Protein aggregates in Huntington's disease

Huntington's disease (HD) is an incurable neurodegenerative disease characterized by abnormal motor movements, personality changes, and early death. HD is caused by a mutation in the IT-15 gene that expands abnormally the number of CAG nucleotide repeats. As a result, the translated protein huntingtin contains disease-causing expansions of glutamines (polyQ) that make it prone to misfold and aggregate. While the gene and mutations that cause HD are known, the mechanisms underlying HD pathogenesis are not. Here we will review the state of knowledge of HD, focusing especially on a hallmark pathological feature-intracellular aggregates of mutant Htt called inclusion bodies (IBs). We will describe the role of IBs in the disease. We speculate that IB formation could be just one component of a broader coping response triggered by misfolded Htt whose efficacy may depend on the extent to which it clears toxic forms of mutant Htt. We will describe how IB formation might be regulated and which factors could determine different coping responses in different subsets of neurons. A differential regulation of IB formation as a function of the cellular context could, eventually, explain part of the neuronal vulnerability observed in HD.     

Reagents that block neuronal death from Huntington's disease also curb oxidative stress

Patients with Huntington's disease suffer severe neuronal loss and signs of oxidative damage in the brain. Previously we found that primary neurons from embryonic cortex of mice bearing the Huntington's disease mutation (140 glutamines inserted into exon 1 of huntingtin) showed higher levels of reactive oxygen species before cell death. Here, we treated mutant neurons with known neuroprotective agents and determined the effects on neuronal survival and levels of reactive oxygen species. Primary neurons were exposed to the neurotrophin, brain derived neurotrophic factor, the antioxidant N-acetyl-cysteine or a specific inhibitor of glycogen synthase kinase 3-β, SB216763. Each reagent increased the survival of the mutant neurons compared with untreated mutant neurons and also reduced the levels of reactive oxygen species to levels of wild-type neurons. These results suggest that reducing the levels of reactive oxygen species may be necessary to protect neurons with the Huntington's disease mutation from cell death.     

An abnormal striatal synaptic plasticity may account for the selective neuronal vulnerability in Huntington's disease

A marked decrease in the activity of mitochondrial complex II (succinate dehydrogenase, SD) has been found in the brains of Huntington's disease (HD) patients. Here we have examined the possibility that SD inhibitors might produce their toxic action by increasing corticostriatal glutamatergic transmission. We report that SD inhibitors produce a durable augmentation of NMDA-mediated corticostriatal excitation (DANCE) in striatal spiny neurons, but not in striatal cholinergic interneurons. DANCE involves increased intracellular calcium, activation of MAP kinase ERK and is critically dependent upon endogenous dopamine (DA) acting via D2-like receptors. This pathological form of corticostriatal synaptic plasticity might play a key role in the regional and cell-type specific neuronal death observed in HD.     

Oxidized lipoproteins may play a role in neuronal cell death in Alzheimer disease

Oxidative stress in the central nervous system (CNS) may cause oxidation of lipoprotein particles. The oxidized lipoproteins may damage cellular and subcellular membranes, leading to tissue injury and cell death. Human low-density lipoprotein (LDL) and very low-density lipoprotein (VLDL) are oxidized by transition metal ions, such as Cu²⁺. Using PC 12 cells, we tested the cytotoxicity of oxidized LDL and VLDL. Cell death was increased in a dose-dependent manner. Antioxidants added to the incubation medium, such as vitamins E or C, or resveratrol showed some protection. Results indicated that oxidized lipoproteins may serve as an oxidative stressor, which may initiate the neuronal cell death leading to the manifestation of Alzheimer disease (AD).     

Rate of functional decline in Huntington's disease. Huntington Study Group

OBJECTIVE: To determine the rate of functional decline in a large cohort of patients with Huntington's disease (HD) followed at 43 sites by the Huntington Study Group (HSG). METHODS: The annual rate of functional decline was measured using the Total Functional Capacity Scale (TFC) and the Independence Scale (IS) in 960 patients with definite HD followed prospectively for a mean of 18.3 months. All patients were rated with the Unified Huntington's Disease Rating Scale (UHDRS). Sample size calculations for hypothetical clinical trials were calculated. Results: A factor analysis of the UHDRS at baseline yielded 15 factors accounting for 77% of the variance. The TFC score declined at a rate of 0.72 units/year (standard error [SE] 0.04) and the IS score declined at a rate of 4.52 units/year (SE 0.23). Lower TFC score at baseline, indicating more severe impairment, was associated with less rapid annual decline in TFC score, perhaps reflecting the floor effect of the scale. The annual rate of decline for 575 patients with baseline TFC scores of 7 to 13 was 0.97 (SE 0.06), was 0.38 (SE 0.08) for 270 patients with baseline TFC scores of 3 to 6, and was 0.06 (SE 0.1) for 101 patients with TFC scores of 0 to 2. In multivariate analysis (n = 960), longer disease duration and better cognitive status at baseline were associated with a less rapid rate of decline in TFC score, whereas depressive symptomatology was the only factor associated with more rapid decline on the IS score. Age at onset of HD, sex, weight, and education did not affect decline on either score. CONCLUSIONS: The comparable rates of decline on the TFC and the IS scores with other published studies suggest that these estimates of functional decline are representative of HD patients who are evaluated at HSG research sites. In longitudinal analysis, longer disease duration and better neuropsychological performance at baseline were associated with a less rapid rate of decline in TFC score, whereas depressive symptomatology at baseline was associated with a more rapid decline in the IS score. These rates of functional decline and the covariates that modify them should be considered in estimating statistical power and designing future therapeutic trials involving HD patients with early or moderately severe disease.     

Lewy body cortical involvement may not always predict dementia in Parkinson's disease

BACKGROUND: The presence of Lewy bodies (LB) in the neocortex and limbic system in patients with Parkinson's disease (PD) is commonly thought to be linked with cognitive impairment. The authors present here a series of patients with diagnosis of PD in life and no significant cognitive impairment who, at necropsy, satisfied the current neuropathological criteria for dementia with Lewy bodies (DLB). METHODS: Two hundred and seventy six brains with PD pathology were examined at the Queen Square Brain Bank in London between 1993 and 1999. The neuropathological diagnosis was PD, but 117 patients also had sufficient LB involvement above the brain stem to satisfy the current neuropathological criteria for DLB (50 patients had a neuropathological picture consistent with the limbic category of DLB and 67 with neocortical DLB). Forty eight cases were excluded who developed early cognitive impairment together with motor features of parkinsonism, 12 cases for lack of detailed clinical history, and 19 cases with coexistent features of advanced Alzheimer's disease changes. Thirty eight patients (13.8% of the total with PD pathology and 32.5 % of the total with DLB pathology) were found where there was no or very late cognitive impairment reported in the clinical records. RESULTS: Selected cases were 24 men and 14 women, with a mean (SD) age at onset of parkinsonian symptoms of 60.1 (10.1) years and a mean disease duration of 15.3 (5.5) years. At some time during the evolution of the disease 21 patients developed different degrees of cognitive impairment (after a mean disease duration of 12.2 (4.8) years). Clinical diagnosis at death was PD in 10 cases and PD with dementia in 11. In the remaining 17 patients no history of cognitive impairment was ever recorded in life and all of them had a clinical diagnosis of PD at death; in this subgroup, nine patients later revealed a neuropathological picture consistent with limbic (or transitional) category of DLB and eight with neocortical DLB. Interestingly, in all these patients the parkinsonian features including the response to dopaminergic drugs were indistinguishable from classic brain stem PD. CONCLUSIONS: The authors demonstrate that the classic pathology of DLB can commonly be seen outside the generally accepted clinical spectrum for DLB and that important factors other than the absolute number of LB in the neocortex and limbic system influence the development of cognitive impairment in PD. Furthermore, the pathology of PD may be indistinguishable from that reported in DLB, suggesting that the two clinicopathological syndromes may be attributable to the same biological abnormality.     

Nuclear and neuropil aggregates in Huntington's disease: relationship to neuropathology.

The data we report in this study concern the types, location, numbers, forms, and composition of microscopic huntingtin aggregates in brain tissues from humans with different grades of Huntington's disease (HD). We have developed a fusion protein antibody against the first 256 amino acids that preferentially recognizes aggregated huntingtin and labels many more aggregates in neuronal nuclei, perikarya, and processes in human brain than have been described previously. Using this antibody and human brain tissue ranging from presymptomatic to grade 4, we have compared the numbers and locations of nuclear and neuropil aggregates with the known patterns of neuronal death in HD. We show that neuropil aggregates are much more common than nuclear aggregates and can be present in large numbers before the onset of clinical symptoms. There are also many more aggregates in cortex than in striatum, where they are actually uncommon. Although the striatum is the most affected region in HD, only 1-4% of striatal neurons in all grades of HD have nuclear aggregates. Neuropil aggregates, which we have identified by electron microscopy to occur in dendrites and dendritic spines, could play a role in the known dendritic pathology that occurs in HD. Aggregates increase in size in advanced grades, suggesting that they may persist in neurons that are more likely to survive. Ubiquitination is apparent in only a subset of aggregates, suggesting that ubiquitin-mediated proteolysis of aggregates may be late or variable.     

Striatal neuronal loss correlates with clinical motor impairment in Huntington's disease

Huntington's disease (HD) is characterized clinically by chorea, motor impairment, psychiatric manifestations, and dementia. Atrophy of the striatum is the neuropathological hallmark of HD, and previous studies have suggested that striatal atrophy correlates more closely with motor impairment than with chorea. Motor impairment, as measured by motor impairment score, correlates with functional disability in HD patients, but chorea does not. In this study, we investigated the relation between neuronal loss and these motor features. We conducted neuropathological and stereologic assessments of neurons in putamen and subthalamic nuclei in HD patients and age‐matched controls. In putamen, we estimated the total number and volume of medium spiny neurons labeled with dopamine‐ and cAMP‐regulated phosphoprotein 32 kDa (DARPP‐32). In subthalamic nuclei, we estimated the total number of neurons on hematoxylin & eosin/luxol fast blue stains. In putamen of HD, immunohistochemistry showed DARPP‐32 neuronal atrophy with extensive disruption of neurites and neuropil; stereologic studies found significant decreases in both the number and size of DARPP‐32 neurons; we also detected a significant reduction of overall putamen volume in HD patients, compared to controls. In subthalamic nuclei, there was a mild, but significant, neuronal loss in the HD group. The loss of neurons in putamen and subthalamic nuclei as well as putaminal atrophy were significantly correlated with severity of motor impairment, but not with chorea. Our findings suggest that neuronal loss and atrophy in striatum and neuronal loss in subthalamic nuclei contribute specifically to the motor impairment of HD, but not to chorea. © 2012 Movement Disorder Society     

Distribution of inclusions in neuronal nuclei and dystrophic neurites in Huntington disease brain

Recently, an N-terminal fragment of huntingtin was localized to neuronal intranuclear inclusions (NII), presumed to cause cellular dysfunction, and to inclusions in dystrophic neurites (IDN) in the neostriatum and neocortex of Huntington disease (HD) patients. In the present immunohistochemical study of autopsy brain of 2 juvenile-onset HD patients, 5 HD patients with adult-onset, and 5 controls, NII and IDN as stained with both N-terminal antiserum to huntingtin and ubiquitin antiserum were detected in the HD neostriatum, neocortex, and allocortex, but not in the HD pallidum, cerebellum, and substantia nigra nor in control brain. The frequency of NII in the HD neocortex was highest in the juvenile patients. Within the allocortex, NII and IDN were found in the entorhinal region, subiculum, and pyramidal cell layer of Ammon's horn. N-terminal huntingtin antiserum also labeled intranuclear granular structures adjacent to the neuronal nuclear membrane in 5 HD patients, one control with idiopathic epilepsy, and one with Alzheimer disease. Our results show that NII formation in HD involves the allocortex in addition to the neostriatum and neocortex. The development of NII in the neocortex and allocortex in HD brain might contribute to the emergence of the cognitive and behavioral symptoms of the disease.     

Genotype does not predict severity of behavioural phenotype in juvenile neuronal ceroid lipofuscinosis (Batten disease)

Aim The primary aim of this investigation was to examine genotype and clinical phenotype differences in individuals with juvenile neuronal ceroid lipofuscinosis (JNCL) who were homozygous for a common disease‐causing deletion or compound heterozygous. The secondary aim was to cross‐validate the Child Behavior Checklist (CBCL) and the Unified Batten Disease Rating Scale (UBDRS), a disease‐specific JNCL rating scale. Method Sixty individuals (28 males, 32 females; mean age 15y 1mo, SD 4y 9mo, range 5y 8mo–31y 1mo) with JNCL completed the UBDRS. Results No significant genotype and clinical phenotype differences were identified when comparing individuals homozygous for the deletion with a heterogeneous group of compound heterozygous individuals. There were significant correlations among related behaviour items and scales on the CBCL and UBDRS (Spearman’s rho ranging from 0.39 [p<0.05] to 0.72 [p<0.01]). Behaviour and physical function ratings were uncorrelated, supporting divergent validity of these two constructs in JNCL. Interpretation Previous reports of genotype and clinical phenotype differences were unsupported in this investigation, which did not find differences between individuals homozygous or heterozygous for the CLN3 deletion. The CBCL, an already validated measure of behaviour problems, appears valid for use in JNCL and cross‐validates well with the UBDRS.     

Caspase-dependent programmed cell death pathways are not activated in generalized seizure-induced neuronal death

Activation of the caspase-dependent cell death pathways has been shown in focal seizures, but whether this occurs in prolonged generalized seizures is not known. We investigated whether the initiator caspase in the extrinsic pathway, caspase-8, or the intrinsic pathway, caspase-9, is activated during the first 24 h following lithium-pilocarpine-induced status epilepticus, when neuronal death is maximal and widespread. The thymuses of rats given methamphetamine were used as positive controls for caspase-3-activated cellular apoptosis. Following methamphetamine treatment, caspase-9 but not caspase-8 was activated in thymocytes. However, 6 or 24 h following status epilepticus, none of 26 brain regions studied showed either caspase-8 or -9 activation by immunohistochemistry, western blotting and enzyme activity assays. Our results provide evidence against the activation of the extrinsic and intrinsic caspase pathways in generalized seizures, which produce morphologically necrotic neurons with internucleosomal DNA cleavage (DNA laddering), a programmed process. In contrast, there is increasing evidence that caspase-independent programmed mechanisms play a prominent role in seizure-induced neuronal death.     

Immunohistochemical study of foci of recent cell death in Huntington's disease

Foci of recent cell death were identified in 10 consecutive cases of Huntington's disease in the caudate nucleus and putamen. Indirect immunohistochemical staining procedures were performed on these areas and controls using fluorescein conjugated anti-globulin and peroxidase-conjugated anti-globulin. Positive results were found when human antiserum to Cytomegalovirus was used. The possibility of abnormal antigen presence within the dying neurons in this disease is discussed.