Data Processing Pipeline for Lipid Profiling of Carotid Atherosclerotic Plaque with Mass Spectrometry Imaging

Abstract

Atherosclerosis is a lipid and inflammation-driven disease of the arteries that is characterized by gradual buildup of plaques in the vascular wall. A so-called vulnerable plaque, consisting of a lipid-rich necrotic core contained by a thin fibrous cap, may rupture and trigger thrombus formation, which can lead to ischemia in the heart (heart attack) or in the brain (stroke). In this study, we present a protocol to investigate the lipid composition of advanced human carotid plaques using matrix-assisted laser desorption ionization (MALDI) mass spectrometry imaging (MSI), providing a framework that should enable the discrimination of vulnerable from stable plaques based on lipid composition. We optimized the tissue preparation and imaging methods by systematically analyzing data from three specimens: two human carotid endarterectomy samples (advanced plaque) and one autopsy sample (early stage plaque). We show a robust data reduction method and evaluate the variability of the endarterectomy samples. We found diacylglycerols to be more abundant in a thrombotic area compared to other plaque areas and could distinguish advanced plaque from early stage plaque based on cholesteryl ester composition. We plan to use this systematic approach to analyze a larger dataset of carotid atherosclerotic plaques. Electronic supplementary material The online version of this article (10.1007/s13361-019-02254-y) contains supplementary material, which is available to authorized users.

A lipid atlas of human carotid atherosclerosis

“…A subset of TAG lipids was also associated with erythrocyte areas in the multivariate model. Previously 18 we described a co-localization between DAGs and thrombus in one of our tissue sections and this finding is thus confirmed in our current dataset. We hypothesize that the co-localization of DAGs and thrombotic plaque elements can be explained by the role that DAGs play in regulating protein kinase C (PKC), an enzyme known to induce an array of pro-atherogenic responses 35 , including impairment of fibrinolysis 36 and platelet activation 37,38 .…”

“…This resulted in a total of 106 MALDI-MSI datasets. MALDI-MSI data was processed using a previously described workflow 18 and lipid images of 194 unique lipids were retained. A list of measured m/z values and the lipids that they represent is provided in Supplementary Table 2.…”

“…MALDI-MSI experiments and data processing were performed according to the methods described in Visscher et al 18 . In short, we desiccated the tissue sections and applied 2,5-dihydroxybenzoic acid (DHB) matrix by sublimation (home-built sublimation system as described in Dekker et al 41 ).…”

“…In this study, we extend the range of atherosclerotic lipids of which spatial distributions have been mapped. We use a previously established MALDI-MSI pipeline for systematic imaging of lipids 18 to visualize the spatial distribution of 194 lipids in 106 tissue sections of 12 carotid atherosclerotic plaques, harvested at carotid endarterectomy surgeries in 12 patients. Analysis of this dataset identifies spatial lipid patterns in advanced atherosclerosis that transcend individual variability.…”

Atheroma-Specific Lipids in ldlr–/– and apoe–/– Mice Using 2D and 3D Matrix-Assisted Laser Desorption/Ionization Mass Spectrometry Imaging

“… 7 In atherosclerotic research, however, MALDI-MSI remains an underexplored technique with only a few studies, which have so far have focused on the optimization of the experimental conditions and data processing. 8 − 10 …”