Associations of cerebrospinal fluid amyloidogenic nanoplaques with cytokines in Alzheimer’s disease

Background:The aggregation of amyloidβ(Aβ)is central in the pathogenesis of Alzheimer’s disease(AD).Recently it has been shown that specifically,larger,Thioflavin T-binding Aβaggregates are associated with increased neuroinflammation and cytokine release.This study was aimed to quantify fibrillary amyloid aggregates,so-called nanoplaques,and investigate their relationship with cytokines in the cerebrospinal fluid(CSF).Methods:CSF was collected from 111 patients assessed for cognitive complaints at the Oslo University Hospital Memory Clinic.The patients were grouped based on their amyloid status.The CSF nanoplaque concentration was quantified with the Thioflavin T-fluorescence correlation spectroscopy(ThT-FCS)assay.The levels of nine cytokines(eotaxin-1,granulocyte stimulating factor,interleukin[IL]-6,IL-7,IL-8,monocyte chemoattractant protein-1,gammainduced protein 10,macrophage inflammatory protein[MIP]-1α,and MIP-1β)were quantified with a magnetic bead-based multiplex assay and read on a Luminex IS 200 instrument.Results:There were 49 amyloid-negative and 62 amyloid-positive patients in the cohort;none of the cytokines differed significantly between the amyloid groups.The increased nanoplaque levels were associated with levels of MIP-1βbelow the lower limit of quantification,and with decreased levels of MIP-1αand IL-8.The associations remained significant when adjusted for age,sex,cognitive function,apolipoproteinε4 status and CSF core biomarker levels.Conclusion:The cytokine levels were not associated with amyloid status in this cohort.The nanoplaque levels were negatively associated with MIP-1β,MIP-1αand IL-8,which is in line with recent findings suggesting that the upregulation of some cytokine markers has a protective role and is negatively associated with AD progression.

Preparation of true solutions of monomeric amyloidogenic protein/peptide:A critical prerequisite for aggregation kinetic study

Our dynamic laser light scattering(LLS) study shows that the current widely used protocols of dissolving amyloidogenic protein/peptide do not really result in a true solution;namely,there always exist a trace amount of interchain aggregates,which greatly affect the association kinetics,partially explaining why different kinetics were reported even for a solution with identical protein and solvent.Recently,using a combination of the conventional dissolution procedure and our newly developed ultra-filtration method,we have developed a novel protocol to prepare a true solution of amyloidogenic protein/peptide without any interchain aggregates.The resultant solutions remain in their monomeric state for at least one week,which is vitally important for further study of the very initial stage of the interchain association under the physiological conditions because more and more evidence suggests that it is those small oligomers rather than large fabric aggregates that are cytotoxic.In addition,this study shows that combining static and dynamic LLS can lead to more physical and microscopic information about the protein association instead of only the size distribution.