Dietary butyrate ameliorates metabolic health associated with selective proliferation of gut Lachnospiraceae bacterium 28-4

Research ArticleEndocrinologyMicrobiology Open Access | 10.1172/jci.insight.166655

Zhuang Li,1,2,3 Enchen Zhou,1,2 Cong Liu,1,2 Hope Wicks,1,2 Sena Yildiz,1,2 Farhana Razack,1,2 Zhixiong Ying,1,2 Sander Kooijman,1,2 Debby P.Y. Koonen,4 Marieke Heijink,5 Sarantos Kostidis,5 Martin Giera,5 Ingrid M.J.G. Sanders,6 Ed J. Kuijper,6,7 Wiep Klaas Smits,6,7 Ko Willems van Dijk,2,8 Patrick C.N. Rensen,1,2,9 and Yanan Wang1,2,9

1Department of Medicine, Division of Endocrinology, and

2Einthoven Laboratory for Experimental Vascular Medicine, Leiden University Medical Center, Leiden, Netherlands.

3Microbiome Medicine Center, Department of Laboratory Medicine, Zhujiang Hospital, Southern Medical University, Guangzhou, China.

4Department of Pediatrics, University Medical Center Groningen, Groningen, Netherlands.

5Center for Proteomics and Metabolomics,

6Department of Medical Microbiology,

7Center for Microbiome Analyses and Therapeutics, and

8Department of Human Genetics, Leiden University Medical Center, Netherlands.

9Med-X Institute, Center for Immunological and Metabolic Diseases, and Department of Endocrinology, the First Affiliated Hospital of Xi’an JiaoTong University, Xi’an Jiaotong University, Xi’an, China.

Address correspondence to: Yanan Wang, Department of Medicine, Division of Endocrinology, Leiden University Medical Center, Albinusdreef 2, 2333 ZA, Leiden, Netherlands. Phone: 317168163; Email: Y.Wang@lumc.nl.

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1Department of Medicine, Division of Endocrinology, and

2Einthoven Laboratory for Experimental Vascular Medicine, Leiden University Medical Center, Leiden, Netherlands.

3Microbiome Medicine Center, Department of Laboratory Medicine, Zhujiang Hospital, Southern Medical University, Guangzhou, China.

4Department of Pediatrics, University Medical Center Groningen, Groningen, Netherlands.

5Center for Proteomics and Metabolomics,

6Department of Medical Microbiology,

7Center for Microbiome Analyses and Therapeutics, and

8Department of Human Genetics, Leiden University Medical Center, Netherlands.

9Med-X Institute, Center for Immunological and Metabolic Diseases, and Department of Endocrinology, the First Affiliated Hospital of Xi’an JiaoTong University, Xi’an Jiaotong University, Xi’an, China.

Address correspondence to: Yanan Wang, Department of Medicine, Division of Endocrinology, Leiden University Medical Center, Albinusdreef 2, 2333 ZA, Leiden, Netherlands. Phone: 317168163; Email: Y.Wang@lumc.nl.

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1Department of Medicine, Division of Endocrinology, and

2Einthoven Laboratory for Experimental Vascular Medicine, Leiden University Medical Center, Leiden, Netherlands.

3Microbiome Medicine Center, Department of Laboratory Medicine, Zhujiang Hospital, Southern Medical University, Guangzhou, China.

4Department of Pediatrics, University Medical Center Groningen, Groningen, Netherlands.

5Center for Proteomics and Metabolomics,

6Department of Medical Microbiology,

7Center for Microbiome Analyses and Therapeutics, and

8Department of Human Genetics, Leiden University Medical Center, Netherlands.

9Med-X Institute, Center for Immunological and Metabolic Diseases, and Department of Endocrinology, the First Affiliated Hospital of Xi’an JiaoTong University, Xi’an Jiaotong University, Xi’an, China.

Address correspondence to: Yanan Wang, Department of Medicine, Division of Endocrinology, Leiden University Medical Center, Albinusdreef 2, 2333 ZA, Leiden, Netherlands. Phone: 317168163; Email: Y.Wang@lumc.nl.

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1Department of Medicine, Division of Endocrinology, and

2Einthoven Laboratory for Experimental Vascular Medicine, Leiden University Medical Center, Leiden, Netherlands.

3Microbiome Medicine Center, Department of Laboratory Medicine, Zhujiang Hospital, Southern Medical University, Guangzhou, China.

4Department of Pediatrics, University Medical Center Groningen, Groningen, Netherlands.

5Center for Proteomics and Metabolomics,

6Department of Medical Microbiology,

7Center for Microbiome Analyses and Therapeutics, and

8Department of Human Genetics, Leiden University Medical Center, Netherlands.

9Med-X Institute, Center for Immunological and Metabolic Diseases, and Department of Endocrinology, the First Affiliated Hospital of Xi’an JiaoTong University, Xi’an Jiaotong University, Xi’an, China.

Address correspondence to: Yanan Wang, Department of Medicine, Division of Endocrinology, Leiden University Medical Center, Albinusdreef 2, 2333 ZA, Leiden, Netherlands. Phone: 317168163; Email: Y.Wang@lumc.nl.

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1Department of Medicine, Division of Endocrinology, and

2Einthoven Laboratory for Experimental Vascular Medicine, Leiden University Medical Center, Leiden, Netherlands.

3Microbiome Medicine Center, Department of Laboratory Medicine, Zhujiang Hospital, Southern Medical University, Guangzhou, China.

4Department of Pediatrics, University Medical Center Groningen, Groningen, Netherlands.

5Center for Proteomics and Metabolomics,

6Department of Medical Microbiology,

7Center for Microbiome Analyses and Therapeutics, and

8Department of Human Genetics, Leiden University Medical Center, Netherlands.

9Med-X Institute, Center for Immunological and Metabolic Diseases, and Department of Endocrinology, the First Affiliated Hospital of Xi’an JiaoTong University, Xi’an Jiaotong University, Xi’an, China.

Address correspondence to: Yanan Wang, Department of Medicine, Division of Endocrinology, Leiden University Medical Center, Albinusdreef 2, 2333 ZA, Leiden, Netherlands. Phone: 317168163; Email: Y.Wang@lumc.nl.

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1Department of Medicine, Division of Endocrinology, and

2Einthoven Laboratory for Experimental Vascular Medicine, Leiden University Medical Center, Leiden, Netherlands.

3Microbiome Medicine Center, Department of Laboratory Medicine, Zhujiang Hospital, Southern Medical University, Guangzhou, China.

4Department of Pediatrics, University Medical Center Groningen, Groningen, Netherlands.

5Center for Proteomics and Metabolomics,

6Department of Medical Microbiology,

7Center for Microbiome Analyses and Therapeutics, and

8Department of Human Genetics, Leiden University Medical Center, Netherlands.

9Med-X Institute, Center for Immunological and Metabolic Diseases, and Department of Endocrinology, the First Affiliated Hospital of Xi’an JiaoTong University, Xi’an Jiaotong University, Xi’an, China.

Address correspondence to: Yanan Wang, Department of Medicine, Division of Endocrinology, Leiden University Medical Center, Albinusdreef 2, 2333 ZA, Leiden, Netherlands. Phone: 317168163; Email: Y.Wang@lumc.nl.

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1Department of Medicine, Division of Endocrinology, and

2Einthoven Laboratory for Experimental Vascular Medicine, Leiden University Medical Center, Leiden, Netherlands.

3Microbiome Medicine Center, Department of Laboratory Medicine, Zhujiang Hospital, Southern Medical University, Guangzhou, China.

4Department of Pediatrics, University Medical Center Groningen, Groningen, Netherlands.

5Center for Proteomics and Metabolomics,

6Department of Medical Microbiology,

7Center for Microbiome Analyses and Therapeutics, and

8Department of Human Genetics, Leiden University Medical Center, Netherlands.

9Med-X Institute, Center for Immunological and Metabolic Diseases, and Department of Endocrinology, the First Affiliated Hospital of Xi’an JiaoTong University, Xi’an Jiaotong University, Xi’an, China.

Address correspondence to: Yanan Wang, Department of Medicine, Division of Endocrinology, Leiden University Medical Center, Albinusdreef 2, 2333 ZA, Leiden, Netherlands. Phone: 317168163; Email: Y.Wang@lumc.nl.

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1Department of Medicine, Division of Endocrinology, and

2Einthoven Laboratory for Experimental Vascular Medicine, Leiden University Medical Center, Leiden, Netherlands.

3Microbiome Medicine Center, Department of Laboratory Medicine, Zhujiang Hospital, Southern Medical University, Guangzhou, China.

4Department of Pediatrics, University Medical Center Groningen, Groningen, Netherlands.

5Center for Proteomics and Metabolomics,

6Department of Medical Microbiology,

7Center for Microbiome Analyses and Therapeutics, and

8Department of Human Genetics, Leiden University Medical Center, Netherlands.

9Med-X Institute, Center for Immunological and Metabolic Diseases, and Department of Endocrinology, the First Affiliated Hospital of Xi’an JiaoTong University, Xi’an Jiaotong University, Xi’an, China.

Address correspondence to: Yanan Wang, Department of Medicine, Division of Endocrinology, Leiden University Medical Center, Albinusdreef 2, 2333 ZA, Leiden, Netherlands. Phone: 317168163; Email: Y.Wang@lumc.nl.

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1Department of Medicine, Division of Endocrinology, and

2Einthoven Laboratory for Experimental Vascular Medicine, Leiden University Medical Center, Leiden, Netherlands.

3Microbiome Medicine Center, Department of Laboratory Medicine, Zhujiang Hospital, Southern Medical University, Guangzhou, China.

4Department of Pediatrics, University Medical Center Groningen, Groningen, Netherlands.

5Center for Proteomics and Metabolomics,

6Department of Medical Microbiology,

7Center for Microbiome Analyses and Therapeutics, and

8Department of Human Genetics, Leiden University Medical Center, Netherlands.

9Med-X Institute, Center for Immunological and Metabolic Diseases, and Department of Endocrinology, the First Affiliated Hospital of Xi’an JiaoTong University, Xi’an Jiaotong University, Xi’an, China.

Address correspondence to: Yanan Wang, Department of Medicine, Division of Endocrinology, Leiden University Medical Center, Albinusdreef 2, 2333 ZA, Leiden, Netherlands. Phone: 317168163; Email: Y.Wang@lumc.nl.

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1Department of Medicine, Division of Endocrinology, and

2Einthoven Laboratory for Experimental Vascular Medicine, Leiden University Medical Center, Leiden, Netherlands.

3Microbiome Medicine Center, Department of Laboratory Medicine, Zhujiang Hospital, Southern Medical University, Guangzhou, China.

4Department of Pediatrics, University Medical Center Groningen, Groningen, Netherlands.

5Center for Proteomics and Metabolomics,

6Department of Medical Microbiology,

7Center for Microbiome Analyses and Therapeutics, and

8Department of Human Genetics, Leiden University Medical Center, Netherlands.

9Med-X Institute, Center for Immunological and Metabolic Diseases, and Department of Endocrinology, the First Affiliated Hospital of Xi’an JiaoTong University, Xi’an Jiaotong University, Xi’an, China.

Address correspondence to: Yanan Wang, Department of Medicine, Division of Endocrinology, Leiden University Medical Center, Albinusdreef 2, 2333 ZA, Leiden, Netherlands. Phone: 317168163; Email: Y.Wang@lumc.nl.

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1Department of Medicine, Division of Endocrinology, and

2Einthoven Laboratory for Experimental Vascular Medicine, Leiden University Medical Center, Leiden, Netherlands.

3Microbiome Medicine Center, Department of Laboratory Medicine, Zhujiang Hospital, Southern Medical University, Guangzhou, China.

4Department of Pediatrics, University Medical Center Groningen, Groningen, Netherlands.

5Center for Proteomics and Metabolomics,

6Department of Medical Microbiology,

7Center for Microbiome Analyses and Therapeutics, and

8Department of Human Genetics, Leiden University Medical Center, Netherlands.

9Med-X Institute, Center for Immunological and Metabolic Diseases, and Department of Endocrinology, the First Affiliated Hospital of Xi’an JiaoTong University, Xi’an Jiaotong University, Xi’an, China.

Address correspondence to: Yanan Wang, Department of Medicine, Division of Endocrinology, Leiden University Medical Center, Albinusdreef 2, 2333 ZA, Leiden, Netherlands. Phone: 317168163; Email: Y.Wang@lumc.nl.

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1Department of Medicine, Division of Endocrinology, and

2Einthoven Laboratory for Experimental Vascular Medicine, Leiden University Medical Center, Leiden, Netherlands.

3Microbiome Medicine Center, Department of Laboratory Medicine, Zhujiang Hospital, Southern Medical University, Guangzhou, China.

4Department of Pediatrics, University Medical Center Groningen, Groningen, Netherlands.

5Center for Proteomics and Metabolomics,

6Department of Medical Microbiology,

7Center for Microbiome Analyses and Therapeutics, and

8Department of Human Genetics, Leiden University Medical Center, Netherlands.

9Med-X Institute, Center for Immunological and Metabolic Diseases, and Department of Endocrinology, the First Affiliated Hospital of Xi’an JiaoTong University, Xi’an Jiaotong University, Xi’an, China.

Address correspondence to: Yanan Wang, Department of Medicine, Division of Endocrinology, Leiden University Medical Center, Albinusdreef 2, 2333 ZA, Leiden, Netherlands. Phone: 317168163; Email: Y.Wang@lumc.nl.

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1Department of Medicine, Division of Endocrinology, and

2Einthoven Laboratory for Experimental Vascular Medicine, Leiden University Medical Center, Leiden, Netherlands.

3Microbiome Medicine Center, Department of Laboratory Medicine, Zhujiang Hospital, Southern Medical University, Guangzhou, China.

4Department of Pediatrics, University Medical Center Groningen, Groningen, Netherlands.

5Center for Proteomics and Metabolomics,

6Department of Medical Microbiology,

7Center for Microbiome Analyses and Therapeutics, and

8Department of Human Genetics, Leiden University Medical Center, Netherlands.

9Med-X Institute, Center for Immunological and Metabolic Diseases, and Department of Endocrinology, the First Affiliated Hospital of Xi’an JiaoTong University, Xi’an Jiaotong University, Xi’an, China.

Address correspondence to: Yanan Wang, Department of Medicine, Division of Endocrinology, Leiden University Medical Center, Albinusdreef 2, 2333 ZA, Leiden, Netherlands. Phone: 317168163; Email: Y.Wang@lumc.nl.

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1Department of Medicine, Division of Endocrinology, and

2Einthoven Laboratory for Experimental Vascular Medicine, Leiden University Medical Center, Leiden, Netherlands.

3Microbiome Medicine Center, Department of Laboratory Medicine, Zhujiang Hospital, Southern Medical University, Guangzhou, China.

4Department of Pediatrics, University Medical Center Groningen, Groningen, Netherlands.

5Center for Proteomics and Metabolomics,

6Department of Medical Microbiology,

7Center for Microbiome Analyses and Therapeutics, and

8Department of Human Genetics, Leiden University Medical Center, Netherlands.

9Med-X Institute, Center for Immunological and Metabolic Diseases, and Department of Endocrinology, the First Affiliated Hospital of Xi’an JiaoTong University, Xi’an Jiaotong University, Xi’an, China.

Address correspondence to: Yanan Wang, Department of Medicine, Division of Endocrinology, Leiden University Medical Center, Albinusdreef 2, 2333 ZA, Leiden, Netherlands. Phone: 317168163; Email: Y.Wang@lumc.nl.

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1Department of Medicine, Division of Endocrinology, and

2Einthoven Laboratory for Experimental Vascular Medicine, Leiden University Medical Center, Leiden, Netherlands.

3Microbiome Medicine Center, Department of Laboratory Medicine, Zhujiang Hospital, Southern Medical University, Guangzhou, China.

4Department of Pediatrics, University Medical Center Groningen, Groningen, Netherlands.

5Center for Proteomics and Metabolomics,

6Department of Medical Microbiology,

7Center for Microbiome Analyses and Therapeutics, and

8Department of Human Genetics, Leiden University Medical Center, Netherlands.

9Med-X Institute, Center for Immunological and Metabolic Diseases, and Department of Endocrinology, the First Affiliated Hospital of Xi’an JiaoTong University, Xi’an Jiaotong University, Xi’an, China.

Address correspondence to: Yanan Wang, Department of Medicine, Division of Endocrinology, Leiden University Medical Center, Albinusdreef 2, 2333 ZA, Leiden, Netherlands. Phone: 317168163; Email: Y.Wang@lumc.nl.

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1Department of Medicine, Division of Endocrinology, and

2Einthoven Laboratory for Experimental Vascular Medicine, Leiden University Medical Center, Leiden, Netherlands.

3Microbiome Medicine Center, Department of Laboratory Medicine, Zhujiang Hospital, Southern Medical University, Guangzhou, China.

4Department of Pediatrics, University Medical Center Groningen, Groningen, Netherlands.

5Center for Proteomics and Metabolomics,

6Department of Medical Microbiology,

7Center for Microbiome Analyses and Therapeutics, and

8Department of Human Genetics, Leiden University Medical Center, Netherlands.

9Med-X Institute, Center for Immunological and Metabolic Diseases, and Department of Endocrinology, the First Affiliated Hospital of Xi’an JiaoTong University, Xi’an Jiaotong University, Xi’an, China.

Address correspondence to: Yanan Wang, Department of Medicine, Division of Endocrinology, Leiden University Medical Center, Albinusdreef 2, 2333 ZA, Leiden, Netherlands. Phone: 317168163; Email: Y.Wang@lumc.nl.

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1Department of Medicine, Division of Endocrinology, and

2Einthoven Laboratory for Experimental Vascular Medicine, Leiden University Medical Center, Leiden, Netherlands.

3Microbiome Medicine Center, Department of Laboratory Medicine, Zhujiang Hospital, Southern Medical University, Guangzhou, China.

4Department of Pediatrics, University Medical Center Groningen, Groningen, Netherlands.

5Center for Proteomics and Metabolomics,

6Department of Medical Microbiology,

7Center for Microbiome Analyses and Therapeutics, and

8Department of Human Genetics, Leiden University Medical Center, Netherlands.

9Med-X Institute, Center for Immunological and Metabolic Diseases, and Department of Endocrinology, the First Affiliated Hospital of Xi’an JiaoTong University, Xi’an Jiaotong University, Xi’an, China.

Address correspondence to: Yanan Wang, Department of Medicine, Division of Endocrinology, Leiden University Medical Center, Albinusdreef 2, 2333 ZA, Leiden, Netherlands. Phone: 317168163; Email: Y.Wang@lumc.nl.

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1Department of Medicine, Division of Endocrinology, and

2Einthoven Laboratory for Experimental Vascular Medicine, Leiden University Medical Center, Leiden, Netherlands.

3Microbiome Medicine Center, Department of Laboratory Medicine, Zhujiang Hospital, Southern Medical University, Guangzhou, China.

4Department of Pediatrics, University Medical Center Groningen, Groningen, Netherlands.

5Center for Proteomics and Metabolomics,

6Department of Medical Microbiology,

7Center for Microbiome Analyses and Therapeutics, and

8Department of Human Genetics, Leiden University Medical Center, Netherlands.

9Med-X Institute, Center for Immunological and Metabolic Diseases, and Department of Endocrinology, the First Affiliated Hospital of Xi’an JiaoTong University, Xi’an Jiaotong University, Xi’an, China.

Address correspondence to: Yanan Wang, Department of Medicine, Division of Endocrinology, Leiden University Medical Center, Albinusdreef 2, 2333 ZA, Leiden, Netherlands. Phone: 317168163; Email: Y.Wang@lumc.nl.

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Published February 22, 2023 - More info

Published in Volume 8, Issue 4 on February 22, 2023
JCI Insight. 2023;8(4):e166655. https://doi.org/10.1172/jci.insight.166655.
© 2023 Li et al. This work is licensed under the Creative Commons Attribution 4.0 International License. To view a copy of this license, visit http://creativecommons.org/licenses/by/4.0/. Published February 22, 2023 - Version history
Received: October 26, 2022; Accepted: January 13, 2023 View PDF Abstract

Short-chain fatty acids, including butyrate, have multiple metabolic benefits in individuals who are lean but not in individuals with metabolic syndrome, with the underlying mechanisms still being unclear. We aimed to investigate the role of gut microbiota in the induction of metabolic benefits of dietary butyrate. We performed antibiotic-induced microbiota depletion of the gut and fecal microbiota transplantation (FMT) in APOE*3-Leiden.CETP mice, a well-established translational model for developing human-like metabolic syndrome, and revealed that dietary butyrate reduced appetite and ameliorated high-fat diet–induced (HFD-induced) weight gain dependent on the presence of gut microbiota. FMT from butyrate-treated lean donor mice, but not butyrate-treated obese donor mice, into gut microbiota–depleted recipient mice reduced food intake, attenuated HFD-induced weight gain, and improved insulin resistance. 16S rRNA and metagenomic sequencing on cecal bacterial DNA of recipient mice implied that these effects were accompanied by the selective proliferation of Lachnospiraceae bacterium 28-4 in the gut as induced by butyrate. Collectively, our findings reveal a crucial role of gut microbiota in the beneficial metabolic effects of dietary butyrate as strongly associated with the abundance of Lachnospiraceae bacterium 28-4.

Introduction

Obesity is becoming a global health concern. Although lifestyle interventions, including calorie restriction (1) and pharmacotherapy, have been shown to be effective in inducing weight loss (2), cessation of intervention generally leads to weight regain. Therefore, intervention strategies aimed at attaining sustained weight loss are still required.

Dietary fiber intake is associated with lower BW, lower incidence of cardiometabolic diseases, and lower mortality from type 2 diabetes (T2D) and coronary heart disease (3). One of the main mechanisms attributed to the cardiometabolic benefits of dietary fiber is the production of short-chain fatty acids (SCFAs), including acetate, propionate, and butyrate, by microbial fermentation (4). Particularly, butyrate was shown to prevent diet-induced obesity (DIO) (5), improve glucose homeostasis, and alleviate insulin resistance in mice (6). Moreover, we previously demonstrated that dietary butyrate prevents high-fat diet–induced (HFD-induced) weight gain in mice mainly via reducing food intake, in addition to modestly increasing energy expenditure by activating brown adipose tissue (BAT) (7). A recent clinical study, however, showed that oral butyrate improves glucose metabolism only in lean individuals but not in subjects with metabolic syndrome (8). Further investigation of the precise molecular targets of butyrate in various metabolic contexts is, therefore, necessary to provide insight into the differential response of individuals with and without obesity upon butyrate intervention, which may lead to the identification of personalized therapeutic strategies to combat obesity and associated cardiometabolic diseases in the clinic.

A groundbreaking human intervention study showed that adding fiber to an isoenergetic diet beneficially alters the gut microbiota by promoting SCFA-producing bacterial strains, which alleviates T2D as evident from the improvement of HbA1c (9). In addition, a recent human study demonstrated that the overall gut microbiota shifts in parallel with glycemic status, suggesting that the variation of gut microbiota is strongly associated with insulin resistance of the host (10). Collectively, these findings indicate that gut microbiota plays a crucial role in maintaining cardiometabolic health.

Here, we addressed the role of gut microbiota in the induction of metabolic benefits of dietary butyrate by using antibiotic-induced microbiota depletion (AIMD) of the gut and fecal microbiota transplantation (FMT) in APOE*3-Leiden.CETP (E3L.CETP) mice, a well-established translational model for developing human-like diet-induced cardiometabolic diseases (11). We reveal that the beneficial metabolic effects of dietary butyrate are crucially dependent on gut microbiota. In particular, FMT from butyrate-treated lean donor mice, but not from butyrate-treated obese donor mice, induces enrichment of Lachnospiraceae bacterium 28-4 in recipient mice, positively correlating with their cardiometabolic health.

Results

Dietary butyrate reduces food intake and attenuates HFD-induced weight gain dependent on gut microbiota. Our previous work suggested that butyrate alters gut microbiota composition, which may contribute to the beneficial effects of butyrate on metabolic health (7). Therefore, we first explored the role of gut microbiota in the metabolic benefits of dietary butyrate. To this end, male E3L.CETP mice that underwent AIMD or received saline (vehicle) were simultaneously fed an HFD without or with sodium butyrate for 6 weeks (Figure 1A). Compared with vehicle, AIMD largely reduced 16S rRNA expression (–95%, P < 0.01) in fresh fecal samples collected after the intervention (Figure 1B), verifying depletion of the bacterial gut microbiota. In the vehicle group, butyrate administration attenuated HFD-induced fat mass gain (–49%, P < 0.05) without affecting lean mass (Figure 1C), as explained by reduced daily (–15%, P < 0.05) and cumulative (–13%, P < 0.05) food intake (Figure 1, D and E). AIMD abolished the effects of butyrate on fat mass gain (Figure 1F) and food intake (Figure 1, G and H), indicating that the induction of the metabolic benefits by dietary butyrate is strictly dependent on the presence of gut microbiota.

Dietary butyrate reduces food intake and attenuates HFD-induced weight gainFigure 1

Dietary butyrate reduces food intake and attenuates HFD-induced weight gain dependent on gut microbiota. (A) Mice underwent AIMD or received saline (vehicle) for 6 weeks while being fed an HFD without or with 5% (weight by weight [w/w], i.e., on average 0.12 g per day per mouse) sodium butyrate. (B) At the end of the treatment, fresh feces were collected and bacterial DNA was quantified by 16S rRNA gene amplification by PCR (n = 8–9). (C and F) Body composition was measured by MRI (n = 8). (D and G) The average food intake per day throughout the whole intervention period was calculated (n = 5). (E and H) The cumulative food intake was calculated (n = 5). Data are shown as means ± SEM; statistical significance between 2 groups was determined with 2-tailed Student’s unpaired t test. For data represented in the line graphs showing the changes over time for a continuous variable, statistical significance between 2 groups at each time point was determined using 2-tailed Student’s unpaired t test. *P < 0.05, **P < 0.01; AIMD vs. vehicle in B or Butyrate vs. Control in CH.

Additionally, we examined the role of gut microbiota in the effects of dietary butyrate on energy expenditure, lipid metabolism, and the activity of BAT, a key regulator in energy hemostasis (12). In the vehicle group, dietary butyrate decreased the respiratory exchange ratio during the night period (–5%, P < 0.05; Supplemental Figure 1A; supplemental material available online with this article; https://doi.org/10.1172/jci.insight.166655DS1), as a result of the increased fat oxidation (+14%, P < 0.05; Supplemental Figure 1B) at the expense of carbohydrate oxidation (–28%, P < 0.01; Supplemental Figure 1C). In addition, dietary butyrate also accelerated the clearance of glycerol tri[3H]oleate-labeled ([3H]TO-labeled) triglyceride-rich lipoprotein-like (TRL-like) particles from the circulation (P < 0.05; Supplemental Figure 1D) and increased the uptake of [3H]TO-derived radioactivity by BAT (+110%, P < 0.05; Supplemental Figure 1E). Consistent with our previous findings (7), butyrate activated BAT and enhanced BAT thermogenic capacity, as evidenced by reducing intracellular lipid content (Supplemental Figure 1, F and I) as well as increasing protein expression of both uncoupling protein-1 (UCP-1) (Supplemental Figure 1, G and I) and tyrosine hydroxylase (TH), a marker of sympathetic nerve activity (Supplemental Figure 1, H and I). In contrast, in the AIMD group, butyrate neither affected the respiratory exchange ratio (Supplemental Figure 1J), fat oxidation, and carbohydrate oxidation rate (Supplemental Figure 1, K and L) nor influenced the [3H]TO clearance from the circulation and the tissue uptake of [3H]TO-derived radioactivity (Supplemental Figure 1, M and N). AIMD also abolished the effects of butyrate on BAT activation (Supplemental Figure 1, O–R).

FMT transplantation from butyrate-treated lean donor mice attenuates HFD-induced weight gain and improves insulin resistance in recipient mice. To further elucidate the causal impact of gut microbiota on the metabolic benefits of dietary butyrate, donor mice were fed an HFD without (Control) or with butyrate (Butyrate) for 12 weeks (prevention strategy), and from week 6 onward, fresh fecal bacteria were isolated from donors and transplanted to gut microbiota–depleted mice upon HFD feeding for 6 weeks (Figure 2A). Compared with mice receiving FMT from control donors, FMT from butyrate-treated donors caused a persistent decrease in BW gain (–54% at 6 weeks, P < 0.01; Figure 2B), accompanied by a decrease in fat mass (–26%, P < 0.05; Figure 2C), as well as a reduction in daily food intake (–15%, P < 0.05; Figure 2D). In addition, FMT from butyrate-treated donors tended to decrease fasting plasma levels of glucose (–11%, P = 0.07; Figure 2E) and insulin (–25%, P = 0.07; Figure 2F) and markedly reduced homeostatic model assessment of insulin resistance (HOMA-IR) (–32%, P < 0.05; Figure 2G) in recipient mice.

FMT from butyrate-treated lean donor mice attenuates HFD-induced weight gaiFigure 2

FMT from butyrate-treated lean donor mice attenuates HFD-induced weight gain and improves insulin resistance in recipient mice. (A) Mice were fed an HFD without or with 5% (w/w) sodium butyrate prevention for 6 weeks. After this, fresh feces were collected weekly and used for FMT to gut microbiota-depleted recipient mice that were fed an HFD for 6 weeks. (B) BW was measured weekly and the BW change was calculated (n = 8). (C) At the end of the experiment, body composition was measured by MRI (n = 8). (D) The average food intake per day throughout the intervention period was calculated (n = 8). (E) Fasting glucose (n = 7–8) and (F) insulin (n = 8) plasma levels were measured. (G) They were then used for calculation of HOMA-IR (n = 7–8). Data are shown as means ± SEM; statistical significance between 2 groups was determined with 2-tailed Student’s unpaired t test; For data represented in the line graphs showing the changes over time for a continuous variable, statistical significance between 2 groups at each time point was determined using 2-tailed Student’s unpaired t test. *P < 0.05, **P < 0.01; Butyrate vs. Control.

Compared with FMT from control donors, FMT from butyrate-treated donors decreased the respiratory exchange ratio in recipient mice (–3%, P < 0.05; Supplemental Figure 2A) accompanied by an unchanged fat oxidation rate (Supplemental Figure 2B) and decreased carbohydrate oxidation rate (–28%, P < 0.05; Supplemental Figure 2C). The clearance of [3H]TO from the circulation (Supplemental Figure 2D) and uptake of [3H]TO-derived radioactivity by various organs including BAT were not altered (Supplemental Figure 2E). In line with this, BAT activity was comparable between both groups, as no differences were observed for intracellular lipid content (Supplemental Figure 2, F and I), UCP-1 expression (Supplemental Figure 2, G and I), and TH expression (Supplemental Figure 2, H and I) within BAT. Collectively, these data reveal that butyrate indirectly, i.e., through modulation of the gut microbiota, causes satiety and attenuates HFD-induced weight gain and insulin resistance.

FMT from butyrate-treated lean donor mice selectively enriches Lachnospiraceae bacterium 28-4 in recipient mice. To unravel the specific effects of butyrate on the composition of the gut microbiota in relation to its metabolic effects on the host, 16S rRNA-Seq as well as metagenomic sequencing were performed on cecal bacterial genomic DNA of all mice receiving FMT from control or butyrate-treated donors.

16S rRNA-Seq analysis revealed changes in the gut microbial ecology in recipient mice. The observed richness of the operational taxonomic unit (OTU) (Figure 3A) and Shannon index (α-diversity, Figure 3B) were not different between recipient groups. In favorable contrast, FMT from butyrate-treated donors induced an apparent difference in composition of gut microbiota in recipient mice, as presented by an increase in abundance of Firmicutes (+28%, P < 0.05) at the expense of predominantly Bacteroidetes (–10%; Figure 3C and Supplemental Table 1), and induced different clustering in principal coordinates analysis (PCoA) plot of unweighted unique fraction metric (UniFrac) distances on the OTU level (β-diversity, Figure 3D).

FMT from butyrate-treated lean donor mice selectively enriches LachnospiracFigure 3

FMT from butyrate-treated lean donor mice selectively enriches Lachnospiraceae bacterium 28-4 in recipient mice. At the end of the FMT study, the recipients’ bacterial genomic DNA was isolated from the cecum content and sequenced. By 16S rRNA gene analysis, (A) the number of observed species (n = 9–10), (B) Shannon diversity (n = 9–10), and (C) the community abundance of gut microbiota on phylum level (n = 9–10) were assessed. (D) The PCoA plot of unweighted UniFrac distances on OTU levels was then calculated (n = 9–10). (E) By analyzing the metagenomic sequencing data, the abundance of gut microbiota (top 30 based on relative abundance) on species level was compared using Wilcoxon’s rank-sum test and its correlation with metabolic outcomes (BW, food intake, glucose, and insulin) were presented in Spearman’s correlation heatmap (n = 5). (F) Correlation of abundance of Lachnospiraceae bacterium 28-4 with BW was analyzed (n = 5). (G) Correlations of abundance of Lachnospiraceae bacterium 28-4 with metabolic outcomes (BW, food intake, glucose, and insulin) were analyzed using RDA (n = 5). Data are shown as box plot with whiskers at min/max in A and B; statistical significance between 2 groups was determined with 2-tailed Student’s unpaired t test in A and B and Wilcoxon’s rank-sum test in E. *P < 0.05, **P < 0.01; Butyrate vs. Control.

Next, metagenomic analysis revealed more distinctive variations in the gut microbiota at the species level. We identified 6,840 species in total, spanning 1,851 genera and 104 phyla (Supplemental Table 2). Among those, 859 species were significantly regulated by FMT from butyrate-treated donors compared with control donors (Supplemental Table 3). In particular, among the top 30 species based on relative abundance, FMT from butyrate-treated donors markedly increased the relative abundance of Lachnospiraceae bacterium 28-4 (+2.9-fold, P < 0.01), while it decreased the relative abundance of Bacteroides sp. CAG:709, Bacteroides sp. CAG:770, Bacteroides sp. CAG:545, Alistipes sp. CAG:435, Flavonifractor plautii, Alistipes sp. CAG:514, and Pseudoflavonifractor capillosus (Figure 3E) in recipient mice. Enrichment with Lachnospiraceae bacterium 28-4

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