Bellina Veronesi

Translocation and potential neurological effects of fine and ultrafine particles a critical update

Particulate air pollution has been associated with respiratory and cardiovascular disease. Evidence for cardiovascular and neurodegenerative effects of ambient particles was reviewed as part of a workshop. The purpose of this critical update is to summarize the evidence presented for the mechanisms involved in the translocation of particles from the lung to other organs and to highlight the potential of particles to cause neurodegenerative effects.Fine and ultrafine particles, after deposition on the surfactant film at the air-liquid interface, are displaced by surface forces exerted on them by surfactant film and may then interact with primary target cells upon this displacement. Ultrafine and fine particles can then penetrate through the different tissue compartments of the lungs and eventually reach the capillaries and circulating cells or constituents, e.g. erythrocytes. These particles are then translocated by the circulation to other organs including the liver, the spleen, the kidneys, the heart and the brain, where they may be deposited. It remains to be shown by which mechanisms ultrafine particles penetrate through pulmonary tissue and enter capillaries. In addition to translocation of ultrafine particles through the tissue, fine and coarse particles may be phagocytized by macrophages and dendritic cells which may carry the particles to lymph nodes in the lung or to those closely associated with the lungs. There is the potential for neurodegenerative consequence of particle entry to the brain. Histological evidence of neurodegeneration has been reported in both canine and human brains exposed to high ambient PM levels, suggesting the potential for neurotoxic consequences of PM-CNS entry. PM mediated damage may be caused by the oxidative stress pathway. Thus, oxidative stress due to nutrition, age, genetics among others may increase the susceptibility for neurodegenerative diseases. The relationship between PM exposure and CNS degeneration can also be detected under controlled experimental conditions. Transgenic mice (Apo E -/-), known to have high base line levels of oxidative stress, were exposed by inhalation to well characterized, concentrated ambient air pollution. Morphometric analysis of the CNS indicated unequivocally that the brain is a critical target for PM exposure and implicated oxidative stress as a predisposing factor that links PM exposure and susceptibility to neurodegeneration.Together, these data present evidence for potential translocation of ambient particles on organs distant from the lung and the neurodegenerative consequences of exposure to air pollutants.

Nanosize titanium dioxide stimulates reactive oxygen species in brain microglia and damages neurons in vitro.

Background Titanium dioxide is a widely used nanomaterial whose photo-reactivity suggests that it could damage biological targets (e.g., brain) through oxidative stress (OS). Objectives Brain cultures of immortalized mouse microglia (BV2), rat dopaminergic (DA) neurons (N27), and primary cultures of embryonic rat striatum, were exposed to Degussa P25, a commercially available TiO2 nanomaterial. Physical properties of P25 were measured under conditions that paralleled biological measures. Findings P25 rapidly aggregated in physiological buffer (800–1,900 nm; 25°C) and exposure media (~ 330 nm; 37°C), and maintained a negative zeta potential in both buffer (–12.2 ± 1.6 mV) and media (–9.1 ± 1.2 mV). BV2 microglia exposed to P25 (2.5–120 ppm) responded with an immediate and prolonged release of reactive oxygen species (ROS). Hoechst nuclear stain was reduced after 24-hr (≥100 ppm) and 48-hr (≥2.5 ppm) exposure. Microarray analysis on P25-exposed BV2 microglia indicated up-regulation of inflammatory, apoptotic, and cell cycling pathways and down-regulation of energy metabolism. P25 (2.5–120 ppm) stimulated increases of intracellular ATP and caspase 3/7 activity in isolated N27 neurons (24–48 hr) but did not produce cytotoxicity after 72-hr exposure. Primary cultures of rat striatum exposed to P25 (5 ppm) showed a reduction of immunohistochemically stained neurons and microscopic evidence of neuronal apoptosis after 6-hr exposure. These findings indicate that P25 stimulates ROS in BV2 microglia and is nontoxic to isolated N27 neurons. However, P25 rapidly damages neurons at low concentrations in complex brain cultures, plausibly though microglial generated ROS.

Nanometer size diesel exhaust particles are selectively toxic to dopaminergic neurons: the role of microglia, phagocytosis, and NADPH oxidase

The contributing role of environmental factors to the development of Parkinson’s disease has become increasingly evident. We report that mesencephalic neuron‐glia cultures treated with diesel exhaust particles (DEP; 0.22 µM) (5–50 µg/ml) resulted in a dose‐dependent decrease in dopaminergic (DA) neurons, as determined by DA‐uptake assay and tyrosine‐hydroxylase immunocytochemistry (ICC). The selective toxicity of DEP for DA neurons was demonstrated by the lack of DEP effect on both GABA uptake and Neu‐N immunoreactive cell number. The critical role of microglia was demonstrated by the failure of neuron‐enriched cultures to exhibit DEP‐induced DA neurotoxicity, where DEP‐induced DA neuron death was reinstated with the addition of microglia to neuron‐enriched cultures. OX‐42 ICC staining of DEP treated neuron‐glia cultures revealed changes in microglia morphology indicative of activation. Intracellular reactive oxygen species and superoxide were produced from enriched‐microglia cultures in response to DEP. Neuron‐glia cultures from NADPH oxidase deficient (PHOX−/−) mice were insensitive to DEP neurotoxicity when compared with control mice (PHOX+/+). Cytochalasin D inhibited DEP‐induced superoxide production in enriched‐microglia cultures, implying that DEP must be phagocytized by microglia to produce superoxide. Together, these in vitro data indicate that DEP selectively damages DA neurons through the phagocytic activation of microglial NADPH oxidase and consequent oxidative insult.

The outdoor air pollution and brain health workshop

Accumulating evidence suggests that outdoor air pollution may have a significant impact on central nervous system (CNS) health and disease. To address this issue, the National Institute of Environmental Health Sciences/National Institute of Health convened a panel of research scientists that was assigned the task of identifying research gaps and priority goals essential for advancing this growing field and addressing an emerging human health concern. Here, we review recent findings that have established the effects of inhaled air pollutants in the brain, explore the potential mechanisms driving these phenomena, and discuss the recommended research priorities/approaches that were identified by the panel.

Neuropeptides and capsaicin stimulate the release of inflammatory cytokines in a human bronchial epithelial cell line.

The role of neuropeptides in initiating and modulating airway inflammation was examined in a human bronchial epithelial cell line (i.e. BEAS-2B). At a range of concentrations, exposure of BEAS-2B cells to Substance P (SP) or calcitonin gene related protein resulted in immediate increases in intracellular calcium ([Ca(2+)](i)), the synthesis of the transcripts for the inflammatory cytokines, IL-6, IL-8 and TNFalpha after 2 h exposure, and the release of their proteins after 6 h exposure. Addition of thiorphan (100 nM), an inhibitor of neutral endopeptidase, enhanced the levels of SP-stimulated cytokine release. Stimulation of IL-6 by SP occurred in a conventional receptor-mediated manner as demonstrated by its differential release by fragments SP 4-11 and SP 1-4 and by the blockage of IL-6 release with the non-peptide, NK-1 receptor antagonist, CP-99 994. In addition to the direct stimulation of inflammatory cytokines, SP (0.5 microM), in combination with TNFalpha (25 units/ml), synergistically stimulated IL-6 release. BEAS-2B cells also responded to the botanical irritant, capsaicin (10 microM) with increases in [Ca(2+)](i) and IL-8 cytokine release after 4 h exposure. The IL-8 release was dependent on the presence of extracellular calcium. Capsaicin-stimulated increases of [Ca(2+)](i) and cytokine release could be reduced to control levels by pre-exposure to capsazepine, an antagonist of capsaicin (i.e. vanilloid) receptor(s) or by deletion of extracellular calcium from the exposure media. The present data indicate that the BEAS-2B human epithelial cell line expresses neuropeptide and capsaicin-sensitive pathways, whose activation results in immediate increases of [Ca(2+)](i) stimulation of inflammatory cytokine transcripts and the release of their cytokine proteins.

NEUROLOGICAL IMPACTS FROM INHALATION OF POLLUTANTS AND THE NOSE-BRAIN CONNECTION

The effects of inhaled particles have focused heavily on the respiratory and cardiovascular systems. Most studies have focused on inhaled metals, whereas less information is available for other particle types regarding the effects on the brain and other extra-pulmonary organs. We review here the key available literature on nanoparticle uptake and transport through the olfactory pathway, the experimental data from animal and in vitro studies, and human epidemiological observations. Nanoparticles (<0.1 μm in one dimension) may easily reach the brain from the respiratory tract via sensory neurons and transport from the distal alveoli into the blood or lymph as free particles or inside phagocytic cells. These mechanisms and subsequent biologic responses may be influenced by the chemical composition of inhaled particles. Animal studies with ambient particulate matter and certain other particles show alterations in neuro-inflammatory markers of oxidative stress and central neurodegeneration. Human observations indicate motor, cognitive, and behavioral changes especially after particulate metal exposure in children. Exposure to co-pollutants and/or underlying disease states could also impact both the biokinetics and effects of airborne particles in the brain. Data are needed from the areas of inhalation, neurology, and metal toxicology in experimental and human studies after inhalation exposure. An increased understanding of the neurotoxicity associated with air pollution exposure is critical to protect susceptible individuals in the workplace and the general population.

Acetylcholinesterase and Neuropathy Target Esterase Inhibitions in Neuroblastoma Cells to Distinguish Organophosphorus Compounds Causing Acute and Delayed Neurotoxicity

The differential inhibition of the target esterases acetylcholinesterase (AChE) and neuropathy target esterase (NTE, neurotoxic esterase) by organophosphorus compounds (OPs) is followed by distinct neurological consequences in exposed subjects. The present study demonstrates that neuroblastoma cell lines (human SH-SY5Y and murine NB41A3) can be used to differentiate between neuropathic OPs (i.e., those inhibiting NTE and causing organophosphorus-induced delayed neuropathy) and acutely neurotoxic OPs (i.e., those highly capable of inhibiting AChE). In these experiments, concentration-response data indicated that the capability to inhibit AChE was over 100x greater than the capability to inhibit NTE for acutely toxic, nonneuropathic OPs (e.g., paraoxon and malaoxon) in both cell lines. Inhibition of AChE was greater than inhibition of NTE, without overlap of the concentration-response curves, for OPs which are more likely to cause acute, rather than delayed, neurotoxic effects in vivo (e.g., chlorpyrifos-oxon, dichlorvos, and trichlorfon). In contrast, concentrations inhibiting AChE and NTE overlapped for neuropathy-causing OPs. For example, apparent IC50 values for NTE inhibition were less than 9.6-fold the apparent IC50 values for AChE inhibition when cells were exposed to the neuropathy-inducing OPs diisopropyl phosphorofluoridate, cyclic tolyl saligenin phosphate, phenyl saligenin phosphate, mipafox, dibutyl dichlorovinyl phosphate, and di-octyl-dichlorovinyl phosphate. In all cases, esterase inhibition occurred at lower concentrations than those needed for cytoxicity. These results suggest that either mouse or human neuroblastoma cell lines can be considered useful in vitro models to distinguish esterase-inhibiting OP neurotoxicants.

Particulate matter, oxidative stress and neurotoxicity.

Particulate matter (PM), a component of air pollution has been epidemiologically associated with sudden deaths, cardiovascular and respiratory illnesses. The effects are more pronounced in patients with pre-existing conditions such as asthma, diabetes or obstructive pulmonary disorders. Clinical and experimental studies have historically focused on the cardiopulmonary effects of PM. However, since PM particles carry numerous biocontaminants that are capable of triggering free radical production and cytokine release, the possibility that PM may affect organs systems sensitive to oxidative stress must be considered. Four independent studies that summarize the neurochemical and neuropathological changes found in the brains of PM exposed animals are described here. These were recently presented at two 2007 symposia sponsored by the Society of Toxicology (Charlotte, NC) and the International Neurotoxicology Association (Monterey, CA).

Effects of subchronic exposures to concentrated ambient particles. VII. Degeneration of dopaminergic neurons in Apo E-/- mice.

Abstract This study reports that subchronic exposure of Tuxedo, NY concentrated ambient particulates (CAPs) produces neuropathological damage in the brains of Apo E-deficient mice (Apo E−/−). These genetically modified mice are characterized by elevated levels of oxidative stress (OS) in the brain. Microscopic examination of coronal sections of the brain, immunocytochemically stained for dopamineric neurons, indicated that neurons from the substantia nigral nucleus compacta were significantly reduced by 29% in CAPs-exposed Apo E−/− mice relative to air-exposed Apo E−/− controls. In addition, statistically significant increases (p < .05) in immunocytochemically stained astrocytes were noted. The dopaminergic neurons of the nucleus compacta are specifically targeted in Parkinsons disease. The present study expands the systems affected by particulate matter to include the brain, and supports an environmental role for the development of neurodegeneration in OS-susceptible individuals.

Fornix transections disrupt the ontogeny of latent inhibition in the rat

In three experiments, we examined the ontogeny of latent inhibition in a conditioned taste-aversion paradigm. In the first experiment, 18-, 25-, and 32-day-old rats received a pairing of a 0.6% Sanka solution with an i.p. injection of LiCl (0.75% b.w., 0.4 M) or physiological saline either after four preexposures to the taste or control treatment without taste preexposure. Impairment of conditioning by taste preexposure (latent inhibition) was evident at 32 days of age, but not at 18 or 25 days of age. In Experiment 2, we investigated the possibility that our failure to observe latent inhibition in 18-day-olds resulted from the particular experimental parameters employed in Experiment 1. In Experiment 2, taste-exposure durations were longer, the retention interval between conditioning and testing was shorter, and deprivation conditions were more typical of those used with preweanling subjects. Again, no evidence of latent inhibition was found. In Experiment 3, we examined the effect of sham or fornix lesions at 18 days of age on latent inhibition at 32 days of age. Lesioned subjects exhibited equivalent conditioned taste aversions but failed to show latent inhibition. Thus, latent inhibition of taste-aversion learning develops between 18 and 32 days of age in the rat, and this development is disrupted by fornix transection during infancy. These findings suggest a role for septohippocampal maturation in the ontogeny of latent inhibition.