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Declining FXR expression coordinates neonatal beta cell mass development with microbial bile acid metabolism maturation in mice

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Abstract

Aims/hypothesis

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Diet switch during weaning induces gut microbiome maturation, accompanied by the formation of adequate functional beta cell mass. Bile acid (BA), an essential microbial metabolite, regulates host glucose homeostasis by binding to its main receptor, farnesoid X receptor (FXR, encoded by NR1H4). However, the precise roles of microbial BA metabolism and FXR signalling in neonatal beta cell development are still unclear.

Methods

Islet FXR levels were determined at different perinatal stages. Postnatal changes in gut microbiome and BA profiles were examined in mice, with changes in germ-free mouse BAs serving as the control. We genetically modified beta cells to sustain FXR expression after birth (using Nr1h4-knockin [βFxrKI] mice) and performed morphological and functional analysis on murine islets. Single-cell RNA-seq and single-cell assay for transposase-accessible chromatin sequencing of islet cells were used to study FXR-mediated downstream regulation in islets. Lineage tracing was performed to evaluate beta cell fate transition. Mendelian randomisation (MR) and human islet proteomics data analysis were applied to study the pathological relevance in human diabetes.

Results

FXR expression in beta cells declined after birth (positive cell proportion, 29.1 ± 3.1% at embryonic day 18.5 vs 4.2 ± 2.4% at 3 weeks postnatal in mice, p<0.001). This physiological change paralleled the ascending of FXR-agonistic BAs derived from gut microbiome maturation (unconjugated BA proportion, 0.9 ± 0.6% at 1 week vs 14.0 ± 5.6% at 3 weeks, p<0.05). βFxrKI mice had limited beta cell mass growth (approximately 70% of the control level at 1 week of age and only 15% of the control level at 8 weeks of age) and developed high blood glucose levels by weaning (random blood glucose, 15.2 ± 1.7 mmol/l in βFxrKI vs 7.7 ± 0.5 mmol/l in control, p<0.001), mainly resulting from elevated cell apoptosis (1.95-, 1.79-, and 3.27-fold increase vs control at 1, 2 and 3 weeks, respectively) and altered beta cell identity. Casp6 was identified as a key downstream target in beta cell FXR. Intervention with antibiotics or a specific caspase-6 (CASP6) inhibitor partially recovered the phenotypes of βFxrKI mice. Further validation in humans showed that islet FXR/CASP6 levels were elevated in individuals with type 2 diabetes (FXR, −0.039 ± 1.257 a.u. in donors without diabetes vs 0.646 ± 1.140 a.u. in donors with diabetes, p=0.0371; CASP6, −1.575 ± 0.307 a.u. in donors without diabetes vs −1.325 ± 0.381 a.u. in donors with diabetes, p=0.011). MR analysis further supported the effect of human islet FXR expression in elevating HbA1c (β=0.006, p<0.001) with lowering fasting insulin level (β=−0.009, p=0.02) and the effect of CASP6 expression in enhancing 2 h glucose (β=0.039, p=0.01).

Conclusions/interpretation

The declining FXR–CASP6 signals in neonatal beta cells could serve as a programmed host response to the maturing gut microbial BA metabolism to maintain normal postnatal beta cell mass development and ensure glycaemic homeostasis in adults.

Data availability

Raw data of scRNA-seq and scATAC-seq are deposited in the Gene Expression Omnibus (GEO) database under the accession number GSE241408. The code used in this Mendelian randomisation study is publicly available at https://github.com/Angela-linyt/Gene_Glu_MR.git.

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Abbreviations

Abx:

Antibiotics

AMPK:

AMP-activated protein kinase

ATAC:

Assay for transposase-accessible chromatin

BA:

Bile acid

CASP6:

Caspase-6

ChIP:

Chromatin immunoprecipitation

Conv:

Conventional environment

DAR:

Differentially accessible region

DEG:

Differentially expressed gene

E18.5:

Embryonic day 18.5

FA:

Folic acid

FOLR1:

Folate receptor 1

FXR:

Farnesoid X receptor

FXRE:

FXR response element

GF:

Germ-free

GO:

Gene Ontology

GSEA:

Gene-set enrichment analysis

GSIS:

Glucose-stimulated insulin secretion

MafA:

Musculoaponeurotic fibrosarcoma oncogene family protein A

MR:

Mendelian randomisation

OCR:

Oxygen consumption rate

OXPHOS:

Oxidative phosphorylation

P1w:

Postnatal 1 week of age

P2w, P3w, P8w:

Postnatal 2, 3, 8 weeks of age

PBA:

Primary bile acid

PDX1:

Pancreatic and duodenal homeobox 1

PP (cells):

Pancreatic polypeptide (cells)

PPY:

Pancreatic polypeptide

qPCR:

Quantitative real-time PCR

SBA:

Secondary bile acid

scATAC-seq:

Single-cell assay for transposase-accessible chromatin sequencing

scRNA-seq:

Single-cell RNA-seq

TUNEL:

TdT-mediated dUTP nick-end labelling

UMAP:

Uniform Manifold Approximation and Projection

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Author notes

  1. Chenyang Fu, Tingting Li, Yiming Hao and Yiting Lin contributed equally.

Authors and Affiliations

  1. Department of Endocrine and Metabolic Diseases, Shanghai Institute of Endocrine and Metabolic Diseases, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China

    Chenyang Fu, Tingting Li, Yiming Hao, Yiting Lin, Yixuan Qiu, Yangyang Jia, Jie Yang, Bei Liu, Duanyi Hua, Qicheng Ni, Jie Zheng, Guang Ning & Yanyun Gu

  2. Shanghai National Clinical Research Center for Metabolic Diseases, Key Laboratory for Endocrine and Metabolic Diseases of the National Health Commission of the PR China, Shanghai National Center for Translational Medicine, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China

    Chenyang Fu, Tingting Li, Yiming Hao, Yiting Lin, Yixuan Qiu, Yangyang Jia, Jie Yang, Bei Liu, Duanyi Hua, Qicheng Ni, Jie Zheng, Guang Ning & Yanyun Gu

  3. Shanghai Jiayin Biotechnology, Shanghai, China

    Chengyang Wang

  4. Department of Biliary-Pancreatic Surgery, Renji Hospital, Shanghai Jiao Tong University School of Medicine, Shanghai, China

    Tao Chen

  5. ULB Center for Diabetes Research, Université Libre de Bruxelles, Brussels, Belgium

    Anthony Piron & Miriam Cnop

  6. Division of Endocrinology, ULB Erasmus Hospital, Brussels University Hospital, Université Libre de Bruxelles, 1070, Brussels, Belgium

    Miriam Cnop

  7. WEL Research Institute, Wavre, Belgium

    Miriam Cnop

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  1. Chenyang Fu
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Corresponding authors

Correspondence to Jie Zheng, Guang Ning or Yanyun Gu.

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Acknowledgements

We thank D. Accili from Colombia University for profound discussion and instructive suggestions, and also thank Q. Wang, X. Wang, J. Wang and R. Liu (Shanghai Institute of Endocrine and Metabolic Diseases, Ruijin Hospital, Shanghai Jiao Tong University School of Medicine) and X. Cheng (State Key Laboratory of Cell Biology, CAS Center for Excellence in Molecular Cell Science, Shanghai Institute of Biochemistry and Cell Biology, Chinese Academy of Sciences). We thank Y. Huang, Q. Hang, and all members of the Core Facility of Basic Medical Sciences of Shanghai Jiao Tong University for their technical support. We thank X. Gao (Shanghai Profleader Biotechnology Co., China) for assisting in ultra-performance LC-MS/MS measurements of BA profiles. We thank XYZGenomics Co. for their assistance with single-cell data analysis.

Data availability

Raw data of scRNA-seq and scATAC-seq are deposited in the Gene Expression Omnibus (GEO) database under the accession number GSE241408.

Code availability

The code used in this Mendelian randomisation study is publicly available at https://github.com/Angela-linyt/Gene_Glu_MR.git.

Funding

This study was funded by grants from the National Natural Science Foundation of China (92157112, 82100835, 32570728, 82570934 ) and grants from National Key Research and Development Program of China (82088102), National Key Research and Development Program of China (2022YFC2505203), Noncommunicable Chronic Diseases–National Science and Technology Major Project (2024ZD0531500, 2024ZD0531502), Innovative research team of high-level local universities in Shanghai, European Union Horizon Health project NEMESIS, Walloon Region strategic axis Fonds de la Recherche Scientifique (FRFS)–Walloon Excellence in Life Sciences and Biotechnology (WELBIO), National Fund for Scientific Research (FNRS) and the Fondation Philippe Wiener–Maurice Anspach (FWA).

Authors’ relationships and activities

The authors declare that there are no relationships or activities that might bias, or be perceived to bias, their work.

Contribution statement

YG, CF and GN designed and conceptualised the study. CF, TL, YH, YL, YQ, CW, YJ, DH, TC and BL conducted experiments. CF, TL, YH, YL, CW, YJ, MC, AP, JZ and YG performed quality control. JY, YQ and CF did mouse husbandry. YG, CF, TL, BL, CW, QN, MC, JZ and GN analysed and discussed the data. CF and YG wrote the first draft of the manuscript. All the authors contributed to data interpretation, critically reviewed and edited the manuscript, and approved the final version. GN, JZ and YG had full access to all the data in the study and accept responsibility for the integrity of the data and the accuracy of the data analyses.

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Fu, C., Li, T., Hao, Y. et al. Declining FXR expression coordinates neonatal beta cell mass development with microbial bile acid metabolism maturation in mice. Diabetologia (2025). https://doi.org/10.1007/s00125-025-06618-w

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