A single injection of gain-of-function mutant PCSK9 adeno-associated virus vector induces cardiovascular calcification in mice with no genetic modification.
Studying atherosclerotic calcification in vivo requires mouse models with genetic modifications. Previous studies showed that injection of recombinant adeno-associated virus vector (AAV) encoding a gain-of-function mutant PCSK9 into mice promotes atherosclerosis. We aimed to study cardiovascular calcification induced by PCSK9 AAV in C57BL/6J mice.



10 week-old C57BL/6J mice received a single injection of AAV encoding mutant mPCSK9 (rAAV8/D377Y-mPCSK9). Ldlr(-/-) mice served as positive controls. Mice consumed a high-fat, high-cholesterol diet for 15 or 20 weeks. Aortic calcification was assessed by fluorescence reflectance imaging (FRI) of a near-infrared calcium tracer.



Serum levels of PCSK9 (0.14 μg/mL to 20 μg/mL, p < 0.01) and total cholesterol (82 mg/dL to 820 mg/dL, p < 0.01) increased within one week after injection and remained elevated for 20 weeks. Atherosclerotic lesion size was similar between PCSK9 AAV and Ldlr(-/-) mice. Aortic calcification was 0.01% ± 0.01 in PCSK9 AAV mice and 15.3% ± 6.1 in Ldlr(-/-) mice at 15 weeks (p < 0.01); by 20 weeks, the PCSK9 AAV mice aortic calcification grew to 12.4% ± 4.9. Tissue non-specific alkaline phosphatase activity was similar in PCSK9 AAV mice and Ldlr(-/-) mice at 15 and 20 weeks, respectively. As example of the utility of this model in testing modulators of calcification in vivo, PCSK9 AAV injection to sortilin-deficient mice demonstrated reduced aortic calcification by 46.3% (p < 0.05) compared to littermate controls.



A single injection of gain-of-function PCSK9 AAV into C57BL/6J mice is a useful tool to study cardiovascular calcification in mice with no genetic manipulation.
Center for Interdisciplinary Cardiovascular Sciences, Cardiovascular Division, Brigham and Women's Hospital, Harvard Medical School, Boston, MA 02115, USA.
Claudia
C
Goettsch
Center for Interdisciplinary Cardiovascular Sciences, Cardiovascular Division, Brigham and Women's Hospital, Harvard Medical School, Boston, MA 02115, USA.
Joshua D
JD
Hutcheson
Center for Interdisciplinary Cardiovascular Sciences, Cardiovascular Division, Brigham and Women's Hospital, Harvard Medical School, Boston, MA 02115, USA.
Sumihiko
S
Hagita
Center for Interdisciplinary Cardiovascular Sciences, Cardiovascular Division, Brigham and Women's Hospital, Harvard Medical School, Boston, MA 02115, USA.
Maximillian A
MA
Rogers
Center for Interdisciplinary Cardiovascular Sciences, Cardiovascular Division, Brigham and Women's Hospital, Harvard Medical School, Boston, MA 02115, USA.
Michael D
MD
Creager
Center for Interdisciplinary Cardiovascular Sciences, Cardiovascular Division, Brigham and Women's Hospital, Harvard Medical School, Boston, MA 02115, USA.
Tan
T
Pham
Center for Interdisciplinary Cardiovascular Sciences, Cardiovascular Division, Brigham and Women's Hospital, Harvard Medical School, Boston, MA 02115, USA.
Jung
J
Choi
Center for Interdisciplinary Cardiovascular Sciences, Cardiovascular Division, Brigham and Women's Hospital, Harvard Medical School, Boston, MA 02115, USA.
Andrew K
AK
Mlynarchik
Center for Interdisciplinary Cardiovascular Sciences, Cardiovascular Division, Brigham and Women's Hospital, Harvard Medical School, Boston, MA 02115, USA.
Brett
B
Pieper
The Lundbeck Foundation Research Center MIND, Danish Research Institute of Translational Neuroscience, Nordic EMBL Partnership for Molecular Medicine, Danish Diabetes Academy, Department of Biomedicine, Aarhus University, 8000, Denmark.
Mads
M
Kjolby
Center for Interdisciplinary Cardiovascular Sciences, Cardiovascular Division, Brigham and Women's Hospital, Harvard Medical School, Boston, MA 02115, USA; Center for Excellence in Vascular Biology, Cardiovascular Division, Brigham and Women's Hospital, Harvard Medical School, Boston, MA 02115, USA.
Masanori
M
Aikawa
Center for Interdisciplinary Cardiovascular Sciences, Cardiovascular Division, Brigham and Women's Hospital, Harvard Medical School, Boston, MA 02115, USA; Center for Excellence in Vascular Biology, Cardiovascular Division, Brigham and Women's Hospital, Harvard Medical School, Boston, MA 02115, USA. Electronic address: eaikawa@partners.org.
Elena
E
Aikawa
D033942:Adaptor Proteins, Vesicular Transport / Q000378:metabolism; D000818:Animals; D050197:Atherosclerosis / Q000378:metabolism; D002114:Calcinosis / Q000473:pathology*; D002784:Cholesterol / Q000378:metabolism; D000229:Dependovirus; D004195:Disease Models, Animal; D005260:Female; D005822:Genetic Vectors; D008099:Liver / Q000378:metabolism; D008297:Male; D051379:Mice; D008810:Mice, Inbred C57BL; D008822:Mice, Transgenic; D008856:Microscopy, Fluorescence; D009154:Mutation; D000071449:Proprotein Convertase 9 / Q000235:genetics*; D043484:Proprotein Convertases / Q000235:genetics; D011973:Receptors, LDL / Q000235:genetics; D000099183:Sortilin
D033942:Adaptor Proteins, Vesicular Transport; D002784:Cholesterol; D000071449:Proprotein Convertase 9; D043484:Proprotein Convertases; D011973:Receptors, LDL; D000099183:Sortilin; C472126:Pcsk9 protein, mouse
AAV-PCSK9; Animal model; Cardiovascular calcification