Examining atherosclerotic lesions in three dimensions at the nanometer scale with cryo-FIB-SEM

Author:

Capua-Shenkar Jenny1,Varsano Neta1,Itzhak Noya-Ruth1,Kaplan-Ashiri Ifat2,Rechav Katya2ORCID,Jin Xueting3,Niimi Manabu4,Fan Jianglin4,Kruth Howard S.3,Addadi Lia1ORCID

Affiliation:

1. Department of Chemical and Structural Biology, Weizmann Institute of Science, 7610001 Rehovot, Israel

2. Department of Chemical Research Support, Weizmann Institute of Science, 7610001 Rehovot, Israel

3. Experimental Atherosclerosis Section, NIH, Bethesda, MD 20892

4. Department of Molecular Pathology, University of Yamanashi, Yamanashi 409-3898, Japan

Abstract

We employed in a correlative manner an unconventional combination of methods, comprising cathodoluminescence, cryo–scanning electron microscopy (SEM), and cryo–focused ion beam (FIB)-SEM, to examine the volumes of thousands of cubed micrometers from rabbit atherosclerotic tissues, maintained in close-to-native conditions, with a resolution of tens of nanometers. Data from three different intralesional regions, at the media–lesion interface, in the core, and toward the lumen, were analyzed following segmentation and volume or surface representation. The media–lesion interface region is rich in cells and lipid droplets, whereas the core region is markedly richer in crystals and has lower cell density. In the three regions, thin crystals appear to be associated with intracellular or extracellular lipid droplets and multilamellar bodies. Large crystals are independently positioned in the tissue, not associated with specific cellular components. This extensive evidence strongly supports the idea that the lipid droplet surfaces and the outer membranes of multilamellar bodies play a role in cholesterol crystal nucleation and growth and that crystal formation occurs, in part, inside cells. The correlative combination of methods that allowed the direct examination of cholesterol crystals and lipid deposits in the atherosclerotic lesions may be similarly used for high-resolution examination of other tissues containing pathological or physiological cholesterol deposits.

Funder

United States-Israel Binational Science Foundation

Publisher

Proceedings of the National Academy of Sciences

Subject

Multidisciplinary

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