Lucio G. Costa

Polybrominated Diphenyl Ethers: Neurobehavioral Effects Following Developmental Exposure

Polybrominated diphenyl ethers (PBDEs), a class of widely used flame retardants, are becoming widespread environmental pollutants, as indicated by studies on sentinel animal species, as well as humans. Of particular concern are the reported increasingly high levels of PBDEs in human milk, as should be given that almost no information is available on their potential effects on developing organisms. In order to address this issue, studies have been conducted in mice and rats to assess the potential neurotoxic effects of perinatal exposure to PBDEs (congeners 47, 99, 153 and the penta-BDE mixture DE-71). Characteristic endpoints of PBDE neurotoxicity are, among others, endocrine disruption (e.g. decreased thyroid hormone levels), alteration in cholinergic system activity (behavioral hyporesponsivity to nicotine challenge), as well as alterations of several behavioral parameters. In particular, the main hallmark of PBDE neurotoxicity is a marked hyperactivity at adulthood. Furthermore, a deficit in learning and memory processes has been found at adulthood in neonatally exposed animals. Some of neurotoxic effects of PBDEs are comparable to those of polychlorinated biphenyls (PCBs), though the latter class of compounds seems to exert a stronger toxic effect. Available information on PBDE neurotoxicity obtained from animal studies and the possibility of neonatal exposure to PBDEs via the mothers milk suggest that these compounds may represent a potential risk for neurobehavioral development in humans.

Polyphenols and health: What compounds are involved?

On the basis of prospective, cross-sectional and intervention studies linking polyphenols to human health, several experimental papers in the literature have tried to evaluate the molecular mechanisms involved in their bioactivity. Polyphenols are reported to in vitro inhibit cancer cell proliferation, reduce vascularisation, protect neurons, stimulate vasodilation and improve insulin secretion, but are often studied as aglycones or as sugar conjugates and at non-physiological concentration. However, it is now well established that polyphenols undergo substantial metabolism after being ingested by humans in dietary relevant amount and that concentrations of plasma metabolites after a normal dietary intake rarely exceed nmol/L. This viewpoint intends to highlight that uncritical judgements made on the basis of the published literature, particularly about toxicity and bioactivity, may sometimes have been misled and misleading and to conclude that i) bioavailability values reported in the literature for phenolic compounds should be strongly reconsidered in the light of the large number of newly identified circulating and excreted metabolites, with particular attention to colonic ring-fission products which are obviously contributing much more than expected to the percentage of their absorption; ii) it is phenolic metabolites, formed in the small intestine and hepatic cells, and low molecular weight catabolic products of the colonic microflora to travel around the human body in the circulatory system or reach body tissues to elicit bioactive effects. Understanding these compounds certainly carries interest for drug-discovery but also for dietary prevention of disease.

Antiglycative and neuroprotective activity of colon-derived polyphenol catabolites

SCOPEnDietary flavonoids and allied phenolic compounds are thought to be beneficial in the control of diabetes and its complications, because of their ability to inhibit oxidative stress, protein glycation and to act as neuroprotectants. Following ingestion by humans, polyphenolic compounds entering the large intestine undergo extensive metabolism by interaction with colonic microbiota and it is metabolites and catabolites of the parent compounds that enter the circulatory system. The aim of this study was to investigate the inhibitory activity of some colonic microbiota-derived polyphenol catabolites against advanced glycation endproducts formation in vitro and to determine their ability, at physiological concentrations, to counteract mild oxidative stress of cultured human neuron cells.nnnMETHODS AND RESULTSnThis study demonstrated that ellagitannin-derived catabolites (urolithins and pyrogallol) are the most effective antiglycative agents, whereas chlorogenic acid-derived catabolites (dihydrocaffeic acid, dihydroferulic acid and feruloylglycine) were most effective in combination in protecting neuronal cells in a conservative in vitro experimental model.nnnCONCLUSIONnSome polyphenolic catabolites, generated in vivo in the colon, were able in vitro to counteract two key features of diabetic complications, i.e. protein glycation and neurodegeneration. These observations could lead to a better control of these events, which are usually correlated with hyperglycemia.

Is decabromodiphenyl ether (BDE-209) a developmental neurotoxicant?

Polybrominated diphenyl ether (PBDE) flame retardants have become ubiquitous environmental pollutants. The relatively higher body burden in toddlers and children has raised concern for their potential developmental neurotoxicity, which has been suggested by animal studies, in vitro experiments, and recent human epidemiological evidence. While lower brominated PBDEs have been banned in several countries, the fully brominated decaBDE (BDE-209) is still utilized, though manufacturers will discontinue production in the U.S.A. in 2013. The recent decision by the U.S. Environmental Protection Agency to base the reference dose (RfD) for BDE-209 on a developmental neurotoxicity study has generated some controversy. Because of its bulky configuration, BDE-209 is poorly absorbed and does not easily penetrate the cell wall. Its acute and chronic toxicities are relatively low, with the liver and the thyroid as the primary targets, though there is some evidence of carcinogenicity. A few animal studies have indicated that BDE-209 may cause developmental neurotoxicity, affecting motor and cognitive domains, as seen for other PBDEs. Limited in vivo and in vitro studies have also evidenced effects of BDE-209 on thyroid hormone homeostasis and direct effects on nervous cells, again similar to what found with other lower brominated PBDEs. In contrast, a recent developmental neurotoxicity study, carried out according to international guidelines, has provided no evidence of adverse effects on neurodevelopment, and this should be considered in a future re-evaluation of BDE-209. While estimated exposure to BDE-209 in children is believed to be several orders of magnitude below the most conservative RfD proposed by the USEPA, questions remain on the extent and relevance of BDE-209 metabolism to lower brominated PBDEs in the environment and in humans.

Low concentrations of the brominated flame retardants BDE-47 and BDE-99 induce synergistic oxidative stress-mediated neurotoxicity in human neuroblastoma cells.

Polybrominated diphenyl ether (PBDE) flame retardants have become widespread environmental contaminants. The highest body burden has been found in toddlers and infants, due to their exposure through breast milk and house dust, and the current concern for potential adverse health effects of PBDEs relates to their developmental neurotoxicity. The mechanisms underlying the neurotoxicity of PBDEs are largely not understood, though there is evidence that PBDEs may elicit oxidative stress. In this study, two different mathematical models were used to evaluate the interaction between BDE-47 and BDE-99 on viability of neuronal cells. The combined exposure to these compounds induced synergistic effects at concentrations of BDE-47 below its threshold doses, and in a wide range of BDE-99 concentrations below its IC(50). In contrast, at concentrations of BDE-47 near its IC(50) value, and in a wide range of BDE-99 concentrations, antagonistic effects were observed. The interactions observed on cell viability were confirmed by an assessment of induction of oxidative stress. The finding that co-exposure to BDE-47 and BDE-99 could induce synergistic neurotoxic effects, in particular at low doses of BDE-47, is of much toxicological interest, as humans are exposed to mixtures of PBDEs, most notably tetra- and penta-BDE congeners.

DOMOIC ACID AS A DEVELOPMENTAL NEUROTOXIN

Domoic acid (DomA) is an excitatory amino acid which can accumulate in shellfish and finfish under certain environmental conditions. DomA is a potent neurotoxin. In humans and in non-human primates, oral exposure to a few mg/kg DomA elicits gastrointestinal effects, while slightly higher doses cause neurological symptoms, seizures, memory impairment, and limbic system degeneration. In rodents, which appear to be less sensitive than humans or non-human primates, oral doses cause behavioral abnormalities (e.g. hindlimb scratching), followed by seizures and hippocampal degeneration. Similar effects are also seen in other species (from sea lions to zebrafish), indicating that DomA exerts similar neurotoxic effects across species. The neurotoxicity of DomA is ascribed to its ability to interact and activate the AMPA/KA receptors, a subfamily of receptors for the neuroexcitatory neurotransmitter glutamate. Studies exploring the neurotoxic effects of DomA on the developing nervous system indicate that DomA elicits similar behavioral, biochemical and morphological effects as in adult animals. However, most importantly, developmental neurotoxicity is seen at doses of DomA that are one to two orders of magnitude lower than those exerting neurotoxicity in adults. This difference may be due to toxicokinetic and/or toxicodynamic differences. Estimated safe doses may be exceeded in adults by high consumption of shellfish contaminated with DomA at the current limit of 20 microg/g. Given the potential higher susceptibility of the young to DomA neurotoxicity, additional studies investigating exposure to, and effects of this neurotoxin during brain development are warranted.

Modulation of Neuritogenesis by Astrocyte Muscarinic Receptors

Astrocytes have been shown to release factors that have promoting or inhibiting effects on neuronal development. However, mechanisms controlling the release of such factors from astrocytes are not well established. Astrocytes express muscarinic receptors whose activation stimulates a robust intracellular signaling, although the role of these receptors in glial cells is not well understood. Acetylcholine and acetylcholine receptors are present in the brain before synaptogenesis occurs and are believed to be involved in neuronal maturation. The present study was undertaken to investigate whether stimulation of muscarinic receptors in astrocytes would modulate neurite outgrowth in hippocampal neurons. Rat hippocampal neurons, co-cultured with rat cortical astrocytes previously exposed to the cholinergic agonist carbachol, displayed longer neurites. The effect of carbachol in astrocytes was due to the activation of M3 muscarinic receptors. Exposure of astrocytes to carbachol increased the expression of the extracellular matrix proteins fibronectin and laminin-1 in these cells. This effect was mediated in part by an increase in laminin-1 and fibronectin mRNA levels and in part by the up-regulation of the production and release of plasminogen activator inhibitor-1, an inhibitor of the proteolytic degradation of the extracellular matrix. The inhibition of fibronectin activity strongly reduced the effect of carbachol on the elongation of all the neurites, whereas inhibition of laminin-1 activity reduced the elongation of minor neurites only. Plasminogen activator inhibitor-1 also induced neurite elongation through a direct effect on neurons. Taken together, these results demonstrate that cholinergic muscarinic stimulation of astrocytes induces the release of permissive factors that accelerate neuronal development.

Mechanisms of Neuroprotection by Quercetin: Counteracting Oxidative Stress and More.

Increasing interest has recently focused on determining whether several natural compounds, collectively referred to as nutraceuticals, may exert neuroprotective actions in the developing, adult, and aging nervous system. Quercetin, a polyphenol widely present in nature, has received the most attention in this regard. Several studies in vitro, in experimental animals and in humans, have provided supportive evidence for neuroprotective effects of quercetin, either against neurotoxic chemicals or in various models of neuronal injury and neurodegenerative diseases. The exact mechanisms of such protective effects remain elusive, though many hypotheses have been formulated. In addition to a possible direct antioxidant effect, quercetin may also act by stimulating cellular defenses against oxidative stress. Two such pathways include the induction of Nrf2-ARE and induction of the antioxidant/anti-inflammatory enzyme paraoxonase 2 (PON2). In addition, quercetin has been shown to activate sirtuins (SIRT1), to induce autophagy, and to act as a phytoestrogen, all mechanisms by which quercetin may provide its neuroprotection.

Neurotoxicants Are in the Air: Convergence of Human, Animal, and In Vitro Studies on the Effects of Air Pollution on the Brain

In addition to increased morbidity and mortality caused by respiratory and cardiovascular diseases, air pollution may also negatively affect the brain and contribute to central nervous system diseases. Air pollution is a mixture comprised of several components, of which ultrafine particulate matter (UFPM; <100u2009nm) is of much concern, as these particles can enter the circulation and distribute to most organs, including the brain. A major constituent of ambient UFPM is represented by traffic-related air pollution, mostly ascribed to diesel exhaust (DE). Human epidemiological studies and controlled animal studies have shown that exposure to air pollution may lead to neurotoxicity. In addition to a variety of behavioral abnormalities, two prominent effects caused by air pollution are oxidative stress and neuroinflammation, which are seen in both humans and animals and are confirmed by in vitro studies. Among factors which can affect neurotoxic outcomes, age is considered the most relevant. Human and animal studies suggest that air pollution (and DE) may cause developmental neurotoxicity and may contribute to the etiology of neurodevelopmental disorders, including autistic spectrum disorders. In addition, air pollution exposure has been associated with increased expression of markers of neurodegenerative disease pathologies.

Gender differences in brain susceptibility to oxidative stress are mediated by levels of paraoxonase-2 expression

Paraoxonase 2 (PON2), a member of a gene family that also includes PON1 and PON3, is expressed in most tissues, including the brain. In mouse brain, PON2 levels are highest in dopaminergic areas (e.g., striatum) and are higher in astrocytes than in neurons. PON2 is primarily located in mitochondria and exerts a potent antioxidant effect, protecting mouse CNS cells against oxidative stress. The aim of this study was to characterize PON2 expression and functions in the brains of male and female mice. Levels of PON2 (protein, mRNA, and lactonase activity) were higher in brain regions and cells of female mice. Astrocytes and neurons from male mice were significantly more sensitive (by 3- to 4-fold) to oxidative stress-induced toxicity than the same cells from female mice. Glutathione levels did not differ between genders. Importantly, no significant gender differences in susceptibility to the same oxidants were seen in cells from PON2(-/-) mice. Treatment with estradiol induced a time- and concentration-dependent increase in the levels of PON2 protein and mRNA in male (4.5-fold) and female (1.8-fold) astrocytes, which was dependent on activation of estrogen receptor-α. In ovariectomized mice, PON2 protein and mRNA were decreased to male levels in brain regions and in liver. Estradiol protected astrocytes from wild-type mice against oxidative stress-induced neurotoxicity, but did not protect cells from PON2(-/-) mice. These results suggest that PON2 is a novel major intracellular factor that protects CNS cells against oxidative stress and confers gender-dependent susceptibility to such stress. The lower expression of PON2 in males may have broad ramifications for susceptibility to diseases involving oxidative stress, including neurodegenerative diseases.