Ketogenic Diet Reduces Midlife Mortality and Improves Memory in Aging Mice

Abstract

Ketogenic diets recapitulate certain metabolic aspects of dietary restriction such as reliance on fatty acid metabolism and production of ketone bodies. We investigated whether an isoprotein ketogenic diet (KD) might, like dietary restriction, affect longevity and healthspan in C57BL/6 male mice. We find that Cyclic KD, KD alternated weekly with the Control diet to prevent obesity, reduces midlife mortality but does not affect maximum lifespan. A non-ketogenic high-fat diet (HF) fed similarly may have an intermediate effect on mortality. Cyclic KD improves memory performance in old age, while modestly improving composite healthspan measures. Gene expression analysis identifies downregulation of insulin, protein synthesis, and fatty acid synthesis pathways as mechanisms common to KD and HF. However, upregulation of PPARα target genes is unique to KD, consistent across tissues, and preserved in old age. In all, we show that a non-obesogenic ketogenic diet improves survival, memory, and healthspan in aging mice.

Keywords: beta-hydroxybutyrate; healthspan; ketogenic diet; longevity.

Figure 1

Cyclic ketogenic diet (switching weekly between KD and control diet) started at 12 mo old reduces mid-life mortality of C57BL/6 males. A, Diet composition. B, Five diet regimens. C, Plasma BHB levels of 4 mo mice on diets for 10 weeks; blood drawn on consecutive weeks during both day and night (N=6). Cyclic diets indicate the food eaten each week. Data that did not differ between weeks or night/day are combined for clarity. D–L Data from main lifespan study. D, Body weight trajectories. E, Overall mean caloric intake. F, Mean weekly weight change, showing weight cycling on cyclic diets. G–I, Survival curves for Cyclic KD (G) and Cyclic HF (H) vs. Control, and for KD vs. HF (I). J–L, Daily chi square tests of differences in survival for Cyclic KD (J) and Cyclic HF (K) vs. Control, and for KD vs. HF (L). Survival statistics below include the final log rank test, the best daily chi square test (J–L), the best daily log rank test, and the frequency of observed differences occurring among random-data monte carlo simulations. Lifespan study, C57BL/6 NIA males starting 12 mo old: N=61 Control, 37 KD, 26 HF, 81 Cyclic KD, 36 Cyclic HF. See also Figures S1 and S2.

Figure 2

Effect of Cyclic KD on memory and other healthspan measures with aging. A, Experimental timeline. All mice ate control diet during Baseline and Aged testing periods. B, Schema of composite scores in L–N. C–D, Memory retrieval tasks of Place Avoidance show better performance in Cyclic KD group. E, Maximum escape velocity following shock in Place Avoidance. F, Old Age Novel Object Recognition test shows improved memory in Cyclic KD group. G–K, Activity and physical function show fewer differences between Cyclic KD and Control groups, but also little age-related decline. L, Normalized composite of four echocardiogram measures using 3 mo old mice as baseline. M–N, 35-item baseline-normalized composite healthspan score (Table S1) shows modestly less age-related decline in Cyclic KD (M), spread across many items (N). Healthspan study, C57BL/6 NIA males starting 12 mo old: N=27 Control, 31 Cyclic KD. See also Figure S3.

Figure 3

KD activates PPARα gene expression pattern distinct from HF. A, Experimental timeline. B–G, RNAseq transcriptome analysis of 1 week on diets in 12 mo old mice. B, Plasma BHB levels for study of immediate diet effects. C–D, Gene expression changes in liver, defined permissively for subsequent pathway analysis (Log2FC>0.38, P<0.02), show many genes down-regulated in common by KD and HF but few genes up-regulated in common. E–G, Top ingenuity upstream regulators associated with KD in liver (E), with the same regulators highlighted for HF in liver (F) and KD in kidney (G). PPARα activation pattern is present in both liver and kidney for KD, and not for HF. H–I, Study of 26 mo old Cyclic KD and Control mice. H, Q-PCR of genes involved in fatty acid synthesis, glucose/insulin signaling, and TOR activity. I, Q-PCR of PPARα target genes. B–G, 12 mo mice on diets for 1 week, collected at night, N = 5 Control, 7 KD, 6 HF. H–I, 26 mo mice on diets for 14 mo, collected at night during control-fed week, N = 8 Control, 8 Cyclic KD. See also Figure S4.