A MALDI Mass Spectrometry Imaging Sample Preparation Method for Venous Thrombosis with Initial Lipid Characterization of Lab-Made and Murine Clots

Venous thromboembolism (VTE) and its complications affect
over
900,000 people in the U.S. annually, with a third of cases resulting
in fatality. Despite such a high incidence rate, venous thrombosis
research has not led to significant changes in clinical treatments,
with standard anti-coagulant therapy (heparin followed by a vitamin
K antagonist) being used since the 1950s. Mechanical thrombectomy
is an alternative strategy for treating venous thrombosis; however,
clinical guidelines for patient selection have not been well-established
or accepted. The effectiveness of both treatments is impacted by the
heterogeneity of the thrombus, including the mechanical properties
of its cellular components and its molecular makeup. A full understanding
of the complex interplay between disease initiation and progression,
biochemical molecular changes, tissue function, and mechanical properties
calls for a multiplex and multiscale approach. In this work, we establish
a protocol for using matrix-assisted laser desorption/ionization (MALDI)
mass spectrometry imaging to characterize spatial heterogeneity of
biomolecules in lab-made blood clots and ex vivo murine
thrombi. In this work, we compared (1) tissue preservation and cryosectioning
methods, (2) various matrixes, 9-aminoacridine hydrochloride monohydrate
(9AA), 2,5-dihydroxybenzoic acid (DHB), and alpha-cyano-4-hydroxycinnamic
acid matrix (CHCA), (3) plasma-rich versus red-blood-cell rich lab-made
blood clots, and (4) lab-made blood clots versus ex vivo murine thrombi. This project is the first step in our work to combine
mass spectrometry imaging with biomechanical testing of blood clots
to improve our understanding of VTE.