High-resolution scanning electron microscopy for the analysis of three-dimensional ultrastructure of clots in acute ischemic stroke.

Background: Characterization of acute ischemic stroke (AIS) clots has typically focused on two-dimensional histological analysis of the thrombus. The three-dimensional (3D) architecture and distribution of components within emboli have not been fully investigated. The aim of this study was to examine the composition and microstructure of AIS clots using histology and serial block-face scanning electron microscopy (SBFSEM).
Methods: As part of the multi-institutional STRIP registry, 10 consecutive AIS emboli were collected from 10 patients treated by mechanical thrombectomy. Histological and immunohistochemical analysis was performed to determine clot composition. SBFSEM was used to assess the ultrastructural organization of the clots and specific features of individual components.
Results: Quantification of Martius Scarlett Blue stain identified fibrin (44.4%) and red blood cells (RBCs, 32.6%) as the main components. Immunohistochemistry showed a mean platelet and von Willebrand factor content of 23.9% and 11.8%, respectively. The 3D organization of emboli varied greatly depending on the region analyzed. RBC-rich areas were composed mainly of tightly packed RBCs deformed into polyhedrocytes with scant fibrin fibers interwoven between cells. The regions with mixed composition showed thick fibrin fibers along with platelets, white blood cells and RBC clusters. Fibrin-rich areas contained dense fibrin masses with sparse RBC. In three cases, the fibrin formed a grid-like or a sponge-like pattern, likely due to thrombolytic treatment. Segmentation showed that fibrin fibers were thinner and less densely packed in these cases.
Conclusions: 3D-SEM provides novel and potentially clinically relevant information on clot components and ultrastructure which may help to inform thrombolytic treatment and medical device design.
Competing Interests: Competing interests: RK reports NIH funding (R01 NS076491, R43 NS110114 and R44 NS107111), is a research consultant for Cerenovus, Insera Therapeutics LLC, Marblehead Medical LLC, MicroVention Inc, MIVI Neuroscience Inc, Neurogami Medical Inc and Triticum Inc, and has stock in Neurosigma Inc (money paid to institution). DFK is President of Marblehead Medical and has a patent pending in balloon catheter technologies, and receives research support from Cerenovus, Insera Therapeutics LLC, Medtronic, MicroVention Inc, MIVI Neuroscience Inc, NeuroSave, Neurogami Medical Inc, Sequent Medical and Insera, and has stock in Neurosigma Inc (money paid to institution). He is on the Scientific Advisory Board of Triticum Inc and previously served on a SAB for Boston Scientific. WB is CMO of Marblehead Medical and has a patent pending in balloon catheter technologies, and he is a consultant for Cerenovus and MicroVention Inc. He reports NIH funding (R01 NS105853).
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