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Intravital Live cell Triggered Imaging System Reveals Monocyte Patrolling and Macrophage Migration in Atherosclerosis

  • Author(s): McArdle, Sara Ashley
  • Advisor(s): Ley, Klaus
  • et al.
Abstract

Atherosclerosis is an underlying cause of two of the leading causes of death in the world, heart attacks and strokes. It is a chronic inflammatory disease characterized by plaque build-up in large and medium arteries. During disease progression, monocytes in the blood invade the arterial wall, where they can differentiate into macrophages and dendritic cells. Together, these cells phagocytose lipids, clear apoptotic cell debris, present antigen to T cells, and produce pro- and anti-inflammatory cytokines. Both monocyte recruitment to the plaque and macrophage function within the plaque require cell motion. The purpose of this work is to study monocyte and macrophage movement in the context of atherosclerosis.

First, the mechanism of how the atypical chemokine receptor Duffy Antigen / Receptor for Chemokines (DARC) influences the progression of atherosclerosis is investigated. DARC alters the localization of chemokines and it was thought that through this it could influence cell recruitment to the arteria wall. It was confirmed that the absence of DARC does promote plaque formation, but a molecular mechanism of action was not discovered.

Next, to study myeloid cell motion more directly, I developed a system for imaging fluorescent leukocytes in atherosclerotic arteries in vivo. The major challenge in intravital imaging of large arteries is the motion artifacts due to the expansion of the arterial wall with the heartbeat. This new system utilizes cardiac triggering and novel image post-processing algorithms to remove these motion artifacts and allow for cell motion quantification. Apoe-/- Cx3cr1GFP/+ Cd11cYFP mice were generated to visualize labeled monocytes and macrophages. Monocytes were observed patrolling the endothelium of atherosclerotic arteries and the motion characteristics were quantified. Macrophages were seen actively probing their local environment as well as migrating through the plaque. Finally, these macrophages were extensively phenotyped by cell surface markers to link observed motion to known subsets and function.

Supplemental videos include a demonstration of the improvement in movie quality with cardiac triggering and image post-processing, examples movies of monocytes patrolling atherosclerotic arteries and macrophages migrating within the plaque and an example 3D reconstruction of atherosclerotic plaque acquired in vivo. Supplemental SNP analysis of the mice used in this work is also provided.

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