Rifaximin stimulates nitrogen detoxification by PXR-independent mechanisms in human small intestinal organoids

BACKGROUND AND AIMS: Recurrent hepatic encephalopathy (HE) is characterized by hyperammonaemia in combination with neuropsychiatric abnormalities and is treated with lactulose and rifaximin. Rifaximin is a pregnane X receptor (PXR) agonist with low systemic and high intestinal bioavailability. The mechanisms by which it alleviates HE are unclear. We used human small intestinal (hSI) organoids to study whether rifaximin, via PXR activation, affects the epithelial biotransformation machinery, and to gain understanding of its low systemic availability. METHODS: We generated PXR knockdown hSI organoids via lentiviral delivery of short hairpin RNAs. Organoids were cultured for 24 h with rifaximin or rifampicin. RNA-sequencing and metabolomics were performed to analyse gene expression and amino acid metabolism. Luminal rifaximin was quantified by photospectrometry. RESULTS: Treatment of wild-type hSI organoids with rifaximin resulted in >twofold differential expression of 131 genes compared to DMSO. These effects were largely PXR independent and related to amino acid metabolism. Rifaximin decreased expression of glutaminase-2 and increased expression of asparagine synthetase and solute carrier 7A11, thereby increasing intracellular glutamine and asparagine concentrations, indicating active ammonia detoxification. Rifaximin was apically excreted into the lumen in an ATP binding cassette B1 (ABCB1)-dependent manner. CONCLUSIONS: Rifaximin-after uptake into enterocytes-stimulates intracellular nitrogen detoxification by PXR-independent mechanisms. Active apical excretion of rifaximin by ABCB1 into the intestinal lumen explains its low systemic bioavailability. Our study implies that rifaximin, next to modulation of the microbiome, has direct effects on ammonia scavenging in the human small intestinal epithelium

Pyruvate metabolism controls chromatin remodeling during CD4+ T cell activation

Upon antigen-specific T cell receptor (TCR) engagement, human CD4 + T cells proliferate and differentiate, a process associated with rapid transcriptional changes and metabolic reprogramming. Here, we show that the generation of extramitochondrial pyruvate is an important step for acetyl-CoA production and subsequent H3K27ac-mediated remodeling of histone acetylation. Histone modification, transcriptomic, and carbon tracing analyses of pyruvate dehydrogenase (PDH)-deficient T cells show PDH-dependent acetyl-CoA generation as a rate-limiting step during T activation. Furthermore, T cell activation results in the nuclear translocation of PDH and its association with both the p300 acetyltransferase and histone H3K27ac. These data support the tight integration of metabolic and histone-modifying enzymes, allowing metabolic reprogramming to fuel CD4 + T cell activation. Targeting this pathway may provide a therapeutic approach to specifically regulate antigen-driven T cell activation

Dissecting the allosteric FXR modulation: A chemical biology approach using guggulsterone as a chemical tool

Guggulsterone is a promiscuous ligand for endocrine and metabolic lipid receptors traditionally used to treat a number of diseases including diabesity, hyperlipidemia, atherosclerosis, and osteoarthritis. Although relatively weak, its activity at the farnesoid X receptor (FXR) is particularly intriguing as guggulsterone acts as an antagonist with a peculiar ability of gene selective modulation. We report here a chemical biology study with the aim to further characterize the biological action of guggulsterone at the FXR and to obtain further insights into the functional role played by noncanonical FXR binding pockets S2 and S3. Our results suggest that the FXR accessory pockets might act as potential targets for small molecules able to modulate the metabolic activation of the receptor without affecting the anti-inflammatory activity thus revealing a new approach for disclosing selective FXR modulators that might bypass potential side-effects from chronic treatments

Dax1 modulates ERα-dependent hypothalamic estrogen sensing in female mice.

Acknowledgements: The Section of Endocrinology and Investigative Medicine is funded by grants from the MRC, NIHR and is supported by the NIHR Biomedical Research Centre Funding Scheme and the NIHR/Imperial Clinical Research Facility. This article presents independent research. W.S.D. is funded by an NIHR Research Professorship and an NIHR Senior Investigator Award. This work was supported by a Sir Henry Dale Fellowship jointly funded by The Wellcome Trust and The Royal Society to BMO (105545/Z/14/Z). Grants from the Tyrolean Science Fund to JMRP agreement F.18896. KR and GSHY are supported by the MRC Metabolic Diseases Unit (MC_UU_00014/1). BYHL is supported by a BBSRC Project Grant (BB/S017593/1). JAT is supported by an NIHR Clinical Lectureship (CL-2019-14-504). The Cambridge Brain Bank is supported by the NIHR Cambridge Biomedical Research Centre. SMM was funded by a Commonwealth Scholarship and WHC by the Ford Physiology Fund Endowment. AM and I-MA are supported by MRC intramural funding and ERC Advanced Grant (787470-IntraGutSex). KGM is supported by BBSRC (BB/W001497/1) and DUK (18/0005886, 20/0006295) project grants. IC was an Academy of Medical Sciences Springboard Fellow (SBF005\1050) during this study, the Scheme was supported by the British Heart Foundation, Diabetes UK, the Global Challenges Research Fund, the Government Department for Business, Energy and Industrial Strategy and the Wellcome Trust. The views expressed are those of the author(s) and not necessarily those of the Wellcome Trust, the NHS, the NIHR or the Department of Health.Funder: The Section of Endocrinology and Investigative Medicine is funded by grants from the MRC, NIHR and is supported by the NIHR Biomedical Research Centre Funding Scheme and the NIHR/Imperial Clinical Research Facility. W.S.D. is funded by an NIHR Research Professorship and an NIHR Senior Investigator Award. This work was supported by a Sir Henry Dale Fellowship jointly funded by The Wellcome Trust and The Royal Society to BMO (105545/Z/14/Z). Grants from the Tyrolean Science Fund to JMRP agreement F.18896. KR and GSHY are supported by the MRC Metabolic Diseases Unit (MC_UU_00014/1). BYHL is supported by a BBSRC Project Grant (BB/S017593/1). JAT is supported by an NIHR Clinical Lectureship (CL-2019-14-504). The Cambridge Brain Bank is supported by the NIHR Cambridge Biomedical Research Centre. SMM was funded by a Commonwealth Scholarship and WHC by the Ford Physiology Fund Endowment. AM and I-MA are supported by MRC intramural funding and ERC Advanced Grant (787470-IntraGutSex).Coupling the release of pituitary hormones to the developmental stage of the oocyte is essential for female fertility. It requires estrogen to restrain kisspeptin (KISS1)-neuron pulsatility in the arcuate hypothalamic nucleus, while also exerting a surge-like effect on KISS1-neuron activity in the AVPV hypothalamic nucleus. However, a mechanistic basis for this region-specific effect has remained elusive. Our genomic analysis in female mice demonstrate that some processes, such as restraint of KISS1-neuron activity in the arcuate nucleus, may be explained by region-specific estrogen receptor alpha (ERα) DNA binding at gene regulatory regions. Furthermore, we find that the Kiss1-locus is uniquely regulated in these hypothalamic nuclei, and that the nuclear receptor co-repressor NR0B1 (DAX1) restrains its transcription specifically in the arcuate nucleus. These studies provide mechanistic insight into how ERα may control the KISS1-neuron, and Kiss1 gene expression, to couple gonadotropin release to the developmental stage of the oocyte

Dax1 modulates ERα-dependent hypothalamic estrogen sensing in female mice.

Coupling the release of pituitary hormones to the developmental stage of the oocyte is essential for female fertility. It requires estrogen to restrain kisspeptin (KISS1)-neuron pulsatility in the arcuate hypothalamic nucleus, while also exerting a surge-like effect on KISS1-neuron activity in the AVPV hypothalamic nucleus. However, a mechanistic basis for this region-specific effect has remained elusive. Our genomic analysis in female mice demonstrate that some processes, such as restraint of KISS1-neuron activity in the arcuate nucleus, may be explained by region-specific estrogen receptor alpha (ERα) DNA binding at gene regulatory regions. Furthermore, we find that the Kiss1-locus is uniquely regulated in these hypothalamic nuclei, and that the nuclear receptor co-repressor NR0B1 (DAX1) restrains its transcription specifically in the arcuate nucleus. These studies provide mechanistic insight into how ERα may control the KISS1-neuron, and Kiss1 gene expression, to couple gonadotropin release to the developmental stage of the oocyte

Dax1 modulates ERα-dependent hypothalamic estrogen sensing in female mice

Abstract Coupling the release of pituitary hormones to the developmental stage of the oocyte is essential for female fertility. It requires estrogen to restrain kisspeptin (KISS1)-neuron pulsatility in the arcuate hypothalamic nucleus, while also exerting a surge-like effect on KISS1-neuron activity in the AVPV hypothalamic nucleus. However, a mechanistic basis for this region-specific effect has remained elusive. Our genomic analysis in female mice demonstrate that some processes, such as restraint of KISS1-neuron activity in the arcuate nucleus, may be explained by region-specific estrogen receptor alpha (ERα) DNA binding at gene regulatory regions. Furthermore, we find that the Kiss1-locus is uniquely regulated in these hypothalamic nuclei, and that the nuclear receptor co-repressor NR0B1 (DAX1) restrains its transcription specifically in the arcuate nucleus. These studies provide mechanistic insight into how ERα may control the KISS1-neuron, and Kiss1 gene expression, to couple gonadotropin release to the developmental stage of the oocyte

Dietary Protein Defends Lean Mass and Maintains the Metabolic Benefits of Glucagon Receptor Agonism in Mice

OBJECTIVE: - Glucagon has long been proposed as a component of multi-agonist obesity therapeutics due to its ability to induce energy expenditure and cause weight loss. However, chronic glucagon-receptor agonism has been associated with a reduction in circulating amino acids and loss of lean mass. Importantly, it is currently not known whether the metabolic benefits of glucagon can be maintained under contexts that allow the defence of lean mass.METHODS: We investigate the metabolic effects of the long-acting glucagon receptor agonist, G108, when administered to obese mice at low-doses, and with dietary protein supplementation.RESULTS: - Dietary protein supplementation can only fully defend lean mass at a low dose of G108 that is sub-anorectic and does not reduce fat mass. However, in this context, G108 is still highly effective at improving glucose tolerance and reducing liver fat in obese mice. Mechanistically, liver RNA-Seq analysis reveals that dietary protein supplementation defends anabolic processes in low-dose G108-treated mice, and its effects on treatment-relevant glucose and lipid pathways are preserved.CONCLUSION: - Glucagon-mediated energy expenditure and weight loss may be mechanistically coupled to hypoaminocidemia and lean mass loss. However, our data suggest that glucagon can treat MAFLD at doses which allow full defence of lean mass given sufficient dietary protein intake. Therefore, proportionate glucagon therapy may be safe and effective in targeting hepatocytes and improving in glycaemia and liver fat.</p

Steroidogenic control of liver metabolism through a nuclear receptor-network

Objective: Coupling metabolic and reproductive pathways is essential for the survival of species. However, the functions of steroidogenic enzymes expressed in metabolic tissues are largely unknown. Methods and results: Here, we show that in the liver, the classical steroidogenic enzyme Cyp17a1 forms an essential nexus for glucose and ketone metabolism during feed-fast cycles. Both gain- and loss-of-function approaches are used to show that hepatic Cyp17a1 is induced by fasting, catalyzes the production of at least one hormone-ligand (DHEA) for the nuclear receptor PPARα, and is ultimately required for maintaining euglycemia and ketogenesis during nutrient deprivation. The feedback-loop that terminates Cyp17a1-PPARα activity, and re-establishes anabolic liver metabolism during re-feeding is mapped to postprandial bile acid-signaling, involving the receptors FXR, SHP and LRH-1. Conclusions: Together, these findings represent a novel paradigm of homeostatic control in which nutritional cues feed-forward on to metabolic pathways by influencing extragonadal steroidogenesis

Steroidogenic control of liver metabolism through a nuclear receptor-network

Objective: Coupling metabolic and reproductive pathways is essential for the survival of species. However, the functions of steroidogenic enzymes expressed in metabolic tissues are largely unknown. Methods and results: Here, we show that in the liver, the classical steroidogenic enzyme Cyp17a1 forms an essential nexus for glucose and ketone metabolism during feed-fast cycles. Both gain- and loss-of-function approaches are used to show that hepatic Cyp17a1 is induced by fasting, catalyzes the production of at least one hormone-ligand (DHEA) for the nuclear receptor PPARα, and is ultimately required for maintaining euglycemia and ketogenesis during nutrient deprivation. The feedback-loop that terminates Cyp17a1-PPARα activity, and re-establishes anabolic liver metabolism during re-feeding is mapped to postprandial bile acid-signaling, involving the receptors FXR, SHP and LRH-1. Conclusions: Together, these findings represent a novel paradigm of homeostatic control in which nutritional cues feed-forward on to metabolic pathways by influencing extragonadal steroidogenesis

Pyruvate metabolism controls chromatin remodeling during CD4+ T cell activation

Summary: Upon antigen-specific T cell receptor (TCR) engagement, human CD4+ T cells proliferate and differentiate, a process associated with rapid transcriptional changes and metabolic reprogramming. Here, we show that the generation of extramitochondrial pyruvate is an important step for acetyl-CoA production and subsequent H3K27ac-mediated remodeling of histone acetylation. Histone modification, transcriptomic, and carbon tracing analyses of pyruvate dehydrogenase (PDH)-deficient T cells show PDH-dependent acetyl-CoA generation as a rate-limiting step during T activation. Furthermore, T cell activation results in the nuclear translocation of PDH and its association with both the p300 acetyltransferase and histone H3K27ac. These data support the tight integration of metabolic and histone-modifying enzymes, allowing metabolic reprogramming to fuel CD4+ T cell activation. Targeting this pathway may provide a therapeutic approach to specifically regulate antigen-driven T cell activation