Somatic mitochondrial DNA mutations in early Parkinson and incidental Lewy body disease.

Somatic mutations in mitochondrial DNA (mtDNA) are hypothesized to play a role in Parkinson disease (PD), but large increases in mtDNA mutations have not previously been found in PD, potentially because neurons with high mutation levels degenerate and thus are absent in late stage tissue. To address this issue, we studied early stage PD cases and cases of incidental Lewy body disease (ILBD), which is thought to represent presymptomatic PD. We show for the first time that mtDNA mutation levels in substantia nigra neurons are significantly elevated in this group of early PD and ILBD cases.

Impaired ß-amyloid secretion in Alzheimer's disease pathogenesis.

A central question in Alzheimer's disease (AD) research is what role ß-amyloid peptide (Aß) plays in synaptic dysfunction. Synaptic activity increases Aß secretion, potentially inhibiting synapses, but also decreases intraneuronal Aß, protecting synapses. We now show that levels of secreted Aß fall with time in culture in neurons of AD-transgenic mice, but not wild-type mice. Moreover, the ability of synaptic activity to elevate secreted Aß and reduce intraneuronal Aß becomes impaired in AD-transgenic but not wild-type neurons with time in culture. We demonstrate that synaptic activity promotes an increase in the Aß-degrading protease neprilysin at the cell surface and a concomitant increase in colocalization with Aß42. Remarkably, AD-transgenic but not wild-type neurons show reduced levels of neprilysin with time in culture. This impaired ability to secrete Aß and reduce intraneuronal Aß has important implications for the pathogenesis and treatment of AD.

High-resolution 3D reconstruction reveals intra-synaptic amyloid fibrils.

ß-Amyloid (Aß) accumulation and aggregation are hallmarks of Alzheimer's disease (AD). High-resolution three-dimensional (HR-3D) volumetric imaging allows for better analysis of fluorescence confocal microscopy and 3D visualization of Aß pathology in brain. Early intraneuronal Aß pathology was studied in AD transgenic mouse brains by HR-3D volumetric imaging. To better visualize and analyze the development of Aß pathology, thioflavin S staining and immunofluorescence using antibodies against Aß, fibrillar Aß, and structural and synaptic neuronal proteins were performed in the brain tissue of Tg19959, wild-type, and Tg19959-YFP mice at different ages. Images obtained by confocal microscopy were reconstructed into three-dimensional volumetric datasets. Such volumetric imaging of CA1 hippocampus of AD transgenic mice showed intraneuronal onset of Aß42 accumulation and fibrillization within cell bodies, neurites, and synapses before plaque formation. Notably, early fibrillar Aß was evident within individual synaptic compartments, where it was associated with abnormal morphology. In dendrites, increasing intraneuronal thioflavin S correlated with decreases in neurofilament marker SMI32. Fibrillar Aß aggregates could be seen piercing the cell membrane. These data support that Aß fibrillization begins within AD vulnerable neurons, leading to disruption of cytoarchitecture and degeneration of spines and neurites. Thus, HR-3D volumetric image analysis allows for better visualization of intraneuronal Aß pathology and provides new insights into plaque formation in AD.

Hashimoto's encephalopathy cases: Chinese experience.

BACKGROUND: Hashimoto's encephalopathy is a poorly understood syndrome consisting of heterogeneous neurological symptoms and high serum antithyroid antibody titers, typically responding to steroids. More clinical series studies are required to characterize the clinical, laboratory and imaging features, and outcomes, especially in the Chinese population. METHODS: We analyzed the clinical, laboratory, and imaging features and outcomes of thirteen consecutive patients with Hashimoto's encephalopathy diagnosed in Xuan Wu Hospital, Beijing from 2005 to 2010 retrospectively. RESULTS: Cognitive impairment (84.6%) and psychiatric symptoms (38.5%) were the most frequent symptoms. Seizures (30.8%) and myoclonus (7.7%) were less common than previously described. Three (23.1%) patients showed abnormal signals in hippocampus or temporal lobe, which were believed related to their memory disorders or seizures. MRI changes showed resolution paralleling clinical improvement in one patient. Among eight patients who received steroid therapy, five patients recovered, one patient improved with residual deficits, and two patients relapsed or had no effect. Among five non-steroid treated patients, three patients experienced stable remission with antiepileptic drugs or general neurotrophic therapy, and two patients experienced continuous deterioration. CONCLUSIONS: Most patients with Hashimoto's encephalopathy showed good response to steroids. Some patients improved without steroid therapy. Considering its reversible course, we recommend that Hashimoto's encephalopathy should always be in the differential diagnosis while evaluating disorders of the central nervous system.

Mitochondrial dysfunction and oxidative stress in neurodegenerative diseases.

Many lines of evidence suggest that mitochondria have a central role in ageing-related neurodegenerative diseases. Mitochondria are critical regulators of cell death, a key feature of neurodegeneration. Mutations in mitochondrial DNA and oxidative stress both contribute to ageing, which is the greatest risk factor for neurodegenerative diseases. In all major examples of these diseases there is strong evidence that mitochondrial dysfunction occurs early and acts causally in disease pathogenesis. Moreover, an impressive number of disease-specific proteins interact with mitochondria. Thus, therapies targeting basic mitochondrial processes, such as energy metabolism or free-radical generation, or specific interactions of disease-related proteins with mitochondria, hold great promise.

Effects of synaptic modulation on beta-amyloid, synaptophysin, and memory performance in Alzheimer's disease transgenic mice.

Accumulation of ß-amyloid (Aß) and loss of synapses are hallmarks of Alzheimer's disease (AD). How synaptic activity relates to Aß accumulation and loss of synapses is a current topic of major interest. Synaptic activation promotes Aß secretion, and chronic reduction of synaptic activity reduced Aß plaques in an AD transgenic mouse model. This suggested beneficial effects of reducing synaptic activity in AD. We now show that reduced synaptic activity causes detrimental effects on synapses and memory despite reducing plaques using two different models of chronic synaptic inhibition: deafferentation of the barrel cortex and administration of benzodiazepine. An interval of prolonged synaptic inhibition exacerbated loss of synaptophysin compared with synaptically more active brain in AD transgenic but not wild-type mice. Furthermore, an interval of benzodiazepine treatment, followed by a washout period, exacerbated memory impairment in AD transgenic mice. Exacerbation of synaptic and behavioral abnormalities occurred in the setting of reduced Aß plaques but elevated intraneuronal Aß immunoreactivity. These data support beneficial effects of synaptic activation on Aß-related synaptic and behavioral impairment in AD.

Behavioral deficits and progressive neuropathology in progranulin-deficient mice: a mouse model of frontotemporal dementia.

Progranulin haploinsufficiency causes frontotemporal dementia with tau-negative, ubiquitin-positive neuronal inclusion pathology. In this study, we showed that progranulin-deficient mice displayed increased depression- and disinhibition-like behavior, as well as deficits in social recognition from a relatively young age. These mice did not have any deficit in locomotion or exploration. Eighteen-month-old progranulin-deficient mice demonstrated impaired spatial learning and memory in the Morris water maze. In addition to behavioral deficits, progranulin-deficient mice showed a progressive development of neuropathology from 12 mo of age, including enhanced activation of microglia and astrocytes and ubiquitination and cytoplasmic accumulation of phosphorylated TDP-43. Thus, progranulin deficiency induced FTD-like behavioral and neuropathological deficits. These mice may serve as an important tool for deciphering underlying mechanisms in frontotemporal dementia.

Synaptic activity reduces intraneuronal Abeta, promotes APP transport to synapses, and protects against Abeta-related synaptic alterations.

A central question in Alzheimer's disease research is what role synaptic activity plays in the disease process. Synaptic activity has been shown to induce beta-amyloid peptide release into the extracellular space, and extracellular beta-amyloid has been shown to be toxic to synapses. We now provide evidence that the well established synaptotoxicity of extracellular beta-amyloid requires gamma-secretase processing of amyloid precursor protein. Recent evidence supports an important role for intraneuronal beta-amyloid in the pathogenesis of Alzheimer's disease. We show that synaptic activity reduces intraneuronal beta-amyloid and protects against beta-amyloid-related synaptic alterations. We demonstrate that synaptic activity promotes the transport of the amyloid precursor protein to synapses using live cell imaging, and that the protease neprilysin is involved in reduction of intraneuronal beta-amyloid with synaptic activity.

Reduction of oxidative stress, amyloid deposition, and memory deficit by manganese superoxide dismutase overexpression in a transgenic mouse model of Alzheimer's disease.

In Alzheimer's disease (AD), oxidative stress is present early and contributes to disease pathogenesis. We previously reported that in Tg19959 transgenic AD mice, partial deficiency of the mitochondrial antioxidant enzyme manganese superoxide dismutase (MnSOD) exacerbated amyloid pathology. We therefore asked whether MnSOD overexpression would prove beneficial against AD pathogenesis, by studying the offspring of Tg19959 mice crossed with MnSOD-overexpressing mice. At 4 mo of age, there was a 2- to 3-fold increase in MnSOD protein levels in Tg19959-MnSOD mice compared to Tg19959 littermates. Tg19959-MnSOD mice also had a 50% increase in catalase protein levels, a 50% decrease in levels of oxidized protein, and a 33% reduction in cortical plaque burden compared to Tg19959 littermates. Spatial memory was impaired and synaptophysin levels were decreased in Tg19959 mice compared to wild-type littermates, but memory and synaptophysin levels were restored to wild-type levels in Tg19959-MnSOD littermates. These benefits occurred without changes in sodium dodecyl sulfate-soluble or formic acid-soluble Abeta pools or Abeta oligomers in Tg19959-MnSOD mice compared to Tg19959 littermates. These data demonstrate that facilitation of the mitochondrial antioxidant response improves resistance to Abeta, slows plaque formation or increases plaque degradation, and markedly attenuates the phenotype in a transgenic AD mouse model.

Triterpenoid CDDO-methylamide improves memory and decreases amyloid plaques in a transgenic mouse model of Alzheimer's disease.

Oxidative stress is one of the earliest events in the pathogenesis of Alzheimer's disease (AD) and can markedly exacerbate amyloid pathology. Modulation of antioxidant and anti-inflammatory pathways represents an important approach for AD therapy. Synthetic triterpenoids have been found to facilitate antioxidant response and reduce inflammation in several models. We investigated the effect of the triterpenoid, 2-Cyano-3,12-Dioxooleana-1,9-Dien-28-Oic acid-MethylAmide (CDDO-MA) in Tg19959 mice, which carry the human amyloid precursor protein with two mutations. These mice develop memory impairments and amyloid plaques as early as 2-3 months of age. CDDO-MA was provided with chow (800 mg/kg) from 1 to 4 months of age. CDDO-MA significantly improved spatial memory retention and reduced plaque burden, Abeta42 levels, microgliosis, and oxidative stress in Tg19959 mice.

Oligomerization of Alzheimer's beta-amyloid within processes and synapses of cultured neurons and brain.

Multiple lines of evidence implicate beta-amyloid (Abeta) in the pathogenesis of Alzheimer's disease (AD), but the mechanisms whereby Abeta is involved remain unclear. Addition of Abeta to the extracellular space can be neurotoxic. Intraneuronal Abeta42 accumulation is also associated with neurodegeneration. We reported previously that in Tg2576 amyloid precursor protein mutant transgenic mice, brain Abeta42 localized by immunoelectron microscopy to, and accumulated with aging in, the outer membranes of multivesicular bodies, especially in neuronal processes and synaptic compartments. We now demonstrate that primary neurons from Tg2576 mice recapitulate the in vivo localization and accumulation of Abeta42 with time in culture. Furthermore, we demonstrate that Abeta42 aggregates into oligomers within endosomal vesicles and along microtubules of neuronal processes, both in Tg2576 neurons with time in culture and in Tg2576 and human AD brain. These Abeta42 oligomer accumulations are associated with pathological alterations within processes and synaptic compartments in Tg2576 mouse and human AD brains.

Somatic mitochondrial DNA mutations in single neurons and glia.

Somatic mitochondrial DNA (mtDNA) point mutations reach high levels in the brain. However, the cell types that accumulate mutations and the patterns of mutations within individual cells are not known. We have quantified somatic mtDNA mutations in 28 single neurons and in 18 single glia from post-mortem human substantia nigra of six control subjects. Both neurons and glia contain mtDNA with somatic mutations. Single neurons harbor a geometric mean (95% CI) of 200.3 (152.9-262.4) somatic mtDNA point mutations per million base pairs, compared to 133.8 (97.5-184.9) for single glia (p=0.0251). If mutations detected multiple times in the same cell are counted only once, the mean mutation level per million base pairs remains elevated in single neurons (146.9; 124.0-174.2) compared to single glia (100.5; 81.5-126.5; p=0.009). Multiple distinct somatic point mutations are present in different cells from the same subject. Most of these mutations are individually present at low levels (less than 10-20% of mtDNA molecules), but with high aggregate mutation levels, particularly in neurons. These mutations may contribute to changes in brain function during normal aging and neurodegenerative disorders.

Somatic mitochondrial DNA mutations in cortex and substantia nigra in aging and Parkinson's disease.

Oxidative damage to mitochondrial DNA (mtDNA) increases with age in the brain and can induce G:C to T:A and T:A to G:C point mutations. Though rare at any particular site, multiple somatic mtDNA mutations induced by oxidative damage or by other mechanisms may accumulate with age in the brain and thus could play a role in aging and neurodegenerative diseases. However, no prior study has quantified the total burden of mtDNA point mutation subtypes in the brain. Using a highly sensitive cloning and sequencing strategy, we find that the aggregate levels of G:C to T:A and T:A to G:C transversions and of all point mutations increase with age in the frontal cortex (FCtx). In the substantia nigra (SN), the aggregate levels of point mutations in young controls are similar to the levels in the SN or FCtx of elderly subjects. Extrapolation from our data suggests an average of 2.7 (FCtx) to 3.2 (SN) somatic point mutations per mitochondrial genome in elderly subjects. There were no significant differences between Parkinson's disease (PD) patients and age-matched controls in somatic mutation levels. These results indicate that individually rare mtDNA point mutations reach a high aggregate burden in FCtx and SN of elderly subjects.

Accumulation of intraneuronal ß-amyloid 42 peptides is associated with early changes in microtubule-associated protein 2 in neurites and synapses.

Pathologic aggregation of ß-amyloid (Aß) peptide and the axonal microtubule-associated protein tau protein are hallmarks of Alzheimer's disease (AD). Evidence supports that Aß peptide accumulation precedes microtubule-related pathology, although the link between Aß and tau remains unclear. We previously provided evidence for early co-localization of Aß42 peptides and hyperphosphorylated tau within postsynaptic terminals of CA1 dendrites in the hippocampus of AD transgenic mice. Here, we explore the relation between Aß peptide accumulation and the dendritic, microtubule-associated protein 2 (MAP2) in the well-characterized amyloid precursor protein Swedish mutant transgenic mouse (Tg2576). We provide evidence that localized intraneuronal accumulation of Aß42 peptides is spatially associated with reductions of MAP2 in dendrites and postsynaptic compartments of Tg2576 mice at early ages. Our data support that reduction in MAP2 begins at sites of Aß42 monomer and low molecular weight oligomer (M/LMW) peptide accumulation. Cumulative evidence suggests that accumulation of M/LMW Aß42 peptides occurs early, before high molecular weight oligomerization and plaque formation. Since synaptic alteration is the best pathologic correlate of cognitive dysfunction in AD, the spatial association of M/LMW Aß peptide accumulation with pathology of MAP2 within neuronal processes and synaptic compartments early in the disease process reinforces the importance of intraneuronal Aß accumulation in AD pathogenesis.

Neuroprotection by cyclodextrin in cell and mouse models of Alzheimer disease.

There is extensive evidence that cholesterol and membrane lipids play a key role in Alzheimer disease (AD) pathogenesis. Cyclodextrins (CD) are cyclic oligosaccharide compounds widely used to bind cholesterol. Because CD exerts significant beneficial effects in Niemann-Pick type C disease, which shares neuropathological features with AD, we examined the effects of hydroxypropyl-ß-CD (HP-ß-CD) in cell and mouse models of AD. Cell membrane cholesterol accumulation was detected in N2a cells overexpressing Swedish mutant APP (SwN2a), and the level of membrane cholesterol was reduced by HP-ß-CD treatment. HP-ß-CD dramatically lowered the levels of Aß42 in SwN2a cells, and the effects were persistent for 24 h after withdrawal. 4 mo of subcutaneous HP-ß-CD administration significantly improved spatial learning and memory deficits in Tg19959 mice, diminished Aß plaque deposition, and reduced tau immunoreactive dystrophic neurites. HP-ß-CD lowered levels of Aß42 in part by reducing ß cleavage of the APP protein, and it also up-regulated the expression of genes involved in cholesterol transport and Aß clearance. This is the first study to show neuroprotective effects of HP-ß-CD in a transgenic mouse model of AD, both by reducing Aß production and enhancing clearance mechanisms, which suggests a novel therapeutic strategy for AD.

Coenzyme Q10 decreases amyloid pathology and improves behavior in a transgenic mouse model of Alzheimer's disease.

Increased oxidative stress is implicated in the pathogenesis of Alzheimer's disease (AD). A large body of evidence suggests that mitochondrial dysfunction and increased reactive oxygen species occur prior to amyloid-ß (Aß) deposition. Coenzyme Q10 (CoQ10), a component of the mitochondrial electron transport chain, is well characterized as a neuroprotective antioxidant in animal models and human trials of Huntington's disease and Parkinson's disease, and reduces plaque burden in AßPP/PS1 mice. We now show that CoQ10 reduces oxidative stress and amyloid pathology and improves behavioral performance in the Tg19959 mouse model of AD. CoQ10 treatment decreased brain levels of protein carbonyls, a marker of oxidative stress. CoQ10 treatment resulted in decreased plaque area and number in hippocampus and in overlying cortex immunostained with an Aß42-specific antibody. Brain Aß42 levels were also decreased by CoQ10 supplementation. Levels of amyloid-ß protein precursor (AßPP) ß-carboxyterminal fragments were decreased. Importantly, CoQ10-treated mice showed improved cognitive performance during Morris water maze testing. Our results show decreased pathology and improved behavior in transgenic AD mice treated with the naturally occurring antioxidant compound CoQ10. CoQ10 is well tolerated in humans and may be promising for therapeutic trials in AD.

Mitochondria and antioxidant targeted therapeutic strategies for Alzheimer's disease.

Oxidative stress and mitochondrial dysfunction are important features present in Alzheimer's disease (AD). They appear early and contribute to disease progression, both in human postmortem AD brains as well as in transgenic AD mouse brains. For this reason, targeting oxidative stress and mitochondria in AD may lead to the development of promising therapeutic strategies. Several exogenous antioxidant compounds have been tested and found beneficial in transgenic AD mice, such as vitamins and spices. However, their efficacy was much more modest in human trials. More recently, new strategies have been elaborated to promote endogenous antioxidant systems. Different pathways involved in oxidative stress response have been identified. Compounds able to upregulate these pathways are being generated and tested in animal models of AD and in human patients. Upregulation of antioxidant gene expression was beneficial in mice, giving hope for future avenues in the treatment of AD and other neurodegenerative disorders.

MicroRNA-related cofilin abnormality in Alzheimer's disease.

Rod-like structures composed of actin and the actin-binding protein cofilin are found in Alzheimer's disease (AD) patients. However, the mechanisms underlying formation of these structures and their pathological consequences are still largely unknown. We found that microRNAs 103 and 107 repress translation of cofilin, and that reduced levels of miR-103 or miR-107 are associated with elevated cofilin protein levels and formation of rod-like structures in a transgenic mouse model of AD. These results suggest that microRNAs may play an important role in cytoskeletal pathology in AD.

N-iminoethyl-L-lysine improves memory and reduces amyloid pathology in a transgenic mouse model of amyloid deposition.

A large body of evidence suggests the importance of inflammation and oxidative or nitrosative stress in Alzheimer's disease (AD) pathogenesis. Inflammatory stimuli upregulate transcription of inducible nitric oxide synthase (iNOS), which can lead to the production of nitric oxide and other reactive nitrogen species. We previously found that genetic deletion of iNOS in mice overexpressing the amyloid precursor protein (APP) and presenilin-1 (PS1) reduced mortality, nitrosative stress, amyloid plaque burden, microgliosis, astrocytosis, and peri-plaque tau phosphorylation. We therefore examined the effects of N6-(1-iminoethyl)-L-lysine (L-NIL), a pharmacological iNOS inhibitor, or d-NIL, its enantiomeric control, in a transgenic mouse model of amyloid deposition. Tg19959 mice carry human APP with two mutations and develop amyloid plaques and memory impairment starting at 3-4 months of age. Mice were given L-NIL or D-NIL in the drinking water from 1 month of age and assessed behaviorally and histopathologically at 8 months of age. We found that L-NIL administration reduced disinhibition in the elevated plus maze, improved spatial memory performance in the Morris water maze, and decreased cortical amyloid deposition as well as microglial activation in 8-month-old Tg19959 mice. These findings are consistent with previous reports demonstrating that iNOS inhibition ameliorates AD pathogenesis.