Abstract WP304: Investigating Thrombus Architecture Using Spatial Transcriptomics

Abstract

Introduction: Spatial transcriptomic (ST) platforms provide high-resolution localization of gene expression profiles within intact tissue, offering insight to cellular phenotype and ultimately tissue function. While cerebral thrombus morphology has been well-characterized, their functional architecture has not. Although this study is insufficiently powered to infer specific pathophysiologic conclusions, our findings offer preliminary insight into the functional segmentation of cerebral thrombi using ST. Methods: Four thrombi retrieved from cerebral arteries of ischemic stroke patients during standard of care thrombectomy were formalin-fixed and paraffin-embedded. Morphology marker antibodies (vimentin, CD45) and RNA probes coupled to photocleavable oligonucleotide tags were applied to thrombus specimens. Regions of interest (ROI) were randomly selected based on visualized cellular clustering within each specimen. ST analysis was performed using NanoString's GeoMx Digital Spatial Profiling platform. Results: Of the 24 ROIs, 0 samples were below the 50% warning, and 18,676 targets were analyzed. 1,127 genes were expressed in 50% of ROIs. 8,793 genes normalized by 3rd quartile, expressed above the Limit of Quantitation in at least 1% of ROIs. Differential expression analysis using a false discovery rate of < 0.05 revealed 54 differentially expressed genes (DEG), all of which were protein-coding using edgeR’s exact test. Conclusions: Although ST does not offer visibility into the presence of fully formed ribosomes, the identification of intact cytoplasmic RNA suggests there is significant transcriptional activity within thrombi. We cautiously present pilot data to support feasibility for spatially-resolved transcriptomic analysis of cerebral thrombi which, upon further technical refinement, may facilitate discovery of new targets for stroke prevention or treatment.