Particle Toxicology - Chapter 19 ppt

19 Air Pollution and Human Brain Pathology: ARole for Air Pollutants in the Pathogenesis of Alzheimer’sDisease Lilian Caldero ´ n-Garciduen ˜ as TheCenter for Structural andFunctional Neurosciences, University of Montana William Reed DepartmentofPediatrics andCenter for Environmental Medicine, University of North CarolinaatChapelHill CONTENTS 19.1 Introduction 331 19.2 Molecular Basis of Alzheimer’s Disease Pathogenesis 332 19.3 Alzheimer’s Disease Pathogenesis and COX2 334 19.4 The Mexico City Environment 334 19.5 COX2 and IL-1b Expression,Ab 42 Accumulation, and Neuropathology in the Brains of Dogs and Humans Exposed to SevereUrban Air Pollution 337 19.6 ClinicalStudies of Mexico City Children 341 19.7 Potential Mechanisms of Air Pollutant-Induced Inflammation and Neurodegeneration 343 19.7.1 Air Pollutant-Induced Systemic Inflammation 343 19.7.2 Transport of PM-AssociatedMetals to the Brain 343 19.7.3 LPS Toxicity 345 19.8 Summary 346 Acknowledgments 346 References 346 19.1 INTRODUCTION Adverse health effects associated with chronic exposures to air pollutants(indoor, outdoor, and occupational settings) are an important issue for millions of people aroundthe world. As the world population becomes older, significant increases in neurodegenerative diseasessuch as Alzheimer’s have been projected over the next decades (Brookmeyer, Gray, and Kawas 1998;Hebert et al. 2003). Alzheimer’s disease (AD) is an irreversible, fatal brain disorder that presently affects 4.5 million people in the United States and it is projected that it will affect between 13 and 16 million by 331 © 2007 by Taylor & Francis Group, LLC 2050 (Brookmeyer, Gray, and Kawas 1998;Hebert et al. 2003). Alzheimer’s patients have amajor medical, social,and economicimpact,thus any factorsthat could modify these projections need to be pursued and integratedinto multidisciplinarystudies of AD. Therole played by the environment in thepathogenesis of AD is unclear (Brown,Lockwood, andSonawane2005).Our findings suggestthat exposures to significant levels of particulate matter and photo-oxidantsmay accelerate the appearance of precursors of Alzheimer’s disease in sentinel animals and in humans. In this chapter, we will review the pathophysiology of AD as it is currently understood and summarize comparativepathology, humanneuropathology,and clinicalstudies of residentsof cities with significant chronicconcentrationsofparticulate matter, endotoxins,ozone,and a myriadofotherair pollutants. We discuss how air pollutants might promote AD indirectly by causing systemic inflammation or directly by causing brain injury following their entry intothe brain via knownpathways. 19.2 MOLECULAR BASIS OF ALZHEIMER’S DISEASE PATHOGENESIS Alzheimer’s brains exhibit two pathological hallmarks: (1) the accumulation of b -amyloid peptides (Ab )inthe extracellular space in the form of neuritic plaques and (2) intraneuronal filamentous tangles (neurofibrillarytangles, NFTs) containing hyperphosphorylated tau protein (reviewed in Selkoe2001). A b peptides are 37–43 amino acid proteolytic fragments of b -amyloid precursor protein(APP), an alternativelyspliced transmembraneproteinexpressed by all cells. Normal neurons primarily express the 695 amino acid form of APP. A b peptides are generated by the proteolytic cleavage of APP by two proteases, b -and g -secretase(Selkoe2004). Neuritic plaques are foci of extracellular A b deposition that are associated with axonal and dendritic injury (Selkoe 2001). Alarge part of the fibrillar A b found in plaques is the 42 amino acid-isoform (Ab 42) that is more hydrophobic and prone to aggregation than otherAb isoforms. Precursor lesions of neuritic plaques are referred to as “diffuse plaques” (Selkoe 2001). Genetic studies of familial AD (FAD), an inherited, early onset form of AD, suggestthat the generation of A b 42 playsacontributory role in AD pathogenesis. Mutations in any of three genes, APP, presenillin-1, and presenillin-2cause aspecific increase in A b 42 generation that is associated presenillin with FAD (Scheuneretal. 1996). All three genes might be expected to regulate A b 42 generation, as APP is the precursor of A b peptides and the presenilins are essential components of g -secretase(reviewedinSelkoe and Kopan 2003). An increaseinAPP gene dose as occursin Down’ssyndrome (3 copies of chromosome 21 rather than 2) is associated with early onset AD as well (Lemere et al. 1996). Finally, amajor risk factor for the development of AD is the epsilon 4 polymorphism of the apolipoprotein Egene (Corder et al. 1993), whose proteinproductenhances A b 42 stability and accumulation (Strittmatteretal. 1993; reviewed in George-Hyslop2000). These findingssupport the amyloid cascade hypothesisofADpathogenesis. Theamyloid cascade hypothesispostulates that increased production of the A b 42 results in its accumulation and oligomerization in limbic and association cortices leading to the gradual depo- sitionofAb 42 oligomers as diffuse plaques and to subtle effects of the oligomers on synaptic efficacy. These early changesare believedtocause microglial and astrocytic activation, prolifera- tive responses in microglia (gliosis), altered neuronal ionic homeostasis, and widespread synaptic dysfunction andneurodegeneration(Selkoe 2001; Clearyetal. 2005).Inthishypothesis, an increaseinthe generation of A b 42 is the essential and self-propagating pathogenic event in AD. An alternative pathogenic mechanism proposes that AD is aconsequenceofafailure of the control of neuronal differentiation (reviewed in Arendt 2003;Nagy 2005;and Webberetal. 2005). In general, brain neurons rest in apostmitotic, differentiated state. However, adaptivebehaviors such as learning and memory require that synapses are continually lost or formed and strengthened or weakenedinaremodeling process that relies upon cellular repair machinery. It is hypothesized that normalsynapticturnovertogether with genetic factorsthatcause instabilityinthe repair Particle Toxicology332 © 2007 by Taylor & Francis Group, LLC machinery or external factors that result in neuronal injury (e.g., oxidative stress and inflammation) or bothcould result in excessivedemandupon therepair machinery andincreasethe risk of dedifferentiation with subsequent cell cycleactivation (Arendt 2003). Alternatively,cell cycle activation in neuronscould be mediated by inappropriate mitogenic stimuli caused by altered expression or mutation of molecular components of signal transduction pathways that regulatethe cell cycle. Although the function of APP is still poorly understood, limited evidence suggests that APP may regulateneuronal survival (Koo and Kopan2004)by signaling to the nucleus,aprocess that probably involves itsproteolytic cleavage by b and g secretases. Thus,overexpressionofAPP andmutations in APPand presenilinsmight disturb neuronal cell cyclearrest initiating AD pathogenesis. Reentry into the cell cycle is believedtobeanatural consequence of aging, however,innormal aging, brain neurons arrest in G1 phase. In contrast, AD neurons appear to undergo DNA replication (S phase)(Yang, Geldmacher, and Herrup 2001)and become trapped G2 as suggested by the aberrant expression of G2-specific cell cycleregulatingproteins in AD brain (Nagy2005). Although the mechanismofarrest in G2 is unclear, it presumablyoccurs becauseneurons are incapable of completing the cell cycle by undergoingmitosis. In this alternate hypothesis, the apparent failure of the G1/S transition checkpointisbelieved to be the essential pathogenicevent in AD. G2 phase is characterizedbythe activation of specific cyclin-dependent kinases (cdk) that ensure the progression through G2 to mitosis. The activation of G2-specific cdks is associated with agradual destabilizationofthe microtubule (MT) cytoskeleton. In neurons, MTs are essential for axonal transport. Thus, cellcycleactivation is aplausible cause of the axonopathy and transport deficits that are observed early in AD pathogenesis. Moreover,transport deficits inhibitanterograde and retrograde transportofAPP, causing the buildupofAPP in the neuronal cell body and an increaseinAb 42 generation (Stokin et al. 2005). Neurons in AD brain upregulate the expression of proteininhibitorsofthe cell cycle, such as glycogen synthase kinase-3(reviewed in Bhat, BuddHaeberlein, andAvila 2004)and cyclin- dependent kinase inhibitors (Arendt et al. 1996;reviewed in Nagy 2005). It is suggested that this phenomenon is caused by the cellular stress incurred by the inabilityofneurons in G2 to undergo mitosis and is presumably intendedtoinhibitprogression to mitosis.However, there are likelyside effects of these changesingeneexpression. For example, glycogensynthase kinase-3could phosphorylate the soluble tau protein, which has been released from depolymerizingMTs, thus promoting developmentofNFTs. Indeed,anumberofstress-activatedkinasesare capable of phosphorylating tau protein (Lovestone andReynolds1997) andmay also promotethe development NFTs. In the alternative hypothesis, the pathogenicevent is irreversible, because the neuronisleft with no mechanism for progressing to mitosis or returning to G0 phase.Rather it progressively degen- erates as axonopathy andtransportdeficits worsen andAb 42 and hyperphosphorylation of tau proteinbuild up eventually forming neuritic plaques and NFTs. Thealternative hypothesis of AD pathogenesisintroduces amechanism by which external events,suchasexposure to airpollutants, couldaffectADpathogenesis.Perturbations of the neuronal microenvironment, such as toxicant-induced oxidativestress or inflammation,could cause enough damage to elicitreentry into the cell cycle. In other words, damage to aneuronal networkthat retains synapticremodeling ability could present amitogenicstimulus and triggerthe development of AD. Alternatively,both toxicant-induced brain oxidative stress and inflammation could accelerate the consequencesofcell cyclereentry by neurons. The “two hit hypothesis” of AD pathogenesis postulates that both cellcycle reentry and oxidative stress are necessary for the development of Alzheimer’s disease (Zhu et al. 2004). This suggestion seems reasonable given that neurons trapped in G2 are probably more vulnerable insults. Thus external factors that cause oxidative stress or inflammation couldsignificantly accelerate AD pathogenesis. Air Pollution and Human Brain Pathology 333 © 2007 by Taylor & Francis Group, LLC 19.3 ALZHEIMER’S DISEASE PATHOGENESIS AND COX2 Vane(1971) showed that the anti-inflammatoryaction of nonsteroidal anti-inflammatory drugs (NSAIDs) dependsontheir ability to inhibit cyclooxygenases, which in turn results in adiminished synthesis of prostaglandins. In the early 1990s, cyclooxygenases were showntoexist as at least two distinct isoforms: COX1 and COX2. COX2has emergedasthe isoform that is primarily responsible for the synthesis of the prostanoids involved in acute and chronic inflammatory states (Hinz and Brune 2002). Underbasal conditions, COX2 has limited constitutive neuronal distribution in the CNS and contributes to synaptic activityand memory consolidation (Breder, Dewitt, and Kraig 1995;Breder and Saper 1996;Minghetti 2004). COX2 expression is induced by various proinflammatorystimuli, such as cytokines, growth factors, and tumor promoters (Hinz and Brune 2002;Minghetti 2004). Age is the most important risk factor for development of AD. The number of peoplewith the disease doubles every 5yearsbeyond age 65. Oxidativestress is one of the crucial factorsparti- cipating in the agingprocess. Cyclooxygenase-derived reactive oxygen species (ROS) generation increases with age, and cyclooxygenase-mediated prostanoid synthesis is oneofthe major sources of ROS in the aging process (Kim et al. 2000). Transgenic mice overexpressing humanCOX2 in hippocampalneurons developneuronalapoptosis andcognitivedeficitsinanage-dependent manner, and are moresusceptibleto b -amyloid toxicity with potentiation of redox impairment (Hoetal. 1999;Bazan 2001;Bazan and Lukiw 2002). COX2expression in hippocampal neurons may be apredictor of early AD (Hoetal. 2001)and chronic increased COX2production in brain may have anumber of consequences, including free radical mediated cellular damage, vascular dysfunction, alterations in cellular metabolism and neuronal cell cycle, and increases in total A b content (Naslund et al. 2000;Strauss et al. 2000;Hoetal. 2001;Xiang et al. 2002a;Scali et al. 2003). COX2influences processing of APP and promotes amyloid plaque deposition in amouse modelofAD(Xiang et al. 2002b). That COX2 playsarole in AD pathogenesis is also supported by epidemiological studies showing an association between long-term use of NSAIDs and areduced risk of developing AD (Aisen 2002), by the protectiveeffects of COX2 inhibitors in models of AD (Giovannini et al. 2003), and by gene expression profiling showing significantly upregulated stress induced proteins, including COX2, in humanbrain (Lukiw 2004). 19.4 THE MEXICO CITY ENVIRONMENT Mexico City represents an extreme of urban growth and environmental pollution (Chowetal. 2004). It is amegacity that covers an area of 2000 km 2 surrounded by aseries of volcanicand discontinuousmountain ranges that limitthe natural ventilation of the basin.The basin has more than 30,000 industrialfacilities and 3.5 million vehicles, with an estimated annual emission of 2.6 million tons of particulate and gaseous air pollutants. The critical air pollutants are ozone (O 3 )and particulate matter (PM). Figure19.1 illustrates 8hO 3 concentrations observed during January 2005 at three monitoring stations. The higherconcentrations of ozone are registered in the SW, down- wind fromthe areas wherethe ozone precursors areproduced.Figure19.2 showsthe annual concentrations of PM 2.5 micrometers or less in aerodynamic diameter (PM 2.5 )observed in NE, SW and downtown Mexico City. Pollutant levels in Mexico City vary within arelatively narrow range throughout the year, so its residents are exposed all year long to significant burden air pollutants. The pollution levels have been sustained or worsenedinthe last 20 years (Bravo and Torres 2002), so exposures of current children and teenagersare truly lifelong, having begun in utero. Moreover, thereisarelatively low mobility of Mexico City residents, so individualstend to be exposedtothe same environment for a prolonged period. Thus Mexico City presents an opportunity to studychronic health effects associ- ated with prolonged year-round exposures to severeair pollution. Particle Toxicology334 © 2007 by Taylor & Francis Group, LLC Merced Xalostoc 0.150 8-h standard 1234567 Local standard time 8910 111213141516171819 202122 2423 0.120 0.090 Ozone 8-h average (ppm) Ozone 8-h average (ppm) 0.060 0.030 0.000 Pedregal 0.150 8-h standard 1234567 Local standard time 8910 111213141516171819 202122 2423 0.120 0.090 0.060 0.030 0.000 Ozone 8-h average (ppm) 0.150 8-h standard 1234567 Local standard time 8910 111213141516171819 202122 24 0 10 20 km 23 0.120 0.090 0.060 0.030 0.000 FIGURE 19.1 Spatial and temporal profile of O 3 air pollution in metropolitan Mexico City. The P 50 ( D ), and the monthly diurnal average of 8hO 3 concentrations ( , ) observed during January 2005 are illustrated at three representative monitoring stations: Xalostoc, located in anortheast industrial area; Merced, located downtown; and Pedregal, aresidential area located in the southwest. The higher concentrations of ozone are registered in Pedregal, downwind of the areas where the ozone precursors are produced. Air Pollution and Human Brain Pathology 335 © 2007 by Taylor & Francis Group, LLC 1 0 50 PM 2.5 ( μ g/m 3 ) 100 150 200 250 300 350 400 Coyoacan Merced San agustin 234567 Local standard time 8910 1112 131415 16171819 202122 24 0 10 20 km 23 1234567 Local standard time 8910111213141516171819 202122 2423 1234567 Local standard time 8910111213141516171819 202122 2423 0 50 PM 2.5 ( μ g/m 3 ) 100 150 200 250 300 350 400 0 50 PM 2.5 ( μ g/m 3 ) 100 150 200 250 300 350 400 FIGURE 19.2 Spatial and temporal profile of PM 2.5 air pollution in metropolitan Mexico City. Typically, the highest PM 2.5 concentrations are observed in San Agustin in the northeast and Merced downtown, with the lowestlevels in Pedregal in the southwest. PM 2.5 annual concentrations are above the annual standard for all three stations. Particle Toxicology336 © 2007 by Taylor & Francis Group, LLC The potentially toxic components of PM air pollution include acids, polyaromatic hydrocar- bons, metals, biological products such as lipopolysaccharide (LPS), and inorganiccarbonparticles. It is suspected that O 3 ,polyaromatichydrocarbons, transition metals,and inorganiccarbon particleshave acommon mechanism of toxicity—the depletion of cellular anti-oxidant defenses (cellularoxidative stress) through the generation of ROS (i.e., hydrogen peroxide, superoxide, hydroxyl radical, and singlet oxygen).However, the mechanism by which ROS are elicitedvaries significantly with the pollutant. The pollutant-induced oxidative stress probably activates stress- response signal transduction pathways by the activation or inactivation of oxidant-sensitive com- ponents of these pathways (e.g., transcription factors and phosphatases). Some pollutants, such as the transition metal vanadium (a potent phosphatase inhibitor itself), may activate these pathways by ROS-independentmechanisms as well. The stress-responsepathways regulate the expression of genes encoding antioxidant synthesis and reducing enzymes,detoxificationenzymes,and they also regulate immune, inflammatory, cell survival, and apoptotic responses.LPS activates many of the same pathways by acting through aspecificreceptorcomplex composed of theLPS binding proteinCD14and aheterodimeric plasma membrane proteincomplexcomposedofToll-like receptor-4 (TLR4) and MD2. Chronicactivation of the stress response pathways culminates in achronic inflammatory response in targeted tissues. We have focused on toxicity due to metals and LPS. The mostabundant metalsinMexico City PM are: Ca, Fe, K, Zn, and Pb, while metalstypically present in motor vehicle exhaust and fuel oil combustion products, Cr, Ni, Vand S, are present in lower concentrations (Chow et al. 2002). LPS content in PM samples show arange of 15.3 to 20.6 ng/mg of PM and SE samples show the highest endotoxin concentrations 59 EU/mg PM (Alfaro-Moreno et al. 2002;Osornio-Vargas et al. 2003). 19.5 COX2 AND IL-1b EXPRESSION, A b 42 ACCUMULATION, AND NEUROPATHOLOGY IN THE BRAINS OF DOGS AND HUMANS EXPOSED TO SEVERE URBAN AIR POLLUTION Healthy Mexico City dogs experience chronic upper and lower respiratory tract inflammation and breakdown of both therespiratory andolfactoryepithelialbarriers. Thebrainsofthese dogs exhibited endothelial and astrocytic upregulation of cycloxygenase-2 (COX2) expression in olfac- tory bulb, frontal cortex,and hippocampus,three critical targets in Alzheimer’s disease (Caldero ´ n-Garciduen ˜ as et al.2003a).There wasalsoactivation of neuronal NF- k B, increased induced nitricoxide synthase (iNOS)expression in cortical endothelial cells as early as 4weeks of age, and breakdownofthe blood brain barrier (BBB) (Caldero ´ n-Garciduen ˜ as et al. 2002). iNOS derived nitric oxide (NO) contributes to thegenerationofperoxynitrite andBBB breakdown (Winkler 2001),leadingtovasogenicedema andsecondary braindamage(Thiel andAudus 2001;Chao et al. 1992). Thus the increase in iNOS expression may be related to the breakdown of theBBB.Furthermore,NOmay also contribute to themaintenance,self-perpetuation, and progression of neurodegenerative processes (Grammasetal. 1997). Olfactory bulband hippocampalapurinic/apyrimidinic sitesingenomic DNA were signi- ficantlyhigherinexposedMexicoCitydogs versuscontrols (Caldero ´ n-Garciduen ˜ as et al. 2003a), suggestive of increased oxidative stress.There was significant DNA damage in the olfac- tory bulb of young Mexico City dogs several months before cortical A b 42 diffuse plaques were detected (Caldero ´ n-Garciduen ˜ as et al. 2003a). In Mexico City dogs olfactory mucosa pathology appeared by 4months of age and thus appears to be aprecursorofthe olfactory bulb pathology (Caldero ´ n-Garciduen ˜ as et al. 2003a). Even young dogs ( ! 1year) showed accumulation of A b 42 in neurons, glial cells and blood vessels,and the presence of A b 42-positive diffuse plaques. Dogs from Mexico City exhibited white matter perivascular gliosisasearly as 3moofage and astrogliosis increased significantly with age (Figure 19.3). Air Pollution and Human Brain Pathology 337 © 2007 by Taylor & Francis Group, LLC These observations were consistent with an acceleration of an Alzheimer’s-like pathologyin apparently healthy dogs. Theforegoing studies usingdogs as sentinels suggested that chronic exposure to severe air pollution might have adverse effects on human brain. To examinethis possibility, we conducted a studyusing autopsy brain samples from Mexican subjects, all lifelong residents of two large cities with severe air pollution, Mexico City and Monterrey, and five small cities with low levels of air pollution. Evidence of chronic respiratory tract inflammation was present in all residents of cities with severeair pollution. COX2mRNA abundance was measured by real-time RT-PCR analysis of total RNA isolated from brain tissues. There was asignificant elevation of COX2 mRNA levels in frontal cortex and hippocampus of the high exposure group (Figure 19.4aand d), along with an elevation of COX2 immunoreactivity in frontal cortex confirmed by quantitative image analysis of COX2immuno- reactivity (IR) (Figure 19.4b). In subjects from the low exposure group, COX2IRwas confined to neuronal cell bodies, whereas subjects from the high exposure group exhibited COX2staining in neuronal cell bodies and dendrites, as well as strong COX2 staining of the endothelium in the frontal cortex(Figure 19.4c). COX2IRwas largely confinedtoneurons in thehippocampus (Figure 19.4f). There was astrong positive association betweenCOX2mRNAlevels and oxidative DNA damage as measured by apurinic/apyrimidinic (AP)sites ( r Z 0.89, p Z 0.001) in frontal cortex (Caldero ´ n-Garciduen ˜ as et al. 2004). The positivecorrelation betweenCOX2 mRNA and AP sites couldbeaconsequence of COX2-mediated prostanoid synthesis, amajor source of ROS that are capable of damaging DNA. The DNA damage in frontal cortex suggests that oxidative stress couldbearelevantand early event.Oxidativedamage is an early event in AD, and it is greatest early in the disease and decreases with disease progression (Nunomuraetal. 2001;Perry et al. 2002). In normal brain expression, the 695 amino acid form of APP (APP695) is much greater than the expression of the 751 amino acid form (APP751). AD brain is characterizedbyareversal in the relative expression of APP isoforms as APP751 expressionincreases dramatically. We measured the APP751/APP695 mRNA ratio by real-time RT-PCR in frontal cortex and hippocampus. There FIGURE 19.3 ( See color insert)Reactive gliosis and astrocytic proliferation in the frontal cortex white matter of healthy Mexico City dogs. Glial cells and proliferating astrocytes were localized in paraffin sections of frontal cortex of Mexico City dogs by immunohistochemistry using fluorescein-labeled anti-glial fibrillary acidic protein (GFAP, green) and phycoerythrin-labeled anti-bromodeoxyuridine (BrdU, red), respectively, and examined by confocal microscopy: (a) 3-year-old male, (b) 5-year-old female, and (c) 14-year-old female. Gliosis worsens with age. Images represent maximum intensity projections, showing the maximum intensity of all layers alongthe viewing direction. The insertsrepresent3Dreconstructions fromthe same data sets. (Pictures were taken by Dr. Barbara Rothen-RutishauserPh.D. Institute of Anatomy, University of Bern, Bern, Switzerland.) Particle Toxicology338 © 2007 by Taylor & Francis Group, LLC was no statistically significant difference between the APP751/APP695 ratios in the high and low exposure groups. However, there was asignificant positive correlation between COX2mRNAin frontal cortex and the APP751/APP695 ratio in the frontal cortex of the high exposure group only (Caldero ´ n-Garciduen ˜ as et al. 2004). A b 42 accumulation in frontal cortex and hippocampus was measured by quantitative immu- nohistochemistry.Ab 42 was detected in the perikaryon of pyramidal frontal cortex neurons and in cortical and white matter astrocytes (Figure 19.5a) and subarachnoid and cortical blood vessels (Figure 19.5b) in subjects from the high exposure group. A b 42 accumulation in the frontal cortex (Figure19.5c)and hippocampus(Figure19.5d) of thehighexposuregroup wassignificantly elevated comparedtothe low exposure group. Three subjects in the high exposure group had rare diffuse A b 42 plaque-like staining in the frontal cortex of (32, 38, and 43 yearsold). The diffuse A b 42 plaques were associated with reactive astrocytes (e.g., Figure 19.5e) or apoptotic nuclei (not shown). None of the three subjects carried the apolipoprotein E 3 4allele (Caldero ´ n- Garciduen ˜ as et al. 2004), arisk factor for the development of Alzheimer’s disease. In afollow-up autopsystudy, theolfactorybulbwas examined forevidenceofCOX2 expression and A b 42 accumulation. In accordancewith findings in dogs, there was asignificant up-regulation of COX2 and IL-1b mRNAexpression and A b 42 accumulation in the olfactory bulb of highlyexposedsubjects comparedtocontrols(Figure 19.6). A b 42 accumulation in olfactory bulb was also seen in arterial smooth muscle cells starting as early as the seconddecade of life (Figure 19.7). (f) (c) 0 1 2 3 4 5 * Low High COX2 /18s rRNA (x 10 2 molec. /fmol) Low High 0.0 0.1 0.2 0.3 * COX2 IR (% pos.) 0 1 2 3 4 * Low High COX2 /18s rRNA (x 10 2 molec. /fmol) Low High 0 1 2 3 4 5 COX2 IR (% pos.) (a) (b) (d) (e) FIGURE 19.4 ( See color insert)COX2 expression in frontal cortex (a–c) and hippocampus (d–f). COX2 mRNA abundancewas measured by RT-PCR andnormalizedfor 18srRNAlevels. COX2 proteinwas localized in sections of paraffin-embedded tissues by IHC and its abundance was measured by quantitative image analysis. COX2 mRNA was significantly elevated in the high exposure group in both frontal cortex (a, p Z 0.009), and hippocampus (d, p Z 0.04). COX2 immunoreactivity (IR) was significantly elevated in the high exposure group in frontal cortex (b, p Z 0.01), but not in hippocampus (e). Means G SEMs are shown in A, B, D, and F. (c) Representative COX IHC in frontal cortex from asubject in the high exposure group showing strong staining of endothelial cells in the capillaries (*), and pyramidal neurons (arrow), while other neurons were negative (arrowheads). ScaleZ 20 m m. (f) Representative COX IHC in dentate gyrus from a subject in the high exposure group showing COX2 positive neurons (arrowheads) and capillaries (short arrow). Scale Z 15 m m. (Caldero ´ n-Garciduen ˜ as,L.etal.,Brain inflammationand Alzheimer’s-likepathology in individuals exposed to severe air pollution, Toxicol. Pathol.,32, 650–658, 2004. With permission.) Air Pollution and Human Brain Pathology 339 © 2007 by Taylor & Francis Group, LLC Themajor neuropathological findingsinexposedhumans included: (1) breakdown of the BBB, as indicated by the presence extravascular red blood cells, hemosiderin-ladenmacrophages, reac- tive astrocytes, and apoptotic nuclei (Figure 19.8aand Figure19.8c), (2) age-related progressive reactive gliosisinthe supratentorial white matter (GFAP-positive cells) (Figure19.8b), and (3) 0 A β 42 IR (% pos.) Low Frontal cortex (c) (d) Hippocampus High Low High 5 10 15 10 * * 7 5 2 0 (a) (b) p p * * * (e) FIGURE 19.5 ( See color insert)Ab 42 accumulation in frontal cortex and hippocampus. A b 42 was localized in sections of paraffin-embedded tissues by IHC. (a) Anti-Ab 42 stained pyramidal neurons (p), astrocytes (arrows) and astrocytic processes (arrowheads) around blood vessels (*). (b) In addition to accumulation in pyramidal neurons, (p) A b 42 was deposited in smooth muscle cells (arrows) in cortical arterioles (*). Adead neuron surrounded by glial cells is indicated (arrowhead). (c and d) Quantitative image analysis of A b 42 IHC showed asignificant increase in A b 42 immunoreactivity (Ab 42 IR) in both frontal cortex (c, * p Z 0.04) and hippocampus (d, * p Z 0.001) in the high exposure group. MeansG SEMs are shown. (e) A b 42 IHC of frontal cortex from a38-year-old subject from Mexico City showing diffuse plaque-like staining with surrounding reactive astrocytes (arrows). Scale Z 20 m m. (Caldero ´ n-Garciduen ˜ as,L.etal., Brain inflammationand Alzheimer’s-like pathology in individualsexposed to severe air pollution, Toxicol. Pathol.,32, 650–658, 2004. With permission.) Low High 0.01 0.1 1 10 COX2 mRNA (×10 6 molec /fmol 18s rRNA) Low High 0.1 1 10 IL-1β mRNA (×10 5 Eq /fmol 18s rRNA) Low High 1 2 4 8 16 32 A β 42 immunoreactivity (arbitrary units) FIGURE 19.6 COX2, IL-1b and A b 42 expression in olfactory bulb of low versus high exposed subjects. COX2 and IL1b mRNA abundances were measured by RT-PCR and normalized for 18 srRNA. A b 42 protein was localized in paraffin-embedded tissues by IHC and its abundance was measured by quantitative image analysis. COX2 and IL-1b mRNA were significantly elevated in the high exposure group ( p Z 0.002 and 0.024, respectively). Highly exposed subjects showed asignificant increase in A b 42 immunoreactivity as well. Particle Toxicology340 © 2007 by Taylor & Francis Group, LLC [...]... damage Mexico City children had more serum IL-10 and IL-6 and less IL-8 and IL-1b than age/gender/socioeconomic matched controls, indicating a shift in ´ ˜ the circulating cytokine profile in favor of anti-inflammatory cytokines (Calderon-Garciduenas et al 2003b) These children also exhibited significantly higher plasma concentrations of endothelin-1 (ET-1) (Calderon-Garciduenas et al 2005) and PGE2, and increased... a b-sheet-like conformational change in Ab resulting in enhanced aggregation (Bossy-Wetzel, Schwarzenbacher, and Lipton 2004) Further, Ab is able to potentiate the ability of metal ions, including Fe, Cu, and Al, to generate ROS (Bondy, Guo-Ross, and Truong 199 8) 19. 7.3 LPS TOXICITY Clinical studies of Mexico City children (Section 19. 6) showed a significant increase in the expression of the LPS-binding... children who died suddenly showed significant white matter blood vessel leakage and perivascular gliosis (Figure 19. 8) that could explain the hyperintense frontal T2-weighted images (insert Figure 19. 9 the MR1 picture) FIGURE 19. 9 T1-weighted coronal brain magnetic resonance image of a 14-year-old girl, a life long resident of Mexico City, showing mild cortical frontal atrophy (arrow) and increased volume... thank Dr Ricardo Torres-Jardon from the Universidad National Autonoma de Mexico for Figures 19. 1 and Figure 19. 2 REFERENCES Aisen, P S., Evaluation of selective COX-2 inhibitors for the treatment of Alzheimer’s disease, J Pain Symptom Manage., 23, S35–S40, 2002 Alfaro-Moreno, E., Martinez, L., Garcia-Cuellar, C., Bonner, J C., Murray, J C., Rosas, I., Rosales, S P., and Osornio-Vargas, A R., Biologic... Toxicol Pathol., 31, 524–538, 2003a ´ ˜ Calderon-Garciduenas, L., Mora-Tiscareno, A., Fordham, L A., Valencia-Salazar, G., Chung, C J., Rodriguez-Alcaraz, A., Paredes, R et al., Respiratory damage in children exposed to urban pollution, Pediatr Pulmonol., 36, 148–161, 2003b ´ ´ ˜ Calderon-Garciduenas, L., Reed, W., Maronpot, R R., Henriquez-Roldan, C., Delgado-Chavez, R., Calderon˜ Garciduenas, A., Dragustinovis,... cortex was stained with hematoxylin and eosin (a) and anti-GFAP (b and c) (a) There are numerous perivascular macrophages with hemosiderin-like granules and free RBC surrounding a blood vessel A neuron is seen in the lower left corner (b) Same 11-year-old child showing reactive gliosis in subcortical frontal white matter (GFAP) (c) A 17-year-old Mexico City male with reactive astrocytes around cortical... Fassbender et al 2004) 19. 7.2 TRANSPORT OF PM-ASSOCIATED METALS TO THE BRAIN It is well documented that metals instilled into the nasal cavity or inhaled ultrafine carbon particles ¨ accumulate in olfactory bulb in rodents (Sunderman 2001; Dorman et al 2002; Oberdorster et al 2004) Transport of metals and ultra-fine particles into cortical areas in rodents has been shown as ¨ well (Henriksson 199 7; Dorman 2002;... Expression of the cyclin-dependent kinase inhibitor p16 in Alzheimer’s disease, Neuroreport, 7, 3047–3049, 199 6 © 2007 by Taylor & Francis Group, LLC Air Pollution and Human Brain Pathology 347 Bazan, N G., COX-2 as a multifunctional neuronal modulator, Nat Med., 7, 414–415, 2001 Bazan, N G and Lukiw, W J., Cyclooxygenase-2 and presenilin-1 gene expression induced by interleukin-1b and amyloid b42 peptide... J Neurochem., 89, 1313–1317, 2004 Bondy, S C., Guo-Ross, S X., and Truong, A T., Promotion of transition metal-induced reactive oxygen species formation by b-amyloid, Brain Res., 799, 91–96, 199 8 Bossy-Wetzel, E., Schwarzenbacher, R., and Lipton, S A., Molecular pathways to neurodegeneration, Nat Med., 10 (suppl.), S2–S9, 2004 Bravo, H A and Torres-Jardon, R., Air pollution levels and trends in the... pathology, Toxicol Sci., 61, 356–367, 2001 ´ ˜ Calderon-Garciduenas, L., Azzarelli, B., Acuna, H., Garcia, R., Gambling, T M., Osnaya, N., Monroy, S et al., Air pollution and brain damage, Toxicol Pathol., 30, 373–389, 2002 ´ ´ ´ ˜ Calderon-Garciduenas, L., Maronpot, R R., Torres-Jardon, R., Henrıquez-Roldan, C., Schoonhoven, R., ´ ˜ Acuna-Ayala, H., Villarreal-Calderon, A et al., DNA damage in nasal and brain . perivascular gliosis(Figure 19. 8)thatcouldexplain thehyperintensefrontal T 2 -weightedimages(insert Figure19.9 the MR1 picture). FIGURE 19. 9 T 1 -weighted coronal brain magnetic resonance image of a14-year-old girl,. of inhaled ultrafine particles to the brain, Inhal. Toxicol.,16, 437–445, 2004. Osornio-Vargas, A. R., Bonner, J. C., Alfaro-Moreno, E., Martinez, L., Garcia-Cuellar, C., Ponce-de-Leon- Rosales, S.,. hemosiderin-ladenmacrophages, reac- tive astrocytes, and apoptotic nuclei (Figure 19. 8aand Figure19.8c), (2) age-related progressive reactive gliosisinthe supratentorial white matter (GFAP-positive