Abstract Body: Funding: The INSIGHT Registry is funded by Penumbra.

Introduction
Thrombi have shown to modify and be modified by their microenvironment throughout the pathogenesis of ischemic stroke. The molecular mechanisms by which this occurs and how it corresponds to symptomatic presentation is unknown. Here, we identify a transcriptomic signature in thrombi associated with increased NIHSS severity at presentation, suggesting a mechanistic role for immune clearance in clinical severity of ischemic stroke.

Methods
The INSIGHT registry is a prospective, multicenter, multi-omic registry focused on elucidating the molecular underpinnings of stroke from analysis of clot and interarterial blood. RNA sequencing of 10,990 genes from 292 thrombi from patients undergoing thrombectomy was aligned, normalized, and residualized using Rsubread, edgeR, sva, and limma/voom. Signed co-expression modules were constructed using WGCNA, and each module eigengene (a measure of a module’s principal component) was correlated with NIHSS at presentation using Pearson’s correlation. Networks were visualized and analyzed in cytoscape. Hub genes were determined by number of undirected edges. Network enrichments were conducted using Gene Ontology biological processes and cellular components.

Results
The mean NIHSS in this cohort was 15.14± 7.45. 3 co-expression modules out of 26 total modules detected in the data significantly inversely correlated with NIHSS at presentation. These networks corresponded to downregulation of neuronal activation (p=0.042), interferon signaling (p=0.039), and T cell activation (p=0.023), respectively. Major hub genes in each module included KCNAB2, a potassium channel associated with epilepsy and implicated in susceptibility to poor stroke outcomes in aged mice brains, SRSF11, associated with cognitive decline, PNISR, associated with interferon signaling, and ZAP70, associated with T-cell differentiation.

Conclusion
The association of immune downregulation with increased NIHSS severity suggests that robust immune clearance may mitigate poor outcomes in ischemic stroke, and that thrombi-mediated modification of the local immune environment may exacerbate symptomatic presentation. Conversely, the association of major hub genes with known cognitive decline suggests that thrombi are modified by the neurovascular unit and neurons of surrounding tissues and post-thrombectomy sequencing may provide insight into the underlying health of the surrounding parenchyma.