The conformational epitope for a new Aß42 protofibril-selective antibody partially overlaps with the peptide N-terminal region.

Aggregation and accumulation of amyloid-ß peptide (Aß) is a key component of Alzheimer's disease (AD). While monomeric Aß appears to be benign, oligomers adopt a biologically detrimental structure. These soluble structures can be detected in AD brain tissue by antibodies that demonstrate selectivity for aggregated Aß. Protofibrils are a subset of soluble oligomeric Aß species and are described as small (< 100 nm) curvilinear assemblies enriched in ß-sheet structure. Our own in vitro studies demonstrate that microglial cells are much more sensitive to soluble Aß42 protofibrils compared to Aß42 monomer or insoluble Aß42 fibrils. Protofibrils interact with microglia, trigger Toll-like receptor signaling, elicit cytokine transcription and expression, and are rapidly taken up by the cells. Because of the importance of this Aß species, we sought to develop an antibody that selectively recognizes protofibrils over other Aß species. Immunization of rabbits with isolated Aß42 protofibrils generated a high-titer anti serum with a strong affinity for Aß42 protofibrils. The antiserum, termed AbSL, was selective for Aß42 protofibrils over Aß42 monomers and Aß42 fibrils. AbSL did not react with amyloid precursor protein and recognized distinct pathological features in AD transgenic mouse brain slices. Competition studies with an Aß antibody that targets residues 1-16 indicated that the conformational epitope for AbSL involved the N-terminal region of protofibrils in some manner. The newly developed antibody may have potential diagnostic and therapeutic uses in AD tissue and patients, and targeting of protofibrils in AD may have beneficial effects. Read the Editorial Highlight for this article on page 621. Cover Image for this issue: doi. 10.1111/jnc.13827.

Searching for autoimmune encephalitis: Beware of normal CSF.

OBJECTIVE: To determine the prevalence of cerebrospinal fluid (CSF) markers associated with inflammation (i.e., elevated white blood cell count, protein concentration, and CSF-specific oligoclonal bands) in patients with early active autoimmune encephalitis (AE). METHODS: CSF characteristics, including WBC count, protein concentration, and oligoclonal banding, were analyzed in patients diagnosed with AE at two tertiary care centers. RESULTS: Ninety-five patients were included in the study. CSF white blood cell counts and protein levels were within normal limits for 27% (CI95%: 19-37) of patients with AE. When results of oligoclonal banding were added, 14% (CI95%: 6-16) of patients with AE had "normal" CSF. The median CSF white blood cell count was 8 cells/mm3 (range: 0-544) and the median CSF protein concentration was 0.42 g/L (range: 0.15-3.92). CONCLUSIONS: White blood cell counts and protein levels were within normal limits in the CSF of a substantial proportion of patients with early active AE. Inclusion of CSF oligoclonal banding identified a higher proportion of patients with an inflammatory CSF profile, especially when CSF was sampled early in the disease process.

Aß42 Protofibrils Interact with and Are Trafficked through Microglial-Derived Microvesicles.

Microvesicles (MVs) and exosomes comprise a class of cell-secreted particles termed extracellular vesicles (EVs). These cargo-holding vesicles mediate cell-to-cell communication and have recently been implicated in neurodegenerative diseases such as Alzheimer's disease (AD). The two types of EVs are distinguished by the mechanism of cell release and their size, with the smaller exosomes and the larger MVs ranging from 30 to 100 nm and 100 nm to 1 µm in diameter, respectively. MV numbers are increased in AD and appear to interact with amyloid-ß peptide (Aß), the primary protein component of the neuritic plaques in the AD brain. Because microglial cells play such an important role in AD-linked neuroinflammation, we sought to characterize MVs shed from microglial cells, better understand MV interactions with Aß, and determine whether internalized Aß may be incorporated into secreted MVs. Multiple strategies were used to characterize MVs shed from BV-2 microglia after ATP stimulation. Confocal images of isolated MVs bound to fluorescently labeled annexin-V via externalized phosphatidylserine revealed a polydisperse population of small spherical structures. Dynamic light scattering measurements yielded MV diameters ranging from 150 to 600 nm. Electron microscopy of resin-embedded MVs cut into thin slices showed well-defined uranyl acetate-stained ring-like structures in a similar diameter range. The use of a fluorescently labeled membrane insertion probe, NBD C6-HPC, effectively tracked MVs in binding experiments, and an Aß ELISA confirmed a strong interaction between MVs and Aß protofibrils but not Aß monomers. Despite the lesser monomer interaction, MVs had an inhibitory effect on monomer aggregation. Primary microglia rapidly internalized Aß protofibrils, and subsequent stimulation of the microglia with ATP resulted in the release of MVs containing the internalized Aß protofibrils. The role of MVs in neurodegeneration and inflammation is an emerging area, and further knowledge of MV interaction with Aß may shed light on extracellular spread and influence on neurotoxicity and neuroinflammation.