ApoE −/−/Lysozyme M EGFP/EGFP mice as a versatile model to study monocyte and neutrophil trafficking in atherosclerosis

Abstract

Objectives Intravital microscopy is a useful tool for studying leukocyte trafficking in atherosclerosis. However, distinction between various subclasses of leukocytes using this technology is lacking. Therefore, we generated ApoE −/−/Lysozyme M EGFP/EGFP mice and investigated whether targeted cell types could be visualized by in vivo microscopy and whether absence of lysozyme M will influence atherosclerosis. Methods We crossed male ApoE −/− mice with mice homozygous for a knock-in mutation of enhanced green fluorescent protein (EGFP) in the lysozyme M locus (Lys EGFP/EGFP) creating ApoE −/−/Lys EGFP/EGFP mice. Mice were sacrificed at the age of 26 weeks. Blood was collected for serum lipid analysis, differential white blood cell count and flow cytometry. Lesion area was determined on en face mounted aortas and sections from aortic roots were stained for immunohistochemistry. Atherosclerotic lesions were also studied by confocal- and intravital microscopy. Results Basic parameters, such as white blood cell count, cholesterol profile, lesion area and plaque composition was unaltered in ApoE −/−/Lys EGFP/EGFP mice compared to ApoE −/− mice. Fluorescent neutrophils and monocytes were clearly visualized by intravital fluorescence and confocal microscopy. Fluorescent cells were distributed primarily in the periphery of atherosclerotic lesions indicating a preference for recruitment in these areas. Conclusions ApoE −/−/Lys EGFP/EGFP mice will serve as a useful model to study leukocyte trafficking in atherosclerosis and how different subsets of leukocytes influence atherogenesis.