Spatial Transcriptomics-correlated Electron Microscopy maps transcriptional and ultrastructural responses to brain injury

Abstract

Understanding the complexity of cellular function within a tissue necessitates the combination of multiple phenotypic readouts. Here, we developed a method that links spatially-resolved gene expression of single cells with their ultrastructural morphology by integrating multiplexed error-robust fluorescence in situ hybridization (MERFISH) and large area volume electron microscopy (EM) on adjacent tissue sections. Using this method, we characterized in situ ultrastructural and transcriptional responses of glial cells and infiltrating T-cells after demyelinating brain injury in male mice. We identified a population of lipid-loaded “foamy” microglia located in the center of remyelinating lesion, as well as rare interferon-responsive microglia, oligodendrocytes, and astrocytes that co-localized with T-cells. We validated our findings using immunocytochemistry and lipid staining-coupled single-cell RNA sequencing. Finally, by integrating these datasets, we detected correlations between full-transcriptome gene expression and ultrastructural features of microglia. Our results offer an integrative view of the spatial, ultrastructural, and transcriptional reorganization of single cells after demyelinating brain injury.

Lesion-remote astrocytes govern microglia-mediated white matter repair

“…Quantification of Ccn1 + astrocytes relative to vehicle (PBS) (n=5-6 mice/group). j , spatial transcriptomic analysis correlating regions of LPC-demyelinated corpus callosum with Ccn1 + astrocytes and microglia exhibiting a WDM molecular profile 67 . k , Ccn1 + astrocytes (white arrowheads) intermingle with IBA1 + microglia/macrophages nodules (yellow arrowheads) neighboring mouse MOG 35-55 EAE spinal cord white matter lesions.…”

“…9 e-g). We also interrogated astrocyte Ccn1 expression and molecular indicators of the WDM state from a recently published spatial transcriptomics dataset from lysolecithin demyelination in the mouse brain 67 . Here we determined, once again, that Ccn1 + astrocytes and WDM microglia are intimately associated in demyelinated regions (Fig.…”

Shared inflammatory glial cell signature after stab wound injury, revealed by spatial, temporal, and cell-type-specific profiling of the murine cerebral cortex

“…Despite the numerous advantages offered by recent techniques, the main hurdle in studying cellular interactions is to accurately identify the relevant cellular populations that shape the injured environment. While available stRNA-seq methods retain spatial information, they either lack the cellular resolution required to identify single cells 64 or are restricted to a subset of the transcriptome, introducing bias in data interpretation 65 . Conversely, several studies based on the scRNA-seq failed to associate the identified clusters to the injury because spatial information about clusters was lost 66 .…”

Label-Free Techniques for Probing Biomolecular Condensates

“…61 Additionally, harnessing the high spatial resolution on structure afforded by the TEM, one can also correlate structural information with spatial transcriptomics approaches. 62 Brillouin Microscopy. Brillouin microscopy (BM) has recently emerged as a powerful and versatile technique for studying the mechanical properties of diverse biological samples.…”

“…Some correlative approaches are being developed to combine the advantages of different techniques, such as with optical microscopy techniques like cryo-correlative light and electron microscopy (cryo-CLEM) . Additionally, harnessing the high spatial resolution on structure afforded by the TEM, one can also correlate structural information with spatial transcriptomics approaches …”