Marta Roche-Molina1, David Sanz-Rosa1, Francisco M Cruz1, Jaime García-Prieto1, Sergio López1, Rocío Abia1, Francisco J G Muriana1, Valentín Fuster1, Borja Ibáñez1, Juan A Bernal2. 1. From the Cardiovascular Development and Repair Department (M.R.-M., F.M.C., J.A.B.), and Epidemiology, Atherothrombosis and Imaging Department (D.S.-R., J.G.-P., V.F., B.I.), Centro Nacional de Investigaciones Cardiovasculares (CNIC), Madrid, Spain; Laboratory of Cellular and Molecular Nutrition, Instituto de la Grasa (CSIC), Seville, Spain (S.L., R.A., F.J.G.M.); The Zena and Michael a Wiener Cardiovascular Institute, Mount Sinai School of Medicine, New York, NY (V.F.); and Cardiovascular Institute, Hospital Clínico San Carlos, Madrid, Spain (B.I.). 2. From the Cardiovascular Development and Repair Department (M.R.-M., F.M.C., J.A.B.), and Epidemiology, Atherothrombosis and Imaging Department (D.S.-R., J.G.-P., V.F., B.I.), Centro Nacional de Investigaciones Cardiovasculares (CNIC), Madrid, Spain; Laboratory of Cellular and Molecular Nutrition, Instituto de la Grasa (CSIC), Seville, Spain (S.L., R.A., F.J.G.M.); The Zena and Michael a Wiener Cardiovascular Institute, Mount Sinai School of Medicine, New York, NY (V.F.); and Cardiovascular Institute, Hospital Clínico San Carlos, Madrid, Spain (B.I.). jabernal@cnic.es.
Abstract
OBJECTIVES: Patients with mutations in the proprotein convertase subtilisin/kexin type 9 (PCSK9) gene have hypercholesterolemia and are at high risk of adverse cardiovascular events. We aimed to stably express the pathological human D374Y gain-of-function mutant form of PCSK9 (PCSK9(DY)) in adult wild-type mice to generate a hyperlipidemic and proatherogenic animal model, achieved with a single systemic injection with adeno-associated virus (AAV). APPROACH AND RESULTS: We constructed an AAV-based vector to support targeted transfer of the PCSK9(DY) gene to liver. After injection with 3.5×10(10) viral particles, mice in the C57BL/6J, 129/SvPasCrlf, or FVB/NCrl backgrounds developed long-term hyperlipidemia with a strong increase in serum low-density lipoprotein. Macroscopic and histological analysis showed atherosclerotic lesions in the aortas of AAV-PCSK9(DY) mice fed a high-fat-diet. Advanced lesions in these high-fat-diet-fed mice also showed evidence of macrophage infiltration and fibrous cap formation. Hepatic AAV-PCSK9(DY) infection did not result in liver damage or signs of immunologic response. We further tested the use of AAV-PCSK9(DY) to study potential genetic interaction with the ApoE gene. Histological analysis of ApoE(-/-) AAV-PCSK9(DY) mice showed a synergistic response to ApoE deficiency, with aortic lesions twice as extensive in ApoE(-/-) AAV-PCSK9(DY)-transexpressing mice as in ApoE(-/-) AAV-Luc controls without altering serum cholesterol levels. CONCLUSIONS: Single intravenous AAV-PCSK9(DY) injection is a fast, easy, and cost-effective approach, resulting in rapid and long-term sustained hyperlipidemia and atherosclerosis. We demonstrate as a proof of concept the synergy between PCSK9(DY) gain-of-function and ApoE deficiency. This methodology could allow testing of the genetic interaction of several mutations without the need for complex and time-consuming backcrosses.
OBJECTIVES:Patients with mutations in the proprotein convertase subtilisin/kexin type 9 (PCSK9) gene have hypercholesterolemia and are at high risk of adverse cardiovascular events. We aimed to stably express the pathological humanD374Y gain-of-function mutant form of PCSK9 (PCSK9(DY)) in adult wild-type mice to generate a hyperlipidemic and proatherogenic animal model, achieved with a single systemic injection with adeno-associated virus (AAV). APPROACH AND RESULTS: We constructed an AAV-based vector to support targeted transfer of the PCSK9(DY) gene to liver. After injection with 3.5×10(10) viral particles, mice in the C57BL/6J, 129/SvPasCrlf, or FVB/NCrl backgrounds developed long-term hyperlipidemia with a strong increase in serum low-density lipoprotein. Macroscopic and histological analysis showed atherosclerotic lesions in the aortas of AAV-PCSK9(DY)mice fed a high-fat-diet. Advanced lesions in these high-fat-diet-fed mice also showed evidence of macrophage infiltration and fibrous cap formation. Hepatic AAV-PCSK9(DY) infection did not result in liver damage or signs of immunologic response. We further tested the use of AAV-PCSK9(DY) to study potential genetic interaction with the ApoE gene. Histological analysis of ApoE(-/-) AAV-PCSK9(DY)mice showed a synergistic response to ApoE deficiency, with aortic lesions twice as extensive in ApoE(-/-) AAV-PCSK9(DY)-transexpressing mice as in ApoE(-/-) AAV-Luc controls without altering serum cholesterol levels. CONCLUSIONS: Single intravenous AAV-PCSK9(DY) injection is a fast, easy, and cost-effective approach, resulting in rapid and long-term sustained hyperlipidemia and atherosclerosis. We demonstrate as a proof of concept the synergy between PCSK9(DY) gain-of-function and ApoE deficiency. This methodology could allow testing of the genetic interaction of several mutations without the need for complex and time-consuming backcrosses.
Authors: Hong Lu; Deborah A Howatt; Anju Balakrishnan; Mark J Graham; Adam E Mullick; Alan Daugherty Journal: Arterioscler Thromb Vasc Biol Date: 2016-07-28 Impact factor: 8.311
Authors: Debapriya Basu; Yunying Hu; Lesley-Ann Huggins; Adam E Mullick; Mark J Graham; Tomasz Wietecha; Shelley Barnhart; Allison Mogul; Katharina Pfeiffer; Andreas Zirlik; Edward A Fisher; Karin E Bornfeldt; Florian Willecke; Ira J Goldberg Journal: Circ Res Date: 2018-01-10 Impact factor: 17.367
Authors: 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 Journal: Arterioscler Thromb Vasc Biol Date: 2018-09 Impact factor: 8.311