Somatic editing of Ldlr with adeno-associated viral-CRISPR is an efficient tool for atherosclerosis research

Kelsey E. Jarrett, Ciaran Lee, Marco De Giorgi, Ayrea Hurley, Baiba K. Gillard, Alexandria M. Doerfler, Ang Li, Henry J. Pownall, Gang Bao, William R. Lagor

Research output: Contribution to journalArticle

10 Scopus citations

Abstract

Objective-Atherosclerosis studies in Ldlr knockout mice require breeding to homozygosity and congenic status on C57BL6/J background, a process that is both time and resource intensive. We aimed to develop a new method for generating atherosclerosis through somatic deletion of Ldlr in livers of adult mice. Approach and Results-Overexpression of PCSK9 (proprotein convertase subtilisin/kexin type 9) is currently used to study atherosclerosis, which promotes degradation of LDLR (low-density lipoprotein receptor) in the liver. We sought to determine whether CRISPR/Cas9 (clustered regularly interspaced short palindromic repeats-associated 9) could also be used to generate atherosclerosis through genetic disruption of Ldlr in adult mice. We engineered adeno-associated viral (AAV) vectors expressing Staphylococcus aureus Cas9 and a guide RNA targeting the Ldlr gene (AAV-CRISPR). Both male and female mice received either (1) saline, (2) AAV-CRISPR, or (3) AAV-hPCSK9 (human PCSK9)-D374Y. A fourth group of germline Ldlr-KO mice was included for comparison. Mice were placed on a Western diet and followed for 20 weeks to assess plasma lipids, PCSK9 protein levels, atherosclerosis, and editing efficiency. Disruption of Ldlr with AAV-CRISPR was robust, resulting in severe hypercholesterolemia and atherosclerotic lesions in the aorta. AAV-hPCSK9 also produced hypercholesterolemia and atherosclerosis as expected. Notable sexual dimorphism was observed, wherein AAV-CRISPR was superior for Ldlr removal in male mice, while AAV-hPCSK9 was more effective in female mice. Conclusions-This all-in-one AAV-CRISPR vector targeting Ldlr is an effective and versatile tool to model atherosclerosis with a single injection and provides a useful alternative to the use of germline Ldlr-KO mice.

Original languageEnglish (US)
Pages (from-to)1997-2006
Number of pages10
JournalArteriosclerosis, Thrombosis, and Vascular Biology
Volume38
Issue number9
DOIs
StatePublished - 2018

Keywords

  • CRISPR-Cas systems
  • atherosclerosis
  • gene editing
  • hypercholesterolemia
  • lipoproteins

ASJC Scopus subject areas

  • Cardiology and Cardiovascular Medicine

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