Abstract
The traditional method to establish a cardiovascular disease model induced by high fat and high cholesterol diets is time consuming and laborious and may not be appropriate in all circumstances. A suitable pig model to study metabolic disorders and subsequent atherosclerosis is not currently available. For this purpose, we applied the CRISPR/Cas9 system to Bama minipigs, targeting apolipoprotein E (ApoE) and low density lipoprotein receptor (LDLR) gene simultaneously. Six biallelic knockout pigs of these two genes were obtained successfully in a single step. No off-target incidents or mosaic mutations were detected by an unbiased analysis. Serum biochemical analyses of gene-modified piglets showed that the levels of low density lipoprotein choleserol (LDL-C), total cholesterol (TC) and apolipoprotein B (APOB) were elevated significantly. This model should prove valuable for the study of human cardiovascular disease and related translational research.
Highlights
Atherosclerosis is primarily caused by the imbalance of blood lipids, cholesterol-containing low density lipoprotein (LDL) particles [1]
apolipoprotein E (ApoE)-/-/low density lipoprotein receptor (LDLR)-/mice have been broadly used in atherosclerosis research; lipoprotein profiles and metabolism in mice are different from humans and pigs [4]
The targeted regions of guide RNA (gRNA) for the ApoE and LDLR genes are shown in Figure 1A and 1B
Summary
Atherosclerosis is primarily caused by the imbalance of blood lipids, cholesterol-containing low density lipoprotein (LDL) particles [1]. ApoE and LDLR gene mutations play a large role in the disease progression. ApoE protein has a great effect on cholesterol metabolism in the liver and is an important component of very low density lipoprotein (VLDL) and chylomicrons. LDLR protein, when located on the surface of hepatocytes, plays a critical role in low density lipoprotein cholesterol (LDL-C) clearance in liver. ApoE-/-/LDLR-/mice have been broadly used in atherosclerosis research; lipoprotein profiles and metabolism in mice are different from humans and pigs [4]. Pig models could reproduce many important atherosclerosis features, such as palque rupture and thrombosis. These features were hardly ever observed in mouse models [5]
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