. 2015;10(3):e0120609.

135.

Strahle

Garton

Bazzi

, et al. Role of hemoglobin and iron in hydrocephalus after neonatal intraventricular hemorrhage. Neurosurgery. 2014;75(6):696–705; discussion 706.

136.

Gao

Hua

Keep

Strahle

. Role of red blood cell lysis and iron in hydrocephalus after intraventricular hemorrhage. J Cereb Blood Flow Metab. 2014;34(6):1070–1075.

137.

Cheng

Juul

. Iron balance in the neonate. NeoReviews. 2011;12(3):148–157.

138.

Buonocore

Perrone

Longini

, et al. Non protein bound iron as early predictive marker of neonatal brain damage. Brain. 2003;126(Pt 5):1224–1230.

139.

Schmidt

Tancredi

Krakowiak

Hansen

Ozonoff

. Maternal intake of supplemental iron and risk of autism spectrum disorder. Am J Epidemiol. 2014;180(9):890–900.

140.

Sidrak

Yoong

Woolfenden

. Iron deficiency in children with global developmental delay and autism spectrum disorder. J Paediatr Child Health. 2014;50(5):356–361.

141.

Chen

, et al. Association between psychiatric disorders and iron deficiency anemia among children and adolescents: a nationwide population-based study. BMC Psychiatry. 2013;13(1):161.

142.

Tseng

Cheng

Chen

, et al. Peripheral iron levels in children with autism spectrum disorders vs controls: a systematic review and meta-analysis. Nutr Res. 2018;50:44–52.

143.

Ming Qian

. Iron misregulation in the brain: a primary cause of neurodegenerative disorders. Lancet Neurol. 2003;2(4):246–253.

144.

Askri

Ouni

Galai

, et al. Intranasal instillation of iron oxide nanoparticles induces inflammation and perturbation of trace elements and neurotransmitters, but not behavioral impairment in rats. Environ Sci Pollut Res Int. 2018;25(17):16922–16932.

145.

Costa-Mallen

Gatenby

Friend

, et al. Brain iron concentrations in regions of interest and relation with serum iron levels in parkinson disease. J Neurol Sci. 2017;378:38–44.

146.

Jin

Wang

Zhao

, et al. Decreased serum ceruloplasmin levels characteristically aggravate nigral iron deposition in parkinson’s disease. Brain. 2011;134(1):50–58.

147.

Logroscino

Marder

Graziano

, et al. Altered systemic iron metabolism in Parkinson’s disease. Neurology. 1997;49(3):714–717.

148.

Chen

Singh

Tay

Walczyk

. Imbalance of iron influx and efflux causes brain iron accumulation over time in the healthy adult rat. Metallomics. 2014;6(8):1417–1426.

149.

Banks

Kastin

Fasold

Barrera

Augereau

. Studies of the slow bidirectional transport of iron and transferrin across the blood-brain barrier. Brain Res Bull. 1988;21(6):881–885.

150.

Dallman

Spirito

. Brain iron in the rat: extremely slow turnover in normal rats may explain long-lasting effects of early iron deficiency. J Nutr. 1977;107(6):1075–1081.

151.

Apostolakis

Kypraiou

. Iron in neurodegenerative disorders: being in the wrong place at the wrong time? Rev Neurosci. 2017;28(8):893–911.

152.

Volpe

Laster

Transsulfuration in fetal and postnatal mammalian liver and brain.cystathionine synthase, its relation to hormonal influences, and cystathionine. Biol Neonate. 1972;20(5):385–403.

153.

Marseglia

Nicotera

Salpietro

, et al. Hyperhomocysteinemia and MTHFR polymorphisms as antenatal risk factors of white matter abnormalities in two cohorts of late preterm and full term newborns. Oxid Med Cell Longev. 2015;2015:543134.

154.

Makhro

Mashkina

Solenaya

, et al. Prenatal hyperhomocysteinemia as a model of oxidative stress of the brain. Bull Exp Biol Med. 2008;146(1):33–35.

155.

Gunnison

. Sulphite toxicity: a critical review of in vitro and in vivo data. Food Cosmet Toxicol. 1981;19(5):667–682.

156.

Hayon

Treinin

Wilf

Electronic spectra, photochemistry, and autoxidation mechanism of the sulfite-bisulfite-pyrosulfite systems. SO2-, SO3-, SO4-, and SO5- radicals. J Am Chem Soc. 1972;94(1):47–57.

157.

Eatough

Major

Ryder

, et al. The formation and stability of sulfite species in aerosols. In: Husar

Lodge

Moore

, eds. Sulfur in the Atmosphere. Oxford, England: Pergamon; 1978:263–271.

158.

Gunnison

Sellakumar

Currie

Snyder

. Distribution, metabolism and toxicity of inhaled sulfur dioxide and endogenously generated sulfite in the respiratory tract of normal and sulfite oxidase-deficient rats. J Toxicol Environ Health. 1987;21(1-2):141–162.

159.

Meng

Zhang

. Levels of sulfite in three organs from mice exposed to sulfur [corrected] dioxide. Inhal Toxicol. 2005;17(6):309–313.

160.

Sang

Hou

Yun

SO(2) inhalation induces protein oxidation, DNA-protein crosslinks and apoptosis in rat hippocampus. Ecotoxicol Environ Saf. 2009;72(3):879–884.

161.

Meng

. Oxidative damage of sulfur dioxide on various organs of mice: sulfur dioxide is a systemic oxidative damage agent. Inhal Toxicol. 2003;15(2):181–195.

162.

Petrelli

Pucci

Bezzi

. Astrocytes and microglia and their potential link with autism spectrum disorders. Front Cell Neurosci. 2016;10:21–21.

163.

Jung

Lin

Hwang

. Air pollution and newly diagnostic autism spectrum disorders: a population-based cohort study in taiwan. PloS one. 2013;8(9):e75510.

164.

Inder

Mocatta

Darlow

Spencer

Volpe

Winterbourn

. Elevated free radical products in the cerebrospinal fluid of VLBW infants with cerebral white matter injury. Pediat Res. 2002;52(2):213.

165.

Schwarz

Sholar

Bilbo

. Sex differences in microglial colonization of the developing rat brain. J Neurochemist. 2012;120(6):948–963.

166.

Ruuskanen

Tuch

Brink

, et al. Concentrations of ultrafine, fine and PM2.5 particles in three european cities. Atmospheric Environment. 2001;35:3729–3738.

167.

Tuch

Brand

Wichmann

Heyder

. Variation of particle number and mass concentration in various size ranges of ambient aerosols in eastern germany. Atmospheric Environment. 1997;31(24):4193–4197.

168.

Knibbs

De Dear

. Exposure to ultrafine particles and PM2.5 in four sydney transport modes. Atmospheric Environment. 2010;44(26):3224–3227.

169.

Ebelt

Brauer

Cyrys

, et al. Air quality in postunification erfurt, east germany: associating changes in pollutant concentrations with changes in emissions. Environ Health Perspect. 2001;109(4):325–333.

170.

Pitz

Kreyling

Hölscher

Cyrys

Wichmann

Heinrich

. Change of the ambient particle size distribution in east germany between 1993 and 1999. Atmospheric Environment. 2001;35(25):4357–4366.

171.

Charlson

. Aerosols: characteristics, behavior and measurement. In: Airborne Particles. Baltimore, MD: University Park Press; 1979:1–19.

172.

Chamanza

Wright

. A review of the comparative anatomy, histology, physiology and pathology of the nasal cavity of rats, mice, dogs and non-human primates. relevance to inhalation toxicology and human health risk assessment. J Comp Pathol. 2015;153(4):287–314.

173.

Dong

Shang

Inthavong

, et al. From the cover: comparative numerical modeling of inhaled nanoparticle deposition in human and rat nasal cavities. Toxicol Sci. 2016;152(2):284–296.

174.

Tian

Shang

Chen

, et al. Correlation of regional deposition dosage for inhaled nanoparticles in human and rat olfactory. Particle Fibre Toxicol. 2019;16(1):6.

175.

Drew

. The case for abandoning therapeutic chelation of copper ions in alzheimer’s disease. Front Neurosci. 2017;11:317.

The Impact of Inhaled Ambient Ultrafine Particulate Matter on Developing Brain: Potential Importance of Elemental Contaminants

Abstract

Keywords

Introduction

Methods and Materials

Animals and Postnatal CAP Exposures

Brain Pathology

Corpus callosum area determination

Myelin basic protein immunostaining and image analysis

Laser Ablation Inductively Coupled Plasma Mass Spectrometry

Atomic Absorption Spectrophotometry

Serum Oxidized Glutathione

Immunohistochemistry

Caspase-3

Silver staining

Statistical Analyses

Results

Concentration Effect Functions for Postnatal UFPs

Assessment of Trace Elements in CAPs Exposures

Assessment of Trace Elements in Brain

Potential Ferroptotic Mechanisms

Oxidized GSH

Nucleus Accumbens Neuronal Cell Death

Discussion

Similarities of Effects of Postnatal CAPs Exposures in Mice to Features and Hypothesized Mechanisms of Autism Spectrum Disorder and Other Neurodevelopmental Disorders in Humans

Olfactory Uptake of Elemental Contaminants of AP

Plausibility of Elemental Contaminants of AP for the Postnatal CAPs-Induced Phenotype

Elevated Brain Fe and S Suggest Potential Ferroptotic and Oxidative Stress-Based Mechanisms for Postnatal CAPs Effects

Male Vulnerability to Postnatal CAPs Exposures and Heterogeneity of Neurodevelopmental Disorders

But Our Air Is Getting Cleaner?

Limitations of Animal Models for Human Risk and Research Needs

Footnotes

Authors’ Note

Declaration of Conflicting Interests

Funding

ORCID iD

References