Potential therapeutic implications of histidine catabolism by the gut microbiota in NAFLD patients with morbid obesity.

The gut microbiota contributes to the pathophysiology of non-alcoholic fatty liver disease (NAFLD). Histidine is a key energy source for the microbiota, scavenging it from the host. Its role in NAFLD is poorly known. Plasma metabolomics, liver transcriptomics, and fecal metagenomics were performed in three human cohorts coupled with hepatocyte, rodent, and Drosophila models. Machine learning analyses identified plasma histidine as being strongly inversely associated with steatosis and linked to a hepatic transcriptomic signature involved in insulin signaling, inflammation, and trace amine-associated receptor 1. Circulating histidine was inversely associated with Proteobacteria and positively with bacteria lacking the histidine utilization (Hut) system. Histidine supplementation improved NAFLD in different animal models (diet-induced NAFLD in mouse and flies, ob/ob mouse, and ovariectomized rats) and reduced de novo lipogenesis. Fecal microbiota transplantation (FMT) from low-histidine donors and mono-colonization of germ-free flies with Enterobacter cloacae increased triglyceride accumulation and reduced histidine content. The interplay among microbiota, histidine catabolism, and NAFLD opens therapeutic opportunities.

p16INK4A-dependent senescence in the bone marrow niche drives age-related metabolic changes of hematopoietic progenitors.

Rapid and effective leukocyte response to infection is a fundamental function of the bone marrow (BM). However, with increasing age, this response becomes impaired, resulting in an increased burden of infectious diseases. Here, we investigate how aging changes the metabolism and function of hematopoietic progenitor cells (HPCs) and the impact of the BM niche on this phenotype. We found that, in response to lipopolysaccharide-induced stress, HPC mitochondrial function is impaired, and there is a failure to upregulate the TCA cycle in progenitor populations in aged animals compared with young animals. Furthermore, aged mesenchymal stromal cells (MSCs) of the BM niche, but not HPCs, exhibit a senescent phenotype, and selective depletion of senescent cells from the BM niche, as well as treatment with the senolytic drug ABT-263, improves mitochondrial function of HPCs when stressed with lipopolysaccharide. In summary, age-related HPC metabolic dysfunction occurs indirectly as a "bystander phenomenon" in the aging BM niche and can be restored by targeting senescent MSCs.

Microbiota Dysbiosis and Gut Barrier Dysfunction Associated with Non-Alcoholic Fatty Liver Disease Are Modulated by a Specific Metabolic Cofactors' Combination.

The gut is a selective barrier that not only allows the translocation of nutrients from food, but also microbe-derived metabolites to the systemic circulation that flows through the liver. Microbiota dysbiosis occurs when energy imbalances appear due to an unhealthy diet and a sedentary lifestyle. Dysbiosis has a critical impact on increasing intestinal permeability and epithelial barrier deterioration, contributing to bacterial and antigen translocation to the liver, triggering non-alcoholic fatty liver disease (NAFLD) progression. In this study, the potential therapeutic/beneficial effects of a combination of metabolic cofactors (a multi-ingredient; MI) (betaine, N-acetylcysteine, L-carnitine, and nicotinamide riboside) against NAFLD were evaluated. In addition, we investigated the effects of this metabolic cofactors' combination as a modulator of other players of the gut-liver axis during the disease, including gut barrier dysfunction and microbiota dysbiosis. Diet-induced NAFLD mice were distributed into two groups, treated with the vehicle (NAFLD group) or with a combination of metabolic cofactors (NAFLD-MI group), and small intestines were harvested from all animals for histological, molecular, and omics analysis. The MI treatment ameliorated gut morphological changes, decreased gut barrier permeability, and reduced gene expression of some proinflammatory cytokines. Moreover, epithelial cell proliferation and the number of goblet cells were increased after MI supplementation. In addition, supplementation with the MI combination promoted changes in the intestinal microbiota composition and diversity, as well as modulating short-chain fatty acids (SCFAs) concentrations in feces. Taken together, this specific combination of metabolic cofactors can reverse gut barrier disruption and microbiota dysbiosis contributing to the amelioration of NAFLD progression by modulating key players of the gut-liver axis.

Control of infection by LC3-associated phagocytosis, CASM, and detection of raised vacuolar pH by the V-ATPase-ATG16L1 axis.

The delivery of pathogens to lysosomes for degradation provides an important defense against infection. Degradation is enhanced when LC3 is conjugated to endosomes and phagosomes containing pathogens to facilitate fusion with lysosomes. In phagocytic cells, TLR signaling and Rubicon activate LC3-associated phagocytosis (LAP) where stabilization of the NADPH oxidase leads to sustained ROS production and raised vacuolar pH. Raised pH triggers the assembly of the vacuolar ATPase on the vacuole membrane where it binds ATG16L1 to recruit the core LC3 conjugation complex (ATG16L1:ATG5-12). This V-ATPase-ATG16L1 axis is also activated in nonphagocytic cells to conjugate LC3 to endosomes containing extracellular microbes. Pathogens provide additional signals for recruitment of LC3 when they raise vacuolar pH with pore-forming toxins and proteins, phospholipases, or specialized secretion systems. Many microbes secrete virulence factors to inhibit ROS production and/or the V-ATPase-ATG16L1 axis to slow LC3 recruitment and avoid degradation in lysosomes.

LC3-associated phagocytosis in bone marrow macrophages suppresses acute myeloid leukemia progression through STING activation.

The bone marrow (BM) microenvironment regulates acute myeloid leukemia (AML) initiation, proliferation, and chemotherapy resistance. Following cancer cell death, a growing body of evidence suggests an important role for remaining apoptotic debris in regulating the immunologic response to and growth of solid tumors. Here, we investigated the role of macrophage LC3-associated phagocytosis (LAP) within the BM microenvironment of AML. Depletion of BM macrophages (BMMs) increased AML growth in vivo. We show that LAP is the predominate method of BMM phagocytosis of dead and dying cells in the AML microenvironment. Targeted inhibition of LAP led to the accumulation of apoptotic cells (ACs) and apoptotic bodies (ABs), resulting in accelerated leukemia growth. Mechanistically, LAP of AML-derived ABs by BMMs resulted in stimulator of IFN genes (STING) pathway activation. We found that AML-derived mitochondrial damage-associated molecular patterns were processed by BMMs via LAP. Moreover, depletion of mitochondrial DNA (mtDNA) in AML-derived ABs showed that it was this mtDNA that was responsible for the induction of STING signaling in BMMs. Phenotypically, we found that STING activation suppressed AML growth through a mechanism related to increased phagocytosis. In summary, we report that macrophage LAP of apoptotic debris in the AML BM microenvironment suppressed tumor growth.

Metabolic Regulation of Macrophages by SIRT1 Determines Activation During Cholestatic Liver Disease in Mice.

BACKGROUND & AIMS: Inflammation is the hallmark of chronic liver disease. Metabolism is a key determinant to regulate the activation of immune cells. Here, we define the role of sirtuin 1 (SIRT1), a main metabolic regulator, in controlling the activation of macrophages during cholestatic liver disease and in response to endotoxin. METHODS: We have used mice overexpressing SIRT1, which we treated with intraperitoneal lipopolysaccharides or induced cholestasis by bile duct ligation. Bone marrow-derived macrophages were used for mechanistic in vitro studies. Finally, PEPC-Boy mice were used for adoptive transfer experiments to elucidate the impact of SIRT1-overexpressing macrophages in contributing to cholestatic liver disease. RESULTS: We found that SIRT1 overexpression promotes increased liver inflammation and liver injury after lipopolysaccharide/GalN and bile duct ligation; this was associated with an increased activation of the inflammasome in macrophages. Mechanistically, SIRT1 overexpression associated with the activation of the mammalian target of rapamycin (mTOR) pathway that led to increased activation of macrophages, which showed metabolic rewiring with increased glycolysis and broken tricarboxylic acid cycle in response to endotoxin in vitro. Activation of the SIRT1/mTOR axis in macrophages associated with the activation of the inflammasome and the attenuation of autophagy. Ultimately, in an in vivo model of cholestatic disease, the transplantation of SIRT1-overexpressing myeloid cells contributed to liver injury and fibrosis. CONCLUSIONS: Our study provides novel mechanistic insights into the regulation of macrophages during cholestatic disease and the response to endotoxin, in which the SIRT1/mTOR crosstalk regulates macrophage activation controlling the inflammasome, autophagy and metabolic rewiring.

Free fatty-acid transport via CD36 drives ß-oxidation-mediated hematopoietic stem cell response to infection.

Acute infection is known to induce rapid expansion of hematopoietic stem cells (HSCs), but the mechanisms supporting this expansion remain incomplete. Using mouse models, we show that inducible CD36 is required for free fatty acid uptake by HSCs during acute infection, allowing the metabolic transition from glycolysis towards ß-oxidation. Mechanistically, high CD36 levels promote FFA uptake, which enables CPT1A to transport fatty acyl chains from the cytosol into the mitochondria. Without CD36-mediated FFA uptake, the HSCs are unable to enter the cell cycle, subsequently enhancing mortality in response to bacterial infection. These findings enhance our understanding of HSC metabolism in the bone marrow microenvironment, which supports the expansion of HSCs during pathogenic challenge.

The Role of the Microbiome in Liver Cancer.

Hepatocellular carcinoma (HCC) is the most common malignancy occuring in the context of chronic liver disease and is one of the main causes of cancer-derived death worldwide. The lack of effective treatments, together with the poor prognosis, underlines the urge to develop novel and multidisciplinary therapeutics. An increasing body of evidence shows that HCC associates with changes in intestinal microbiota abundance and composition as well as with impaired barrier function, leading to the release of bacteria and their metabolites to the liver. These factors trigger a cascade of inflammatory responses contributing to liver cirrhosis and constituting an ideal environment for the progression of HCC. Interestingly, the use of bacteriotherapy in human and preclinical studies of chronic liver disease and HCC has been shown to successfully modify the microbiota composition, reducing overall inflammation and fibrosis. In this review, we explore the existing knowledge on the characterisation of the intestinal microbial composition in humans and experimental murine chronic liver disease and HCC, as well as the use of antibiotics and bacteriotherapy as therapeutic options.

Revisiting the Role of Natural Killer Cells in Non-Alcoholic Fatty Liver Disease.

Non-Alcoholic Fatty Liver Disease (NAFLD) is the most common form of chronic liver disease. The histological spectrum of NAFLD ranges from simple steatosis to chronic inflammation and liver fibrosis during Non-Alcoholic Steatohepatitis (NASH). The current view is that innate immune mechanisms represent a key element in supporting hepatic inflammation in NASH. Natural Killer (NK) cells are lymphoid cells and a component of the innate immune system known to be involved in NASH progression. Increasing evidence has shed light on the differential function of circulating and tissue-resident NK cells, as well as on the relevance of metabolism and the microenvironment in regulating their activity. Here, we revisit the complex role of NK cells as regulators of NASH progression as well as potential therapeutic approaches based on their modulation.

Much ado about nothing? Off-target amplification can lead to false-positive bacterial brain microbiome detection in healthy and Parkinson's disease individuals.

BACKGROUND: Recent studies suggested the existence of (poly-)microbial infections in human brains. These have been described either as putative pathogens linked to the neuro-inflammatory changes seen in Parkinson's disease (PD) and Alzheimer's disease (AD) or as a "brain microbiome" in the context of healthy patients' brain samples. METHODS: Using 16S rRNA gene sequencing, we tested the hypothesis that there is a bacterial brain microbiome. We evaluated brain samples from healthy human subjects and individuals suffering from PD (olfactory bulb and pre-frontal cortex), as well as murine brains. In line with state-of-the-art recommendations, we included several negative and positive controls in our analysis and estimated total bacterial biomass by 16S rRNA gene qPCR. RESULTS: Amplicon sequencing did detect bacterial signals in both human and murine samples, but estimated bacterial biomass was extremely low in all samples. Stringent reanalyses implied bacterial signals being explained by a combination of exogenous DNA contamination (54.8%) and false positive amplification of host DNA (34.2%, off-target amplicons). Several seemingly brain-enriched microbes in our dataset turned out to be false-positive signals upon closer examination. We identified off-target amplification as a major confounding factor in low-bacterial/high-host-DNA scenarios. These amplified human or mouse DNA sequences were clustered and falsely assigned to bacterial taxa in the majority of tested amplicon sequencing pipelines. Off-target amplicons seemed to be related to the tissue's sterility and could also be found in independent brain 16S rRNA gene sequences. CONCLUSIONS: Taxonomic signals obtained from (extremely) low biomass samples by 16S rRNA gene sequencing must be scrutinized closely to exclude the possibility of off-target amplifications, amplicons that can only appear enriched in biological samples, but are sometimes assigned to bacterial taxa. Sequences must be explicitly matched against any possible background genomes present in large quantities (i.e., the host genome). Using close scrutiny in our approach, we find no evidence supporting the hypothetical presence of either a brain microbiome or a bacterial infection in PD brains. Video abstract.

Multi-Omics Integration Highlights the Role of Ubiquitination in CCl4-Induced Liver Fibrosis.

Liver fibrosis is the excessive accumulation of extracellular matrix proteins that occurs in chronic liver disease. Ubiquitination is a post-translational modification that is crucial for a plethora of physiological processes. Even though the ubiquitin system has been implicated in several human diseases, the role of ubiquitination in liver fibrosis remains poorly understood. Here, multi-omics approaches were used to address this. Untargeted metabolomics showed that carbon tetrachloride (CCl4)-induced liver fibrosis promotes changes in the hepatic metabolome, specifically in glycerophospholipids and sphingolipids. Gene ontology analysis of public deposited gene array-based data and validation in our mouse model showed that the biological process "protein polyubiquitination" is enriched after CCl4-induced liver fibrosis. Finally, by using transgenic mice expressing biotinylated ubiquitin (bioUb mice), the ubiquitinated proteome was isolated and characterized by mass spectrometry in order to unravel the hepatic ubiquitinated proteome fingerprint in CCl4-induced liver fibrosis. Under these conditions, ubiquitination appears to be involved in the regulation of cell death and survival, cell function, lipid metabolism, and DNA repair. Finally, ubiquitination of proliferating cell nuclear antigen (PCNA) is induced during CCl4-induced liver fibrosis and associated with the DNA damage response (DDR). Overall, hepatic ubiquitome profiling can highlight new therapeutic targets for the clinical management of liver fibrosis.

Intestinal Microbiome-Macrophage Crosstalk Contributes to Cholestatic Liver Disease by Promoting Intestinal Permeability in Mice.

BACKGROUND AND AIMS: Mounting evidence supports an association between cholestatic liver disease and changes in the composition of the microbiome. Still, the role of the microbiome in the pathogenesis of this condition remains largely undefined. APPROACH AND RESULTS: To address this, we have used two experimental models, administering alpha-naphtylisocyanate or feeding a 0.1% 3,5-diethoxycarbonyl-1,4-dihydrocollidine diet, to induce cholestatic liver disease in germ-free mice and germ-free mice conventionalized with the microbiome from wild-type, specific pathogen-free animals. Next, we have inhibited macrophage activation by depleting these cells using clodronate liposomes and inhibiting the inflammasome with a specific inhibitor of NOD-, LRR-, and pyrin domain-containing protein 3. Our results demonstrate that cholestasis, the accumulation of bile acids in the liver, fails to promote liver injury in the absence of the microbiome in vivo. Additional in vitro studies supported that endotoxin sensitizes hepatocytes to bile-acid-induced cell death. We also demonstrate that during cholestasis, macrophages contribute to promoting intestinal permeability and to altered microbiome composition through activation of the inflammasome, overall leading to increased endotoxin flux into the cholestatic liver. CONCLUSIONS: We demonstrate that the intestinal microbiome contributes to cholestasis-mediated cell death and inflammation through mechanisms involving activation of the inflammasome in macrophages.

TGF-ß2 silencing to target biliary-derived liver diseases.

OBJECTIVE: TGF-ß2 (TGF-ß, transforming growth factor beta), the less-investigated sibling of TGF-ß1, is deregulated in rodent and human liver diseases. Former data from bile duct ligated and MDR2 knockout (KO) mouse models for human cholestatic liver disease suggested an involvement of TGF-ß2 in biliary-derived liver diseases. DESIGN: As we also found upregulated TGFB2 in liver tissue of patients with primary sclerosing cholangitis (PSC) and primary biliary cholangitis (PBC), we now fathomed the positive prospects of targeting TGF-ß2 in early stage biliary liver disease using the MDR2-KO mice. Specifically, the influence of TgfB2 silencing on the fibrotic and inflammatory niche was analysed on molecular, cellular and tissue levels. RESULTS: TgfB2-induced expression of fibrotic genes in cholangiocytes and hepatic stellate cellswas detected. TgfB2 expression in MDR2-KO mice was blunted using TgfB2-directed antisense oligonucleotides (AON). Upon AON treatment, reduced collagen deposition, hydroxyproline content and αSMA expression as well as induced PparG expression reflected a significant reduction of fibrogenesis without adverse effects on healthy livers. Expression analyses of fibrotic and inflammatory genes revealed AON-specific regulatory effects on Ccl3, Ccl4, Ccl5, Mki67 and Notch3 expression. Further, AON treatment of MDR2-KO mice increased tissue infiltration by F4/80-positive cells including eosinophils, whereas the number of CD45-positive inflammatory cells decreased. In line, TGFB2 and CD45 expression correlated positively in PSC/PBC patients and localised in similar areas of the diseased liver tissue. CONCLUSIONS: Taken together, our data suggest a new mechanistic explanation for amelioration of fibrogenesis by TGF-ß2 silencing and provide a direct rationale for TGF-ß2-directed drug development.

Fibrosis and the intestinal microbiome; a focus on chronic liver disease.

The role of the microbiome in progression of liver disease is an exciting area of research that is advancing rapidly supported by the development of next-generation sequencing and bioinformatics tools that simultaneously identify the composition and function of the microbiome. Changes in the microbiome are associated with pathogenesis of chronic liver disease; specifically, changes in microbiome composition predict disease severity and specific microbial signatures can be used to distinguish between mild disease, advanced fibrosis and cirrhosis. Future work combining functional metagenomic analysis with preclinical mechanistic studies will be key to advancing our understanding of how the microbiome affects the pathogenesis of different chronic liver disease aetiologies and to identify personalised therapeutics based on modulation of the microbiome and its function.

ROS-mediated PI3K activation drives mitochondrial transfer from stromal cells to hematopoietic stem cells in response to infection.

Hematopoietic stem cells (HSCs) undergo rapid expansion in response to stress stimuli. Here we investigate the bioenergetic processes which facilitate the HSC expansion in response to infection. We find that infection by Gram-negative bacteria drives an increase in mitochondrial mass in mammalian HSCs, which results in a metabolic transition from glycolysis toward oxidative phosphorylation. The initial increase in mitochondrial mass occurs as a result of mitochondrial transfer from the bone marrow stromal cells (BMSCs) to HSCs through a reactive oxygen species (ROS)-dependent mechanism. Mechanistically, ROS-induced oxidative stress regulates the opening of connexin channels in a system mediated by phosphoinositide 3-kinase (PI3K) activation, which allows the mitochondria to transfer from BMSCs into HSCs. Moreover, mitochondria transfer from BMSCs into HSCs, in the response to bacterial infection, occurs before the HSCs activate their own transcriptional program for mitochondrial biogenesis. Our discovery demonstrates that mitochondrial transfer from the bone marrow microenvironment to HSCs is an early physiologic event in the mammalian response to acute bacterial infection and results in bioenergetic changes which underpin emergency granulopoiesis.

Cholestasis induced by bile duct ligation promotes changes in the intestinal microbiome in mice.

Increasing evidence point to the relevance of intestinal disfunction and changes in the microbiome composition during chronic liver disease. More specifically, recent studies have highlighted that cholestatic diseases associate with a reduction in the microbiome diversity in patients. Still, the dynamics of the changes in the microbiome composition observed, as well as their implication in contributing to the pathogenesis of this disease remain largely undefined. Hence, experimental mouse models resembling the human pathogenesis are crucial to move forward our understanding on the mechanisms underpinning cholestatic disease and to enable the development of effective therapeutics. Our results show that the bile duct ligation (BDL) experimental model of cholestasis leads to rapid and significant changes in the microbiome diversity, with more than 100 OTUs being significantly different in faecal samples obtained from WT mice at 3 days and 7 days after BDL when compared to control animals. Changes in the microbial composition in mice after BDL included the enrichment of Akkermansia, Prevotella, Bacteroides and unclassified Ruminococcaceae in parallel with a drastic reduction of the presence of Faecalibacterium prausnitzii. In conclusion, our results support that bile duct ligation induces changes in the microbiome that partly resemble the gut microbial changes observed during human cholestatic disease.

The ATG5-binding and coiled coil domains of ATG16L1 maintain autophagy and tissue homeostasis in mice independently of the WD domain required for LC3-associated phagocytosis.

Macroautophagy/autophagy delivers damaged proteins and organelles to lysosomes for degradation, and plays important roles in maintaining tissue homeostasis by reducing tissue damage. The translocation of LC3 to the limiting membrane of the phagophore, the precursor to the autophagosome, during autophagy provides a binding site for autophagy cargoes, and facilitates fusion with lysosomes. An autophagy-related pathway called LC3-associated phagocytosis (LAP) targets LC3 to phagosome and endosome membranes during uptake of bacterial and fungal pathogens, and targets LC3 to swollen endosomes containing particulate material or apoptotic cells. We have investigated the roles played by autophagy and LAP in vivo by exploiting the observation that the WD domain of ATG16L1 is required for LAP, but not autophagy. Mice lacking the linker and WD domains, activate autophagy, but are deficient in LAP. The LAP-/- mice survive postnatal starvation, grow at the same rate as littermate controls, and are fertile. The liver, kidney, brain and muscle of these mice maintain levels of autophagy cargoes such as LC3 and SQSTM1/p62 similar to littermate controls, and prevent accumulation of SQSTM1 inclusions and tissue damage associated with loss of autophagy. The results suggest that autophagy maintains tissue homeostasis in mice independently of LC3-associated phagocytosis. Further deletion of glutamate E230 in the coiled-coil domain required for WIPI2 binding produced mice with defective autophagy that survived neonatal starvation. Analysis of brain lysates suggested that interactions between WIPI2 and ATG16L1 were less critical for autophagy in the brain, which may allow a low level of autophagy to overcome neonatal lethality. Abbreviations: CCD: coiled-coil domain; CYBB/NOX2: cytochrome b-245: beta polypeptide; GPT/ALT: glutamic pyruvic transaminase: soluble; LAP: LC3-associated phagocytosis; LC3: microtubule-associated protein 1 light chain 3; MEF: mouse embryonic fibroblast; NOD: nucleotide-binding oligomerization domain; NADPH: nicotinamide adenine dinucleotide phosphate; RUBCN/Rubicon: RUN domain and cysteine-rich domain containing Beclin 1-interacting protein; SLE: systemic lupus erythematosus; SQSTM1/p62: sequestosome 1; TLR: toll-like receptor; TMEM: transmembrane protein; TRIM: tripartite motif-containing protein; UVRAG: UV radiation resistance associated gene; WD: tryptophan-aspartic acid; WIPI: WD 40 repeat domain: phosphoinositide interacting.

Fine-Tuning of Sirtuin 1 Expression Is Essential to Protect the Liver From Cholestatic Liver Disease.

Cholestasis comprises aetiologically heterogeneous conditions characterized by accumulation of bile acids in the liver that actively contribute to liver damage. Sirtuin 1 (SIRT1) regulates liver regeneration and bile acid metabolism by modulating farnesoid X receptor (FXR); we here investigate its role in cholestatic liver disease. We determined SIRT1 expression in livers from patients with cholestatic disease, in two experimental models of cholestasis, as well as in human and murine liver cells in response to bile acid loading. SIRT1-overexpressing (SIRToe ) and hepatocyte-specific SIRT1-KO (knockout) mice (SIRThep-/- ) were subjected to bile duct ligation (BDL) and were fed with a 0.1% DDC (3,5-diethoxycarbonyl-1,4-dihydrocollidine) diet to determine the biological relevance of SIRT1 during cholestasis. The effect of NorUDCA (24-norursodeoxycholic acid) was tested in BDL/SIRToe mice. We found that SIRT1 was highly expressed in livers from cholestatic patients, mice after BDL, and Mdr2 knockout mice (Mdr2-/- ) animals. The detrimental effects of SIRT1 during cholestasis were validated in vivo and in vitro. SIRToe mice showed exacerbated parenchymal injury whereas SIRThep-/- mice evidenced a moderate improvement after BDL and 0.1% DDC feeding. Likewise, hepatocytes isolated from SIRToe mice showed increased apoptosis in response to bile acids, whereas a significant reduction was observed in SIRThep-/- hepatocytes. Importantly, the decrease, but not complete inhibition, of SIRT1 exerted by norUDCA treatment correlated with pronounced improvement in liver parenchyma in BDL/SIRToe mice. Interestingly, both SIRT1 overexpression and hepatocyte-specific SIRT1 depletion correlated with inhibition of FXR, whereas modulation of SIRT1 by NorUDCA associated with restored FXR signaling. Conclusion: SIRT1 expression is increased during human and murine cholestasis. Fine-tuning expression of SIRT1 is essential to protect the liver from cholestatic liver damage.