Optimization of imaging mass cytometry, a single-cell spatial proteomics technology, for the study of Alzheimer disease

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Project Summary This proposal aims to fill a critical gap in applying high-dimensional single-cell spatial proteomics technology - imaging mass cytometry (IMC) - to study neurons and glia in the human brains. IMC is an emerging high-plex technology which combines immunohistochemistry (IHC) staining using metal isotope-labeled antibodies with laser ablation and mass-spectrometry-based detection to produce high-dimensional images. IMC allows the detection of 40-100 protein markers simultaneously on tissue sections and enables a variety of distinct cell types to be analyzed concurrently at a spatially-resolved single-cell resolution. IMC has been applied to cancer, diabetes, immunology, and infectious disease research, identifying functionally distinct immune cell subpopulations associated with disease progression, treatment outcomes, and biomarkers for disease prognosis. As the most common neurodegenerative disease without a disease-modifying treatment, the application of IMC to Alzheimer disease (AD) could provide novel insights into the disease pathogenesis and targets for the development of effective therapeutic. This spatially-resolved single-cell resolution proteomic technology is critical for investigating the impact of AD pathology on different cell types in a spatial context. The extracellular beta- amyloid (Aβ) plaques and the intracellular neurofibrillary tangles (NFT) are hallmarks of AD pathology. Aβ plaques have been linked to selective neuronal death in AD and are surrounded by activated astrocytes and microglia. The burden and location of NFT correlate with neuronal loss, disease severity, and clinical course. However, the molecular mechanisms underlying AD pathologies in relationship to cellular and gene expression changes and neurodegeneration in the human brain remain unclear, which significantly hampers the development of effective treatment strategies. The main challenge of applying IMC to AD research is the lack of a well-designed, validated high-quality antibody panel suited for the brain tissue. This proposal will fill this gap by developing a 37-antibody panel for the analysis of snap-frozen human brain tissues using IMC which will include both the markers to identify all the major cell types in the central nervous system and biomarkers of AD pathologies. In Aim 1, we will design and optimize the immunodetection conditions for a 37-antibody panel for use on postmortem human brain tissue sections and apply the technology to the frontal cortex tissues of AD and matched control cases. In Aim 2, we will develop and optimize a computational pipeline for brain tissue-specific IMC data analysis. This technology could be used to obtain comprehensive spatial information of brain microenvironment and cell-cell interactions among the glia and neurons, which will advance our understanding of how the spatial distribution of Aβ plaques and NFTs affect glial activation and neuronal death. This optimized methodology will provide the neuroscience community a powerful IMC neuroimaging tool for a broad range of research fields such as aging, neurodegeneration, and brain development.