Kathleen S. Montine

Dendritic degeneration in neostriatal medium spiny neurons in Parkinson disease

Dysfunction of neostriatal medium spiny neurons (MSNs) is hypothesized to underlie late-stage motor complications of Parkinson disease (PD). The authors demonstrate shortened dendrite length of MSNs that was similar in four regions of neostriatum in late-stage PD. In contrast, MSN dendrite spine degeneration was unevenly distributed with the greatest loss in caudal putamen. The authors propose that these structural changes in MSN may contribute to late-stage motor complications of PD.

4-hydroxy-2-nonenal pyrrole adducts in human neurodegenerative disease.

Increasing age and inheritance of the ∈4 allele of apolipoprotein E (APOE4) are significant risk factors for sporadic and late onset familial Alzheimer disease (AD); however, the mechanisms by which either leads to AD are unknown. Numerous studies have associated advancing age with increased indices of oxidative challenge to brain, and with still further increased oxidative damage to relevant brain regions in AD patients. A major consequence of oxidative damage to brain is lipid peroxidation with production of the neurotoxic metabolite 4-hydroxy-2-nonenal (HNE). HNE reacts with protein to yield several adducts, including a pyrrole adduct that forms irreversibly in biological systems. Previously, we have shown in a small number of AD and control patients that HNE pyrrole adduct antiserum is immunoreactive with neurofibrillary tangles (NFT), and that this reactivity was significantly associated with inheritance of APOE4. Others have confirmed this pattern of immunoreactivity in AD brain but did not observe an association with APOE4. Herein, we have expanded the study group to 19 AD patients homozygous for APOE4 or APOE3, as well as 30 patients with other neurodegenerative diseases, including diffuse Lewy body disease, Picks disease, progressive supranuclear palsy, Parkinsons disease, and human immunodeficiency virus-1 encephalitis. HNE pyrrole adduct immunoreactivity on NFT in AD patients was strongly associated with APOE4 homozygosity. With the exception of rare immunoreactive Pick bodies in one case of Picks disease, no other structure was recognized by HNE pyrrole adduct antiserum in this series of patients. We propose that there is a significant difference between the interaction of apoE3 and apoE4 with lipid peroxidation in the brains of AD patients.

Distribution of reducible 4-hydroxynonenal adduct immunoreactivity in Alzheimer disease is associated with APOE genotype

Two major risk factors for late-onset familial and sporadic Alzheimer disease (AD), a leading cause of dementia worldwide, are increasing age and inheritance of the ϵ4 allele of the apolipoprotein E gene (APOE4). Several isoform-specific effects of apoE have been proposed; however, the mechanisms by which apoE isofoems influence the pathogenesis of AD are unknown. Also associated with AD is increased lipid peroxidation in the regions of the brain most damaged by disease. 4-hydroxynonenal (HNE), the most potent neurotoxic product of lipid peroxidation, is thought to be deleterious to cells through reactions with protein nucleophiles. We tested the hypothesis that accumulation of the most common forms of HNE-protein adducts, borohydride-reducible adducts, is associated with AD and examined whether there was a relationship to APOE. Our results demonstrated that reducible HNE adducts were increased in the hippocampus, entorhinal cortex, and temporal cortex of patients with AD. Furthermore, our data showed that the pattern of reducible HNE adduct accumulation was related to APOE genotype; AD patients homozygous for APOE4 had pyramidal neuron cytoplasmic accumulation of reducible HNE adducts, while AD APOE3 homozygotes had both pyramidal neuron and astrocyte accumulation of reducible HNE adducts. This is in contrast to our previous observations that a distinct HNE protein adduct, the pyrrole adduct, accumulates on neurofibrillary tangles in AD patients. We conclude that APOE genotype influences the cellular distribution of increased reducible HNE adduct accumulation in AD.

Proteomic analysis of neurofibrillary tangles in Alzheimer disease identifies GAPDH as a detergent-insoluble paired helical filament tau binding protein

We performed proteomic analysis of neurofibrillary tangles (NFTs) obtained by laser capture microdissection from pyramidal neurons in hippocampal sector CA1 in patients with Alzheimer disease (AD) using liquid chromatography (LC)‐mass spectrometry (MS)/MS. We discovered a total of 155 proteins in laser captured NFTs, 72 of which were identified by multiple unique peptides. Of these 72 proteins, 63 had previously unknown association with NFTs; one of these was glyceraldehyde‐3‐phosphate dehydrogenase (GAPDH). We validated by immunohistochemistry that GAPDH colocalized with the majority of NFTs as well as plaque‐like structures in AD brain and was co‐immunoprecipitated by antibodies to abnormal forms of tau in AD, but not tau from AD temporal cortex. Characterization of GAPDH showed that it, along with phosphorylated tau and Aβ peptides, was present in detergent‐insoluble fractions from AD temporal cortex but not from age‐matched controls. These data are the first proteomic investigation of NFTs. Moreover, our results validate this approach by demonstrating that GAPDH, a glycolytic and microtubule binding protein, not only co‐localized to NFTs and immunoprecipitated with PHF‐tau, but also is one of the few proteins known to undergo conversion to a detergent‐insoluble form in AD.

Cerebrospinal Fluid Aβ42, Tau, and F2-Isoprostane Concentrations in Patients With Alzheimer Disease, Other Dementias, and in Age-Matched Controls

Abstract Objective.—To test the hypothesis that quantification of cerebrospinal fluid (CSF) F2-isoprostanes (F2-IsoPs), in vivo biomarkers of free radical damage, along with CSF Aβ42 and tau levels...

Ecology of the Aging Human Brain

BACKGROUND Alzheimer disease, cerebral vascular brain injury, and isocortical Lewy body disease (LBD) are the major contributors to dementia in community- and population-based studies. OBJECTIVE To estimate the prevalence of clinically silent forms of these diseases in cognitively normal (CN) adults. DESIGN Autopsy study. SETTING Community- and population based. PARTICIPANTS A total of 1672 brain autopsies from the Adult Changes in Thought study, Honolulu-Asia Aging Study, Nun Study, and Oregon Brain Aging Study, of which 424 met the criteria for CN. MAIN OUTCOME MEASURES Of these, 336 cases had a comprehensive neuropathologic examination of neuritic plaque density, Braak stage for neurofibrillary tangles, LB distribution, and number of cerebral microinfarcts. RESULTS Forty-seven percent of CN cases had moderate or frequent neuritic plaque density; of these, 6% also had Braak stage V or VI for neurofibrillary tangles. Fifteen percent of CN cases had medullary LBD; 8% also had nigral and 4% isocortical LBD. The presence of any cerebral microinfarcts was identified in 33% and of high-level cerebral microinfarcts in 10% of CN individuals. Overall, the burden of lesions in each individual and their comorbidity varied widely within each study but were similar across studies. CONCLUSIONS These data show an individually varying complex convergence of subclinical diseases in the brain of older CN adults. Appreciating this ecology should help guide future biomarker and neuroimaging studies and clinical trials that focus on community- and population-based cohorts.

Microglial EP2 is critical to neurotoxicity from activated cerebral innate immunity

Prostaglandin (PG) E2 acts via four functionally antagonistic E‐prostanoid (EP) receptors that are expressed on multiple cell types in the nervous system; these are designated EP1–4. We showed previously that EP2 null mice are protected from CD14‐dependent neuronal damage in vivo following intracerebroventricular (ICV) injection of lipopolysaccharide (LPS). Clear interpretation of this neuroprotective outcome is limited because EP2 is expressed on glia and neurons. We tested the hypothesis that microglial EP2 is required for paracrine neurotoxicity following activation of innate immunity, using primary murine microglia and neuron co‐cultures. We demonstrated that microglial EP2 was necessary for lipopolysaccharide (LPS)‐activated microglia‐mediated neurotoxicity, as well as induction of inducible nitric oxide synthase (iNOS) and cyclooxygenase 2 (COX‐2). Genetic deletion of microglial iNOS, pharmacological suppression of COX‐2 activity, or addition of exogenous superoxide dismutase (SOD) and catalase in the presence of EP2 also abolished neurotoxicity. This loss of paracrine neurotoxicity by EP2−/− microglia occurred in the absence of reduced cytokine levels. We conclude that microglial EP2 is critical to innate immunity‐mediated paracrine damage to neurons involving COX‐2 and iNOS. EP2 should be considered as a therapeutic target for suppression of microglial innate immunity‐mediated damage in neurodegenerative diseases.

Pharmacologic suppression of neuronal oxidative damage and dendritic degeneration following direct activation of glial innate immunity in mouse cerebrum

Activation of glial innate immunity is widely proposed to contribute to a number of degenerative and destructive diseases of brain. However, the precise role of activated innate immunity has been difficult to define in vivo because of multiple simultaneous pathogenic processes and responses to injury that confound interpretation of results from complex models of disease. Here, we used the model of intracerebroventricular (ICV) injection of lipopolysaccharide (LPS) to test the hypothesis that directly activated glial innate immunity leads to neurodegeneration in cerebrum and to establish the molecular determinants of and neuroprotectants from such innate immunity‐mediated neuronal damage. Our results showed that ICV LPS induced delayed, reversible oxidative damage to cerebral neuronal membranes as measured by F4‐neuroprostanes that was coincident with degeneration of the hippocampal pyramidal neuron dendritic system, but not neuron death, in adult mice. Both neuronal oxidative damage and dendritic degeneration were NF‐κB and iNOS dependent and were completely suppressed by ibuprofen and α‐tocopherol, but not naproxen or γ‐tocopherol. These results prove that activation of glial innate immunity can lead to neurodegeneration independent of other pathologic processes, closely associate oxidative damage to neuronal membranes with degeneration of the dendritic system, and provide a possible explanation for the varying efficacy of neuroprotectants that have been suggested in observational studies of dementia.

Prostaglandin E2 receptor subtype 2 (EP2) regulates microglial activation and associated neurotoxicity induced by aggregated α-synuclein

BackgroundThe pathogenesis of idiopathic Parkinsons disease (PD) remains elusive, although evidence has suggested that neuroinflammation characterized by activation of resident microglia in the brain may contribute significantly to neurodegeneration in PD. It has been demonstrated that aggregated α-synuclein potently activates microglia and causes neurotoxicity. However, the mechanisms by which aggregated α-synuclein activates microglia are not understood fully.MethodsWe investigated the role of prostaglandin E2 receptor subtype 2 (EP2) in α-synuclein aggregation-induced microglial activation using ex vivo, in vivo and in vitro experimental systems.ResultsResults demonstrated that ablation of EP2(EP2-/-) significantly enhanced microglia-mediated ex vivo clearance of α-synuclein aggregates (from mesocortex of Lewy body disease patients) while significantly attenuating neurotoxicity and extent of α-synuclein aggregation in mice treated with a parkinsonian toxicant 1-methyl-4-phenyl-1,2,3,6-tetrahydropyridine. Furthermore, we report that reduced neurotoxicity by EP2-/- microglia could be attributed to suppressed translocation of a critical cytoplasmic subunit (p47-phox) of NADPH oxidase (PHOX) to the membranous compartment after exposure to aggregated α-synuclein.ConclusionThus, it appears that microglial EP2 plays a critical role in α-synuclein-mediated neurotoxicity.

Effects of reactive gamma-ketoaldehydes formed by the isoprostane pathway (isoketals) and cyclooxygenase pathway (levuglandins) on proteasome function.

Oxidative stress can impair proteasome function, both of which are features of neurodegenerative diseases. Inhibition of proteasome function leads to protein accumulation and cell death. We discovered recently the formation of highly reactive γ‐ketoaldehydes, isoketals (IsoKs), and neuroketals (NeuroKs) as products of the isoprostane and neuroprostane pathways of free radical‐induced lipid peroxidation that are analogous to cyclooxygenase‐derived levuglandins (LGs). Because aldehydes that are much less reactive than IsoKs have been shown to inhibit proteasome function, we explored the ability of the proteasome to degrade IsoK‐adducted proteins/peptides and the effect of IsoK and IsoK‐adducted proteins/peptides on proteasome function. Adduction of IsoK to model proteasome substrates significantly reduced their rate of degradation by the 20S proteasome. The ability of IsoK to inhibit proteasome function directly was observed only at very high concentrations. However, at much lower concentrations, an IsoK‐adducted protein (ovalbumin) and peptide (Aβ1–40) significantly inhibited chymotrypsin‐like activity of the 20S proteasome. Moreover, incubation of IsoK with P19 neuroglial cultures dose‐dependently inhibited proteasome activity (IC50 = 330 nM) and induced cell death (LC50 = 670 nM). These findings suggest that IsoKs/NeuroKs/LGs can inhibit proteasome activity and, if overproduced, may have relevance to the pathogenesis of neurodegenerative diseases.