Per capita fructose consumption has increased 100-fold over the last century1. Epidemiological studies suggest that excessive fructose consumption, and especially consumption of sweet drinks, is associated with hyperlipidaemia, non-alcoholic fatty liver disease, obesity and diabetes2,3,4,5,6,7. Fructose metabolism begins with its phosphorylation by the enzyme ketohexokinase (KHK), which exists in two alternatively spliced forms8. The more active isozyme, KHK-C, is expressed most strongly in the liver, but also substantially in the small intestine9,10 where it drives dietary fructose absorption and conversion into other metabolites before fructose reaches the liver11,12,13. It is unclear whether intestinal fructose metabolism prevents or contributes to fructose-induced lipogenesis and liver pathology. Here we show that intestinal fructose catabolism mitigates fructose-induced hepatic lipogenesis. In mice, intestine-specific KHK-C deletion increases dietary fructose transit to the liver and gut microbiota and sensitizes mice to fructose’s hyperlipidaemic effects and hepatic steatosis. In contrast, intestine-specific KHK-C overexpression promotes intestinal fructose clearance and decreases fructose-induced lipogenesis. Thus, intestinal fructose clearance capacity controls the rate at which fructose can be safely ingested. Consistent with this, we show that the same amount of fructose is more strongly lipogenic when drunk than eaten, or when administered as a single gavage, as opposed to multiple doses spread over 45 min. Collectively, these data demonstrate that fructose induces lipogenesis when its dietary intake rate exceeds the intestinal clearance capacity. In the modern context of ready food availability, the resulting fructose spillover drives metabolic syndrome. Slower fructose intake, tailored to intestinal capacity, can mitigate these consequences.
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The data that support the findings of this study are available from the corresponding author (J.D.R.) upon request. Source data are provided with this paper.
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This work was supported by a grant from the DRC Regional Metabolomics Core (no. P30 DK19525 to J.D.R.), National Institutes of Health Pioneer grant (no. 1DP1DK113643 to J.D.R.) and grant no. DK107667 to Z.A. C.J. was a postdoctoral fellow of the American Diabetes Association (no. 1–17-PDF-076). We thank members of the Arany and Rabinowitz laboratories for scientific discussions.
J.D.R. is a consultant and receives research funding from Pfizer and is an advisor of Colorado Research Partners and cofounder of VL54. All other authors declare no conflicts of interest.
Peer review information Primary Handling Editor: Christoph Schmitt.
Publisher’s note Springer Nature remains neutral with regard to jurisdictional claims in published maps and institutional affiliations.
Extended Data Fig. 1 Intestine-specific Khk-C ablation increases fructose spillover to the colonic microbiota.
a, Khk-C floxed mice were generated by inserting two loxP sequences (black arrowheads) on both sides of a Khk-C specific exon (orange). The mice were then crossed with the Villin-Cre mice to generate intestine-specific Khk-C KO mice. b, qPCR (left) and western blots (right) show ablation of Khk-C in the jejunum but not in the liver (N = 6 mice for WT, 4 mice for KO). c, Khk-A mRNA is induced in the jejunum of KO mice (N = 6 mice for WT, 4 mice for KO). d, e, Mice received 1:1 mixture of 13C-fructose and unlabeled glucose (1 g/kg each) via oral gavage and the labeled F1P in duodenum and jejunum were measured in d and e, respectively (N = 3 mice for each time point). The AUC is shown on the right for the jejunum in e. f, AUC of labeled fructose in the cecal content over 1 h after gavage (N = 3 mice per group). g, Concentrations of total labeled carbons in acetate, propionate and butyrate in the cecal content 1 h or 2 h after gavage (N = 4, 3, 5, 5 mice). Data are mean ± standard error. Numbers in graphs indicate P-values by two-sided Student’s t-test. Source data
Mice received normal water or 10% sucrose water for 8 weeks. a, Fructolytic gene expression in the jejunum (N = 4, 6, 7, 12 mice). b, Metabolic parameters. P-values were calculated by two-sided Student’s t-test without adjustment for multiple comparisons. c, Body weights (N = 5 mice per group, except N = 4 mice for male KO). d, Epididymal fat pad weights (N = 5 mice per group, except N = 4 mice for male KO). Data are mean ± standard error.
a, Mice received 1:1 mixture of fructose and glucose (either hexose 13C-labeled, 2 g/kg each) via oral gavage. After 6 h, the labeling of saponified palmitate in serum, liver and jejunum was compared (N = 6 mice per group) IC, Ion count. b, Mice received normal water or 10% sucrose water for 8 weeks. Mice received 50% D2O in the drinking water overnight. The next morning, circulating fatty acid labeling was measured. Data are reported as the total number of deuterium atoms assimilated into the indicated saponified fatty acid (product of fatty acid concentration x average number of deuteriums per fatty acid molecule) (N = 7, 8, 4, 7 mice). c, d, Fractional contribution of fructose carbons into lipogenic acetyl-CoA pools. The acetyl group labeling is inferred based on the labeled palmitate mass isotopic distribution measured 6 h after 13C-fructose gavage (with unlabeled glucose, 2 g/kg each) in c (N = 6, 6, 8, 9 mice), or 24 h after ad lib drinking fructose and glucose mixture (15% each, either hexose 13C-labeled) in d (N = 4, 5, 4, 5 mice). Data are mean ± standard error. Numbers in graphs indicate P-values by two-sided Student’s t-test.
Extended Data Fig. 4 Intestine-specific Khk-C ablation worsens hyperlipidemia after chronic sucrose drinking.
Mice received 10% sucrose water for 1 month or 3 months. Heatmap shows circulating lipid profiles measured by LC-MS. Each lipid species was normalized by the average of all mice (N = 6, 3, 8, 6 mice). TG, triglyceride; DG, diglyceride.
Extended Data Fig. 5 Intestine-specific Khk-C ablation worsens hepatic steatosis after chronic sucrose drinking.
Mice received normal water (NW) or 10% sucrose water (Suc) for 5 months. a, Heatmap shows hepatic lipid profiles measured by LC-MS. Each lipid species was normalized by the average of all mice (N = 4, 5, 8, 7 mice). b, Sucrose drinking increases saturated triglycerides, consistent with increased lipogenesis. Note that the trend is stronger in the intestine-specific Khk-C KO mice. Y-axis is a log scale. Numbers on the x-axis (for example, 48:1) mean the total number of carbons (for example, 48) and the number of double bonds (for example, 1) in triglyceride, with more unsaturated triglycerides having more double bonds (N = 4, 5, 8, 7 mice). Data are mean ± standard error.
Extended Data Fig. 6 Hepatic gene expression profiles of intestine-specific Khk-C KO mice after chronic sucrose drinking.
a–c, Mice received normal water or 10% sucrose water for 5 months. Then, the livers were isolated at 9 AM and the mRNA levels of the indicated genes were measured (N = 6, 6, 11, 15 mice). Data are mean ± standard error. Numbers in graphs indicate P-values by two-sided Student’s t-test.
Extended Data Fig. 7 Intestine-specific Khk-C overexpression increases intestinal fructose catabolism.
a, Khk-C inducible overexpressing transgenic (TG) mice were generated by cloning a Khk-C coding sequence (orange) under the TET-on promoter (green). Then, the mice were crossed with mice that harbored Villin-Cre driver (blue) and rtTA with a stop-codon (gray) in the ROSA26 genomic region. Mice were fed with doxycycline in the drinking water to induce deletion of the stop-codon and subsequent Khk-C overexpression in the Villin-expressing intestinal epithelial cells. b, qPCR (left) and western blots (right) show Khk-C induction in the jejunum but not in the liver (N = 4 mice per group). The exposure time for Khk-C blot in the jejunum was decreased to prevent signal saturation. c, Khk-A mRNA levels were measured in the liver or jejunum (N = 4 mice per group). d, e, Mice received 1:1 mixture of 13C-fructose and unlabeled glucose (1 g/kg each) via oral gavage and the levels of labeled F1P and glycerate in the jejunum were measured in d, e, respectively (N = 3 mice for each time point). AUC is shown on the right. Data are mean ± standard error. Numbers in graphs indicate P-values by two-sided Student’s t-test. Source data
Mice received normal water or sucrose water for 3 months. a, Body weights (N = 5, 6 mice). b, Heatmap shows circulating lipid profiles measured by LC-MS. Each lipid species was normalized by the average of all mice (N = 3, 6, 4, 5 mice). c, Daily water consumption (N = 6, 5, 4, 5 mice). Data are mean ± standard error. Number in the graph indicates a P-value by two-sided Student’s t-test.
Extended Data Fig. 9 Intestine-specific Khk-C TG overexpression induces cecal fructose catabolism and reduces lipogenesis.
a–d, Mice received normal water or sucrose water for 3 months. Mice received 1:1 mixture of 13C-fructose and unlabeled glucose (1 g/kg) via oral gavage and the AUC of cecal labeled F1P was measured in a (N = 3 mice per group). Representative hematoxylin & eosin stained jejunum sections from two independent experiments are shown in b. Mice received 1:1 mixture of 13C-fructose and unlabeled glucose (1 g/kg) via oral gavage and the AUC of labeled fructose in the cecal content was measured in c (N = 4, 3 mice). The mRNA levels of fructolytic genes in the cecum was measured in d (N = 7, 6 mice). e-g, Mice received 1:1 mixture of fructose and glucose (2 g/kg each) via oral gavage twice a day for five days. The mRNA levels of fructolytic genes in the jejunum was measured in e (N = 5, 4, 5, 6 mice). On the first day (day 1) or the last day (day 5), the mice received 1:1 mixture of 13C-fructose and unlabeled glucose (2 g/kg each) via oral gavage and the 13C-labeling of the indicated circulating saponified fatty acids was measured after 6 h in f (N = 10 mice for WT, 6 mice for TG). Alternatively, on day 5, mice received 100% D2O via intra-peritoneal injection and the deuterium labeling of the indicated circulating saponified fatty acids was measured after 6 h in g (N = 5 mice per group). Data are mean ± standard error. Numbers in graphs indicate P-values by two-sided Student’s t-test.
a, Mice received 1:1 mixture of 13C-fructose and unlabeled glucose (2 g/kg) via oral gavage. The same total quantity of fructose was administered as a single high-dose or divided doses over 45 min. Then, for each group, intestinal fructose catabolism was quantitated, using the labeled metabolite concentration differences between the portal and systemic blood (N = 3 mice per group). b, Experimental design for fructose drinking (10% sucrose drinking water) or eating (10% sucrose hydrogel). Mice were provided the liquid or solid containing sucrose for 7 days. On day 7, the water (in either the sucrose drinking water or the sucrose hydrogel) was 25% D2O. This served two purposes: (i) enabling precise tracking of total consumption based on circulating D2O enrichment and (ii) lipogenesis measurement. On day 8, blood was harvested. c, d, The mice that received the sweetened hydrogel showed less lipogenesis than the mice that received the sweetened drinking water. Deuterium labeling of circulating water was indistinguishable in c (N = 9, 6 mice), indicating identical total fructose consumption. Deuterium labeling of circulating saponified fatty acids was greater in the liquid group in d (N = 9, 6 mice). Data are mean ± standard error. Numbers in graphs indicate P-values by two-sided Student’s t-test.
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Jang, C., Wada, S., Yang, S. et al. The small intestine shields the liver from fructose-induced steatosis. Nat Metab 2, 586–593 (2020). https://doi.org/10.1038/s42255-020-0222-9
Nature Metabolism (2020)