Lipid Heterogeneity between Astrocytes and Neurons Revealed by Single‐Cell MALDI‐MS Combined with Immunocytochemical Classification

Abstract

Transcriptomics characterizes cells based on their potential molecular repertoire whereas direct mass spectrometry (MS) provides information on the actual compounds present within cells.S ingle-cell matrix-assisted laser desorption/ionization (MALDI) MS can measure the chemical contents of individual cells but spectra are difficult to correlate to conventional cell types,l imiting the metabolic information obtained. We present ap rotocol that combines MALDI-MS with immunocytochemistry to assayoverathousand individual rat brain cells.The approach entwines the wealth of knowledge obtained by immunocytochemical profiling with mass spectral information on the predominant lipids within eachcell. While many lipid species showed ahigh degree of similarity between neurons and astrocytes,s eventeen significantly differed between the two cell types,including four phosphatidylethanolamines elevated in astrocytes and nine phosphatidylcholines in neurons.

Exploring the Fundamental Structures of Life: Non‐Targeted, Chemical Analysis of Single Cells and Subcellular Structures

“…[91] In that work, MALDI-MS was used first to prescreen cells for peptides indicative of aspecific cell type,a nd CE-MS was used to analyze metabolite profiles of cells representative of ac ell type.A sa nother example,b y combining high-throughput single-cell measurements with immunocytochemistry to distinguish astrocytes from neurons, specific lipid profiles were determined for each cell type. [125] We expect multimodal measurements to remain ar apidly developing approach for increasing chemical coverage. [126] By coupling single-cell MS techniques with complementary targeted and non-targeted techniques,s uch as single-cell transcriptomics,s pectroscopy,a nd staining,e ven more information can be garnered.…”

Morphometric Cell Classification for Single‐Cell MALDI‐Mass Spectrometry Imaging

“…It has therefore become an invaluable tool in biological research for the label‐free imaging of otherwise inaccessible molecular classes, such as metabolites, lipids, or drugs, in tissues . Despite advances in sample preparation and instrumentation that enabled MALDI‐MSI to retrieve chemical information at single‐cell resolution a very limited number of high‐spatial‐resolution MALDI‐MSI biomedical studies has been reported …”

“…[2,3] Despite advances in sample preparation and instrumentation that enabled MALDI-MSI to retrieve chemical information at single-cell resolution a very limited number of high-spatial-resolution MALDI-MSI biomedical studies has been reported. [4][5][6][7] Detailed histological tissue annotations to contextualize the obtained molecular signals from individual cells are considered a major limitation for comprehensive biomedical research using MALDI-MSI. Typically, the MSI-analyzed tissue section is histologically stained, co-registered to the MALDI-MSI data, and annotated by a pathologist.…”

NMR Methods for Determining Lipid Turnover via Stable Isotope Resolved Metabolomics

“…With the help of stable isotope labeling, it is possible to follow the fate of individual atoms from precursors into downstream metabolites. However, assessing stable isotope incorporation into complex lipids, especially at low enrichment, has long been a challenge due to the complexity of lipid structures and the vast number of lipid species [11,12]. Nevertheless, modern mass spectrometers are capable of resolving different isotopologues in stable isotope experiments designed to assess lipid turnover in a wide variety of biological systems including tracer studies in vivo [13][14][15][16][17][18][19].…”