High-Resolution Imaging of Intravascular Atherogenic Inflammation in Live Mice
Rationale: 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 athero-susceptible mice in vivo.
Methods and Results: We have devised a procedure to mechanically stabilize the mouse carotid artery 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 athero-susceptible 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 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 two-photon microscopy to visualize the three-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.
- Intravital microscopy
- two-photon microscopy
- intravascular imaging
- carotid artery
- Received September 10, 2013.
- Revision received December 18, 2013.
- Accepted December 23, 2013.