A Periodic Diet that Mimics Fasting Promotes Multi-System Regeneration, Enhanced Cognitive Performance, and Healthspan

Abstract

Prolonged fasting (PF) promotes stress resistance, but its effects on longevity are poorly understood. We show that alternating PF and nutrient-rich medium extended yeast lifespan independently of established pro-longevity genes. In mice, 4 days of a diet that mimics fasting (FMD), developed to minimize the burden of PF, decreased the size of multiple organs/systems, an effect followed upon re-feeding by an elevated number of progenitor and stem cells and regeneration. Bi-monthly FMD cycles started at middle age extended longevity, lowered visceral fat, reduced cancer incidence and skin lesions, rejuvenated the immune system, and retarded bone mineral density loss. In old mice, FMD cycles promoted hippocampal neurogenesis, lowered IGF-1 levels and PKA activity, elevated NeuroD1, and improved cognitive performance. In a pilot clinical trial, three FMD cycles decreased risk factors/biomarkers for aging, diabetes, cardiovascular disease, and cancer without major adverse effects, providing support for the use of FMDs to promote healthspan.

Copyright © 2015 Elsevier Inc. All rights reserved.

Figures

Figure 1. Periodic FMD promotes a lean bodyweight, improves health-span and promotes tissue regeneration

A) Periodic fasting (PF, alternating cycles of SDC media and water) prolongs lifespan in wild type (WT) S. Cerevisiae (DBY746), in rim15Δ and B)msn2Δ msn4Δ gis1Δ DBY746 mutants. C) PF induces cellular stress resistance against hydrogen peroxide in S. Cerevisiae (DBY746). D) Mouse body weight profile. Dotted lines represent FMD cycles. E) Consumed kcal/g of bodyweight. F) Total adipose tissue (TAT), G) Subcutaneous adipose tissue (SAT), H) Visceral adipose tissue (VAT) and I) Lean body mass at 28 months of age. N= 3/group. J) – K) Representative images of the SAT (gray) and VAT (red) in the lumbar L3 region. L) Kidney M) Heart and N) Liver weight as % change. N= 8-10/group. O) Liver H&E staining of control (1, 2) and FMD mouse at the end of the FMD regimen (3) or 24 hours after re-feeding (4). Unorganized cells (arrow) indicate liver repopulation. 1, 3: 40X magnification; 2, 4: 20X magnification. P) Hepatic proliferative index (Ki67+) after 1, 3 and 7 days of refeeding compared to control. N= 3-4/group. Q) Pax7 and R) p62 protein expression level, N= 3-4/group. S) Tissue mineral density (mg Hydroxyapatite/cm3) of the femur. N= 5/group. All data are expressed as the mean ± SEM.

Figure 2. Periodic FMD cycle reduce and delay cancer, rejuvenate the hematopoietic system and induce mesenchymal stem/progenitor cells

A) Hepatic lymphomatous nodules (bar= 400 microns). B) Lymphoma in the renal medulla (bar= 100 microns), C) in a mesenteric lymph node (bar= 100 microns) and D) in the spleen (bar= 100 microns). E) Hepatic lymphoma containing atypical cells with abnormal DNA (circle) and mitosis (arrows, bar= 100 microns). Subcutaneous fibrosarcoma in relationship to F) the epidermis and with invasion into G) the skeletal muscle tissue. H) Cytological details (bar= 100 microns). I) Autopsy-confirmed neoplasms. J) Lymphoma incidence. K) Neoplasms in relationship to the onset (arrow) of the FMD diet. L) Number of animals with 0 to more than 5 abnormal lesions determined at autopsy. M) Inflammatory incidence. N) Dermatitis incidence in %. Images show progression of dermatitis. O) – T) Complete blood counts. N= 7-12/group. O) White blood cells, P) Lymphoid: myeloid ratio. Q) Platelets, R) Red blood cells and S) Hemoglobin. Other CBC parameters are summarized in Table S3 and Figure S8. T) lin−Scal-1+CD45− mesenchymal stem/progenitor cells (MSPC) in bone marrow cells from control mature (M, 8–10 month), old (O, 20.5 month), and FMD mice 7 days after refeeding (FMD-RF; 20.5 month). N= 4-5/group. All data are expressed as the mean ± SEM.

Figure 3. Periodic FMD cycle improve motor coordination, hippocampal dependent learning, short- and long-term memory

A) Best Rotarod performance score at 23 months. N= 18/group. B) Rotarod performance as linear regression for each cohort (dashed lines). N= 18/group. C) Spontaneous alternation behavior (SAB) at 23 months. N= 11/group. D) Recognition index at 23 months in the novel object recognition task. E) Exploration time of the old vs. novel object (New, dashed bar). N= 8/group. F) – I) Success rate, latency, error number and deviation from escape box in the Barnes maze at 23 months (N= 7-12/group). J) – K) Strategies used to locate escape box. All data are expressed as the mean ± SEM.

Figure 4. Periodic FMD cycle promotes adult neurogenesis

A) Hippocampal immunohistochemistry of control (top row) and FMD (bottom row, see Methods for details) fed 23 months-old animals for BrdU (left panel, green), DCX (middle panel, red) and BrdU+ DCX+ (right panel). B) Age-dependent BrdU+ cell counts in sub-granular zone of the dentate gyrus (DG) (N= 4/group) C) BrdU+ cells in the DG at the end of the FMD (N= 4/group). D) DCX+ staining in the DG in 23 months-old animals (N= 4/group). E) Percentage of double-positive BrdU+ DCX+ cells in the DG (N= 4/group). F) Hippocampal IGF-1 level after FMD (N= 3/group). G) IGF-1R mRNA level in the DG (N= 3/group). H) PKA activity level in the DG (N= 5/group). I) NeuroD1 mRNA level in the DG (N= 3/group). All data are expressed as the mean ± SEM.

Figure 5. Periodic FMD cycle increase median lifespan but do not affect maximum lifespan

A) Kaplan-Meier survival curve for control and FMD cohort (N= 46 and 29, respectively). B) Overview for onset of death, 75%-, median-, 25%- and maximum lifespan in months with % change. C) Cumulative incidence rates of deaths associated with neoplasia. D) Cumulative incidence rates of deaths not associated with neoplasia. E) Overview over the date of death not associated with neoplasms. The change from the 4 day FMD to 3 day FMD is indicated by the green shaded area at 26.6 month. The stop of the 3 day FMD and switch to the ad lib control diet after 6 FMD cycles is indicated by the white shaded area. Numbers over the red squares indicate the number of animals deceased on the particular date, asterisk indicates that death during the FMD regime or within 3 days of refeeding. All data are expressed as the mean ± SEM.

Figure 6. Effects of a human adapted FMD regimen in a pilot clinical trial

A) Subjects were randomized to either the Fasting Mimicking Diet (FMD) or a control group. Subjects in the FMD cohort consumed the FMD for 5 consecutive days every month for 3 months and returned to normal diet in-between FMDs. Control subjects continued their normal diet. Measurements were performed prior to the diet (Baseline), immediately after the first FMD cycle (FMD) and during the recovery period after the 3rd cycle (FMD-RF). Subjects in the control group were evaluated within the same time frame as the FMD-RF subjects (End). B) Glucose (N= 19). C) β-hydroxybutyrate (FMD N= 19, Control N= 18). D) IGF-I (FMD N= 19, Control N= 18) and E) IGFBP-I (FMD N= 19, Control N= 17). F) Body weight (N= 19). G) Trunk fat (FMD N= 18, Control N= 19) and H) Lean body mass evaluated by dual energy x-ray absorptiometry. I) C-reactive protein (CRP; FMD N= 19, Control N= 18) levels of all subjects (left panels) and subjects in the average or high risk group for heart disease (N= 8; right panel). J) Percentage of lin−CD184+CD45− mesenchymal stem/progenitor cells (MSPC) in the peripheral blood mono-nucleated cell population (FMD N= 16, Control N= 14). All data are expressed as the mean ± SEM.