Four weeks after ligation, mice were sacrificed. The arterial tree was perfused with PBS containing 0.1 mg/ml nitroprusside (Sigma, St Louis, MO) and subsequently with 1% paraformaldehyde via a catheter inserted into the left cardiac ventricle. Right carotid artery cross-sections (4 μm thick) were cut at 100 μm intervals. At each level of 100 μm, a cross-section was stained with Lawson solution and used to determine intima, lumen and total vessel area as well as elastic lamina fragmentation. The relative intimal collagen content, i.e. the percentage of total intima area that stained positive for Sirius red, was determined under a microscope coupled to a computerized morphometry system (Quantimet 570, Leica). Morphometric analyses were performed by a blinded investigator (LB, intra-observer variability was < 10%) using a computerized morphometry system (Quantimet 570, Leica). Serum cholesterol level was determined with the CHOD-PAP method (Roche Diagnostics).
Donners M.M., Bai L., Lutgens S.P., Wijnands E., Johnson J., Schurgers L.J., Liu C.L., Daemen M.J., Cleutjens K.B., Shi G.P., Biessen E.A, & Heeneman S. (2016). Cathepsin K Deficiency Prevents the Aggravated Vascular Remodeling Response to Flow Cessation in ApoE-/- Mice. PLoS ONE, 11(9), e0162595.
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