Pathological Crystallization of Cholesterol in Atherosclerosis

Human atherosclerotic tissues will be obtained from the Department of Vascular Surgery, Sheba Medical Center. Tissues will be taken from donors undergoing routine endarterectomy procedures.We chose carotid artery as the source for the atheromas, due to its high prevalence and its considerable size.All speciemnts will be studied in Cryo-Scanning Electron Microscopy (Cryo-SEM). Tissues will be observed in hydrated vitrified state, after High Pressure Freezing (HPF) and freeze fracture, as well established in the WIS group. Samples will be observed at -120°C. Cathode-Luminescence (CL) detects a luminescence signal induced by electrons in cholesterol crystals and CE condensed states. It will be implemented herein under cryogenic conditions. Sample preparation is as for cryo-SEM. Cryo-FIB-SEM block face serial imaging allows the acquisition of serial images over large volumes (tens of cubed micrometers), at a relatively high imaging resolution (up to 5nm3 voxel size). Volumes of up to 40x40x40 ml will be imaged with a resolution of up to 10nm. The data will be minimally processed and aligned using FIJI, reconstructed and manually segmented using Avizo Lite 9.2.0 software.