Yu-Xin Xu1, Valeska Redon2, Haojie Yu3, William Querbes4, James Pirruccello1, Abigail Liebow4, Amy Deik5, Kevin Trindade2, Xiao Wang6, Kiran Musunuru6, Clary B Clish5, Chad Cowan7, Kevin Fizgerald4, Daniel Rader2, Sekar Kathiresan8. 1. Center for Genomic Medicine and Cardiovascular Research Center, Massachusetts General Hospital, Boston, MA 02114, USA. 2. Institute for Translational Medicine and Therapeutics, Perelman School of Medicine, University of Pennsylvania, 11-125 Translational Research Center, 3400 Civic Center Blvd, Building 421, Philadelphia, PA 19104-5158, USA. 3. Department of Stem Cell and Regenerative Biology, Harvard Stem Cell Institute, Harvard University, Cambridge, MA 02138, USA. 4. Alnylam Pharmaceuticals, 300 Third Street, 3rd Floor, Cambridge, MA 02142, USA. 5. Broad Institute of Harvard and MIT, Cambridge, MA 02142, USA. 6. Cardiovascular Institute, Department of Medicine, Perelman School of Medicine, University of Pennsylvania, Philadelphia 19104, USA. 7. Department of Stem Cell and Regenerative Biology, Harvard Stem Cell Institute, Harvard University, Cambridge, MA 02138, USA; Center for Regenerative Medicine, Massachusetts General Hospital, Boston, MA 02114, USA. 8. Center for Genomic Medicine and Cardiovascular Research Center, Massachusetts General Hospital, Boston, MA 02114, USA. Electronic address: skathiresan1@partners.org.
Abstract
BACKGROUND AND AIMS: Angiopoietin-like 3 (ANGPTL3) has emerged as a key regulator of lipoprotein metabolism in humans. Homozygous loss of ANGPTL3 function causes familial combined hypolipidemia characterized by low plasma levels of triglycerides (TG), high-density lipoprotein cholesterol (HDL-C), and low-density lipoprotein cholesterol (LDL-C). While known effects of ANGPTL3 in inhibiting lipoprotein lipase and endothelial lipase contribute to the low TG and HDL-C, respectively, the basis of low LDL-C remains unclear. Our aim was to explore the role of ANGPTL3 in modulating plasma LDL-C. METHODS: We performed RNAi-mediated gene silencing of ANGPTL3 in five mouse models and in human hepatoma cells. We validated results by deleting ANGPTL3 gene using the CRISPR/Cas9 genome editing system. RESULTS: RNAi-mediated Angptl3 silencing in mouse livers resulted in very low TG, HDL-C and LDL-C, a pattern similar to the human phenotype. The effect was observed in wild-type and obese mice, while in hCETP/apolipoprotein (Apo) B-100 double transgenic mice, the silencing decreased LDL-C and TG, but not HDL-C. In a humanized mouse model (Apobec1-/- carrying human ApoB-100 transgene) deficient in the LDL receptor (LDLR), Angptl3 silencing had minimum effect on LDL-C, suggesting the effect being linked to LDLR. This observation is supported by an additive effect on LDL-C between ANGPTL3 and PCSK9 siRNAs. ANGPTL3 gene deletion induced cellular long-chain TG and ApoB-100 accumulation with elevated LDLR and LDLR-related protein (LRP) 1 expression. Consistent with this, ANGPTL3 deficiency by gene deletion or silencing reduced nascent ApoB-100 secretion and increased LDL/VLDL uptake. CONCLUSIONS: Reduced secretion and increased uptake of ApoB-containing lipoproteins may contribute to the low LDL-C observed in mice and humans with genetic ANGPTL3 deficiency.
BACKGROUND AND AIMS: Angiopoietin-like 3 (ANGPTL3) has emerged as a key regulator of lipoprotein metabolism in humans. Homozygous loss of ANGPTL3 function causes familial combined hypolipidemia characterized by low plasma levels of triglycerides (TG), high-density lipoprotein cholesterol (HDL-C), and low-density lipoprotein cholesterol (LDL-C). While known effects of ANGPTL3 in inhibiting lipoprotein lipase and endothelial lipase contribute to the low TG and HDL-C, respectively, the basis of low LDL-C remains unclear. Our aim was to explore the role of ANGPTL3 in modulating plasma LDL-C. METHODS: We performed RNAi-mediated gene silencing of ANGPTL3 in five mouse models and in humanhepatoma cells. We validated results by deleting ANGPTL3 gene using the CRISPR/Cas9 genome editing system. RESULTS: RNAi-mediated Angptl3 silencing in mouse livers resulted in very low TG, HDL-C and LDL-C, a pattern similar to the human phenotype. The effect was observed in wild-type and obesemice, while in hCETP/apolipoprotein (Apo) B-100 double transgenic mice, the silencing decreased LDL-C and TG, but not HDL-C. In a humanized mouse model (Apobec1-/- carrying humanApoB-100 transgene) deficient in the LDL receptor (LDLR), Angptl3 silencing had minimum effect on LDL-C, suggesting the effect being linked to LDLR. This observation is supported by an additive effect on LDL-C between ANGPTL3 and PCSK9 siRNAs. ANGPTL3 gene deletion induced cellular long-chain TG and ApoB-100 accumulation with elevated LDLR and LDLR-related protein (LRP) 1 expression. Consistent with this, ANGPTL3deficiency by gene deletion or silencing reduced nascent ApoB-100 secretion and increased LDL/VLDL uptake. CONCLUSIONS:Reduced secretion and increased uptake of ApoB-containing lipoproteins may contribute to the low LDL-C observed in mice and humans with genetic ANGPTL3 deficiency.
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