A Ketogenic Diet Is Protective Against Atherosclerosis in Apolipoprotein E Knockout Mice

Abstract

Abstract Objectives A ketogenic diet (KD) positively impacts cardiovascular disease (CVD) risk factors yet its effect on atherosclerosis, the main cause of CVD, is elusive. We hypothesize that, when compared to a high-fat diet (HF), KD protects from atherosclerosis. Methods Seven-week-old male Apoe−/− mice, a model for human atherosclerosis, were fed ad libitum (%kcal) KD (81-fat, 1-carbohydrate, 18-protein; n = 4) or HF (40-fat, 42-carbohydrate, 18-protein; n = 5). After 4, 8 and 12 weeks, plasma was collected and used to (1) quantify beta-hydroxybutyrate levels (OH-But) by a colorimetric assay; or (2) assess systemic inflammation, a key feature associated with atherosclerosis, using a panel of inflammatory cytokines; or (3) explore diet-driven changes in levels of atherosclerosis-relevant metabolites using a targeted metabolomic approach by triple quadrupole mass spectrometry. At the endpoint, mice were euthanized and their perfusion-fixed aortas were subjected to 3-D analysis by magnetic resonance imaging to quantify the extent of atherosclerosis. Data were reconstructed using Matlab and segmented to obtain atherosclerotic plaque volumes using Avizo 9.0. Results The inflammatory cytokines were significantly (P < 0.05, Student's t-test) elevated in HF-mice compared to KD-mice after 12 weeks; MIP-1α, (25 ± 13 vs 13 ± 4 pg/mol), MCP-1 (165 ± 67 vs 69 ± 23 pg/mol), TNF-α(51 ± 17 vs 35 ± 5 pg/mol), and IL-1β (2 ± 2 vs 0.6 ± 0.1 pg/mol). OH-But levels were always significantly higher in KD-mice than in HF-mice, thus confirming the presence of ketosis in KD-mice. Significant changes in the plasma metabolome of KD-mice, when compared to HF-mice, were present for dimethylated arginines, gut-derived metabolites, certain bile acids, some acylcarnitines and ceramides, certain sphingolipids and cholesterol esters, and constituents of the major intracellular antioxidant glutathione. Lastly, aortic plaque burden was significantly decreased in KD mice (2.77 ± 1.02%) than in HF mice (13.94 ± 3.72%). Conclusions KD was associated with decreased inflammation, changes in several metabolic intermediates and an atheroprotective effect based on MRI analysis. Funding Sources Huck Institutes of the Life Sciences; Graduate Program in Nutritional Sciences.