Abstract Body: INTRODUCTION. Cerebral microbleeds (CMBs) are age-related manifestations of microangiopathies predisposing patients to hemorrhagic stroke (HS) and cognitive decline. Little work has currently been done to elucidate mechanisms in the pathophysiology of CMBs. Herein, we report transcriptomic signatures of CMBs, as well as important location/etiological and sex-specific differences, to glean mechanistic insights in human autopsy tissue and a murine E4FAD model.

METHODS. Fourteen human CMB lesions were identified at autopsy by a neuropathologist (n=9 lobar and n=5 deep brain lesions). In addition, 6 CMB lesions were identified from E4FAD mice (n=3 male and n=3 female) sacrificed at 8 months. For each lesion, contralateral control brain tissue was also obtained from the same brain. Serial sections were obtained for each lesional and control region. The first section was stained for non-heme iron to localize CMB and control regions. The second section was then used for paired GeoMx bulk spatial transcriptomics. Downstream differential expression analyses were completed using DeSeq2, with enriched pathway analyses using Ingenuity pathway analysis (IPA).

RESULTS. Spatial transcriptomics identified 781 differentially expressed genes (DEGs) in human CMB and 448 in E4FAD mouse lesions compared with controls (p<0.05, FDR corrected). Pathway enrichment identified 110 homologous pathways between human and E4FAD mouse CMBs, related to vascular permeability, senescence and neuroinflammation (p<0.05, FDR corrected). Furthermore, lobar compared to deep microbleeds showed 52 IPA pathways related to similar functions, particularly in relation to extracellular matrix deposition (p<0.05, FDR corrected). Notable differences identified etiologic-specific pathways, such as arteriolosclerosis and amyloid processing. Male compared with female mouse CMBs identified 12 common mechanisms involved in blood brain barrier dysfunction and neuroinflammation with differences related to sex-specific neurotransmitter and hormonal variances (p<0.05, FDR corrected).

CONCLUSIONS. This is the first report to address transcriptomic changes within the CMB microenvironment, while addressing mechanistic differences by sex and lesional location. Further studies shall aim to delineate mechanisms that lead CMBs to develop into larger HS. The results also pave the way for identification of mechanistically linked circulating molecules in biomarker discovery as well as for potential therapeutic targets.