Xianfeng Zeng

All Publications

• Gut bacterial nutrient preferences quantified in vivo CELL Zeng, X., Xing, X., Gupta, M., Keber, F. C., Lopez, J. G., Lee, Y. J., Roichman, A., Wang, L., Neinast, M. D., Donia, M. S., Wuhr, M., Jang, C., Rabinowitz, J. D. 2022; 185 (18): 3441-+

  Abstract

  Great progress has been made in understanding gut microbiomes' products and their effects on health and disease. Less attention, however, has been given to the inputs that gut bacteria consume. Here, we quantitatively examine inputs and outputs of the mouse gut microbiome, using isotope tracing. The main input to microbial carbohydrate fermentation is dietary fiber and to branched-chain fatty acids and aromatic metabolites is dietary protein. In addition, circulating host lactate, 3-hydroxybutyrate, and urea (but not glucose or amino acids) feed the gut microbiome. To determine the nutrient preferences across bacteria, we traced into genus-specific bacterial protein sequences. We found systematic differences in nutrient use: most genera in the phylum Firmicutes prefer dietary protein, Bacteroides dietary fiber, and Akkermansia circulating host lactate. Such preferences correlate with microbiome composition changes in response to dietary modifications. Thus, diet shapes the microbiome by promoting the growth of bacteria that preferentially use the ingested nutrients.

  View details for DOI 10.1016/j.cell.2022.07.020

  View details for Web of Science ID 000852676300001

  View details for PubMedID 36055202

  View details for PubMedCentralID PMC9450212

• Comprehensive quantification of metabolic flux during acute cold stress in mice. Cell metabolism Bornstein, M. R., Neinast, M. D., Zeng, X., Chu, Q., Axsom, J., Thorsheim, C., Li, K., Blair, M. C., Rabinowitz, J. D., Arany, Z. 2023; 35 (11): 2077-2092.e6

  Abstract

  Cold-induced thermogenesis (CIT) is widely studied as a potential avenue to treat obesity, but a thorough understanding of the metabolic changes driving CIT is lacking. Here, we present a comprehensive and quantitative analysis of the metabolic response to acute cold exposure, leveraging metabolomic profiling and minimally perturbative isotope tracing studies in unanesthetized mice. During cold exposure, brown adipose tissue (BAT) primarily fueled the tricarboxylic acid (TCA) cycle with fat in fasted mice and glucose in fed mice, underscoring BAT's metabolic flexibility. BAT minimally used branched-chain amino acids or ketones, which were instead avidly consumed by muscle during cold exposure. Surprisingly, isotopic labeling analyses revealed that BAT uses glucose largely for TCA anaplerosis via pyruvate carboxylation. Finally, we find that cold-induced hepatic gluconeogenesis is critical for CIT during fasting, demonstrating a key functional role for glucose metabolism. Together, these findings provide a detailed map of the metabolic rewiring driving acute CIT.

  View details for DOI 10.1016/j.cmet.2023.09.002

  View details for PubMedID 37802078

• NAD precursors cycle between host tissues and the gut microbiome CELL METABOLISM Chellappa, K., McReynolds, M. R., Lu, W., Zeng, X., Makarov, M., Hayat, F., Mukherjee, S., Bhat, Y. R., Lingala, S. R., Shima, R. T., Descamps, H. C., Cox, T., Ji, L., Jankowski, C., Chu, Q., Davidson, S. M., Thaiss, C. A., Migaud, M. E., Rabinowitz, J. D., Baur, J. A. 2022; 34 (12): 1947-+

  Abstract

  Nicotinamide adenine dinucleotide (NAD) is an essential redox cofactor in mammals and microbes. Here we use isotope tracing to investigate the precursors supporting NAD synthesis in the gut microbiome of mice. We find that dietary NAD precursors are absorbed in the proximal part of the gastrointestinal tract and not available to microbes in the distal gut. Instead, circulating host nicotinamide enters the gut lumen and supports microbial NAD synthesis. The microbiome converts host-derived nicotinamide into nicotinic acid, which is used for NAD synthesis in host tissues and maintains circulating nicotinic acid levels even in the absence of dietary consumption. Moreover, the main route from oral nicotinamide riboside, a widely used nutraceutical, to host NAD is via conversion into nicotinic acid by the gut microbiome. Thus, we establish the capacity for circulating host micronutrients to feed the gut microbiome, and in turn be transformed in a manner that enhances host metabolic flexibility.

  View details for DOI 10.1016/j.cmet.2022.11.004

  View details for Web of Science ID 000901818900009

  View details for PubMedID 36476934

  View details for PubMedCentralID PMC9825113

• Ketogenic diet and chemotherapy combine to disrupt pancreatic cancer metabolism and growth MED Yang, L., TeSlaa, T., Ng, S., Nofal, M., Wang, L., Lan, T., Zeng, X., Cowan, A., McBride, M., Lu, W., Davidson, S., Liang, G., Oh, T., Downes, M., Evans, R., Von Hoff, D., Guo, J., Han, H., Rabinowitz, J. D. 2022; 3 (2): 119-+

  Abstract

  Ketogenic diet is a potential means of augmenting cancer therapy. Here, we explore ketone body metabolism and its interplay with chemotherapy in pancreatic cancer.Metabolism and therapeutic responses of murine pancreatic cancer were studied using KPC primary tumors and tumor chunk allografts. Mice on standard high-carbohydrate diet or ketogenic diet were treated with cytotoxic chemotherapy (nab-paclitaxel, gemcitabine, cisplatin). Metabolic activity was monitored with metabolomics and isotope tracing, including 2H- and 13C-tracers, liquid chromatography-mass spectrometry, and imaging mass spectrometry.Ketone bodies are unidirectionally oxidized to make NADH. This stands in contrast to the carbohydrate-derived carboxylic acids lactate and pyruvate, which rapidly interconvert, buffering NADH/NAD. In murine pancreatic tumors, ketogenic diet decreases glucose's concentration and tricarboxylic acid cycle contribution, enhances 3-hydroxybutyrate's concentration and tricarboxylic acid contribution, and modestly elevates NADH, but does not impact tumor growth. In contrast, the combination of ketogenic diet and cytotoxic chemotherapy substantially raises tumor NADH and synergistically suppresses tumor growth, tripling the survival benefits of chemotherapy alone. Chemotherapy and ketogenic diet also synergize in immune-deficient mice, although long-term growth suppression was only observed in mice with an intact immune system.Ketogenic diet sensitizes murine pancreatic cancer tumors to cytotoxic chemotherapy. Based on these data, we have initiated a randomized clinical trial of chemotherapy with standard versus ketogenic diet for patients with metastatic pancreatic cancer (NCT04631445).

  View details for DOI 10.1016/j.medj.2021.12.008

  View details for Web of Science ID 000758830400011

  View details for PubMedID 35425930

  View details for PubMedCentralID PMC9004683

• Spatially resolved isotope tracing reveals tissue metabolic activity NATURE METHODS Wang, L., Xing, X., Zengl, X., Jackson, S., TeSlaa, T., Al-Dalahmah, O., Samarah, L. Z., Goodwin, K., Yang, L., McReynolds, M. R., Li, X., Wolff, J. J., Rabinowitz, J. D., Davidson, S. M. 2022; 19 (2): 223-+

  Abstract

  Isotope tracing has helped to determine the metabolic activities of organs. Methods to probe metabolic heterogeneity within organs are less developed. We couple stable-isotope-labeled nutrient infusion to matrix-assisted laser desorption ionization imaging mass spectrometry (iso-imaging) to quantitate metabolic activity in mammalian tissues in a spatially resolved manner. In the kidney, we visualize gluconeogenic flux and glycolytic flux in the cortex and medulla, respectively. Tricarboxylic acid cycle substrate usage differs across kidney regions; glutamine and citrate are used preferentially in the cortex and fatty acids are used in the medulla. In the brain, we observe spatial gradations in carbon inputs to the tricarboxylic acid cycle and glutamate under a ketogenic diet. In a carbohydrate-rich diet, glucose predominates throughout but in a ketogenic diet, 3-hydroxybutyrate contributes most strongly in the hippocampus and least in the midbrain. Brain nitrogen sources also vary spatially; branched-chain amino acids contribute most in the midbrain, whereas ammonia contributes in the thalamus. Thus, iso-imaging can reveal the spatial organization of metabolic activity.

  View details for DOI 10.1038/s41592-021-01378-y

  View details for Web of Science ID 000752252500001

  View details for PubMedID 35132243

• Circulating metabolite homeostasis achieved through mass action NATURE METABOLISM Li, X., Hui, S., Mirek, E. T., Jonsson, W. O., Anthony, T. G., Lee, W., Zeng, X., Jang, C., Rabinowitz, J. D. 2022; 4 (1): 141-+

  Abstract

  Homeostasis maintains serum metabolites within physiological ranges. For glucose, this requires insulin, which suppresses glucose production while accelerating its consumption. For other circulating metabolites, a comparable master regulator has yet to be discovered. Here we show that, in mice, many circulating metabolites are cleared via the tricarboxylic acid cycle (TCA) cycle in linear proportionality to their circulating concentration. Abundant circulating metabolites (essential amino acids, serine, alanine, citrate, 3-hydroxybutyrate) were administered intravenously in perturbative amounts and their fluxes were measured using isotope labelling. The increased circulating concentrations induced by the perturbative infusions hardly altered production fluxes while linearly enhancing consumption fluxes and TCA contributions. The same mass action relationship between concentration and consumption flux largely held across feeding, fasting and high- and low-protein diets, with amino acid homeostasis during fasting further supported by enhanced endogenous protein catabolism. Thus, despite the copious regulatory machinery in mammals, circulating metabolite homeostasis is achieved substantially through mass action-driven oxidation.

  View details for DOI 10.1038/s42255-021-00517-1

  View details for Web of Science ID 000744999800001

  View details for PubMedID 35058631

  View details for PubMedCentralID PMC9244777

• Serine catabolism generates liver NADPH and supports hepatic lipogenesis NATURE METABOLISM Zhang, Z., TeSlaa, T., Xu, X., Zeng, X., Yang, L., Xing, G., Tesz, G. J., Clasquin, M. F., Rabinowitz, J. D. 2021; 3 (12): 1608-+

  Abstract

  Carbohydrate can be converted into fat by de novo lipogenesis, a process upregulated in fatty liver disease. Chemically, de novo lipogenesis involves polymerization and reduction of acetyl-CoA, using NADPH as the electron donor. The feedstocks used to generate acetyl-CoA and NADPH in lipogenic tissues remain, however, unclear. Here we show using stable isotope tracing in mice that de novo lipogenesis in adipose is supported by glucose and its catabolism via the pentose phosphate pathway to make NADPH. The liver, in contrast, derives acetyl-CoA for lipogenesis from acetate and lactate, and NADPH from folate-mediated serine catabolism. Such NADPH generation involves the cytosolic serine pathway in liver running in the opposite direction to that observed in most tissues and tumours, with NADPH made by the SHMT1-MTHFD1-ALDH1L1 reaction sequence. SHMT inhibition decreases hepatic lipogenesis. Thus, liver folate metabolism is distinctively wired to support cytosolic NADPH production and lipogenesis. More generally, while the same enzymes are involved in fat synthesis in liver and adipose, different substrates are used, opening the door to tissue-specific pharmacological interventions.

  View details for DOI 10.1038/s42255-021-00487-4

  View details for Web of Science ID 000723498100001

  View details for PubMedID 34845393

  View details for PubMedCentralID PMC8721747

• Metabolite discovery through global annotation of untargeted metabolomics data NATURE METHODS Chen, L., Lu, W., Wang, L., Xing, X., Chen, Z., Teng, X., Zeng, X., Muscarella, A. D., Shen, Y., Cowan, A., McReynolds, M. R., Kennedy, B. J., Lato, A. M., Campagna, S. R., Singh, M., Rabinowitz, J. D. 2021; 18 (11): 1377-+

  Abstract

  Liquid chromatography-high-resolution mass spectrometry (LC-MS)-based metabolomics aims to identify and quantify all metabolites, but most LC-MS peaks remain unidentified. Here we present a global network optimization approach, NetID, to annotate untargeted LC-MS metabolomics data. The approach aims to generate, for all experimentally observed ion peaks, annotations that match the measured masses, retention times and (when available) tandem mass spectrometry fragmentation patterns. Peaks are connected based on mass differences reflecting adduction, fragmentation, isotopes, or feasible biochemical transformations. Global optimization generates a single network linking most observed ion peaks, enhances peak assignment accuracy, and produces chemically informative peak-peak relationships, including for peaks lacking tandem mass spectrometry spectra. Applying this approach to yeast and mouse data, we identified five previously unrecognized metabolites (thiamine derivatives and N-glucosyl-taurine). Isotope tracer studies indicate active flux through these metabolites. Thus, NetID applies existing metabolomic knowledge and global optimization to substantially improve annotation coverage and accuracy in untargeted metabolomics datasets, facilitating metabolite discovery.

  View details for DOI 10.1038/s41592-021-01303-3

  View details for Web of Science ID 000712400000005

  View details for PubMedID 34711973

  View details for PubMedCentralID PMC8733904

• Quantitative Fluxomics of Circulating Metabolites CELL METABOLISM Hui, S., Cowan, A. J., Zeng, X., Yang, L., TeSlaa, T., Li, X., Bartman, C., Zhang, Z., Jang, C., Wang, L., Lu, W., Rojas, J., Baur, J., Rabinowitz, J. D. 2020; 32 (4): 676-+

  Abstract

  Mammalian organs are nourished by nutrients carried by the blood circulation. These nutrients originate from diet and internal stores, and can undergo various interconversions before their eventual use as tissue fuel. Here we develop isotope tracing, mass spectrometry, and mathematical analysis methods to determine the direct sources of circulating nutrients, their interconversion rates, and eventual tissue-specific contributions to TCA cycle metabolism. Experiments with fifteen nutrient tracers enabled extensive accounting for both circulatory metabolic cycles and tissue TCA inputs, across fed and fasted mice on either high-carbohydrate or ketogenic diet. We find that a majority of circulating carbon flux is carried by two major cycles: glucose-lactate and triglyceride-glycerol-fatty acid. Futile cycling through these pathways is prominent when dietary content of the associated nutrients is low, rendering internal metabolic activity robust to food choice. The presented in vivo flux quantification methods are broadly applicable to different physiological and disease states.

  View details for DOI 10.1016/j.cmet.2020.07.013

  View details for Web of Science ID 000582325100018

  View details for PubMedID 32791100

  View details for PubMedCentralID PMC7544659

• The small intestine shields the liver from fructose-induced steatosis NATURE METABOLISM Jang, C., Wada, S., Yang, S., Gosis, B., Zeng, X., Zhang, Z., Shen, Y., Lee, G., Arany, Z., Rabinowitz, J. D. 2020; 2 (7): 586-+

  Abstract

  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 diabetes2-7. 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 liver11-13. 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.

  View details for DOI 10.1038/s42255-020-0222-9

  View details for Web of Science ID 000551962800006

  View details for PubMedID 32694791

  View details for PubMedCentralID PMC8020332

• Dietary fructose feeds hepatic lipogenesis via microbiota-derived acetate NATURE Zhao, S., Jang, C., Liu, J., Uehara, K., Gilbert, M., Izzo, L., Zeng, X., Trefely, S., Fernandez, S., Carrer, A., Miller, K. D., Schug, Z. T., Snyder, N. W., Gade, T. P., Titchenell, P. M., Rabinowitz, J. D., Wellen, K. E. 2020; 579 (7800): 586-+

  Abstract

  Consumption of fructose has risen markedly in recent decades owing to the use of sucrose and high-fructose corn syrup in beverages and processed foods1, and this has contributed to increasing rates of obesity and non-alcoholic fatty liver disease2-4. Fructose intake triggers de novo lipogenesis in the liver4-6, in which carbon precursors of acetyl-CoA are converted into fatty acids. The ATP citrate lyase (ACLY) enzyme cleaves cytosolic citrate to generate acetyl-CoA, and is upregulated after consumption of carbohydrates7. Clinical trials are currently pursuing the inhibition of ACLY as a treatment for metabolic diseases8. However, the route from dietary fructose to hepatic acetyl-CoA and lipids remains unknown. Here, using in vivo isotope tracing, we show that liver-specific deletion of Acly in mice is unable to suppress fructose-induced lipogenesis. Dietary fructose is converted to acetate by the gut microbiota9, and this supplies lipogenic acetyl-CoA independently of ACLY10. Depletion of the microbiota or silencing of hepatic ACSS2, which generates acetyl-CoA from acetate, potently suppresses the conversion of bolus fructose into hepatic acetyl-CoA and fatty acids. When fructose is consumed more gradually to facilitate its absorption in the small intestine, both citrate cleavage in hepatocytes and microorganism-derived acetate contribute to lipogenesis. By contrast, the lipogenic transcriptional program is activated in response to fructose in a manner that is independent of acetyl-CoA metabolism. These data reveal a two-pronged mechanism that regulates hepatic lipogenesis, in which fructolysis within hepatocytes provides a signal to promote the expression of lipogenic genes, and the generation of microbial acetate feeds lipogenic pools of acetyl-CoA.

  View details for DOI 10.1038/s41586-020-2101-7

  View details for Web of Science ID 000520406700003

  View details for PubMedID 32214246

  View details for PubMedCentralID PMC7416516

• Metabolite Exchange between Mammalian Organs Quantified in Pigs CELL METABOLISM Jang, C., Hui, S., Zeng, X., Cowan, A. J., Wang, L., Chen, L., Morscher, R. J., Reyes, J., Frezza, C., Hwang, H., Imai, A., Saito, Y., Okamoto, K., Vaspoli, C., Kasprenski, L., Zsido, G. A., Gorman, J. H., Gorman, R. C., Rabinowitz, J. D. 2019; 30 (3): 594-+

  Abstract

  Mammalian organs continually exchange metabolites via circulation, but systems-level analysis of this shuttling process is lacking. Here, we compared, in fasted pigs, metabolite concentrations in arterial blood versus draining venous blood from 11 organs. Greater than 90% of metabolites showed arterial-venous differences across at least one organ. Surprisingly, the liver and kidneys released not only glucose but also amino acids, both of which were consumed primarily by the intestine and pancreas. The liver and kidneys exhibited additional unexpected activities: liver preferentially burned unsaturated over more atherogenic saturated fatty acids, whereas the kidneys were unique in burning circulating citrate and net oxidizing lactate to pyruvate, thereby contributing to circulating redox homeostasis. Furthermore, we observed more than 700 other cases of tissue-specific metabolite production or consumption, such as release of nucleotides by the spleen and TCA intermediates by pancreas. These data constitute a high-value resource, providing a quantitative atlas of inter-organ metabolite exchange.

  View details for DOI 10.1016/j.cmet.2019.06.002

  View details for Web of Science ID 000484370400017

  View details for PubMedID 31257152

  View details for PubMedCentralID PMC6726553

• Extracellular vesicles derived from ODN-stimulated macrophages transfer and activate Cdc42 in recipient cells and thereby increase cellular permissiveness to EV uptake SCIENCE ADVANCES Zhang, Y., Jin, X., Liang, J., Guo, Y., Sun, G., Zeng, X., Yin, H. 2019; 5 (7): eaav1564

  Abstract

  Endosomal Toll-like receptors (TLRs) mediate intracellular innate immunity via the recognition of DNA and RNA sequences. Recent work has reported a role for extracellular vesicles (EVs), known to transfer various nucleic acids, in uptake of TLR-activating molecules, raising speculation about possible roles of EVs in innate immune surveillance. Whether EV-mediated uptake is a general mechanism, however, was unresolved; and the molecular machinery that might be involved was unknown. We show that, when macrophages are stimulated with the TLR9 agonist CpG oligodeoxynucleotides (ODN), the secreted EVs transport ODN into naïve macrophages and induce the release of chemokine TNF-α. In addition, these EVs transfer Cdc42 into recipient cells, resulting in further enhancement of their cellular uptake. Transport of ODN and Cdc42 from TLR9-activated macrophages to naïve cells via EVs exerts synergetic effects in propagation of the intracellular immune response, suggesting a general mechanism of EV-mediated uptake of pathogen-associated molecular patterns.

  View details for DOI 10.1126/sciadv.aav1564

  View details for Web of Science ID 000478770400017

  View details for PubMedID 31355328

  View details for PubMedCentralID PMC6656539

• Small Molecule and Peptide Recognition of Protein Transmembrane Domains BIOCHEMISTRY Zeng, X., Wu, P., Yao, C., Liang, J., Zhang, S., Yin, H. 2017; 56 (15): 2076-2085

  Abstract

  Membrane proteins play vital roles in cell signaling, molecular transportation, and cell adhesion. The interactions of transmembrane domains are much less well understood than those of their water-soluble counterparts, and they have been deemed "undruggable" despite their important biological functions such as protein anchoring, signal transduction, and ligand recognition. Nevertheless, continual developments in this area have revealed useful probes for investigating and regulating these membrane proteins. This review summarizes and evaluates the strategies available for discovering small molecules and peptides that recognize the protein transmembrane domains of membrane proteins, with a particular focus on rational design and library screening.

  View details for DOI 10.1021/acs.biochem.6b00909

  View details for Web of Science ID 000399858600002

  View details for PubMedID 28353343

• Pyrimidine Triazole Thioether Derivatives as Toll-Like Receptor5 (TLR5)/Flagellin Complex Inhibitors CHEMMEDCHEM Yan, L., Liang, J., Yao, C., Wu, P., Zeng, X., Cheng, K., Yin, H. 2016; 11 (8): 822-826

  Abstract

  Protein-protein interactions have been regarded as "undruggable" despite their importance in many biological processes. The complex formed between host toll-like receptor 5 (TLR5) and flagellin, a globular protein that is the main component of a bacterial flagellum, plays a vital role in a number of pathogen defenses, immunological diseases and cancers. Through high-throughput screening, we identified two hits with a common pharmacophore, which were used to successfully develop a series of small-molecule probes as novel inhibitors of flagellin binding to TLR5. In a multitude of assays, 4-((4-benzyl-5-(pyridin4yl)-4H-1,2,4-triazol-3-yl)thio)pyrido[3',2':4,5]thieno[3,2-d]pyrimidine (TH1020) was identified as a potent antagonist of TLR5 signaling with promising activity (IC50 =0.85±0.12 μm) and specificity. Furthermore, TH1020 was shown to repress the expression of downstream TNF-α signaling pathways mediated by the TLR5/flagellin complex formation. Based on molecular docking simulation, TH1020 is suggested to compete with flagellin and disrupt its association with TLR5. TH1020 provides a much-needed molecular probe for studying this important protein-protein interaction and a lead compound for identifying novel therapeutics targeting TLR5.

  View details for DOI 10.1002/cmdc.201500471

  View details for Web of Science ID 000374693500009

  View details for PubMedID 26634412