Abstract WP262: Three-dimensional Characterization Of Cerebral Microvasculature And Microhemorrhages

Abstract

Introduction: Cerebral microhemorrhages (CMH) are the pathologic substrate of MRI-demonstrable cerebral microbleeds, and are thought to occur when integrity of cerebral microvessels is compromised, leading to deposits of hemosiderin/iron accumulating within brain tissue. The presence of CMH is associated with cognitive decline and ischemic and hemorrhagic stroke. Hypertension (HTN) is a major risk factor for cerebral microbleeds and has been linked to a more significant microbleed burden in patients. Despite the clinical significance of HTN and cerebral microbleeds, our understanding of HTN-induced microbleed formation remains limited, and the vascular origin of CMH remains obscure. Methods: We used a perfusion-based vascular label with tissue clearing to enable three-dimensional visualization of CMH with the surrounding microvascular network. Adult (18-month-old) C57BL/6J mice were administered angiotensin II to induce HTN (n=10) or PBS (n=10). Lectin-DyLight-649 was injected retro-orbitally to bind to the endothelial walls of all blood vessels in a mouse. Brains were cut into 1-mm thick sections, stained with Prussian blue to label CMH, cleared using iDISCO, and imaged via confocal microscopy. MATLAB and neuTube were used to quantify vessel diameters. Results: Using this methodology, we were able to collect three-dimensional transmission images of CMH that were colocalized with three-dimensional fluorescence images of the surrounding microvasculature. The average vessel diameter of the 5 nearest vessels to 17 different CMH was found to be 4.22±0.81 μm, which corresponds to capillary-size vessels. Conclusions: These data support a capillary origin for cerebral microhemorrhages in this mouse model. The capillary origin implies a role for the blood-brain barrier in the development of CMH and cerebral microbleeds in this model of HTN and aging. Three-dimensional imaging has substantial utility for improved understanding of CMH origin.