Two Specific Sulfatide Species Are Dysregulated during Renal Development in a Mouse Model of Alport Syndrome

Gessel, Megan Murray; Spraggins, Jeffrey M.; Voziyan, Paul A.; Abrahamson, Dale R.; Caprioli, Richard M.; Hudson, Billy G.

doi:10.1002/lipd.12171

Cited by 11 publications

(6 citation statements)

References 39 publications

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“…1 Diseases of the kidney, such as diabetic kidney disease (DKD), can alter glomeruli, leading to podocyte loss, expansion of the mesangium, and thickening of the GBM. [2][3][4] Determining the distribution of biomolecules among these cell types in healthy glomeruli is necessary to better understand cellular changes in diseased states. 5 Imaging technologies are emerging to address the challenge of defining molecular characteristics of tissue features with increasing specificity.…”

Section: Introductionmentioning

confidence: 99%

RevealingIn SituMolecular Profiles of Glomerular Cell Types and Substructures with Integrated Imaging Mass Spectrometry and Multiplexed Immunofluorescence Microscopy

Esselman,

Moser,

Tideman

et al. 2024

Preprint

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Glomeruli filter blood through the coordination of podocytes, mesangial cells, fenestrated endothelial cells, and the glomerular basement membrane. Cellular changes, such as podocyte loss, are associated with pathologies like diabetic kidney disease (DKD). However, little is known regarding thein situmolecular profiles of specific cell types and how these profiles change with disease. Matrix-assisted laser desorption/ionization imaging mass spectrometry (MALDI IMS) is well-suited for untargeted tissue mapping of a wide range of molecular classes. Additional imaging modalities can be integrated with MALDI IMS to associate these biomolecular distributions to specific cell types. Herein, we demonstrate an integrated workflow combining MALDI IMS and multiplexed immunofluorescence (MxIF) microscopy. High spatial resolution MALDI IMS (5 µm pixel size) was used to determine lipid distributions within human glomeruli, revealing intra-glomerular lipid heterogeneity. Mass spectrometric data were linked to specific glomerular cell types through new methods that enable MxIF microscopy to be performed on the same tissue section following MALDI IMS without sacrificing signal quality from either modality. A combination of machine-learning approaches was assembled, enabling cell-type segmentation and identification based on MxIF data followed by the mining of cell type or cluster-associated MALDI IMS signatures using classification models and interpretable machine learning. This allowed the automated discovery of spatially specific biomarker candidates for glomerular substructures and cell types. Overall, the work presented here establishes a toolbox for probing molecular signatures of glomerular cell types and substructures within tissue microenvironments and provides a framework that applies to other kidney tissue features and organ systems.

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Section: Introductionmentioning

confidence: 99%

RevealingIn SituMolecular Profiles of Glomerular Cell Types and Substructures with Integrated Imaging Mass Spectrometry and Multiplexed Immunofluorescence Microscopy

Esselman,

Moser,

Tideman

et al. 2024

Preprint

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show abstract

“…The filtration process occurs at the start of the nephron, the glomerulus, which is a spherical, multicellular functional tissue unit (FTU) with a diameter of ∼200 μm. , A glomerulus contains important substructures such as fenestrated endothelium, glomerular basement membrane, and podocytes, among others. Modifications in the glomeruli’s ability to perform filtration is at the center of various kidney diseases such as Alport syndrome and diabetic nephropathy. − The former can cause glomerular damage, which leads to alterations in glomerular substructures, such as the basement membrane. Grove et al demonstrated an increase in lysophospholipid and ganglioside species in glomeruli of mice with diabetic nephropathy .…”

Section: Introductionmentioning

confidence: 99%

Microscopy-Directed Imaging Mass Spectrometry for Rapid High Spatial Resolution Molecular Imaging of Glomeruli

Esselman,

Patterson,

Migas

et al. 2023

J. Am. Soc. Mass Spectrom.

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The glomerulus is a multicellular functional tissue unit (FTU) of the nephron that is responsible for blood filtration. Each glomerulus contains multiple substructures and cell types that are crucial for their function. To understand normal aging and disease in kidneys, methods for high spatial resolution molecular imaging within these FTUs across whole slide images is required. Here we demonstrate a workflow using microscopy-driven selected sampling to enable 5 μm pixel size matrix-assisted laser desorption/ionization imaging mass spectrometry (MALDI IMS) of all glomeruli within whole slide human kidney tissues. Such high spatial resolution imaging entails large numbers of pixels, increasing the data acquisition times. Automating FTU-specific tissue sampling enables high-resolution analysis of critical tissue structures, while concurrently maintaining throughput. Glomeruli were automatically segmented using coregistered autofluorescence microscopy data, and these segmentations were translated into MALDI IMS measurement regions. This allowed high-throughput acquisition of 268 glomeruli from a single whole slide human kidney tissue section. Unsupervised machine learning methods were used to discover molecular profiles of glomerular subregions and differentiate between healthy and diseased glomeruli. Average spectra for each glomerulus were analyzed using Uniform Manifold Approximation and Projection (UMAP) and k-means clustering, yielding 7 distinct groups of differentiated healthy and diseased glomeruli. Pixel-wise k-means clustering was applied to all glomeruli, showing unique molecular profiles localized to subregions within each glomerulus. Automated microscopy-driven, FTU-targeted acquisition for high spatial resolution molecular imaging maintains high-throughput and enables rapid assessment of whole slide images at cellular resolution and identification of tissue features associated with normal aging and disease.

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“…The biological functions of sulfatides is vast and in the brain, sulfatides are highly enriched in myelin . With regard to MSI, sulfatides have been mapped in brain tissues to understand oligodendrocyte development as well as in renal tissues of a model of Alport Syndrome . Thus, continued mapping of these crucial lipids is of the utmost importance in biological specimens such as brain tissue.…”

Section: Introductionmentioning

confidence: 99%

DBDA Matrix Increases Ion Abundance of Fatty Acids and Sulfatides in MALDI-TOF and Mass Spectrometry Imaging Studies

Khamidova

Pergande

Pathmasiri

et al. 2023

J. Am. Soc. Mass Spectrom.

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MALDI-TOF MS is a powerful tool to analyze biomolecules, owing to its soft ionization nature that generally results in simple spectra of singly charged ions. Implementation of the technology in the imaging mode provides a means to spatially map analytes in situ. Recently, a new matrix, DBDA (N1,N4-dibenzylidenebenzene-1,4-diamine) was reported to facilitate the ionization of free fatty acids in negative ion mode. Building on this finding, we sought to implement DBDA for MALDI mass spectrometry imaging studies in brain tissue and successfully map oleic acid, palmitic acid, stearic acid, docosahexaenoic acid, and arachidonic acid using mouse brain sections. Moreover, we hypothesized that DBDA would provide superior ionization for sulfatides, a class of sulfolipids with multiple biological functions. Herein, we also demonstrate that DBDA is ideal for MALDI mass spectrometry imaging of fatty acids and sulfatides in brain tissue sections. Additionally, we show enhanced ionization of sulfatides using DBDA compared with three different traditionally used MALDI matrices. Together these results provide new opportunities for studies to measure sulfatides by MALDI-TOF MS.

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Two Specific Sulfatide Species Are Dysregulated during Renal Development in a Mouse Model of Alport Syndrome

Cited by 11 publications

References 39 publications

RevealingIn SituMolecular Profiles of Glomerular Cell Types and Substructures with Integrated Imaging Mass Spectrometry and Multiplexed Immunofluorescence Microscopy

RevealingIn SituMolecular Profiles of Glomerular Cell Types and Substructures with Integrated Imaging Mass Spectrometry and Multiplexed Immunofluorescence Microscopy

Microscopy-Directed Imaging Mass Spectrometry for Rapid High Spatial Resolution Molecular Imaging of Glomeruli

DBDA Matrix Increases Ion Abundance of Fatty Acids and Sulfatides in MALDI-TOF and Mass Spectrometry Imaging Studies

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