Deep Learning-Based Image Classification in Differentiating Tufted Astrocytes, Astrocytic Plaques, and Neuritic Plaques.

This study aimed to develop a deep learning-based image classification model that can differentiate tufted astrocytes (TA), astrocytic plaques (AP), and neuritic plaques (NP) based on images of tissue sections stained with phospho-tau immunohistochemistry. Phospho-tau-immunostained slides from the motor cortex were scanned at 20× magnification. An automated deep learning platform, Google AutoML, was used to create a model for distinguishing TA in progressive supranuclear palsy (PSP) from AP in corticobasal degeneration (CBD) and NP in Alzheimer disease (AD). A total of 1500 images of representative tau lesions were captured from 35 PSP, 27 CBD, and 33 AD patients. Of those, 1332 images were used for training, and 168 images for cross-validation. We tested the model using 100 additional test images taken from 20 patients of each disease. In cross-validation, precision and recall for each individual lesion type were 100% and 98.0% for TA, 98.5% and 98.5% for AP, and 98.0% and 100% for NP, respectively. In a test set, all images of TA and NP were correctly predicted. Only eleven images of AP were predicted to be TA or NP. Our data indicate the potential usefulness of deep learning-based image classification methods to assist in differential diagnosis of tauopathies.

Label-free vibrational imaging of different Aß plaque types in Alzheimer's disease reveals sequential events in plaque development.

The neuropathology of Alzheimer's disease (AD) is characterized by hyperphosphorylated tau neurofibrillary tangles (NFTs) and amyloid-beta (Aß) plaques. Aß plaques are hypothesized to follow a development sequence starting with diffuse plaques, which evolve into more compact plaques and finally mature into the classic cored plaque type. A better molecular understanding of Aß pathology is crucial, as the role of Aß plaques in AD pathogenesis is under debate. Here, we studied the deposition and fibrillation of Aß in different plaque types with label-free infrared and Raman imaging. Fourier-transform infrared (FTIR) and Raman imaging was performed on native snap-frozen brain tissue sections from AD cases and non-demented control cases. Subsequently, the scanned tissue was stained against Aß and annotated for the different plaque types by an AD neuropathology expert. In total, 160 plaques (68 diffuse, 32 compact, and 60 classic cored plaques) were imaged with FTIR and the results of selected plaques were verified with Raman imaging. In diffuse plaques, we detect evidence of short antiparallel ß-sheets, suggesting the presence of Aß oligomers. Aß fibrillation significantly increases alongside the proposed plaque development sequence. In classic cored plaques, we spatially resolve cores containing predominantly large parallel ß-sheets, indicating Aß fibrils. Combining label-free vibrational imaging and immunohistochemistry on brain tissue samples of AD and non-demented cases provides novel insight into the spatial distribution of the Aß conformations in different plaque types. This way, we reconstruct the development process of Aß plaques in human brain tissue, provide insight into Aß fibrillation in the brain, and support the plaque development hypothesis.

Amyloid polymorphisms constitute distinct clouds of conformational variants in different etiological subtypes of Alzheimer's disease.

The molecular architecture of amyloids formed in vivo can be interrogated using luminescent conjugated oligothiophenes (LCOs), a unique class of amyloid dyes. When bound to amyloid, LCOs yield fluorescence emission spectra that reflect the 3D structure of the protein aggregates. Given that synthetic amyloid-ß peptide (Aß) has been shown to adopt distinct structural conformations with different biological activities, we asked whether Aß can assume structurally and functionally distinct conformations within the brain. To this end, we analyzed the LCO-stained cores of ß-amyloid plaques in postmortem tissue sections from frontal, temporal, and occipital neocortices in 40 cases of familial Alzheimer's disease (AD) or sporadic (idiopathic) AD (sAD). The spectral attributes of LCO-bound plaques varied markedly in the brain, but the mean spectral properties of the amyloid cores were generally similar in all three cortical regions of individual patients. Remarkably, the LCO amyloid spectra differed significantly among some of the familial and sAD subtypes, and between typical patients with sAD and those with posterior cortical atrophy AD. Neither the amount of Aß nor its protease resistance correlated with LCO spectral properties. LCO spectral amyloid phenotypes could be partially conveyed to Aß plaques induced by experimental transmission in a mouse model. These findings indicate that polymorphic Aß-amyloid deposits within the brain cluster as clouds of conformational variants in different AD cases. Heterogeneity in the molecular architecture of pathogenic Aß among individuals and in etiologically distinct subtypes of AD justifies further studies to assess putative links between Aß conformation and clinical phenotype.

[Expression of cytokine IL-1α and S100ß in different types of plaques in Alzheimer's disease].

OBJECTIVE: To study the significance of cytokine IL-1α and S100ß expression in formation and evolution of different types of plaques in Alzheimer's disease. METHODS: Thirty-four autopsy cases of Alzheimer's disease encountered during the period from 1982 to 2008 were retrieved from the archival files of Department of Pathology, Beijing Hospital. Tissue blocks were taken from hippocampus for dual immunostaining for IL-1α/Aß and S100ß/Aß. RESULTS: Immunohistochemical studied for IL-1α/Aß and S100ß/Aß delineated four different types of senile plaques: diffuse non-neuritic plaques, diffuse neuritic plaques, dense-core neuritic plaques and dense-core non-neuritic plaques. The numbers of IL-1α-positive microglias and S100ß-positive astrocytes associated with diffuse neuritic plaques were (7.29 ± 3.04) per mm(2) and (6.49 ± 2.20) per mm(2), respectively. In contrast, the numbers of IL-1α-positive microglias and S100ß-positive astrocytes associated with diffuse non-neuritic plaques, dense-core neuritic plaques and dense-core non-neuritic plaques were (3.24 ± 1.53) per mm(2) and (4.14 ± 1.77) per mm(2), (2.09 ± 1.37) per mm(2) and (2.25 ± 0.83) per mm(2), and (1.38 ± 0.90) per mm(2) and (0.58 ± 0.36) per mm(2), respectively. The numbers of IL-1α-positive microglias and S100ß-positive astrocytes associated with diffuse neuritic plaques were significantly higher than those of the other three types of plaques (P < 0.05). CONCLUSION: The IL-1α-positive microglias and S100ß-positive astrocytes may be of certain significance in transformation of diffuse non-neuritic plaques to diffuse neuritic plaques in Alzheimer's disease.

Morphologically distinct types of amyloid plaques point the way to a better understanding of Alzheimer's disease pathogenesis.

The details of the sequence of pathological events leading to neuron death in Alzheimer's disease (AD) are not known. Even the formation of amyloid plaques, one of the major histopathological hallmarks of AD, is not clearly understood; both the origin of the amyloid and the means of its deposition remain unclear. It is still widely considered, however, that amyloid plaques undergo gradual growth in the interstitial space of the brain via continual extracellular deposition of amyloid beta peptides at "seeding sites," and that these growing plaques encroach progressively on neurons and their axons and dendritic processes, eventually leading to neuronal death. Actually, histopathological evidence to support this mechanism is sparse and of uncertain validity. The fact that the amyloid deposits in AD brains that are collectively referred to as plaques are of multiple types and that each seems to have a different origin often is overlooked. We have shown experimentally that many of the so-called "diffuse amyloid plaques," which lack associated inflammatory cells, are either the result of leaks of amyloid from blood vessels at focal sites of blood-brain barrier breaches or are artifacts resulting from grazing sections through the margins of dense core plaques. In addition, we have provided experimental evidence that neuronal death via necrosis leaves a residue that takes the form of a spheroid "cloud" of amyloid, released by cell lysis, surrounding a dense core that often contains neuronal nuclear material. Support for a neuronal origin for these "dense core amyloid plaques" includes their ability to attract inflammatory cells (microglia and immigrant macrophages) and that they contain nuclear and cytoplasmic components that are somewhat resistant to proteolysis by lysosomes released during neuronal cell lysis. We discuss here the clinical and therapeutic importance of recognizing that amyloid deposition occurs both within neurons (intracellular) and in the interstitial (extracellular) space of the brain. For dense core plaques, we propose that the latter location largely follows from the former. This scenario suggests that blocking intraneuronal amyloid deposition should be a primary therapeutic target. This strategy also would be effective for blocking the gradual compromise of neuronal function resulting from this intraneuronal deposition, and the eventual death and lysis of these amyloid-burdened neurons that leads to amyloid release and the appearance of dense core amyloid plaques in the interstitium of AD brains.

The development of amyloid beta protein deposits in the aged brain.

The deposition of amyloid beta protein (Abeta) in the human brain and the generation of neurofibrillary tangles are the histopathological hallmarks of Alzheimer's disease. Accumulation of Abeta takes place in senile plaques and in cerebrovascular deposits as a result of an imbalance between Abeta production and clearance. This Review describes the different types of Abeta deposits, which can be distinguished by their morphology and by the hierarchical involvement of distinct areas of the brain in Abeta deposition. The role of intracellular Abeta in Abeta deposition and the mechanism of Abeta toxicity are also discussed.

Type-specific evolution of amyloid plaque and angiopathy in APPsw mice.

To clarify how Abeta deposits start in the brain, we examined the early to late stages of senile plaques and amyloid angiopathy in APPsw mice. All types of human senile plaques were observed in the mouse brains. The premature forms of cored plaques appeared first in the cerebral cortex of mice at 7-8 months old. Then, amyloid angiopathy emerged, followed by diffuse plaques consisting of Abeta1-42. Modifications of the N-terminus of Abeta were late phase phenomena. The premature forms of cored plaques were composed of central Abeta1-40 amyloid cores, surrounding amorphous Abeta1-42 deposits, and accumulation of Abeta1-42 in some peripheral cells. These cells were incorporated in amyloid cores, and these plaques developed to large cored plaques composed of Abeta1-40 and Abeta1-42. The size and number of cored plaques were increased with age. These findings indicate different evolution paths for cored plaques and diffuse plaques, and suggest the presence of a pathway that initiates with the intracellular accumulation of Abeta1-42 and leads to the development of classic plaques in human brain tissues.

Part of membrane-bound Abeta exists in rafts within senile plaques in Tg2576 mouse brain.

To clarify whether rafts are the site of abnormal amyloid beta protein (Abeta) deposition, we examined the ultrastructural localization of both flotillin-1 (pre-embedding) and Abeta (post-embedding) in Tg2576 mouse brains. After observing the exact areas of senile plaques by reflection contrast microscopy, we observed these same plaques under an electron microscope. Membrane-bound Abeta was predominantly observed on plasma membranes of small processes in diffuse plaques. Non-fibrillar and fibrillar Abeta was increased in primitive plaques, and the fibrillar form was predominant in mature plaques. The number of flotillin-1-positive rafts per field in mature plaques was prominently less than those outside of the plaques, in diffuse plaques and in primitive plaques. The colocalization of flotillin-1 with Abeta42 appeared approximately 10% of flotillin-1-positive rafts within senile plaques, while there was no colocalization found outside of the plaques. This study ultrastructurally demonstrated that part of membrane-bound Abeta exists in lipid rafts within senile plaques, and suggests that rafts could be one of the sites for initial Abeta deposition.

Generation of prion transmission barriers by mutational control of amyloid conformations.

Self-propagating beta-sheet-rich protein aggregates are implicated in a wide range of protein-misfolding phenomena, including amyloid diseases and prion-based inheritance. Two properties have emerged as common features of amyloids. Amyloid formation is ubiquitous: many unrelated proteins form such aggregates and even a single polypeptide can misfold into multiple forms--a process that is thought to underlie prion strain variation. Despite this promiscuity, amyloid propagation can be highly sequence specific: amyloid fibres often fail to catalyse the aggregation of other amyloidogenic proteins. In prions, this specificity leads to barriers that limit transmission between species. Using the yeast prion [PSI+], we show in vitro that point mutations in Sup35p, the protein determinant of [PSI+], alter the range of 'infectious' conformations, which in turn changes amyloid seeding specificity. We generate a new transmission barrier in vivo by using these mutations to specifically disfavour subsets of prion strains. The ability of mutations to alter the conformations of amyloid states without preventing amyloid formation altogether provides a general mechanism for the generation of prion transmission barriers and may help to explain how mutations alter toxicity in conformational diseases.

In vivo imaging of reactive oxygen species specifically associated with thioflavine S-positive amyloid plaques by multiphoton microscopy.

Amyloid-beta, the primary constituent of senile plaques in Alzheimer's disease, is hypothesized to cause neuronal damage and cognitive failure, but the mechanisms are unknown. Using multiphoton imaging, we show a direct association between amyloid-beta deposits and free radical production in vivo in live, transgenic mouse models of Alzheimer's disease and in analogous ex vivo experiments in human Alzheimer tissue. We applied two fluorogenic compounds, which become fluorescent only after oxidation, before imaging with a near infrared laser. We observed fluorescence associated with dense core plaques, but not diffuse plaques, as determined by subsequent addition of thioflavine S and immunohistochemistry for amyloid-beta. Systemic administration of N-tert-butyl-alpha-phenylnitrone, a free radical spin trap, greatly reduced oxidation of the probes. These data show directly that a subset of amyloid plaques produces free radicals in living, Alzheimer's models and in human Alzheimer tissue. Antioxidant therapy neutralizes these highly reactive molecules and may therefore be of therapeutic value in Alzheimer's disease.

MAP-2 immunolabeling can distinguish diffuse from dense-core amyloid plaques in brains with Alzheimer's disease.

Alzheimer's disease (AD) neuropathology is characterized by the presence of diffuse and dense-core (neuritic) amyloid plaques in specific areas of the brain. The origin of these plaques and the relationship between them is poorly understood. Current methods to identify clearly these types of plaques in the AD brains are largely dependent upon morphological characteristics. Dense-core amyloid plaques in the entorhinal cortex and hippocampus of AD brains might arise from the lysis of neurons overburdened by excessive intracellular deposition of amyloid beta1-42 (Abeta42) peptide. The local release of active lysosomal enzymes, which persist within these plaques, might degrade most of the released intracellular proteins, leaving behind only those that are resistant to proteolytic digestion, such as ubiquitin, tau, neurofilament proteins and amyloid. To test the possibility that proteins that are sensitive to proteolysis may be degraded selectively in plaques, we used immunohistochemistry to examine the distribution of microtubule-associated protein-2 (MAP-2), a protein localized primarily in neuronal dendrites and known to be sensitive to proteolysis. Uniform MAP-2 immunolabeling was detected throughout the somatodendritic compartment of neurons in age-matched control cortical brain tissues as well as throughout areas of Abeta42-positive diffuse plaques in AD brains. In contrast, analysis of serial sections revealed that MAP-2 was absent from Abeta42-positive dense-core plaques in AD brains. Our results indicate that this differential MAP-2 immunolabeling pattern among plaques may be employed as a reliable and sensitive method to distinguish dense-core plaques from diffuse plaques within AD brain tissue. Furthermore, this biochemical distinction indicates that dense-core and diffuse plaques are formed by different mechanisms.

Fractal analysis of senile plaque observed in various animal species.

In the present study, the fractal dimension (FD), a concept to determine morphological complexity, was applied to morphological estimation of animal and human senile plaque using a computer-aided method. The FDs of mature plaque in a 17-year-old dog were significantly higher than those of diffuse plaque in 11- to 16-year-old dogs. In both types of plaque, the FD tended to increase as the size expanded and there was a significant difference between the slope values of the approximate line for diffuse and mature plaque. In humans, there was also a significant difference in FD value between diffuse and mature plaque. No significant differences were observed between the two types of plaque in a bear or a cynomolgus monkey. The FD of feline diffuse plaque was significantly lower than that of a camel, bear and monkey. These results indicated that the diffuse and mature plaque of the dog might form in a different manner, and similar events may occur in human senile plaque formation. In addition, specific shapes and different FD values of the diffuse plaque among animals suggested that the original conditions for plaque formation would be different.

Beta-amyloid plaques: stages in life history or independent origin?

Several types of discrete beta-amyloid (Abeta) deposit or senile plaque have been identified in the brains of individuals with Alzheimer's disease and Down's syndrome. The majority of these plaques can be classified into four morphological types: diffuse, primitive, classic and compact. Two hypotheses have been proposed to account for these plaques. Firstly, that the diffuse, primitive, classic and compact plaques develop in sequence and represent stages in the life history of a single plaque type. Secondly, that the different Abeta plaques develop independently and therefore, unique factors are involved in the formation of each type. To attempt to distinguish between these hypotheses, the morphology, ultrastructure, composition, and spatial distribution in the brain of the four types of plaque were compared. Although some primitive plaques may develop from diffuse plaques, the evidence suggests that a unique combination of factors is involved in the pathogenesis of each plaque type and, therefore, supports the hypothesis that the major types of Abeta plaque develop independently.

Frequencia da mutacao delta-mtDNA 4977 e acumulo de placas neuriticas e emaranhados neurofibrilares no giro para-hipocampal humano, no envelhecimento normal e na doenca de Alzheimer / Delta mtDNA 4977 mutational frequence and neuritic plaques and neurofibrillary tangles in human parahippocampal gyrus, in normal aging and Alzheimer's disease

A presenca da mutacao-delecao mtDNA no giro para-hipocampal humano foi investigada em 95 pacientes autopsiados de tres series de origens geograficas distintas, Alemanha, Brasil e Japao, incluindo 70 pacientes sem doencas neuropsiquiatricas e 25 pacientes portadores da doenca de Alzheimer. Somente a serie alema, caracterizada por maiores proporcoes de neuronios medios e grandes, e alta incidencia de placas neuriticas e emaranhados neurofibrilares no giro para-hipocampal, apresentou a delta-mtDNA em niveis detectaveis pela reacao de cadeia da polimerase (PCR). As series brasileira e japonesa, caracterizadas por menores proporcoes de neuronios medios e grandes e baixa incidencia de placas e emaranhados, nao apresentaram niveis detectaveis da alfa-mtDNA. A frequencia f da alfa-mtDNA foi tres vezes menor no grupo de pacientes portadores da doenca de Alzheimer (f=0,12) que no grupo controle (f=0,37) (p=0,03)...