Chemical Imaging of Evolving Amyloid Plaque Pathology and Associated Aβ Peptide Aggregation in a Transgenic Mouse Model of Alzheimer's Disease

One of the major hallmarks of Alzheimer's disease (AD) pathology is the formation of extracellular amyloid β (Aβ) plaques. While Aβ has been suggested to be critical in inducing and, potentially, driving the disease, the molecular basis of AD pathogenesis is still under debate. Extracellular Aβ plaque pathology manifests itself upon aggregation of distinct Aβ peptides, resulting in morphologically different plaque morphotypes, including mainly diffuse and cored senile plaques. As plaque pathology precipitates long before any clinical symptoms occur, targeting the Aβ aggregation processes provides a promising target for early interventions. However, the chain of events of when, where and what Aβ species aggregate and form plaques remains unclear. The aim of the current study was to investigate the potential of MALDI-IMS as a tool to study the evolving pathology in transgenic mouse models for AD. To that end, we used an emerging, chemical imaging modality - MALDI imaging mass spectrometry - that allows for delineating Aβ aggregation with specificity at the single plaque level. We identified that plaque formation occurs first in cortical regions and that these younger plaques contain higher levels of 42 amino acid-long Aβ (Aβ1-42). Plaque maturation was found to be characterized by a relative increase in deposition of Aβ1-40, which was associated with the appearance of a cored morphology of the plaques. Finally, other C-terminally truncated Aβ species (Aβ1-38 and Aβ1-39) exhibited a similar aggregation pattern as Aβ1-40, suggesting that these species have similar aggregation characteristics. These results suggest that initial plaque formation is seeded by Aβ1-42; a process that is followed by plaque maturation upon deposition of Aβ1-40 as well as deposition by other C-terminally modified Aβ species

Sensitive detection of Aβ protofibrils by proximity ligation - relevance for Alzheimer's disease

<p>Abstract</p> <p>Background</p> <p>Protein aggregation plays important roles in several neurodegenerative disorders. For instance, insoluble aggregates of phosphorylated tau and of Aβ peptides are cornerstones in the pathology of Alzheimer's disease. Soluble protein aggregates are therefore potential diagnostic and prognostic biomarkers for their cognate disorders. Detection of the aggregated species requires sensitive tools that efficiently discriminate them from monomers of the same proteins. Here we have established a proximity ligation assay (PLA) for specific and sensitive detection of Aβ protofibrils via simultaneous recognition of three identical determinants present in the aggregates. PLA is a versatile technology in which the requirement for multiple target recognitions is combined with the ability to translate signals from detected target molecules to amplifiable DNA strands, providing very high specificity and sensitivity.</p> <p>Results</p> <p>For specific detection of Aβ protofibrils we have used a monoclonal antibody, mAb158, selective for Aβ protofibrils in a modified PLA, where the same monoclonal antibody was used for the three classes of affinity reagents required in the assay. These reagents were used for detection of soluble Aβ aggregates in solid-phase reactions, allowing detection of just 0.1 pg/ml Aβ protofibrils, and with a dynamic range greater than six orders of magnitude. Compared to a sandwich ELISA setup of the same antibody the PLA increases the sensitivity of the Aβ protofibril detection by up to 25-fold. The assay was used to measure soluble Aβ aggregates in brain homogenates from mice transgenic for a human allele predisposing to Aβ aggregation.</p> <p>Conclusions</p> <p>The proximity ligation assay is a versatile analytical technology for proteins, which can provide highly sensitive and specific detection of Aβ aggregates - and by implication other protein aggregates of relevance in Alzheimer's disease and other neurodegenerative disorders.</p

Engineered antibodies: new possibilities for brain PET?

International audienceAlmost 50 million people worldwide are affected by Alzheimer's disease (AD), the most common neurodegenerative disorder. Development of disease-modifying therapies would benefit from reliable, non-invasive positron emission tomography (PET) biomarkers for early diagnosis, monitoring of disease progression, and assessment of therapeutic effects. Traditionally, PET ligands have been based on small molecules that, with the right properties, can penetrate the blood-brain barrier (BBB) and visualize targets in the brain. Recently a new class of PET ligands based on antibodies have emerged, mainly in applications related to cancer. While antibodies have advantages such as high specificity and affinity, their passage across the BBB is limited. Thus, to be used as brain PET ligands, antibodies need to be modified for active transport into the brain. Here, we review the development of radioligands based on antibodies for visualization of intrabrain targets. We focus on antibodies modified into a bispecific format, with the capacity to undergo transferrin receptor 1 (TfR1)-mediated transcytosis to enter the brain and access pathological proteins, e.g. amyloid-beta. A number of such antibody ligands have been developed, displaying differences in brain uptake, pharmacokinetics, and ability to bind and visualize the target in the brain of transgenic mice. Potential pathological changes related to neurodegeneration, e.g. misfolded proteins and neuroinflammation, are suggested as future targets for this novel type of radioligand. Challenges are also discussed, such as the temporal match of radionuclide half-life with the ligand's pharmacokinetic profile and translation to human use. In conclusion, brain PET imaging using bispecific antibodies, modified for receptor-mediated transcytosis across the BBB, is a promising method for specifically visualizing molecules in the brain that are difficult to target with traditional small molecule ligands

Scandinavian society for the study of diabetes Abstracts Seventh Meeting

Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/46039/1/125_2005_Article_BF01219478.pd

High Efficiency of Plating of the Thermophilic Sulfur-Dependent Archaebacterium Sulfolobus acidocaldarius

A procedure for plating Sulfolobus acidocaldarius using Gelrite as the gelling agent is presented. The technique uses a supporting gel of 0.8% (wt/vol) Gelrite and an overlay soft gel of 0.4% (wt/vol) Gelrite, in which the colonies are grown. The plating efficiency was essentially 100%