Synthetic Low Density Lipoprotein Receptor Knockout Mouse Model to Study Atherosclerosis Regression

Regression of atherosclerotic plaques occurs when plasma total cholesterol (TC) is markedly reduced. Commonly used animal models of regression include a surgical model of transplantation of atherosclerotic arteries from Apoe -/- or Ldlr -/- mice to C57BL/6J mice, and the ‘Reversa mouse’ in which hypercholesterolemia due to deletion of LDL receptor (LDLR) and over-expression of apoB100 is conditionally reversed. These models require either challenging surgeries or time-consuming breeding strategies. We utilized a synthetic method to create and then reverse hypercholesterolemia and atherosclerosis by transient knockdown of hepatic LDLR using antisense oligonucleotides (ASOs). C57BL6/J mice on Western diet were treated once a week for 16 weeks with intraperitoneal injections of LDLR ASO. TC increased to ~600 mg/dl in two weeks, and remained high. After 16 weeks, one group of mice was sacrificed for baseline analysis and the remaining mice were treated with the sense ASO to antagonize any residual LDLR ASO activity. The mice were then either kept on Western diet or switched to chow diet. Within a week, TC decreased to ~150 mg/dl in both regression groups and mice were sacrificed after another week. LDLR mRNA and protein analysis in the baseline group showed that ASO-mediated LDLR deletion was efficient. Similar analysis in both regression groups at the end of the study confirmed recovery of hepatic LDLR expression. Mice in the baseline group had visible plaques in the aortic arch and brachiocephalic artery (BCA). Analysis of the BCA showed that lesion size, visualized by Movat’s staining, was not significantly different between the baseline and regression groups. BCA lesions stained for MAC-2, a marker for macrophages, showed reduced macrophage content in both regression groups but more significantly in the Western diet group. Our data show that lesion remodeling and regression, in terms of inflammatory cell content, occur in our model. These changes occur relatively quickly as we analyzed lesions after two weeks of plasma cholesterol reversal. In conclusion, we have developed a synthetic LDLR hepatic knockout mouse model of atherosclerosis regression that can be used for any genetically modified mouse strain and obviates the need for extensive mouse breeding.