Abstract 5567: The Ldlr −/− Mouse as a Novel Model of Aortic Calcification: Application of Micro-Computerized Tomography

Abstract

BACKGROUND : Patients with familial hypercholesterolemia (FH) due to loss-of-function mutations in the low-density lipoprotein receptor (LDLR) gene exhibit an age-dependent and LDLR gene dosage effect on aortic calcification. We have previously documented that subjects with homozygous FH show significant aortic calcification by their teens, an effect delayed by 20 years in heterozygotes and observed much later in control subjects. The degree of calcification is independent of initial and life long burden of low-density lipoprotein cholesterol concentration. This clinical observation suggests a potential role for the LDLR in osteoblast signalling pathways. In order to better study this observation, we established an animal model of aortic calcification and a novel technique; based on micro-computerized tomography (CT) to examine vascular calcifications in mice. METHOD AND RESULT: We examined C57BL/6 wild-type (wt) and Ldlr −/− mice at 12 and 18 months. The Ldlr −/− mice develop spontaneous atherosclerosis in normal chow-fed older mice. Vascular calcification has not been previously examined in this model. Under isoflurane anesthesia, the aorta was perfused in-situ with saline. The aorta was then carefully dissected and fixed ex-vivo . The tissue was embedded in paraffin block and calcium content was determined by digital x-ray imaging, dual X-ray densitometry, and micro-CT scanning of the entire aorta and aortic valve. The paraffin blocks were then sectioned and Alizarin Red stain was used to further quantify calcified tissue. Analysis of 12 and 18 month wt and Ldlr −/− mice confirmed our observations in human, with aortic calcifications identified at areas of shear stress. CONCLUSIONS: The Ldlr −/− mouse model develops aortic calcification even when fed a normal chow diet. Calcifications at areas of shear stress suggest that calcific aortic stenosis and arterial wall calcifications may share many similar pathophysiological features. This animal model of aortic calcifications, and the CT imaging technique used in the present study, may provide a better understanding of the potential links between FH and calcium deposition in the vascular wall. This model has the potential to study aortic valve calcifications as well.