Fig. 6. Late dietary intervention by switching to ERF improves health metrics and extends the lifespan of Titan mice.

a Switching standard breeding feed (SBF) to energy-reduced feed (ERF) at 12 weeks resulted in a persistent average weight loss in Titan siblings (n = 23 per group). Mann–Whitney-U (MWU)-test was calculated for day 140, p = 0.0011. b Compared to age-matched control Titan mice, ERF-fed Titan mice had a lower percentage of intra-abdominal fat at 21 weeks (n = 7 per group). Paired Wilcoxon-test, p = 0.0313. c Late ERF feeding at 12 weeks of age decreased the levels of plasma cholesterol (pval.adj = 0.00013), high-density lipoprotein (HDL) (pval.adj = 0.0013), glucose (pval.adj = 0.043), leptin (pval.adj = 0.043), glycerol (pval.adj = 0.0165), and non-esterified fatty acids (NEFA) (pval.adj = 0.00132) in 21-week-old mice (n = 10 per group). MWU-test, by multiple correction to calculate p values. d Quantification of polyacetylation of histone 4 4–17 following ERF feeding. p values, paired MWU-test (H4K5K8) = 0.8125, (H4K12K16) = 0.0469 (H4-rest di-ac) = 0.1563, (H4K5K8K12) = 0.9375, (H4K5K8K16) = 0.0781, (H4K5K12K16) = 0.0313, (H4K8K12K16) = 0.0781, (H4 tetra-ac) = 0.375 (n = 7 per group). e RT-PCR comparing gene expression of candidate genes from ERF- and SBF-fed Titan mice siblings at 21 weeks of age (n = 7 per group). Paired t-test was performed followed by multiple correction to calculate adjusted p values; pval.adj (Dmdg) = 0.032, (Gnmt) = 0.012, (Bhmt) = 0.736, (Cyp7b1) = 0.008, (Elovl5) = 0.736, (Cyp2c37) = 0.012, (Acaca) = 0.0587, (Cth) = 0.012. f Switching to ERF feeding at 12 weeks of age (vertical black arrowhead) increased the lifespan of both Titan and control mice (Log-rank test; p = 0.0087, χ² = 6.892 Titan and p = 0.162, χ² = 1.955 control). The ages at 25, 75, and 90% death and the median lifespan are presented in Supplementary Table 1. *p < 0.05, **p < 0.01, ***p < 0.001. Error bars indicate SEM.