Noninvasive Antemortem Detection of Retinal Prions by a Fluorescent Tracer.

BACKGROUND: Neurodegenerative diseases are widespread yet challenging to diagnose and stage antemortem. As an extension of the central nervous system, the eye harbors retina ganglion cells vulnerable to degeneration, and visual symptoms are often an early manifestation of neurodegenerative disease. OBJECTIVE: Here we test whether prion protein aggregates could be detected in the eyes of live mice using an amyloid-binding fluorescent probe and high-resolution retinal microscopy. METHODS: We performed retinal imaging on an experimental mouse model of prion-associated cerebral amyloid angiopathy in a longitudinal study. An amyloid-binding fluorophore was intravenously administered, and retinal imaging was performed at timepoints corresponding to early, mid-, and terminal prion disease. Retinal amyloid deposits were quantified and compared to the amyloid load in the brain. RESULTS: We report that by early prion disease (50% timepoint), discrete fluorescent foci appeared adjacent to the optic disc. By later timepoints, the fluorescent foci surrounded the optic disc and tracked along retinal vasculature. CONCLUSION: The progression of perivascular amyloid can be directly monitored in the eye by live imaging, illustrating the utility of this technology for diagnosing and monitoring the progression of cerebral amyloid angiopathy.

Mutant three-repeat tau expression initiates retinal ganglion cell death through Caspase-2.

The microtubule-associated protein tau is implicated in multiple degenerative diseases including retinal diseases such as glaucoma; however, the way tau initiates retinopathy is unclear. Previous retinal assessments in mouse models of tauopathy suggest that mutations in four-repeat (4R) tau are associated with disease-induced retinal dysfunction, while shifting tau isoform ratio to favor three-repeat (3R) tau production enhanced photoreceptor function. To further understand how alterations in tau expression impact the retina, we analyzed the retinas of transgenic mice overexpressing mutant 3R tau (m3R tau-Tg), a model known to exhibit Pick's Disease pathology in the brain. Analysis of retinal cross-sections from young (3 month) and adult (9 month) mice detected asymmetric 3R tau immunoreactivity in m3R tau-Tg retina, concentrated in the retinal ganglion and amacrine cells of the dorsal retinal periphery. Accumulation of hyperphosphorylated tau was detected specifically in the detergent insoluble fraction of the adult m3R tau-Tg retina. RNA-seq analysis highlighted biological pathways associated with tauopathy that were uniquely altered in m3R tau-Tg retina. The upregulation of transcript encoding apoptotic protease caspase-2 coincided with increased immunostaining in predominantly 3R tau positive retinal regions. In adult m3R tau-Tg, the dorsal peripheral retina of the adult m3R tau-Tg exhibited decreased cell density in the ganglion cell layer (GCL) and reduced thickness of the inner plexiform layer (IPL) compared to the ventral peripheral retina. Together, these data indicate that mutant 3R tau may mediate toxicity in retinal ganglion cells (RGC) by promoting caspase-2 expression which results in RGC degeneration. The m3R tau-Tg line has the potential to be used to assess tau-mediated RGC degeneration and test novel therapeutics for degenerative diseases such as glaucoma.

Neuroprotective Effects of Chronic Resveratrol Treatment and Exercise Training in the 3xTg-AD Mouse Model of Alzheimer's Disease.

To date, there is no cure or effective treatment for Alzheimer's disease (AD), a chronic neurodegenerative condition that affects memory, language, and behavior. AD is characterized by neuroinflammation, accumulation of brain amyloid-beta (Aß) oligomers and neurofibrillary tangles, increased neuronal apoptosis, and loss of synaptic function. Promoting regular exercise and a diet containing polyphenols are effective non-pharmacological approaches that prevent the progression of neurodegenerative diseases. In this study, we measured various conformational toxic species of Aß and markers of inflammation, apoptosis, endolysosomal degradation, and neuroprotection after 5 months of exercise training (ET), resveratrol (Resv) treatment, or combination treatment in the 3xTg-AD mouse model of AD. Our main results indicate that Resv decreased neuroinflammation and accumulation of Aß oligomers, increased levels of neurotrophins, synaptic markers, silent information regulator, and decreased markers of apoptosis, autophagy, endolysosomal degradation and ubiquitination in the brains of 3xTg-AD mice. ET improved some markers related to neuroprotection, but when combined with Resv treatment, the benefits achieved were as effective as Resv treatment alone. Our results show that the neuroprotective effects of Resv, ET or Resv and ET are associated with reduced toxicity of Aß oligomers, suppression of neuronal autophagy, decreased apoptosis, and upregulation of key growth-related proteins.

Vitamin D fortification of foods in India: present and past scenario.

India is a densely populated country known for its traditional, cultural and lingual diversity. In India Deficiency of vitamin D is seen in both the genders and among all the age groups. Micronutrient deficiencies are steadily adding to the increasing burden of health related co-morbidities with low dietary calcium and magnesium intake in Indian population. Despite lots of sunshine, vitamin D insufficiency is widespread in India according to the age and regions (50-90%). In order to increase vitamin D intake with normal diet, the food industry will have to find a more effective strategy to improve general health conditions. The breadth of widely used foods, including milk, cheese, margarine, dairy products, and various breakfast drinks, can improve the condition of vitamin D deficiency in Asian countries such as India. The requirements for calculating the necessary micronutrients and vitamin D fortification of foods and drinks, successful strategies should be developed and emphasized. There is need to improve the effectiveness of various fruit drinks through fortification of vitamin D, which can reduce vitamin D deficiency in the general population as well as in different age groups.

Effects and Underlying Mechanisms of Bioactive Compounds on Type 2 Diabetes Mellitus and Alzheimer's Disease.

Type 2 diabetes mellitus is a complicated metabolic disorder characterized by hyperglycemia and glucose intolerance. Alzheimer's disease is a progressive brain disorder characterized by a chronic loss of cognitive and behavioral function. Considering the shared characteristics of both diseases, common therapeutic and preventive agents may be effective. Bioactive compounds such as polyphenols, vitamins, and carotenoids found in vegetables and fruits can have antioxidant and anti-inflammatory effects. These effects make them suitable candidates for the prevention or treatment of diabetes and Alzheimer's disease. Increasing evidence from cell or animal models suggest that bioactive compounds may have direct effects on decreasing hyperglycemia, enhancing insulin secretion, and preventing formation of amyloid plaques. The possible underlying molecular mechanisms are described in this review. More studies are needed to establish the clinical effects of bioactive compounds.

High Fat With High Sucrose Diet Leads to Obesity and Induces Myodegeneration.

Skeletal muscle utilizes both free fatty acids (FFAs) and glucose that circulate in the blood stream. When blood glucose levels acutely increase, insulin stimulates muscle glucose uptake, oxidation, and glycogen synthesis. Under these conditions, skeletal muscle preferentially oxidizes glucose while the oxidation of fatty acids (FAs) oxidation is reciprocally decreased. In metabolic disorders associated with insulin resistance, such as diabetes and obesity, both glucose uptake, and utilization muscle are significantly reduced causing FA oxidation to provide the majority of ATP for metabolic processes and contraction. Although the causes of this metabolic inflexibility or disrupted "glucose-fatty acid cycle" are largely unknown, a diet high in fat and sugar (HFS) may be a contributing factor. This metabolic inflexibility observed in models of obesity or with HFS feeding is detrimental because high rates of FA oxidation in skeletal muscle can lead to the buildup of toxic metabolites of fat metabolism and the accumulation of pro-inflammatory cytokines, which further exacerbate the insulin resistance. Further, HFS leads to skeletal muscle atrophy with a decrease in myofibrillar proteins and phenotypically characterized by loss of muscle mass and strength. Overactivation of ubiquitin proteasome pathway, oxidative stress, myonuclear apoptosis, and mitochondrial dysfunction are some of the mechanisms involved in muscle atrophy induced by obesity or in mice fed with HFS. In this review, we will discuss how HFS diet negatively impacts the various physiological and metabolic mechanisms in skeletal muscle.

Prophylactic Active Tau Immunization Leads to Sustained Reduction in Both Tau and Amyloid-ß Pathologies in 3xTg Mice.

Amyloid-ß (Aß) and tau pathologies are intertwined in Alzheimer's disease, and various immunotherapies targeting these hallmarks are in clinical trials. To determine if tau pathology influences Aß burden and to assess prophylactic benefits, 3xTg and wild-type mice received tau immunization from 2-6 months of age. The mice developed a high IgG titer that was maintained at 22 months of age. Pronounced tau and Aß pathologies were primarily detected in the subiculum/CA1 region, which was therefore the focus of analysis. The therapy reduced histopathological tau aggregates by 70-74% overall (68% in males and 78-86% in females), compared to 3xTg controls. Likewise, western blot analysis revealed a 41% clearance of soluble tau (38-76% in males and 48% in females) and 42-47% clearance of insoluble tau (47-58% in males and 49% in females) in the immunized mice. Furthermore, Aß burden was reduced by 84% overall (61% in males and 97% in females). These benefits were associated with reductions in microgliosis and microhemorrhages. In summary, prophylactic tau immunization not only prevents tau pathology but also Aß deposition and related pathologies in a sustained manner, indicating that tau pathology can promote Aß deposition, and that a short immunization regimen can have a long-lasting beneficial effect.

Affinity of Tau antibodies for solubilized pathological Tau species but not their immunogen or insoluble Tau aggregates predicts in vivo and ex vivo efficacy.

BACKGROUND: A few tau immunotherapies are now in clinical trials with several more likely to be initiated in the near future. A priori, it can be anticipated that an antibody which broadly recognizes various pathological tau aggregates with high affinity would have the ideal therapeutic properties. Tau antibodies 4E6 and 6B2, raised against the same epitope region but of varying specificity and affinity, were tested for acutely improving cognition and reducing tau pathology in transgenic tauopathy mice and neuronal cultures. RESULTS: Surprisingly, we here show that one antibody, 4E6, which has low affinity for most forms of tau acutely improved cognition and reduced soluble phospho-tau, whereas another antibody, 6B2, which has high affinity for various tau species was ineffective. Concurrently, we confirmed and clarified these efficacy differences in an ex vivo model of tauopathy. Alzheimer's paired helical filaments (PHF) were toxic to the neurons and increased tau levels in remaining neurons. Both toxicity and tau seeding were prevented by 4E6 but not by 6B2. Furthermore, 4E6 reduced PHF spreading between neurons. Interestingly, 4E6's efficacy relates to its high affinity binding to solubilized PHF, whereas the ineffective 6B2 binds mainly to aggregated PHF. Blocking 4E6's uptake into neurons prevented its protective effects if the antibody was administered after PHF had been internalized. When 4E6 and PHF were administered at the same time, the antibody was protective extracellularly. CONCLUSIONS: Overall, these findings indicate that high antibody affinity for solubilized PHF predicts efficacy, and that acute antibody-mediated improvement in cognition relates to clearance of soluble phospho-tau. Importantly, both intra- and extracellular clearance pathways are in play. Together, these results have major implications for understanding the pathogenesis of tauopathies and for development of immunotherapies.

Structural differences between amyloid beta oligomers.

In Alzheimer's disease, soluble Aß oligomers are believed to play important roles in the disease pathogenesis, and their levels correlate with cognitive impairment. We have previously shown that Aß oligomers can be categorized into multiple structural classes based on their reactivity with conformation-dependent antibodies. In this study, we analyzed the structures of Aß40 oligomers belonging to two of these classes: fibrillar and prefibrillar oligomers. We found that fibrillar oligomers were similar in structure to fibrils but were less stable towards denaturation while prefibrillar oligomers were found to be partially disordered. These results are consistent with previously proposed structures for both oligomer classes while providing additional structural information.

Internalization of tau antibody and pathological tau protein detected with a flow cytometry multiplexing approach.

INTRODUCTION: Tau immunotherapy has emerged as a promising approach to clear tau aggregates from the brain. Our previous findings suggest that tau antibodies may act outside and within neurons to promote such clearance. METHODS: We have developed an approach using flow cytometry, a human neuroblastoma cell model overexpressing tau with the P301L mutation, and paired helical filament (PHF)-enriched pathologic tau to effectively screen uptake and retention of tau antibodies in conjunction with PHF. RESULTS: The flow cytometry approach correlates well with Western blot analysis to detect internalized antibodies in naïve and transfected SH-SY5Y cells (r2 = 0.958, and r2 = 0.968, P = .021 and P = .016, respectively). In transfected cells, more antibodies are taken up/retained as pathologic tau load increases, both under co-treated conditions and when the cells are pretreated with PHF before antibody administration (r2 = 0.999 and r2 = 0.999, P = .013 and P = .011, respectively). DISCUSSION: This approach allows rapid in vitro screening of antibody uptake and retention in conjunction with pathologic tau protein before more detailed studies in animals or other more complex model systems.

Intracellular amyloid and the neuronal origin of Alzheimer neuritic plaques.

Genetic analysis of familial forms of Alzheimer's disease (AD) causally links the proteolytic processing of the amyloid precursor protein (APP) and AD. However, the specific type of amyloid and mechanisms of amyloid pathogenesis remain unclear. We conducted a detailed analysis of intracellular amyloid with an aggregation specific conformation dependent monoclonal antibody, M78, raised against fibrillar Aß42. M78 immunoreactivity colocalizes with Aß and the carboxyl terminus of APP (APP-CTF) immunoreactivities in perinuclear compartments at intermediate times in 10month 3XTg-AD mice, indicating that this represents misfolded and aggregated protein rather than normally folded APP. At 12months, M78 immunoreactivity also accumulates in the nucleus. Neuritic plaques at 12months display the same spatial organization of centrally colocalized M78, diffuse chromatin and neuronal nuclear NeuN staining surrounded by peripheral M78 and APP-CTF immunoreactivity as observed in neurons, indicating that neuritic plaques arise from degenerating neurons with intracellular amyloid immunoreactivity. The same staining pattern was observed in neuritic plaques in human AD brains, showing elevated intracellular M78 immunoreactivity at intermediate stages of amyloid pathology (Braak A and B) compared to no amyloid pathology and late stage amyloid pathology (Braak 0 and C, respectively). These results indicate that intraneuronal protein aggregation and amyloid accumulation is an early event in AD and that neuritic plaques are initiated by the degeneration and death of neurons by a mechanism that may be related to the formation of extracellular traps by neutrophils.

Nonhuman amyloid oligomer epitope reduces Alzheimer's-like neuropathology in 3xTg-AD transgenic mice.

Accumulation of beta-amyloid (Aß) is an important pathological event in Alzheimer's disease (AD). It is now well known that vaccination against fibrillar Aß prevents amyloid accumulation and preserves cognitive function in transgenic mouse models. To study the effect of vaccination against generic oligomer epitopes, Aß oligomers, islet amyloid polypeptide oligomers, random peptide oligomer (3A), and Aß fibrils were used to vaccinate 3xTg-AD, which develop a progressive accumulation of plaques and cognitive impairment. Subcutaneous administration of these antigens markedly reduced total plaque load (Aß burden) and improved cognitive function in the 3xTg-AD mouse brains as compared to controls. We demonstrated that vaccination with this nonhuman amyloid oligomer generated high titers of specifically antibodies recognizing Aß oligomers, which in turn inhibited accumulation of Aß pathology in mice. In addition to amyloid plaques, another hallmark of AD is tau pathology. It was found that there was a significant decline in the level of hyper-phosphorylated tau following vaccination. We have previously shown that immunization with 3A peptide improves cognitive function and clears amyloid plaques in Tg2576 mice, which provides a novel strategy of AD therapy. Here, we have shown that vaccination with 3A peptide in 3xTg-AD mice not only clears amyloid plaques but also extensively clears abnormal tau in brain.

Systemic vaccination with anti-oligomeric monoclonal antibodies improves cognitive function by reducing Aß deposition and tau pathology in 3xTg-AD mice.

Alzheimer's disease (AD) is a devastating disorder that is clinically characterized by a comprehensive cognitive decline. Accumulation of the amyloid-beta (Aß) peptide plays a pivotal role in the pathogenesis of AD. In AD, the conversion of Aß from a physiological soluble monomeric form into insoluble fibrillar conformation is an important event. The most toxic form of Aß is oligomers, which is the intermediate step during the conversion of monomeric form to fibrillar form. There are at least two types of oligomers: oligomers that are immunologically related to fibrils and those that are not. In transgenic AD animal models, both active and passive anti-Aß immunotherapies improve cognitive function and clear the parenchymal accumulation of amyloid plaques in the brain. In this report we studied effect of immunotherapy of two sequence-independent non-fibrillar oligomer specific monoclonal antibodies on the cognitive function, amyloid load and tau pathology in 3xTg-AD mice. Anti-oligomeric monoclonal antibodies significantly reduce the amyloid load and improve the cognition. The clearance of amyloid load was significantly correlated with reduced tau hyperphosphorylation and improvement in cognition. These results demonstrate that systemic immunotherapy using oligomer-specific monoclonal antibodies effectively attenuates behavioral and pathological impairments in 3xTg-AD mice. These findings demonstrate the potential of using oligomer specific monoclonal antibodies as a therapeutic approach to prevent and treat Alzheimer's disease.

Immunogenicity, efficacy, safety, and mechanism of action of epitope vaccine (Lu AF20513) for Alzheimer's disease: prelude to a clinical trial.

The Alzheimer's disease (AD) process is understood to involve the accumulation of amyloid plaques and tau tangles in the brain. However, attempts at targeting the main culprits, neurotoxic Aß peptides, have thus far proven unsuccessful for improving cognitive function. Recent clinical trials with passively administrated anti-Aß antibodies failed to slow cognitive decline in mild to moderate AD patients, but suggest that an immunotherapeutic approach could be effective in patients with mild AD. Using an AD mouse model (Tg2576), we tested the immunogenicity (cellular and humoral immune responses) and efficacy (AD-like pathology) of clinical grade Lu AF20513 vaccine. We found that Lu AF20513 induces robust "non-self" T-cell responses and the production of anti-Aß antibodies that reduce AD-like pathology in the brains of Tg2576 mice without inducing microglial activation and enhancing astrocytosis or cerebral amyloid angiopathy. A single immunization with Lu AF20513 induced strong humoral immunity in mice with preexisting memory T-helper cells. In addition, Lu AF20513 induced strong humoral responses in guinea pigs and monkeys. These data support the translation of Lu AF20513 to the clinical setting with the aims of: (1) inducing therapeutically potent anti-Aß antibody responses in patients with mild AD, particularly if they have memory T-helper cells generated after immunizations with conventional tetanus toxoid vaccine, and (2) preventing pathological autoreactive T-cell responses.

Presynaptic membrane receptor in human brain.

Myasthenia gravis (MG) is an autoimmune disease that results from antibody mediated damage of Acetylcholine receptor (AChR) at the neuromuscular junction. The autoimmune character of MG and pathogenic role of AChR antibodies have been established by several workers i.e., the demonstration of anti-AChR antibodies in about 90 % of MG patients. It has been demonstrated that patients with MG also have antibodies against a second protein named presynaptic membrane receptor (PsmR), which is identified by utilizing ß-Bgtx, a ligand which binds to PsmR. Using ß-Bgtx Sepharose 4B affinity matrix, the PsmR was purified from different regions of human cadaver brain by affinity chromatography. Purified receptor was characterized both by biochemical and immunological procedures. PsmR purified from different regions of the brain shows a specific activity of 0.37 ± 0.01, 0.39 ± 0.02 and 0.43 ± 0.005 nM/ µg of protein in Parietal lobe, Occipital lobe and Frontal lobe respectively. The affinity purified PsmR from the brain of 87 and 68 kd (parietal lobe, occipital lobe and frontal lobe) shows immunoreactivity with myasthenic sera. These findings suggest that PsmR from brain is another antigen against which autoantibodies are developed in Myasthenia gravis patients. Upon treatment with various enzymes we concluded that PsmR from brain is a glycoprotein in which the immunoreactivity resides in the carbohydrate as well as the peptide epitopes. In conclusion the PsmR is another antigen against which autoantibodies are formed in different regions of brain. These can be used as a diagnostic tool for detecting antibodies in the sera or cerebrospinal fluid of MG patients.

Vaccination with a non-human random sequence amyloid oligomer mimic results in improved cognitive function and reduced plaque deposition and micro hemorrhage in Tg2576 mice.

BACKGROUND: It is well established that vaccination of humans and transgenic animals against fibrillar Aß prevents amyloid accumulation in plaques and preserves cognitive function in transgenic mouse models. However, autoimmune side effects have halted the development of vaccines based on full length human Aß. Further development of an effective vaccine depends on overcoming these side effects while maintaining an effective immune response. RESULTS: We have previously reported that the immune response to amyloid oligomers is largely directed against generic epitopes that are common to amyloid oligomers of many different proteins and independent of a specific amino acid sequence. Here we have examined whether we can exploit this generic immune response to develop a vaccine that targets amyloid oligomers using a non-human random sequence amyloid oligomer. In order to study the effect of vaccination against generic oligomer epitopes, a random sequence oligomer (3A) was selected as it forms oligomers that react with the oligomer specific A11 antibody. Oligomer mimics from 3A peptide, Aß, islet amyloid polypeptide (IAPP), and Aß fibrils were used to vaccinate Tg2576 mice, which develop a progressive accumulation of plaques and cognitive impairment. Vaccination with the 3A random sequence antigen was just as effective as vaccination with the other antigens in improving cognitive function and reducing total plaque load (Aß burden) in the Tg2576 mouse brains, but was associated with a much lower incidence of micro hemorrhage than Aß antigens. CONCLUSION: These results shows that the amyloid Aß sequence is not necessary to produce a protective immune response that specifically targets generic amyloid oligomers. Using a non-human, random sequence antigen may facilitate the development of a vaccine that avoids autoimmune side effects.

Swedish Alzheimer mutation induces mitochondrial dysfunction mediated by HSP60 mislocalization of amyloid precursor protein (APP) and beta-amyloid.

Alzheimer disease (AD) is a complex disorder that involves numerous cellular and subcellular alterations including impairments in mitochondrial homeostasis. To better understand the role of mitochondrial dysfunction in the pathogenesis of AD, we analyzed brains from clinically well-characterized human subjects and from the 3xTg-AD mouse model of AD. We find Aß and critical components of the γ-secretase complex, presenilin-1, -2, and nicastrin, accumulate in the mitochondria. We used a proteomics approach to identify binding partners and show that heat shock protein 60 (HSP60), a molecular chaperone localized to mitochondria and the plasma membrane, specifically associates with APP. We next generated stable neural cell lines expressing human wild-type or Swedish APP, and provide corroborating in vitro evidence that HSP60 mediates translocation of APP to the mitochondria. Viral-mediated shRNA knockdown of HSP60 attenuates APP and Aß mislocalization to the mitochondria. Our findings identify a novel interaction between APP and HSP60, which accounts for its translocation to the mitochondria.

Effect of synthetic aß peptide oligomers and fluorinated solvents on Kv1.3 channel properties and membrane conductance.

The impact of synthetic amyloid ß (1-42) (Aß(1-42)) oligomers on biophysical properties of voltage-gated potassium channels Kv 1.3 and lipid bilayer membranes (BLMs) was quantified for protocols using hexafluoroisopropanol (HFIP) or sodium hydroxide (NaOH) as solvents prior to initiating the oligomer formation. Regardless of the solvent used Aß(1-42) samples contained oligomers that reacted with the conformation-specific antibodies A11 and OC and had similar size distributions as determined by dynamic light scattering. Patch-clamp recordings of the potassium currents showed that synthetic Aß(1-42) oligomers accelerate the activation and inactivation kinetics of Kv 1.3 current with no significant effect on current amplitude. In contrast to oligomeric samples, freshly prepared, presumably monomeric, Aß(1-42) solutions had no effect on Kv 1.3 channel properties. Aß(1-42) oligomers had no effect on the steady-state current (at -80 mV) recorded from Kv 1.3-expressing cells but increased the conductance of artificial BLMs in a dose-dependent fashion. Formation of amyloid channels, however, was not observed due to conditions of the experiments. To exclude the effects of HFIP (used to dissolve lyophilized Aß(1-42) peptide), and trifluoroacetic acid (TFA) (used during Aß(1-42) synthesis), we determined concentrations of these fluorinated compounds in the stock Aß(1-42) solutions by (19)F NMR. After extensive evaporation, the concentration of HFIP in the 100× stock Aß(1-42) solutions was ∼1.7 µM. The concentration of residual TFA in the 70× stock Aß(1-42) solutions was ∼20 µM. Even at the stock concentrations neither HFIP nor TFA alone had any effect on potassium currents or BLMs. The Aß(1-42) oligomers prepared with HFIP as solvent, however, were more potent in the electrophysiological tests, suggesting that fluorinated compounds, such as HFIP or structurally-related inhalational anesthetics, may affect Aß(1-42) aggregation and potentially enhance ability of oligomers to modulate voltage-gated ion channels and biological membrane properties.

CD45 deficiency drives amyloid-ß peptide oligomers and neuronal loss in Alzheimer's disease mice.

Converging lines of evidence indicate dysregulation of the key immunoregulatory molecule CD45 (also known as leukocyte common antigen) in Alzheimer's disease (AD). We report that transgenic mice overproducing amyloid-ß peptide (Aß) but deficient in CD45 (PSAPP/CD45(-/-) mice) faithfully recapitulate AD neuropathology. Specifically, we find increased abundance of cerebral intracellular and extracellular soluble oligomeric and insoluble Aß, decreased plasma soluble Aß, increased abundance of microglial neurotoxic cytokines tumor necrosis factor-α and interleukin-1ß, and neuronal loss in PSAPP/CD45(-/-) mice compared with CD45-sufficient PSAPP littermates (bearing mutant human amyloid precursor protein and mutant human presenilin-1 transgenes). After CD45 ablation, in vitro and in vivo studies demonstrate an anti-Aß phagocytic but proinflammatory microglial phenotype. This form of microglial activation occurs with elevated Aß oligomers and neural injury and loss as determined by decreased ratio of anti-apoptotic Bcl-xL to proapoptotic Bax, increased activated caspase-3, mitochondrial dysfunction, and loss of cortical neurons in PSAPP/CD45(-/-) mice. These data show that deficiency in CD45 activity leads to brain accumulation of neurotoxic Aß oligomers and validate CD45-mediated microglial clearance of oligomeric Aß as a novel AD therapeutic target.

Conformation dependent monoclonal antibodies distinguish different replicating strains or conformers of prefibrillar Aß oligomers.

BACKGROUND: Age-related neurodegenerative diseases share a number of important pathological features, such as accumulation of misfolded proteins as amyloid oligomers and fibrils. Recent evidence suggests that soluble amyloid oligomers and not the insoluble amyloid fibrils may represent the primary pathological species of protein aggregates. RESULTS: We have produced several monoclonal antibodies that specifically recognize prefibrillar oligomers and do not recognize amyloid fibrils, monomer or natively folded proteins. Like the polyclonal antisera, the individual monoclonals recognize generic epitopes that do not depend on a specific linear amino acid sequence, but they display distinct preferences for different subsets of prefibrillar oligomers. Immunological analysis of a number of different prefibrillar Aß oligomer preparations show that structural polymorphisms exist in Aß prefibrillar oligomers that can be distinguished on the basis of their reactivity with monoclonal antibodies. Western blot analysis demonstrates that the conformers defined by the monoclonal antibodies have distinct size distributions, indicating that oligomer structure varies with size. The different conformational types of Aß prefibrillar oligomers can serve as they serve as templates for monomer addition, indicating that they seed the conversion of Aß monomer into more prefibrillar oligomers of the same type. CONCLUSIONS: These results indicate that distinct structural variants or conformers of prefibrillar Aß oligomers exist that are capable of seeding their own replication. These conformers may be analogous to different strains of prions.