Abstract 14505: Generation of Atherosclerosis in Mice via Somatic Deletion of Ldlr With AAV-CRISPR
Introduction: Genetic studies of atherosclerosis in mice are both time-consuming and expensive. Mice knocked out for a gene of interest must be extensively backcrossed to C57BL/6J, and then bred to Ldlr or Apoe knockout backgrounds. The Clustered Regularly-Interspaced Short Palindromic Repeats/Cas9 (CRISPR/Cas9) genome editing system is a powerful new approach for gene disruption in cells and model organisms.
Hypothesis: In this study, we hypothesized that somatic disruption of Ldlr with CRISPR/Cas9 could generate atherosclerosis in adult mice. In addition, we hypothesized that we could efficiently disrupt an additional liver-expressed gene to investigate its impact on atherosclerosis, using Apob as proof-of-concept.
Methods: Adeno-Associated Viral (AAV) vectors based on serotype 8 were used to deliver small guide RNAs (gRNA) to adult Cas9 transgenic mice. The mice received 1) a nontargeting gRNA, 2) a gRNA targeting Ldlr, or 3) gRNAs to both Ldlr and Apob. Mice were placed on a Western diet and followed for twenty weeks for changes in plasma cholesterol and susceptibility to atherosclerosis.
Results: Disruption of Ldlr with AAV-CRISPR was robust, resulting in severe hypercholesterolemia (control: 350 +/- 18.7 mg/dl vs. Ldlr: 728 +/- 174 mg/dl) and atherosclerotic lesions in the aorta (control: 0.0% lesion vs. Ldlr: 2.21% +/- 2.10%). Mice receiving gRNAs to both Ldlr and Apob had an identical degree of Ldlr disruption, but dramatically lower cholesterol (125 +/- 27.3 mg/dl), profound hepatic steatosis, and a complete prevention of atherosclerosis.
Conclusions: These data show that the AAV-CRISPR system is a valuable and immediately useful time-saving tool for atherosclerosis studies, and can also be used to test the involvement of liver-expressed candidate genes in this disease.
Author Disclosures: K.E. Jarrett: None. Y. Yeh: None. R. Gupta: None. M. Zhang: None. C.M. Lee: None. P. Rodriguez: None. H.J. Pownall: None. G. Bao: None. J.F. Martin: None. W.R. Lagor: None.
- © 2016 by American Heart Association, Inc.