Showing papers by "John Q. Trojanowski published in 2003"

Initiation and synergistic fibrillization of tau and alpha-synuclein.

Abstract: Alpha-synuclein (alpha-syn) and tau polymerize into amyloid fibrils and form intraneuronal filamentous inclusions characteristic of neurodegenerative diseases. We demonstrate that alpha-syn induces fibrillization of tau and that coincubation of tau and alpha-syn synergistically promotes fibrillization of both proteins. The in vivo relevance of these findings is grounded in the co-occurrence of alpha-syn and tau filamentous amyloid inclusions in humans, in single transgenic mice that express A53T human alpha-syn in neurons, and in oligodendrocytes of bigenic mice that express wild-type human alpha-syn plus P301L mutant tau. This suggests that interactions between alpha-syn and tau can promote their fibrillization and drive the formation of pathological inclusions in human neurodegenerative diseases.

Cerebrospinal Fluid Tau and β-Amyloid: How Well Do These Biomarkers Reflect Autopsy-Confirmed Dementia Diagnoses?

Abstract: Background Tau and β-amyloid (Aβ) are proposed diagnostic biomarkers for Alzheimer disease (AD). Previous studies report their relationship to clinical diagnoses of AD and other dementias. To understand their value as predictors of disease-specific patholody, levels determined during life must be correlated with definitive diagnoses in mixed dementia groups and cognitively normal subjects. Objectives To correlate antemortem cerebrospinal fluid (CSF) tau and Aβ levels with definitive dementia diagnosis in a diverse group of patients; to calculate statistics for CSF tau and Aβ. Design Prospective study. Setting Ten clinics experienced in the diagnosis of neurodegenerative dementias. Patients One hundred six patients with dementia and 4 cognitively normal subjects with a definitive diagnosis, and 69 clinically diagnosed cognitively normal subjects. Main Outcome Measures Correlation of CSF tau and Aβ with final diagnosis. Results Mean tau level was 612 pg/mL for the 74 patients with AD, 272 pg/mL for 10 patients with frontal dementia, 282 pg/mL for 3 patients with dementia with Lewy bodies, and 140 pg/mL for 73 cognitively normal control subjects. Tau was less than 334 pg/mL for 20 patients with AD. Aβ 42 was reduced in patients with AD (61 fmol/mL) compared with patients with frontal dementia (133 fmol/mL) and control subjects (109 fmol/mL), but not compared with patients with dementia with Lewy bodies (14 fmol/mL) or prion disease (60 fmol/mL). Conclusions Elevated CSF tau levels are associated with AD pathology and can help discriminate AD from other dementing disorders. However, some patients with AD have a level less than the mean ± 2 SDs of the cognitively normal cohort.

Neurodegeneration and defective neurotransmission in a Caenorhabditis elegans model of tauopathy

Abstract: Frontotemporal dementia with parkinsonism chromosome 17 type (FTDP-17) is caused by mutations in MAPT, the gene encoding tau. FTDP-17 begins with executive function deficits and other abnormal behaviors, which progress to dementia. Neurodegenerative changes include accumulation of aggregated tau as neuronal and glial fibrillary tangles. Aggregated tau is seen in numerous other neurodegenerative diseases, including Alzheimer's disease (AD). We expressed normal and FTDP-17 mutant human tau (mutations P301L and V337M) in Caenorhabditis elegans to model tauopathy disorders. Tau pan-neuronal expression caused progressive uncoordinated locomotion (Unc), characteristic of nervous system defects in worms. Subsequently, insoluble tau accumulates and both soluble and insoluble tau is phosphorylated at many of the sites hyperphosphorylated in FTDP-17, AD, and other tauopathies. Substantial neurodegeneration, seen as bulges and gaps in nerve cords followed by loss of neurons, occurs after insoluble tau begins to accumulate. Axons show vacuoles, membranous infoldings, and whorls with associated amorphous tau accumulations and abnormal tau-positive aggregates. FTDP-17 mutation lines had a more severe Unc phenotype, accumulated more insoluble tau at a younger age, were more resistant to cholinergic inhibitors, and had more severe axonal degeneration when compared with lines expressing normal tau. The Unc phenotype is caused by a presynaptic defect. Postsynaptic transmission is intact. This transgenic model will enable mechanistic dissection of tau-induced neurodegeneration and identification of genes and compounds that inhibit pathological tau formation.

Amyloid-beta is found in drusen from some age-related macular degeneration retinas, but not in drusen from normal retinas.

Abstract: Purpose: Age-related macular degeneration (AMD) is the most common cause of irreversible vision loss in the elderly. Increased understanding of the pathogenesis is necessary. Amyloid-beta (Aβ), a major extracellular deposit in Alzheimer’s disease plaques, has recently been found in drusen, the hallmark extracellular deposit in AMD. The goal of this study was to characterize the distribution and frequency of Aβ deposits in drusen from AMD and normal post mortem human retinas to gain additional insight about the potential role of A β in AMD pathogenesis. Methods: Immunocytochemistry was performed with three Aβ antibodies on sections from 9 normal and 9 AMD (3 early, 3 geographic atrophy, 3 exudative AMD) retinas. Five sections from each eye were evaluated. Aβ positive deposits in drusen were identified using epifluorescence and confocal microscopy. Antibodies were pre-adsorbed with Aβ peptide to verify specificity. Some sections were stained with PAS-hematoxylin to aid in evaluation of morphology. Results: To test and optimize immunocytochemistry, Aβ was detected in amyloid plaques from Alzheimer’s brains. Aβ label was blocked by pre-adsorption of antibody with Aβ peptide, verifying specificity. Four of the 9 AMD retinas and none of the 9 normal retinas had Aβ positive drusen. Two of the early AMD eyes had a few Aβ positive drusen, each with a few Aβ-containing vesicles, and 2 of the geographic atrophy (GA) eyes had many Aβ positive drusen with many Aβ containing vesicles. Conclusions: Aβ was present in 4 of 9 AMD eyes. Within these eyes, Aβ localized to a subset of drusen. None of the 9 normal eyes surveyed, some of which had small drusen, were A beta positive. Aβpositive vesicles were most numerous in GA eyes at the edges of atrophy, the region at risk for further degeneration. These results suggest that A β in drusen correlates with the location of degenerating photoreceptors and retinal pigment epithelium (RPE) cells. Further work will be necessary to determine whether Aβ deposition in drusen may contribute to or result from retinal degeneration.

Nitration of Tau Protein Is Linked to Neurodegeneration in Tauopathies

Abstract: Oxidative and nitrative injury is implicated in the pathogenesis of Alzheimer's disease (AD) and Down syndrome (DS), but no direct evidence links this type of injury to the formation of neurofibrillary tau lesions. To address this, we generated a monoclonal antibody (mAb), n847, which recognizes nitrated tau and α-synuclein. n847 detected nitrated tau in the insoluble fraction of AD, corticobasal degeneration (CBD), and Pick's disease (PiD) brains by Western blots. Immunohistochemistry (IHC) showed that n847 labeled neurons in the hippocampus and neocortex of AD and DS brains. Double-label immunofluorescence with n847 and an anti-tau antibody revealed partial co-localization of tau and n847 positive tangles, while n847 immmunofluorescence and Thioflavin-S double-staining showed that a subset of n847-labeled neurons were Thioflavin-S-positive. In addition, immuno-electron microscopy revealed that tau-positive filaments in tangle-bearing neurons were also labeled by n847 and IHC of other tauopathies showed that some of glial and neuronal tau pathologies in CBD, progressive supranuclear palsy, PiD, and frontotemporal dementia with parkinsonism linked to chromosome 17 also were n847-positive. Finally, nitrated and Thioflavin-S-positive tau aggregates were generated in a oligodendrocytic cell line after treatment with peroxynitrite. Taken together, these findings imply that nitrative injury is directly linked to the formation of filamentous tau inclusions.

Distinct cleavage patterns of normal and pathologic forms of alpha-synuclein by calpain I in vitro.

Abstract: Parkinson's disease (PD) is characterized by fibrillary neuronal inclusions called Lewy bodies (LBs) consisting largely of alpha-synuclein (alpha-syn), the protein mutated in some patients with familial PD. The mechanisms of alpha-syn fibrillization and LB formation are unknown, but may involve aberrant degradation or turnover. We examined the ability of calpain I to cleave alpha-syn in vitro. Calpain I cleaved wild-type alpha-syn predominantly after amino acid 57 and within the non-amyloid component (NAC) region. In contrast, calpain I cleaved fibrillized alpha-syn primarily in the region of amino acid 120 to generate fragments like those that increase susceptibility to dopamine toxicity and oxidative stress. Further, while calpain I cleaved wild-type alpha-syn after amino acid 57, this did not occur in mutant A53T alpha-syn. This paucity of proteolysis could increase the stability of A53T alpha-syn, suggesting that calpain I might protect cells from forming LBs by specific cleavages of soluble wild-type alpha-syn. However, once alpha-syn has polymerized into fibrils, calpain I may contribute to toxicity of these forms of alpha-syn by cleaving at aberrant sites within the C-terminal region. Elucidating the role of calpain I in the proteolytic processing of alpha-syn in normal and diseased brains may clarify mechanisms of neurodegenerative alpha-synucleinopathies.

Protein accumulation in traumatic brain injury.

Abstract: Traumatic brain injury (TBI) is one of the most devastating diseases in our society, accounting for a high percentage of mortality and disability. A major consequence of TBI is the rapid and long-term accumulation of proteins. This process largely reflects the interruption of axonal transport as a result of extensive axonal injury. Although many proteins are found accumulating after TBI, three have received particular attention; beta-amyloid precursor protein and its proteolytic products, amyloid-beta (Abeta) peptides, neurofilament proteins, and synuclein proteins. Massive coaccumulations of all of these proteins are found in damaged axons throughout the white matter after TBI. Additionally, these proteins form aggregates in other neuronal compartments and in brain parenchyma after brain trauma. Interestingly, TBI is also an epigenetic risk factor for developing neurodegenerative disorders, such as Alzheimer's disease and Parkinson's disease. Here, the similarities and differences of these accumulations with pathologies of neurodegenerative diseases will be explored. In addition, the potential deleterious roles of protein accumulations on functional outcome and progressive neurodegeneration following TBI will be examined.

Ubiquitination of α-Synuclein Is Not Required for Formation of Pathological Inclusions in α-Synucleinopathies

Abstract: α-Synucleinopathies, including Parkinson's disease, dementia with Lewy bodies, and multiple system atrophy, are neurodegenerative disorders in which abnormal inclusions containing α-synuclein accumulate in selectively vulnerable neurons and glia. In this report, immunohistochemistry demonstrates ubiquitin in subsets of α-synuclein inclusions in dementia with Lewy bodies and multiple system atrophy. Biochemistry demonstrates that α-synuclein in the sodium dodecyl sulfate-soluble fractions of diseased brains is ubiquitinated, with mono- and di-ubiquitinated species predominating over polyubiquitinated forms. Similar immunohistochemical and biochemical characteristics were observed in an A53T mutant human α-synuclein transgenic mouse model of neurodegenerative α-synucleinopathies. Furthermore, in vitro ubiquitination of α-synuclein fibrils recapitulated the pattern of α-synuclein ubiquitination observed in human disease and the A53T α-synuclein mouse model. These results suggest that ubiquitination of α-synuclein is not required for inclusion formation and follows the fibrillization of α-synuclein.

A comparison of amyloid fibrillogenesis using the novel fluorescent compound K114.

Abstract: Proteinaceous inclusions with amyloidogenic properties are a common link between many neurodegenerative diseases, including Alzheimer's disease and Parkinson's disease. Histological and in vitro studies of amyloid fibrils have advanced the understanding of protein aggregation, and provided important insights into pathogenic mechanisms of these neurodegenerative brain amyloidoses. The classical amyloid dyes Congo Red (CR) and thioflavin T and S, have been used extensively to detect amyloid inclusions in situ. These dyes have also been utilized to monitor the maturation of amyloid fibrils assembled from monomer subunits in vitro. Recently, the compound (trans,trans)-1-bromo-2,5-bis-(3- hydroxycarbonyl-4-hydroxy)styrylbenzene (BSB), derived from the structure of CR, was shown to bind to a wide range of amyloid inclusions in situ. More importantly it was also used to label brain amyloids in live animals. Herein, we show that an analogue of BSB, (trans,trans)-1-bromo-2,5-bis-(4-hydroxy)styrylbenzene (K114), recognizes amyloid lesions, and has distinctive properties which allowed the quantitative monitoring of the formation of amyloid fibrils assembled from the amyloid-β peptide, α-synuclein, and tau.

Interactions of amyloidogenic proteins

Abstract: The various protein deposits of brain amyloidosis share common ultrastructural, biophysical, and histological properties. These amyloidogenic deposits can be composed of distinct proteins, which are conceptually associated with different neurodegenerative diseases. Amyloidogenic proteins are typically soluble monomeric precursors, which undergo remarkable conformation changes associated with the polymerization into 8− to 10−nm wide fibrils, which culminate in the formation of amyloid aggregates. Some amyloidogenic inclusions are extracellular, such as senile plaques of Alzheimer’s disease, which are composed of amyloid β (Aβ) peptides. However, intracytoplasmic amyloid aggregates, such as neurofibrillary tangles in Alzheimer’s disease and Lewy bodies in Parkinson’s disease, are composed of the proteins tau and α-synuclein, respectively. The mounting awareness that the latter proteins are directly linked to the etiology of spectrum of neurodegenerative diseases has resulted in the coining of the terms “tauopathies” and “synucleinopathies.” However, emerging evidence for the overlap in the pathological and clinical features of patients with brain amyloidosis suggests that they may be linked mechanistically. Recently, it was demonstrated that α-synuclein, which has the ability to readily form amyloid in vitro without the need of other co-factors, can initiate tau amyloid formation. Following this initiation step, α-synuclein and tau can synergize the polymerization of each other. Furthermore, increased levels of Aβ peptides in brain can promote the formation of intracellular tau and α-synuclein amyloid aggregates, although the mechanism for this process is still unclear. These results indicate that the formation of amyloid composed of different proteins can affect each other directly or indirectly, likely contributing to the overlap in clinical and pathological features.

Neuron-specific age-related decreases in dopamine receptor subtype mRNAs.

Abstract: Dopaminergic neurotransmission in the CNS is involved in a number of functions, most notably cognition, affect, and motor control. Increased life expectancy, resulting from recent medical advances, has led to a need for furthering understanding of neurobiology of aging and causes of age-related disabilities in order to improve quality of life (Drachman, 1997). Region-specific changes in the functional integrity of the dopaminergic system have been associated with impairment of motor and cognitive function in humans resulting from age. For example, unbiased estimation studies demonstrate age-related losses in sub-stantia nigra pars compacta (SNpc) neurons, a major source of forebrain dopaminergic projections (Fearnley and Lees, 1991; Ma et al., 1999b). Furthermore, immunohistochemistry and gene expression studies reveal down-regulation of the dopamine (DA) transporter in the SNpc of aged humans and monkeys (Bannon et al., 1992; Bannon and Whitty, 1997; Emborg et al., 1998; Ma et al., 1999a). A loss of DA itself has been demonstrated in normal aging as well (Goldman-Rakic and Brown, 1981; Fearnley and Lees, 1991). Moreover, age-related decreases in D1 and D2 DA receptor subtypes have been demonstrated within the aged human forebrain, including the striatum and hippocampus, by using in vivo imaging techniques combined with DA-selective ligands (Rinne et al., 1993; Volkow et al., 1996; Kaasinen et al., 2000) and in post-mortem studies (Severson et al., 1982; Rinne et al., 1993; Joyce et al., 1998). Thus, pre- and postsynaptic alterations in nigrostriatal dopaminergic circuitry exist in the aging brain and may account for alterations in motor function (Fearnley and Lees, 1991; Emborg et al., 1998). The hippocampal formation is another brain region that receives dopaminergic projections from the mesencephalon, specifically the vental tegmentum (Scatton et al., 1980; Verney et al., 1985), and is also vulnerable to aging. Previous studies in rats have demonstrated significant age-related decreases in DA levels (Godefroy et al., 1989; Miguez et al., 1999) and DA receptors (Amenta et al., 2001) in the hippocampus of aged rats. Parallel alterations in humans have been demonstrated in brain regions associated with cognition during aging, including the hippocampus and temporal neocortex (Rinne, 1987; Seeman et al., 1987; Camps et al., 1989; Cortes et al., 1989; Rinne et al., 1990; Kaasinen et al., 2000; Inoue et al., 2001). At present, few primary data exist regarding the cellular specificity of age-related decline of DA receptor expression within the human temporal lobe. Primary difficulties in evaluating DA receptor subtypes in temporal lobe and other cortical regions include a moderate DA receptor subtype density and the paucity of high-affinity/receptor-selective ligands for D1–D5 DA receptors. Alternatively, use of gene expression technologies provides the means to evaluate selectively all DA receptor subtypes in discrete brain regions. For example, Meador-Woodruff and colleagues (Meador-Woodruff, 1994; Meador-Woodruff et al., 1996) have demonstrated the presence of mRNAs encoding the five known DA receptors in the human hippocampal formation by using in situ hybridization. These studies suggest a heterogeneous distribution within subfields of the hippocampal formation and laminae of the temporal lobe. However, the relative abundance of DA receptor mRNAs in specific neuronal populations making up these subregions remains elusive. In situ hybridization allows the analysis of a given mRNA in a single neuron, yet the sensitivity may not allow the analysis of low-abundance mRNAs or the means to evaluate numerous transcripts within the same tissue section. Moreover, reliance on regional assessment of gene expression emphasizes transcripts contained in the majority of neuronal and glial populations and/or transcripts in highest abundance in the region, which may not adequately reflect alterations in gene expression in target neuronal populations. Single-cell molecular biological procedures allow precise localization of changes in gene expression within brain regions (Eberwine et al., 1992; Surmeier et al., 1996; Ginsberg et al., 1999, 2000; Hemby et al., 2002). In the present study, single-cell gene expression procedures were used to assess the relative abundance of DA receptor mRNA levels in CA1 pyramidal neurons and entorhinal cortex (EC) layer II stellate neurons from human post-mortem brain tissue, permitting precise dissection and detailed molecular characterization of specific neuronal populations that are the principal neuronal conduits of information to the hippocampal-entorhinal circuit.

Genetically modified NT2N human neuronal cells mediate long-term gene expression as CNS grafts in vivo and improve functional cognitive outcome following experimental traumatic brain injury

Abstract: Human Ntera-2 (NT2) cells can be differentiated in vitro into well-characterized populations of NT2N neurons that engraft and mature when transplanted into the adult CNS of rodents and humans. They have shown promise as treatments for neurologic disease, trauma, and ischemic stroke. Although these features suggest that NT2N neurons would be an excellent platform for ex vivo gene therapy in the CNS, stable gene expression has been surprisingly difficult to achieve in these cells. In this report we demonstrate stable, efficient, and nontoxic gene transfer into undifferentiated NT2 cells using a pseudotyped lentiviral vector encoding the human elongation factor 1-α promoter and the reporter gene eGFP. Expression of eGFP was maintained when the NT2 cells were differentiated into NT2N neurons after treatment with retinoic acid. When transplanted into the striatum of adult nude mice, transduced NT2N neurons survived, engrafted, and continued to express the reporter gene for long-term time points in vivo. Furthermore, transplantation of NT2N neurons genetically modified to express nerve growth factor significantly attenuated cognitive dysfunction following traumatic brain injury in mice. These results demonstrate that defined populations of genetically modified human NT2N neurons are a practical and effective platform for stable ex vivo gene delivery into the CNS.

Subicular Dendritic Arborization in Alzheimer's Disease Correlates with Neurofibrillary Tangle Density

Abstract: Intracellular accumulation of PHFtau in Alzheimer's disease (AD) disrupts the neuronal cytoskeleton and other neuronal machinery and contributes to axonal and dendritic degeneration, and neuronal death. Furthermore, amyloid-beta (Abeta) has been reported to be toxic to neurons and neurites. While loss of presynaptic elements is an established feature of AD, the nature and extent of dendritic degeneration has been infrequently studied. We investigated MAP2-immunoreactive dendrites using a novel method of high-throughput quantification and also measured cortical thickness and the densities of NeuN-immunoreactive neurons, PHFtau neurofibrillary tangles (NFTs), and Abeta plaque burden in the subiculum in AD and elderly controls. Corrected for atrophy, the "dendritic arborization index" was significantly reduced by up to 66% in all three layers of the subiculum. Laminar thickness was reduced by an average 33% and there was a marked reduction in neuron density of approximately 50%. As expected, NFTs and Abeta plaques were significantly increased in AD. Dendritic arborization indices negatively correlated with NFT densities while no significant correlations were found with Abeta plaque densities. The pattern of dendritic loss in the subiculum and the correlations with NFT densities respectively suggest that deafferentation and intrinsic neurofibrillary degeneration both may contribute to dendritic loss in AD.

Enhanced brain levels of 8,12-iso-iPF2α-VI differentiate AD from frontotemporal dementia

Abstract: Objective: To quantify the isoprostane 8,12-iso-iPF2α-VI in brain tissues obtained from patients with AD, patients with frontotemporal dementia (FTD), and controls. Background: Enhanced brain oxidative stress with secondary damage to cellular macromolecules may play a role in the pathogenesis of AD and FTD. The isoprostane 8,12-iso-iPF2α-VI is a specific and sensitive marker of in vivo lipid peroxidation and is increased in AD. Methods: Levels of this isoprostane were determined by gas chromatography/negative ion chemical ionization mass spectrometry. Results: Levels of 8,12-iso-iPF2α-VI were markedly elevated in both frontal and temporal cortex of AD brains compared to the corresponding areas of FTD and controls. No significant difference in brain 8,12-iso-iPF2α-VI levels for any regions considered was observed between FTD and controls. Conclusions: Lipid peroxidation is a feature of AD, but not FTD. The generation of 8,12-iso-iPF2α-VI in the brain is not a general or final common pathway of neurodegeneration, but may be relatively specific for disease processes in AD and not FTD.

Reduction of Detyrosinated Microtubules and Golgi Fragmentation Are Linked to Tau-Induced Degeneration in Astrocytes

Abstract: Several human neurodegenerative diseases are associated with abnormal accumulations of aggregated tau proteins and glial degeneration in astrocytes, but the mechanism whereby tau proteins cause astrocytic degeneration is unclear. Here, we analyzed the biological consequences of overexpressing the longest human tau isoform in primary cultures of rat astrocytes using adenoviral-mediated gene transfer. Significantly, we found specific decreases in stable detyrosinated [glutamate (Glu)] microtubules (MTs) with concomitant increases in tubulin biosynthesis and the accumulation of acetylated, tyrosinated, α- and β-tubulin. The consequences of this selective reduction in stable Glu-MTs included contemporaneous decreases in kinesin levels, collapse of the intermediate filament network, progressive disruption of kinesin-dependent trafficking of organelles, fragmentation of the Golgi apparatus that culminated in atrophy, and non-apoptotic death of astrocytes. These results suggest that reduced stable Glu-MTs is a primary consequence of tau accumulation that initiates mechanisms underlying astrocyte dysfunction and death in human neurodegenerative glial tauopathies.

Proteasome Inhibition Stabilizes Tau Inclusions in Oligodendroglial Cells that Occur after Treatment with Okadaic Acid

Abstract: Tau-positive inclusions in oligodendrocytes are consistent neuropathological features of corticobasal degeneration, progressive supranuclear palsy, and frontotemporal dementias with Parkinsonism linked to chromosome 17. Here we show by immunohistochemistry that tau-positive oligodendroglial inclusion bodies also contain the small heat-shock protein (HSP) αB-crystallin but not HSP70. To study the molecular mechanisms underlying inclusion body formation, we engineered an oligodendroglia cell line (OLN-t40) to overexpress the longest human tau isoform. Treatment of OLN-t40 cells with okadaic acid (OA), an inhibitor of protein phosphatase 2A, caused tau hyperphosphorylation and a decrease in the binding of tau to microtubules. Simultaneously, tau-positive aggregates that also stained with the amyloid-binding dye thioflavin-S as well as with antibodies to tau and αB-crystallin were detected. However, they were only transiently expressed and were degraded within 24 hr. When the proteasomal apparatus was inhibited by carbobenzoxy-l-leucyl-l-leucyl-l-leucinal (MG-132) after OA treatment, the aggregates were stabilized and were still detectable after 18 hr in the absence of OA. Incubation with MG-132 alone inhibited tau proteolysis and led to the induction of HSPs, including αB-crystallin and to its translocation to the perinuclear region, but did not induce the formation of thioflavin-S-positive aggregates. Hence, although tau hyperphosphorylation induced by protein phosphatase inhibition contributes to pathological aggregate formation, only hyperphosporylation of tau followed by proteasome inhibition leads to stable fibrillary deposits of tau similar to those observed in neurodegenerative diseases.

Expression of α-, β-, and γ-synuclein in glial tumors and medulloblastomas

Abstract: α-, β- and γ-synuclein are highly homologous proteins that are found predominantly in neurons. Abnormal accumulation of synucleins has been associated with diseases of the central nervous system particularly Parkinson's disease. Immunoreactivity of α-synuclein is demonstrated in brain tumors with neuronal differentiation and in schwannomas, whereas γ-synuclein has been demonstrated in breast and ovarian carcinomas. The immunoreactivity of synucleins has not been described in glial tumors. Immunoreactivity of synucleins in glial cells in culture and in pathological conditions, however, suggests that synucleins may be expressed by glial tumors. We studied the expression of α-, β-, and γ-synuclein in 84 human brain tumors (24 ependymomas, 31 astrocytomas, 8 oligodendrogliomas, and 21 medulloblastomas) by immunohistochemistry. Our study demonstrates immunoreactivity for γ-synuclein in high-grade glial tumors; immunoreactivity is found in all anaplastic ependymomas but in only 33% of ependymomas and 16% of myxopapillary ependymomas. Immunoreactivity for γ-synuclein is noted in 63% of glioblastomas but not in other astrocytic tumors. Of medulloblastomas, 76% have immunoreactivity for either α- or β-synuclein or both; no immunoreactivity for γ-synuclein is seen in medulloblastomas.

An English Kindred With a Novel Recessive Tauopathy and Respiratory Failure

Abstract: We present the clinicopathological features of two siblings from a consanguineous marriage who presented with respiratory hypoventilation and died 10 days and 4 years later, respectively. This disorder showed extensive tau neuropathology, and both had a novel homozygous S352L tau gene mutation. This is the first description of a pathologically proved young-onset tauopathy with apparent recessive inheritance.

Selective reduction of soluble Tau proteins in sporadic and familial frontotemporal dementias: an international follow-up study

Abstract: Recently, biochemical criteria were proposed to complement histological criteria for the diagnosis of dementia lacking distinctive histopathology (DLDH), the most common pathological variant of frontotemporal dementias (FTDs), based on evidence of a selective reduction of soluble tau proteins in brains from a large cohort of sporadic DLDH and hereditary FTD (HDDD2 family) patients. To ensure that these findings are not unique to the populations included in the initial report, we extended the previous work by analyzing 22 additional DLDH brains from the United States and international centers. Our biochemical analyses here confirmed the previous findings by demonstrating substantial reductions in soluble brain tau in gray and white matter of 14 cases and moderate reductions in 6 cases of DLDH. We also analyzed brain samples from an additional affected HDDD2 family member, and remarkably, unlike other previously studied members of this kindred, this patient's brain contained substantial amounts of pathological or insoluble tau. These findings confirm and extend the definition of DLDH as a sporadic or familial "tau-less" tauopathy with reduced levels of soluble brain tau and no insoluble tau or fibrillary tau inclusions, and the data also underline the phenotypic heterogeneity of HDDD2, which parallels the phenotypic heterogeneity of other hereditary neurodegenerative FTD tauopathies caused by tau gene mutations.

Alzheimer's disease, neuropeptides, neuropeptidase, and amyloid-beta peptide metabolism.

Abstract: Amyloid-beta peptide (Abeta), the pathogenic agent of Alzheimer's disease (AD), is a physiological metabolite in the brain. We have focused our attention and effort on elucidating the unresolved aspect of Abeta metabolism: proteolytic degradation. Among a number of Abeta-degrading enzyme candidates, we used a novel in vivo paradigm to identify a member of the neutral endopeptidase family, neprilysin, as the major Abeta catabolic enzyme. Neprilysin deficiency results in defects in the metabolism of endogenous Abeta 40 and 42 in a gene dose-dependent manner. Our observations suggest that even partial down-regulation of neprilysin activity, which could be caused by aging, can contribute to AD development by promoting Abeta accumulation. Moreover, we discuss the fact that an aging-dependent decline of neprilysin activity, which leads to elevation of Abeta concentrations in the brain, is a natural process that precedes AD pathology. In this Perspective, we hypothesize that neprilysin down-regulation has a role in sporadic AD (SAD) pathogenesis, and we propose that this knowledge be used for developing preventive and therapeutic strategies through use of a G protein-coupled receptor (GPCR).

Detection of amyloid plaques by radioligands for Aβ40 and Aβ42

Abstract: Alzheimer’s disease (AD) is linked to increased brain deposition of amyloid-β (Aβ) peptides in senile plaques (SPs), and recent therapeutic efforts have focused on inhibiting the production or enhancing the clearance of Aβ in brain. However, it has not been possible to measure the burden of SPs or assess the effect of potential therapies on brain Aβ levels in patients. Toward that end, we have developed a novel radioligand, [125I]TZDM, which binds Aβ fibrils with high affinity, crosses the blood-brain barrier (BBB), and labels amyloid plaques in vivo. Compared to a styrylbenzene probe, [125I]IMSB, [125I]TZDM showed a 10-fold greater brain penetration and labeled plaques with higher sensitivity for in vivo imaging. However, this ligand also labels white matter, which contributes to undesirable high background regions of the brain. Interestingly, parallel to their differential binding characteristics onto fibrils composed of 40 (Aβ40)- or 42 (Aβ42)-amino-acid-long forms of Aβ peptides, these radioligands displayed differential labeling of SPs in AD brain sections under our experimental conditions. It was observed that [125I]IMSB labeled SPs containing Aβ40, amyloid angiopathy (AA), and neurofibrillary tangles, whereas [125I]TZDM detected only SPs and Aβ42-positive AA. Since increased production and deposition of Aβ42 relative to Aβ40 may be crucial for the generation of SPs, [125I]TZDM and related derivatives may be more attractive probes for in vivo plaque labeling. Further structural modifications of TZDM to lower the background labeling will be needed to optimize the plaque-labeling property.

Bilateral subthalamic nucleus deep brain stimulation for advanced PD: correlation of intraoperative MER and postoperative MRI with neuropathological findings.

Abstract: This postmortem study correlated intraoperative subthalamic nucleus (STN) deep brain stimulation (DBS) placement and postoperative magnetic resonance imaging (MRI) with autopsy findings in a patient who died suddenly 4 days postoperatively from a pulmonary embolism. The study demonstrates that (1) MRI stereotactic localization combined with microelectrode recording (MER) is an accurate way to target STN; (2) multiple MER tracts do not cause significant injury to the brain; and (3) postoperative MRI accurately demonstrates location of the DBS electrodes.

Iodinated tracers for imaging amyloid plaques in the brain.

Abstract: Excessive amounts of protein deposits, beta-amyloid (Abeta) plaques, are commonly detected in the postmortem brains of Alzheimer's disease (AD) patients. These Abeta plaques are believed to play an important role in the pathogenesis of the disease. Development of Abeta plaque-specific imaging agents for detecting and monitoring the changes of Abeta plaque deposition in living brains are reported. These agents, if successfully developed, may serve as potential biomarkers for the disease. Several iodinated derivatives based on variety of core structures are labeled with radioiodine as single photon emission computed tomography (SPECT) imaging agents. Thioflavin (or benzothiazole) derivatives displayed excellent in vitro characteristics with high binding affinities for Abeta aggregates (in subnanomolar to nanomolar range) and excellent plaque labeling of AD brain sections. However, the in vivo kinetic properties appeared unfavorable for further development. A novel series of imidazo-pyridine derivative, such as 2-(4'-dimethylaminophenyl)-6-iodo-imidazo[1,2-a]pyridine (IMPY), exhibited highly desirable in vivo properties. Additional structural modification resulting in stilbene derivatives displayed good binding affinities for Abeta aggregates. In addition, fluorene compounds with a rigid tricyclic structure showed in vitro and in vivo characteristics as potential SPECT imaging agents. Criteria for selection of radioiodinated tracers with suitable in vivo properties to detect amyloid plaques are discussed.

Tau and 14-3-3 in glial cytoplasmic inclusions of multiple system atrophy.

Abstract: Multiple system atrophy (MSA) is a neurodegenerative disease characterized by the presence of glial cytoplasmic inclusions (GCIs), which are comprised of fibrils of the protein α-synuclein (α-syn). Increasing evidence indicate that the formation of these lesions leads to cellular dysfunction and degeneration. The events that result in the formation of GCIs remain poorly understood. It is possible that changes in the cytoplasmic milieu, perhaps the aberrant expression of α-syn-interacting proteins, can promote the polymerization of α-syn. The presence of the microtubule-binding protein, tau, in GCIs has been reported in some studies, but these findings have not been consistent, and these studies were performed prior to the availability of the more sensitive methods of detecting GCIs using anti-α-syn antibodies. Recently, 14-3-3 proteins, putative α-syn-interacting partners, have been reported in Lewy bodies, which also are pathological inclusions comprised of α-syn. In this study the presence of tau and 14-3-3 proteins in GCIs of 21 patients with MSA was investigated. For the majority of cases, tau and 14-3-3 proteins were detected only in a subset of GCIs. In some cases none of the GCIs contained 14-3-3 or tau. When present in GCIs, tau was in a hypophosphorylated state as demonstrated with phosphorylation-specific antibodies. α-syn fibrillogenesis without 14-3-3 or tau appears to be sufficient for GCI formation, although it is possible that the accumulation of multi-functional proteins, like 14-3-3, in GCIs contribute to the disruption of cellular homeostasis.

Synphilin in normal human brains and in synucleinopathies: studies with new antibodies.

Abstract: Mutations in the gene encoding alpha-synuclein (alpha-syn) have recently been linked to rare hereditary forms of Parkinson's disease. A yeast two-hybrid screen with alpha-synuclein (alpha-syn) identified synphilin as an alpha-syn-interacting protein, potentially implicating synphilin in the pathogenesis of synucleinopathies. Co-transfection of synphilin and the central (NAC) region of alpha-syn in HEH293 cells resulted in synuclein inclusions. Furthermore, synphilin immunoreactivity has been observed in Lewy bodies (LBs) and glial cytoplasmic inclusions of synucleinopathies. To further characterize synphilin, we utilized two new anti-synphilin antibodies for biochemical and immunohistochemical studies in normal and disease brain tissues. In normal brain tissue, synphilin localized predominantly to large neurons, such as substantia nigra neurons, hippocampal pyramidal and cerebellar Purkinje cells. However, in a few pathological cases synphilin immunoreactivity was present in glial cells and a small percentage of cortical and nigral LBs. In brain extracts, synphilin was observed primarily as a 90-kDa band but protein bands of 50 and 65 kDa were also present in both soluble (high salt) and lipid (Triton X-100) fractions. Additionally, less abundant higher molecular mass species, including a 120-kDa band of similar size to that of synphilin expressed in transiently transfected cells were recovered in 8 M urea-solubilized pellets after sequential extraction of brain tissue with buffers of increasing strengths. The presence of the synphilin of higher molecular mass was detected regardless of alpha-syn pathology and may represent an immature form of synphilin. Thus, although synphilin may be an alpha-syn-interacting protein present in some alpha-syn lesions, it still remains to be determined whether synphilin plays a critical role in mechanisms of brain degeneration in human synucleinopathies.