Investigating structural heterogeneities in amyloid aggregates with multiscale infrared spectroscopic imaging

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Project Summary The misfolding and aggregation of specific proteins into fibrillar amyloid deposits is the pathological hallmark of a wide class of diseases and neurodegenerative disorders, which represent a major public health concern worldwide. However, the precise role of amyloid aggregates in disease onset and progression is not well established. The exact aggregation states that cause neurotoxicity and relationship between distribution of secondary structures in amyloid aggregates and cognitive decline remains unclear. The aim of this project is to investigate the heterogeneities in protein secondary structure in amyloid aggregates both in-vitro and ex-vivo and identify their relationship with disease progression. Our approach relies on utilizing state-of-the-art spatially resolved infrared spectroscopy, namely nanoscale infrared (IR) spectroscopy, super-resolution IR microscopy and confocal IR spectroscopic imaging to map protein secondary structures in amyloid deposits. We aim to integrate these three techniques and develop a spatially and spectrally adaptive IR imaging approach that will enable multiscale measurement of spectral data in tissues. The above three techniques will be further augmented by Raman microscopy. Taken together, this approach will offer multimodal, multiscale structural and chemical insights on amyloid aggregates, from their distribution in tissues and chemical subtypes to structural variations within single fibrils. We will focus on studying amyloid aggregates and their heterogeneities in Alzheimer's disease (AD) to develop and optimize our methods, and subsequently aim to extend these strategies to investigate amyloid aggregates in other diseases such as Parkinson's disease, Breast Cancer and type-II diabetes. The hypothesis underlying this effort is that structural heterogeneities of amyloid deposits, and not just specific fibrillar structures, are correlated with disease progression. We will utilize photothermal AFM-IR, a technique that augments IR spectroscopy with Atomic Force Microscopy, to obtain nanoscale aggregate-specific spectra. Seeded growth from tissue extracts from AD patients will enable probing the differences in aggregation pathways and structural polymorphisms associated with different disease stages. Amyloid aggregates in tissues will be investigated through confocal IR and photothermal IR microscopies, which will allow for identifying amyloid deposits in tissues and then investigating individual plaques with high sub-diffraction spatial resolution. The tissue spectral data will be analyzed to classify amyloid deposits based on their secondary structure distributions, and the correlation between distinct classes of deposits and disease stages will be explored. The unique aspect of this approach is that it uses cutting edge technologies in spatially resolved IR spectroscopy and imaging to investigate a problem that is central to the molecular pathology of a wide range of diseases and development of therapeutic strategies.