title: High-resolution imaging of intravascular atherogenic inflammation in live mice.
abstract: BACKGROUND
  The inflammatory processes that initiate and propagate atherosclerosis remain poorly understood, largely because defining the intravascular behavior of immune cells has been technically challenging. Respiratory and pulsatile movements have hampered in vivo visualization of leukocyte accumulation in athero-prone arteries at resolutions achieved in other tissues.
  
  
  OBJECTIVE
  To establish and to validate a method that allows high-resolution imaging of inflammatory leukocytes and platelets within the carotid artery of atherosusceptible mice in vivo.
  
  
  RESULTS
  We have devised a procedure to stabilize the mouse carotid artery mechanically without altering blood dynamics, which dramatically enhances temporal and spatial resolutions using high-speed intravital microscopy in multiple channels of fluorescence. By applying this methodology at different stages of disease progression in atherosusceptible mice, we first validated our approach by assessing the recruitment kinetics of various leukocyte subsets and platelets in athero-prone segments of the carotid artery. The high temporal and spatial resolution allowed the dissection of both the dynamic polarization of and the formation of subcellular domains within adhered leukocytes. We further demonstrate that the secondary capture of activated platelets on the plaque is predominantly mediated by neutrophils. Finally, we couple this procedure with triggered 2-photon microscopy to visualize the 3-dimensional movement of leukocytes in intimate contact with the arterial lumen.
  
  
  CONCLUSIONS
  The improved imaging of diseased arteries at subcellular resolution presented here should help resolve many outstanding questions in atherosclerosis and other arterial disorders.
authors.affiliation: From the Department of Epidemiology, Atherothrombosis, and Imaging, Centro Nacional de Investigaciones Cardiovasculares, Madrid, Spain (R.C., J.M.G.-G., V.S., C.S.-R., P.M.-S., A.H., V.A.); Institute for Cardiovascular Prevention, Ludwig-Maximilians-University Munich and DZHK (German Centre for Cardiovascular Research), partner site Munich Heart Alliance, Munich, Germany (R.T.A.M., C.W., O.S.); Cardiovascular Research Institute Maastricht, Maastricht, The Netherlands (R.T.A.M., C.W.); and Academic Medical Center, University of Amsterdam, Amsterdam, The Netherlands (O.S.).
authors.forename: Raphael
authors.initials: R
authors.lastname: Chèvre
authors.forename: Jose María
authors.initials: JM
authors.lastname: González-Granado
authors.forename: Remco T A
authors.initials: RT
authors.lastname: Megens
authors.forename: Vinatha
authors.initials: V
authors.lastname: Sreeramkumar
authors.forename: Carlos
authors.initials: C
authors.lastname: Silvestre-Roig
authors.forename: Pedro
authors.initials: P
authors.lastname: Molina-Sánchez
authors.forename: Christian
authors.initials: C
authors.lastname: Weber
authors.forename: Oliver
authors.initials: O
authors.lastname: Soehnlein
authors.forename: Andrés
authors.initials: A
authors.lastname: Hidalgo
authors.forename: Vicente
authors.initials: V
authors.lastname: Andrés
mesh_terms: D000818:Animals; D001057:Apolipoproteins E / Q000235:genetics; D050197:Atherosclerosis / Q000235:genetics / Q000276:immunology / Q000503:physiopathology; D001792:Blood Platelets / Q000276:immunology; D002340:Carotid Artery Diseases / Q000235:genetics / Q000276:immunology* / Q000503:physiopathology*; D017536:Carotid Artery, Common / Q000276:immunology / Q000503:physiopathology; D005260:Female; D049452:Green Fluorescent Proteins / Q000235:genetics; D036904:Leukocyte Rolling / Q000276:immunology; D007962:Leukocytes / Q000276:immunology; D008297:Male; D051379:Mice; D018345:Mice, Knockout; D008856:Microscopy, Fluorescence / Q000379:methods*; D022423:Myeloid Cells / Q000276:immunology; D009504:Neutrophils / Q000276:immunology; D012039:Regional Blood Flow / Q000502:physiology; D014657:Vasculitis / Q000235:genetics / Q000276:immunology* / Q000503:physiopathology*
chemical_list: D001057:Apolipoproteins E; D049452:Green Fluorescent Proteins
keywords: atherosclerosis; blood platelets; carotid arteries; neutrophils