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Fasted FFA levels were similar in all groups (Table 2). To assess whether dietary history influenced whole-body energy metabolism, indirect calorimetric measurements were performed. Respiratory exchange ratio, as an indicator of metabolic substrate usage, showed a significant difference, as expected, between LF- and HF-fed mice at 4 weeks, with a higher carbohydrate oxidation rate and lower fat oxidation rate for the LF-fed mice (Table 3). This was also observed at 12 weeks (Table 3). There was no effect of time (from 4 to 12 weeks) on find more substrate utilization for L versus LL groups and H versus HH groups (data not shown). Relative cumulative frequency curves of RER for LH and HH, as well as for HL and LL groups, align perfectly, which fits the observation that the RER represents current dietary intake independent of dietary history (Fig. 4). Matching fuel oxidation (RER) to fuel availability is defined as a measure GPX4 for metabolic flexibility (Lin et al. 2011) and, in particular for the HL group, this suggests that their metabolic flexibility was preserved despite previous HF consumption. Energy expenditure remained similar between all groups at 4 weeks, even though the HF-fed mice showed an increased BW. At 12 weeks, the LF-consuming groups, LL and HL, were equal in their mean EE, but both HF-consuming groups, LH and HH, showed a significant increase in EE. Again, no significant difference in EE was observed between the mice that were continuously fed the same diet and the mice that switched diets (Table 3). Interestingly, adiposity (Fig. 2C and D) showed a significant difference between LF-fed groups and HF-fed groups, suggesting that fat mass could be responsible for the observed difference in EE. When corrected for BW by covariance analysis, the differences in EE by diet group were no longer significant. Unfortunately, we lack carcass analyses, so we cannot fully exclude a contribution of (small) differences in lean mass, if at all present. In this study, we examined whether previous purified dietary intake patterns have a lasting effect on a newly achieved energy balance and BW. We studied one and two changes in diet and observed that energy intake was dependent on the present diet consumed and not influenced by a previous diet. Body weight, WAT mass and histology, serum learn more metabolic parameters, EE and RER levels depended on the present diet being consumed, without any effect of a previous diet. Our study, although more rigorous in the sense that we used two dietary changes, confirmed previous findings (Parekh et al. 1998), in that no effect of dietary history was seen when mice were fed purified diets with identical ingredients. This set-up minimizes possible other effects of diets, such as differences in growth due to differences in the amount or composition of amino acids, micronutrients or bioactive food components.