Intraneuronal Inclusion Bodies Research Articles

Role of platelet in Parkinson's disease: Insights into pathophysiology & theranostic solutions.

Parkinson's disease (PD) is the second-most-common neurodegenerative disease characterized by motor and non-motor dysfunctions, which currently affects about 10 million people worldwide. Gradual death and progressive loss of dopaminergic neurons in the pars compacta region of substantia nigra result in striatal dopamine deficiency in PD. Specific mutation with further aggregation of α-synuclein in the intraneuronal inclusion bodies is considered the neuropathological hallmark of this disease. PD is often associated with various organelle dysfunctions inside a dopaminergic neuron, including mitochondrial damage, proteasomal impairment, and production of reactive oxygen species, thus causing subsequent neuronal death. Apart from several genetic and non-genetic risk factors, emerging research establishes an association between cardiovascular diseases, including coronary heart disease, myocardial infarction, congestive heart failure, and ischemic stroke with PD. The majority of these cardiovascular diseases have an origin from atherosclerosis, where endothelial dysfunction following thrombus formation is significantly regulated by blood platelet. This non-nucleated cell fragment expresses not only neuron-specific molecules and receptors but also several PD-specific biomarkers such as α-synuclein, parkin, PTEN-induced kinase-1, tyrosine hydroxylase, dopamine transporter, thus making platelet a suitable peripheral model for PD. Besides its similarity with a dopaminergic neuron, platelet structural alterations, as well as functional abnormalities, are also evident in PD. However, the molecular mechanism behind platelet dysfunction is still elusive and quite controversial. This state-of-the-art review describes the detailed mechanism of platelet impairment in PD, addressing the novel platelet-associated therapeutic drug candidates for plausible PD management.

Distinctive Gross Presentation in Free-Ranging White-Tailed Deer (Odocoileus virginianus) with Rabies.

The white-tailed deer (Odocoileus virginianus) is a popular game species in North America and often lives in close proximity to humans and domestic animals. Deer with neurologic signs are of high interest to the general public and wildlife managers because of disease and safety concerns. Our aim was to describe diagnostic findings from free-ranging white-tailed deer diagnosed with rabies from across the eastern US from 2000 to 2021, with emphasis on gross lesions in the skin and soft tissue overlying the skull. We reviewed diagnostic reports of white-tailed deer cases submitted to the Southeastern Cooperative Wildlife Disease Study for those diagnosed with rabies from 2000 to 2021. Rabies virus infection was confirmed by immunohistochemistry or fluorescent antibody test of brain, or both. Nine adult deer from five states were diagnosed with rabies, including seven (78%) females and two (22%) males. Three (33%) deer were found dead, and six (67%) were humanely dispatched for abnormal behavior. Six deer heads were examined grossly and had lesions, including forehead or periorbital alopecia, cutaneous erythema, abrasions and ulcers, and subcutaneous edema. Histologic examination was performed for eight of nine cases, all of which had intraneuronal eosinophilic inclusion (Negri) bodies in cerebrum, cerebellum, or both. Most (6/8; 75%) had perivascular lymphoplasmacytic encephalitis. Rabies should be considered a differential diagnosis in deer with this pattern of head lesions, suggestive of head rubbing or head pressing.

α-mangostin derivative 4e as a PDE4 inhibitor promote proteasomal degradation of alpha-synuclein in Parkinson's disease models through PKA activation

BackgroundParkinson's disease (PD) is a multi-factorial neurodegenerative disease affecting motor function of patients. The hall markers of PD are dopaminergic neuron loss in the midbrain and the presence of intra-neuronal inclusion bodies mainly composed of aggregation-prone protein alpha-synuclein (α-syn). Ubiquitin-proteasome system (UPS) is a multi-step reaction process responsible for more than 80% intracellular protein degradation. Impairment of UPS function has been observed in the brain tissue of PD patients. PDE4 inhibitors have been shown to activate cAMP-PKA pathway and promote UPS activity in Alzheimer's disease model. α-mangostin is a natural xanthonoid with broad biological activities, such as antioxidant, antimicrobial and antitumour activities. Structure-based optimizations based on α-mangostin produced a potent PDE4 inhibitor, 4e. Herein, we studied whether 4e could promote proteasomal degradation of α-syn in Parkinson's disease models through PKA activation. MethodscAMP Assay was conducted to quantify cAMP levels in samples. Model UPS substrates (Ub-G76V-GFP and Ub-R-GFP) were used to monitor UPS-dependent activity. Proteasome activity was investigated by short peptide substrate, Suc-LLVY-AMC, cleavage of which by the proteasome increases fluorescence sensitivity. Tet-on WT, A30P, and A53T α-syn-inducible PC12 cells and primary mouse cortical neurons from A53T transgenic mice were used to evaluate the effect of 4e against α-syn in vitro. Heterozygous A53T transgenic mice were employed to assess the effect of 4e on the clearance of α-syn in vivo, and further validations were applied by western blotting and immunohistochemistry. ResultsTaken together, α-mangostin derivative 4e, a PDE4 inhibitor, efficiently activated the cAMP/PKA pathway in neuronal cells, and promoted UPS activity as evidenced by enhanced degradation of UPS substrate Ub-G76V-GFP and Ub-R-GFP, as well as elevated proteasomal enzyme activity. Interestingly, 4e dramatically accelerated degradation of inducibly-expressed WT and mutant α-syn in PC12 cells, in a UPS dependent manner. Besides, 4e consistently activated PKA in primary neuron and A53T mice brain, restored UPS inhibition and alleviated α-syn accumulation in the A53T mice brain. Conclusions4e is a natural compound derived highly potent PDE4 inhibitor. We revealed its potential effect in promoting UPS activity to degrade pathogenic proteins associated with PD.

Exploring the Release of Toxic Oligomers from α-Synuclein Fibrils with Antibodies and STED Microscopy.

α-Synuclein (αS) is an intrinsically disordered and highly dynamic protein involved in dopamine release at presynaptic terminals. The abnormal aggregation of αS as mature fibrils into intraneuronal inclusion bodies is directly linked to Parkinson’s disease. Increasing experimental evidence suggests that soluble oligomers formed early during the aggregation process are the most cytotoxic forms of αS. This study investigated the uptake by neuronal cells of pathologically relevant αS oligomers and fibrils exploiting a range of conformation-sensitive antibodies, and the super-resolution stimulated emission depletion (STED) microscopy. We found that prefibrillar oligomers promptly penetrate neuronal membranes, thus resulting in cell dysfunction. By contrast, fibril docking to the phospholipid bilayer is accompanied by αS conformational changes with a progressive release of A11-reactive oligomers, which can enter into the neurons and trigger cell impairment. Our data provide important evidence on the role of αS fibrils as a source of harmful oligomers, which resemble the intermediate conformers formed de novo during aggregation, underling the dynamic and reversible nature of protein aggregates responsible for α-synucleinopathies.

The release of toxic oligomers from \u03b1-synuclein fibrils induces dysfunction in neuronal cells

The self-assembly of α-synuclein (αS) into intraneuronal inclusion bodies is a key characteristic of Parkinson’s disease. To define the nature of the species giving rise to neuronal damage, we have investigated the mechanism of action of the main αS populations that have been observed to form progressively during fibril growth. The αS fibrils release soluble prefibrillar oligomeric species with cross-β structure and solvent-exposed hydrophobic clusters. αS prefibrillar oligomers are efficient in crossing and permeabilize neuronal membranes, causing cellular insults. Short fibrils are more neurotoxic than long fibrils due to the higher proportion of fibrillar ends, resulting in a rapid release of oligomers. The kinetics of released αS oligomers match the observed kinetics of toxicity in cellular systems. In addition to previous evidence that αS fibrils can spread in different brain areas, our in vitro results reveal that αS fibrils can also release oligomeric species responsible for an immediate dysfunction of the neurons in the vicinity of these species.

Probing the Origin of the Toxicity of Oligomeric Aggregates of α-Synuclein with Antibodies.

The aggregation of α-synuclein, a protein involved in neurotransmitter release at presynaptic terminals, is associated with a range of highly debilitating neurodegenerative conditions, most notably Parkinson’s disease. Intraneuronal inclusion bodies, primarily composed of α-synuclein fibrils, are the major histopathological hallmarks of these disorders, although small oligomeric assemblies are believed to play a crucial role in neuronal impairment. We have probed the mechanism of neurotoxicity of α-synuclein oligomers isolated in vitro using antibodies targeting the N-terminal region of the protein and found that the presence of the antibody resulted in a substantial reduction of the damage induced by the aggregates when incubated with primary cortical neurons and neuroblastoma cells. We observed a similar behavior in vivo using a strain of C. elegans overexpressing α-synuclein, where the aggregation process itself is also partially inhibited as a result of incubation with the antibodies. The similar effects of the antibodies in reducing the toxicity of the aggregated species formed in vitro and in vivo provide evidence for a common origin of cellular impairment induced by α-synuclein aggregates.

Prion-like propagation of human brain-derived alpha-synuclein in transgenic mice expressing human wild-type alpha-synuclein.

IntroductionParkinson’s disease (PD) and multiple system atrophy (MSA) are neurodegenerative diseases that are characterized by the intracellular accumulation of alpha-synuclein containing aggregates. Recent increasing evidence suggests that Parkinson’s disease and MSA pathology spread throughout the nervous system in a spatiotemporal fashion, possibly by prion-like propagation of alpha-synuclein positive aggregates between synaptically connected areas. Concurrently, intracerebral injection of pathological alpha-synuclein into transgenic mice overexpressing human wild-type alpha-synuclein, or human alpha-synuclein with the familial A53T mutation, or into wild-type mice causes spreading of alpha-synuclein pathology in the CNS. Considering that wild-type mice naturally also express a threonine at codon 53 of alpha-synuclein, it has remained unclear whether human wild-type alpha-synuclein alone, in the absence of endogenously expressed mouse alpha-synuclein, would support a similar propagation of alpha-synuclein pathology in vivo.ResultsHere we show that brain extracts from two patients with MSA and two patients with probable incidental Lewy body disease (iLBD) but not phosphate-buffered saline induce prion-like spreading of pathological alpha-synuclein after intrastriatal injection into mice expressing human wild-type alpha-synuclein. Mice were sacrificed at 3, 6, and 9 months post injection and analyzed neuropathologically and biochemically. Mice injected with brain extracts from patients with MSA or probable iLBD both accumulated intraneuronal inclusion bodies, which stained positive for phosphorylated alpha-synuclein and appeared predominantly within the injected brain hemisphere after 6 months. After 9 months these intraneuronal inclusion bodies had spread to the contralateral hemisphere and more rostral and caudal areas. Biochemical analysis showed that brains of mice injected with brain extracts from patients with MSA and probable iLBD contained hyperphosphorylated alpha-synuclein that also seeded aggregation of recombinant human wild-type alpha-synuclein in a Thioflavin T binding assay.ConclusionsOur results indicate that human wild-type alpha-synuclein supports the prion-like spreading of alpha-synuclein pathology in the absence of endogenously expressed mouse alpha-synuclein in vivo.Electronic supplementary materialThe online version of this article (doi:10.1186/s40478-015-0254-7) contains supplementary material, which is available to authorized users.

Ubiquitin-proteasome system involvement in Huntington's disease.

Huntington’s disease (HD) is a genetic autosomal dominant neurodegenerative disease caused by the expansion of a CAG repeat in the huntingtin (htt) gene. This triplet expansion encodes a polyglutamine stretch (polyQ) in the N-terminus of the high molecular weight (348-kDa) and ubiquitously expressed protein htt. Normal individuals have between 6 and 35 CAG triplets, while expansions longer than 40 repeats lead to HD. The onset and severity of the disease depend on the length of the polyQ tract: the longer the polyglutamine stretch (polyQ) is, the earlier the disease begins and the more severe the symptoms are. One of the main histopathological hallmarks of HD is the presence of intraneuronal proteinaceous inclusion bodies, whose prominent and invariant feature is the presence of ubiquitin (Ub); therefore, they can be detected with anti-ubiquitin and anti-proteasome antibodies. This, together with the observation that mutations in components of the ubiquitin–proteasome system (UPS) give rise to some neurodegenerative diseases, suggests that UPS impairment may be causative of HD. Even though the link between disrupted Ub homeostasis and protein aggregation to HD is undisputed, the functional significance of these correlations and their mechanistic implications remains unresolved. Moreover, there is no consistent evidence documenting an accompanying decrease in levels of free Ub or disruption of Ub pool dynamics in neurodegenerative disease or models thus suggesting that the Ub-conjugate accumulation may be benign and just underlie lesion in 26S function. In this chapter we will elaborate on the different studies that have been performed using different experimental approaches, in order to shed light to this matter.

Inorganic Stressors of Ubiquitin

Many neurodegenerative proteinopathies are characterized by ubiquitin (Ub)-containing intraneuronal inclusion bodies. Recent reports have shown that Ub is able to bind Cu(II) and Zn(II), the dyshomeostasis of which is a hallmark of neurodegeneration. Here we use complementary techniques like potentiometry, circular dichroism-visible, and electron spin resonance to unveil the Ub/metal species that form, at neutral pH, their binding constants and structural features. Next, we show that both Zn(II) and Cu(II) ions hinder the interactions between Ub and Ub-conjugating E2 enzymes and inhibit significantly both Lys48 and Lys63 self-polyubiquitination reactions in a cell-free medium. The effects of Zn(II) and Cu(II) on Lys63 and Lys48 polyUb chain synthesis are compatible with the hypothesis that metal binding to His68 modifies the Ile44 hydrophobic patch of Ub and makes the protein less available for polyUb. These findings contribute to further arguments for a close relationship between metal dyshomeostasis and abnormal protein degradative pathways in the upstream events, triggering neurodegeneration.

P62/SQSTM1-Dependent Autophagy of Lewy Body-Like α-Synuclein Inclusions

α-Synuclein is the main component of Lewy bodies, the intraneuronal inclusion bodies characteristic of Parkinson’s disease. Although α-synuclein accumulation is caused by inhibition of proteasome and autophagy-lysosome, the degradation of α-synuclein inclusions is still unknown. Formation of Lewy body-like inclusions can be replicated in cultured cells by introducing α-synuclein fibrils generated in vitro. We used this cell culture model to investigate the autophagy of α-synuclein inclusions and impaired mitochondria. The intracellular α-synuclein inclusions immediately underwent phosphorylation and ubiquitination. Simultaneously they were encircled by an adaptor protein p62/SQSTM1 and directed to the autophagy-lysosome pathway in HEK293 cell line. Most phospho-α-synuclein-positive inclusions were degraded in 24 h, however, lysosomal dysfunction with bafilomycin A1 significantly affected their clearance. Moreover, inhibition of autophagy by Atg-5 siRNA treatment reduced the incorporation of α-synuclein inclusions into LC3-positive autophagosomes. Knockdown experiments demonstrated the requirement of p62 for α-synuclein autophagy. These results demonstrate that α-synuclein inclusions are preferred targets for p62-dependent autophagy. Next, we investigated the autophagic clearance of impaired mitochondria in α-synuclein inclusion-containing cells. Impaired mitochondria were almost completely eliminated after mitochondrial uncoupling even in the presence of α-synuclein inclusions, suggesting that mitochondrial clearance is not prevented by α-synuclein inclusions in HEK293 cells.

Membrane Interactions of Oligomeric Alpha-Synuclein: Potential Role in Parkinsons Disease

alpha-Synuclein is a small neuronal protein that has been implicated to play an important role in Parkinson's disease. Genetic mutations and multiplications in the alpha-synuclein gene can cause familial forms of the disease. In aggregated fibrillar form, alpha-synuclein is the main component of Lewy bodies, the intraneuronal inclusion bodies characteristic of Parkinson's disease. The loss of functional dopaminergic neurons in Parkinson's disease may be caused by a gain in toxic function of the protein. Elucidating if this gain of toxic function is related to the aggregation of alpha-synuclein may be vital in understanding Parkinson's disease. Although there are many ideas on how alpha-synuclein could be involved in the disease, this review will focus on the amyloid pore hypothesis. This hypothesis assumes that aggregation intermediates or oligomers are more likely to be toxic than monomeric or fibrillar forms of the protein. Oligomeric species are thought to exercise their toxicity through permeabilization of cellular membranes. Membrane pore formation by an oligomeric intermediate might play a role in other neurodegenerative disorders in which protein aggregation and amyloid formation play a role, such as Alzheimer's disease. We will discuss the role of this hypothesis in Parkinson's disease.

Imaging amyloid in Parkinson's disease dementia and dementia with Lewy bodies with positron emission tomography

Although Parkinson's disease with later dementia (PDD) and dementia with Lewy bodies (DLB) are pathologically characterized by the presence of intraneuronal Lewy inclusion bodies, amyloid deposition is also associated to varying degrees with both these disorders. Fibrillar amyloid load can now be quantitated in vivo with positron emission tomography (PET) using imaging biomarkers. Here the reported findings of 11C-PIB PET studies concerning the amyloid load associated with PD and its influence on dementia are reviewed. It is concluded that the presence of amyloid acts to accelerate the dementia process in Lewy body disorders, though has little influence on its nature. Anti-amyloid strategies could be a relevant approach for slowing dementia in a number of DLB and PDD cases.

Frontotemporal lobar degeneration—a coming of age

Frontotemporal lobar degeneration (FTLD) Wrst made its mark in 1892 with initial descriptions by Arnold Pick on the clinical features of six patients with behavioural disorder and language problems, a syndrome which he called amnestic aphasia [20]. Pick also emphasised the focal nature of the macroscopic atrophy, which involved particularly the temporal lobes. Post-mortem brain examination of such patients did not occur until 1911, when Alzheimer noted the absence of senile plaques and neuroWbrillary tangles but revealed the curious presence of swollen or ballooned neurones and rounded, argyrophilic, intraneuronal inclusion bodies within the cerebral cortex and hippocampus [1]—a histopathological picture quite distinct from that of Alzheimer’s disease, for which the Wrst pathological description had itself been given only 4 years earlier. Just as Pick had entitled Alzheimer’s descriptions of plaques and tangles, ‘Alzheimer’s disease’, Onari and Spatz [19] later designated Pick’s cases as ‘Picks disease’, and the swollen cells became known as Pick cells, and the inclusion bodies, Pick bodies. However, what in pathological terms constituted Pick’s disease became increasingly unclear in following years, and all cases of frontotemporal atrophy were to become designated in such a manner, even when the pathognomonic Pick bodies and Pick cells were absent. This broader view of ‘Pick’s disease’ was strengthened by Constantinidis et al. [6], in which the disorder was grouped into three categories, depending on the appearance of the neuropathological changes. Group A cases were ‘classic’ with Pick bodies and Pick cells. Group B cases were associated with cortical atrophy and swollen cells, and probably today diagnosed as corticobasal degeneration. Group C cases had neither of these features, and might now be viewed as progressive subcortical gliosis. DiYculties in distinguishing and classifying histologic subtypes of FTLD paradoxically led to an increasing focus on the deWnition of the clinical symptoms. In 1994, the Manchester–Lund criteria for frontotemporal dementia [4] provided descriptions of the characteristic clinical features, which led to improved recognition of cases, and the realization that FTLD was a relatively common disorder among younger persons with dementia. The Lund–Manchester criteria were further reWned in 1998 by Neary et al. [16], which provided clinical descriptions of the three common presentations of FTLD: frontotemporal dementia (FTD) a social conduct disorder with disinhibition, progressive nonXuent aphasia (PNFA), and semantic dementia (SD) a Xuent aphasia with impairment of semantic verbal memory. Nonetheless, while our understanding of the clinical syndromes underlying FTLD was advancing, in pathogenetic terms, FTLD largely remained on the shelf of pathological curiosities, compared to its more illustrious ‘cousin’, Alzheimer’s disease where major advances in the biology for the major pathological proteins, -amyloid and tau, were being made. This remained so until geneticists, buoyed up by the successful identiWcation of mutations in the amyloid precursor protein (APP) [10] and presenilin (PSEN) [5] genes responsible for early onset familial Alzheimer’s disease, turned their attentions to certain other families with early onset of dementia. Some of these families had in fact been thought to have suVered from Alzheimer’s disease, and had caused some degree of consternation when D. Mann (&) Greater Manchester Neuroscience Center, University of Manchester, Stott Lane, Salford, M6 8HD, UK e-mail: david.mann@man.ac.uk

The aggravating role of the ubiquitin–proteasome system in neurodegenerative disease

Intraneuronal inclusion bodies are key pathological features of most age-related neurodegenerative disorders including Parkinson's disease and Alzheimer's disease. These inclusions are commonly characterized both by the presence of ubiquitinated proteins and the sequestration of components of the UPS (ubiquitin-proteasome system). Unfortunately, as we age, the efficiency of the UPS declines, suggesting that the presence of ubiquitinated proteins and UPS components in inclusions may reflect unsuccessful attempts by the (failing) UPS to remove the aggregating proteins. Whether the physical presence of inclusions causes cell death or, conversely, whether they are non-toxic and their presence reflects a cellular protective mechanism remains highly controversial. Animal and in vitro model systems that allow detailed characterization of the inclusions and their effects on the cell have been developed by us and others. Identification of the mechanisms involved in inclusion formation is already aiding the development of novel therapeutic strategies to prevent or alleviate aggregate-associated neurodegenerative diseases.

Nocardia otitidiscaviarum (GAM-5) induces parkinsonian-like alterations in mouse.

Parkinson's disease, a major neurodegenerative disorder in humans whose etiology is unknown, may be associated with some environmental factors. Nocardia otitidiscaviarum (GAM-5) isolated from a patient with an actinomycetoma produced signs similar to Parkinson's disease following iv injection into NMRI mice. NMRI mice were infected intravenously with a non-lethal dose of 5 x 10(6) colony forming units of N. otitidiscaviarum (GAM-5). Fourteen days after bacterial infection, most of the 60 mice injected exhibited parkinsonian features characterized by vertical head tremor, akinesia/bradykinesia, flexed posture and postural instability. There was a peak of nocardial growth in the brain during the first 24 h followed by a decrease, so that by 14 days nocardiae could no longer be cultured. At 24 h after infection, Gram staining showed nocardiae in neurons in the substantia nigra and occasionally in the brain parenchyma in the frontal and parietal cortex. At 21 days post-infection, tyrosine hydroxylase immunolabeling showed a 58% reduction of tyrosine hydroxylase in the substantia nigra, and a 35% reduction of tyrosine hydroxylase in the ventral tegmental region. Dopamine levels were reduced from 110 +/- 32.5 to 58 +/- 16.5 ng/mg protein (47.2% reduction) in brain from infected mice exhibiting impaired movements, whereas serotonin levels were unchanged (191 +/- 44 protein in control and 175 +/- 39 ng/mg protein in injected mice). At later times, intraneuronal inclusion bodies were observed in the substantia nigra. Our observations emphasize the need for further studies of the potential association between Parkinson's disease or parkinsonism-like disease and exposure to various nocardial species.

Neuroserpin: a selective inhibitor of tissue-type plasminogen activator in the central nervous system.

Neuroserpin is a member of the serine proteinase inhibitor (serpin) gene family that reacts preferentially with tissue-type plasminogen activator (tPA) and is primarily localized to neurons in regions of the brain where tPA is also found. Outside of the central nervous system (CNS) tPA is predominantly found in the blood where its primary function is as a thrombolytic enzyme. However, tPA is also expressed within the CNS where it has a very different function, promoting events associated not only with synaptic plasticity but also with cell death in a number of settings, such as cerebral ischemia and seizures. Neuroserpin is released from neurons in response to neuronal depolarization and plays an important role in the development of synaptic plasticity. Following the onset of cerebral ischemia there is an increase in both tPA activity and neuroserpin expression in the area surrounding the necrotic core (ischemic penumbra), and treatment with neuroserpin following ischemic stroke or overexpression of the neuroserpin gene results in a significant decrease in the volume of the ischemic area as well as in the number of apoptotic cells. TPA activity and neuroserpin expression are also increased in specific areas of the brain by seizures, and treatment with neuroserpin slows the progression of seizure activity throughout the CNS and results in significant neuronal survival in the hippocampus. Mutations in human neuroserpin result in a form of autosomal dominant inherited dementia which is characterized by the presence of intraneuronal inclusion bodies and is known as Familial Encephalopathy with Neuroserpin Inclusion Bodies.

Intraneuronal inclusion bodies: From Negri bodies to proteasomal dysfunction

Rabies, a fatal viral encephalitis which infects domestic and wild animals, is an old recognized disease, but still widely distributed, especially in developing countries. The occurrence of characteristic inclusion bodies in neurons of spinal ganglia and brains of animals infected by rabies was detected by Adelchi Negri in Camillo Golgi’s laboratory and officially reported in 1903. This finding provided a breakthrough in the diagnosis of rabies. Influenced by Golgi’s studies on malaria, Negri proposed that the intracellular bodies he had observed corresponded to a parasite, which he believed to represent the pathogenetic agent of rabies. The causative agents of the disease are now known to be neurotropic RNA viruses which spread into the nervous system via retrograde axonal transport. The Negri bodies have been ascribed to accumulation of viral nucleoproteins and cellular organelles, but their nature and composition is still enigmatic, and research on the pathogenesis of rabies is hampered nowadays by lack of funding. On the other hand, interest in intraneuronal inclusions (presently considered as intracellular foci of sequestration of aggregated proteins) has recently exploded in relation to neurodegenerative diseases. In this context, failure in the clearance of proteins via the ubiquitin-proteasome pathway is raising increasing interest. This pathway involves an enzymatic cascade through which ubiquitin molecules are attached to the protein substrate, which is then degraded by the proteasome complex. Such proteolytic process, important in a variety of basic cellular functions, is the primary mechanism responsible for the elimination of damaged and misfolded proteins. Recent findings indicated that a cellular response designed to reduce the toxicity of misfolded proteins can be exploited by viruses to concentrate structural proteins at virus assembly sites. In addition, the ubiquitin-proteasome pathway was recently found to be involved in the budding process of rabies virus. Interest in the Negri bodies could thus be revived in the context of molecular and cellular studies at the forefront of neurosciences.

Dementia with Lewy bodies.

Lewy bodies are intracytoplasmic, eosinophilic intraneuronal inclusion bodies [1]. They contain phosphorylated and non-phosphorylated microfilaments, β-crystalline, ubiquitin and proteinases [2]. They are therefore indicators of cell response to stress and markers for the degradation and excretion of abnormal proteins from the cell. Although Lewy bodies in the substantia nigra were identified as a key neuropathological element of Parkinson’s disease (PD) early in the century, the importance of cortical Lewy bodies as a feature of some cases of dementia has only recently become evident. A series of single case reports and small case series appeared in the literature between the 1960s and the mid-1980s, suggesting that Lewy bodies in the cerebral cortex could be associated with a rare form of dementia [3, 4, 5, 6]. Hospital-based postmortem series reporting consecutive dementia cases have subsequently identified cortical Lewy bodies in at least 10% of dementia sufferers [7, 8, 9, 10].

Role for the ubiquitin-proteasome system in Parkinson's disease and other neurodegenerative brain amyloidoses.

Many neurodegenerative brain amyloidoses, including Alzheimer's and Parkinson's disease, are characterized by selective neuronal loss together with the appearance of intraneuronal ubiquitin-positive proteinaceous aggregates or inclusion bodies. These features usually result from the abnormal accumulation and processing of mutant, misfolded, or damaged intracellular proteins. It has recently become clear that both genetic factors and aberrant proteolytic degradation may therefore play a major role in neuronal degeneration. Indeed, the linkage of two genes directly involved in the ubiquitin-proteasome system (UPS) in familial Parkinson's disease clearly indicates a central role for the UPS in neurodegeneration, and thus Parkinson's disease is considered the prototypical disorder associated with UPS dysfunction. In this review, we provide an overview of the key genes/proteins implicated in the abnormal UPS-mediated proteolytic processing of unwanted proteins observed in neurodegenerative brain amyloidoses. We also provide an outline of the various components and pathways involved in the normal cellular functioning of the UPS and discuss the mechanisms by which UPS dysfunction can compromise neuronal integrity. A more complete understanding of the UPS and its relationship to the neurodegenerative process will undoubtedly provide tremendous insight into the molecular pathogenesis of amyloidogenic neurodegenerative disorders and will allow the development of novel rational therapies for treating these disorders.

Staging of the intracerebral inclusion body pathology associated with idiopathic Parkinson's disease (preclinical and clinical stages)

The synucleinopathy known as idiopathic Parkinson's disease (IPD) is a multi-system disorder in the course of which only a few predisposed nerve cell types in specific regions of the human brain become progressively involved. The underlying neuropathological process (formation of proteinaceous intraneuronal inclusion bodies) intracerebrally begins in clearly defined induction sites and advances in a topographically predictable sequence. Components of the autonomic, limbic, and motor systems sustain especially heavy damage. During the presymptomatic stages 1 and 2, the IPD-related inclusion body pathology remains confined to the medulla oblongata and olfactory bulb. In stages 3 and 4, the substantia nigra and other nuclear grays of the midbrain and basal forebrain are the focus of initially subtle and, then, severe changes. The illness reaches its symptomatic phase. In end-stages 5 and 6, the pathological process encroaches upon the telencephalic cortex. IPD manifests itself in all of its dimensions, which under the influence of the supervening cortical pathology are subject to increasing complexity.