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Farnesoid X receptor (FXR) belongs to the nuclear receptor superfamily with its endogenous ligands bile acids. Mice with whole body FXR deficiency develop liver tumors spontaneously, but the underlying mechanism is unclear. Moreover, it is unknown whether FXR deficiency in liver alone serves as a tumor initiator or promoter during liver carcinogenesis. This study aims to evaluate the effects of hepatocyte-specific FXR deficiency (FXRhep−/−) in liver tumor formation. The results showed that FXRhep−/− mice did not show spontaneous liver tumorigenesis with aging (up to 24 mo of age). Therefore FXRhep−/− mice were fed a bile acid (cholic acid)-containing diet alone or along with a liver tumor initiator, diethylnitrosamine (DEN). Thirty weeks later, no tumors were found in wild-type or FXRhep−/− mice without any treatment or with DEN only. However, with cholic acid, while only some wild-type mice developed tumors, all FXRhep−/− mice presented with severe liver injury and tumors. Interestingly, FXRhep−/− mouse livers increased basal expression of tumor suppressor p53 protein, apoptosis, and decreased basal cyclin D1 expression, which may prevent tumor development in FXRhep−/− mice. However, cholic acid feeding reversed these effects in FXRhep−/− mice, which is associated with an increased cyclin D1 and decreased cell cycle inhibitors. More in-depth analysis indicates that the increased in cell growth might result from disturbance of the MAPK and JAK/Stat3 signaling pathways. In conclusion, this study shows that hepatic FXR deficiency may only serve as a tumor initiator, and increased bile acids is required for tumor formation likely by promoting cell proliferation.

The liver is unique in regenerative potential, which could recover the lost mass and function after injury from ischemia and resection. The underlying molecular mechanisms of liver regeneration have been extensively studied in the past using the partial hepatectomy (PH) model in rodents, where 2/3 PH is carried out by removing two lobes. The whole process of liver regeneration is complicated, orchestrated event involving a network of connected interactions, which still remain fully elusive. Bile acids (BAs) are ligands of farnesoid X receptor (FXR), a nuclear receptor of ligand-activated transcription factor. FXR has been shown to be highly involved in liver regeneration. BAs and FXR not only interact with each other but also regulate various downstream targets independently during liver regeneration. Moreover, recent findings suggest that tissue-specific FXR also contributes to liver regeneration significantly. These novel findings suggest that FXR has much broader role than regulating BA, cholesterol, lipid and glucose metabolism. Therefore, these researches highlight FXR as an important pharmaceutical target for potential use of FXR ligands to regulate liver regeneration in clinic. This review focuses on the roles of BAs and FXR in liver regeneration and the current underlying molecular mechanisms which contribute to liver regeneration., Graphical abstract This review focuses on the roles of bile acids (BAs) and farnesoid X receptor (FXR) in liver regeneration and the current underlying molecular mechanisms which contribute to liver regeneration.

Fibroblast growth factor (FGF) 15 (human homolog, FGF19) is an endocrine FGF highly expressed in the small intestine of mice. Emerging evidence suggests that FGF15 is critical for regulating hepatic functions; however, the role of FGF15 in liver regeneration is unclear. This study assessed whether liver regeneration is altered in FGF15 knockout (KO) mice following 2/3 partial hepatectomy (PHx). The results showed that FGF15 KO mice had marked mortality, with the survival rate influenced by genetic background. Compared with wild-type mice, the KO mice displayed extensive liver necrosis and marked elevation of serum bile acids and bilirubin. Furthermore, hepatocyte proliferation was reduced in the KO mice because of impaired cell cycle progression. After PHx, the KO mice had weaker activation of signaling pathways that are important for liver regeneration, including signal transducer and activator of transcription 3, nuclear factor-κB, and mitogen-activated protein kinase. Examination of the KO mice at early time points after PHx revealed a reduced and/or delayed induction of immediate-early response genes, including growth-control transcription factors that are critical for liver regeneration. In conclusion, the results suggest that FGF15 deficiency severely impairs liver regeneration in mice after PHx. The underlying mechanism is likely the result of disrupted bile acid homeostasis and impaired priming of hepatocyte proliferation.

Farnesoid X receptor (FXR, Nr1h4) is a ligand-activated transcription factor belonging to the nuclear receptor superfamily. FXR is essential in maintaining bile acid (BA) homeostasis, and FXR−/− mice develop cholestasis, inflammation, and spontaneous liver tumors. The signal transducer and activator of transcription 3 (STAT3) is well known to regulate liver growth, and STAT3 is feedback inhibited by its target gene, the suppressor of cytokine signaling 3 (SOCS3). Strong activation of STAT3 was detected in FXR−/− mouse livers. However, the mechanism of STAT3 activation with FXR deficiency remains elusive. Wild-type (WT) and FXR−/− mice were used to detect STAT3 pathway activation in the liver. In vivo BA feeding or deprivation was used to determine the role of BAs in STAT3 activation, and in vitro molecular approaches were used to determine the direct transcriptional regulation of SOCS3 by FXR. STAT3 was activated in FXR−/− but not WT mice. BA feeding increased, but deprivation by cholestyramine reduced, serum inflammatory markers and STAT3 activation. Furthermore, the Socs3 gene was determined as a direct FXR target gene. The elevated BAs and inflammation, along with reduced SOCS3, collectively contribute to the activation of the STAT3 signaling pathway in the liver of FXR−/− mice. This study suggests that the constitutive activation of STAT3 may be a mechanism of liver carcinogenesis in FXR−/− mice.

Farnesoid X receptor (FXR, Nr1h4) and small heterodimer partner (SHP, Nr0b2) are nuclear receptors that are critical to liver homeostasis. Induction of SHP serves as a major mechanism of FXR in suppressing gene expression. Both FXR(-/-) and SHP(-/-) mice develop spontaneous hepatocellular carcinoma (HCC). SHP is one of the most strongly induced genes by FXR in the liver and is a tumor suppressor, therefore, we hypothesized that deficiency of SHP contributes to HCC development in the livers of FXR(-/-) mice and therefore, increased SHP expression in FXR(-/-) mice reduces liver tumorigenesis. To test this hypothesis, we generated FXR(-/-) mice with overexpression of SHP in hepatocytes (FXR(-/-)/SHP(Tg)) and determined the contribution of SHP in HCC development in FXR(-/-) mice. Hepatocyte-specific SHP overexpression did not affect liver tumor incidence or size in FXR(-/-) mice. However, SHP overexpression led to a lower grade of dysplasia, reduced indicator cell proliferation and increased apoptosis. All tumor-bearing mice had increased serum bile acid levels and IL-6 levels, which was associated with activation of hepatic STAT3. In conclusion, SHP partially protects FXR(-/-) mice from HCC formation by reducing tumor malignancy. However, disrupted bile acid homeostasis by FXR deficiency leads to inflammation and injury, which ultimately results in uncontrolled cell proliferation and tumorigenesis in the liver.

Farnesoid X receptor (Fxr) is a ligand-activated nuclear receptor critical for liver function. Reports indicate that the functions of Fxr in the liver may overlap with those of hepatocyte nuclear factor 4α (Hnf4α), but studies of their precise genome-wide interaction to regulate gene transcription in the liver are lacking. Thus, we compared the genome-wide binding of Fxr and Hnf4α in the liver of mice and characterized their cooperative activity on binding to and activating target gene transcription.Genome-wide ChIP-Seq data of Fxr and Hnf4α in mouse liver were analyzed by MACS, BEDTools, and DAVID. Co-immunoprecipitation, ChIP-qPCR, and luciferase assays were done to test for protein-protein interaction and cooperative binding.ChIP-seq analysis showed nearly 50% binding sites of Fxr and Hnf4α in mouse liver overlap and Hnf4α bound to shared target sites upstream and in close proximity to Fxr. Moreover, genes co-bound by Fxr and Hnf4α are enriched in complement and coagulation cascades and drug metabolism. A direct Fxr-Hnf4α protein interaction dependent on Fxr activity was detected and transcriptional assays suggest that Hnf4α can increase Fxr transcriptional activity. Conversely, binding assays showed Hnf4α can be either Fxr-dependent or -independent at different shared binding sites.Our results showed that Fxr cooperates with Hnf4α in the liver to modulate gene transcription. This study provides the first evidence on a genome-wide scale of both cooperative and independent interactions between Fxr and Hnf4α in regulating gene transcription in the liver.

The farnesoid X receptor (FXR) is a bile acid-activated transcription factor belonging to the nuclear receptor superfamily. FXR deficiency in mice results in cholestasis, metabolic disorders, and tumorigenesis in liver and intestine. FXR is known to contribute to pathogenesis by regulating gene transcription; however, changes in the post-transcriptional modification of proteins associated with FXR modulation have not been determined. In the current study, proteomic analysis of the livers of wild-type (WT) and FXR knockout (FXR-KO) mice treated with a FXR synthetic ligand or vehicle was performed. The results identified five proteins as novel FXR targets. Since FXR deficiency in mice leads to liver tumorigenesis, poly (ADP-ribose) polymerase family, member 1 (Parp1) that is important for DNA repair, was validated in the current study by quantitative real-time PCR, and 1- and 2-dimensional gel electrophoresis/western blot. The results showed that Parp1 mRNA levels were not altered by FXR genetic status or by agonist treatment. However, total Parp1 protein levels were increased in FXR-KO mice as early as 3 month old. Interestingly, total Parp1 protein levels were increased in WT mice in an age-dependent manner (from 3 to 18 months), but not in FXR-KO mice. Finally, activation of FXR in WT mice resulted in reduction of phosporylated Parp1 protein in the liver without affecting total Parp1 protein levels. In conclusion, this study reveals that FXR genetic status and agonist treatment affects basal levels and phosphorylation state of Parp1, respectively. These alterations, in turn, may be associated with the hepatobiliary alterations observed in FXR-KO mice and participate in FXR agonist-induced protection in the liver.

Background Farnesoid X Receptor (FXR) is a member of the nuclear receptor superfamily and is a ligand-activated transcription factor essential for maintaining liver and intestinal homeostasis. FXR is protective against carcinogenesis and inflammation in liver and intestine as demonstrated by the development of inflammation and tumors in the liver and intestine of FXR knock-out mice. However, mechanisms for the protective effects of FXR are not completely understood. This study reports a novel role of FXR in regulating expression of Sqstm1, which encodes for p62 protein. p62 plays an important role in maintaining cellular homeostasis through selective autophagy and activating signal transduction pathways, such as NF-κB to support cell survival and caspase-8 to initiate apoptosis. FXR regulation of Sqstm1 may serve as a protective mechanism. Methods and Results This study showed that FXR bound to the Sqstm1 gene in both mouse livers and ileums as determined by chromatin immunoprecipitation. In addition, FXR activation enhanced transcriptional activation of Sqstm1 in vitro. However, wild-type mice treated with GW4064, a synthetic FXR ligand, showed that FXR activation induced mRNA and protein expression of Sqstm1/p62 in ileum, but not in liver. Interestingly, FXR-transgenic mice showed induced mRNA expression of Sqstm1 in both liver and ileum compared to wild-type mice. Conclusions Our current study has identified a novel role of FXR in regulating the expression of p62, a key factor in protein degradation and cell signaling. Regulation of p62 by FXR indicates tissue-specific and gene-dosage effects. Furthermore, FXR-mediated induction of p62 may implicate a protective mechanism of FXR.

Farnesoid X receptor (FXR) is a nuclear receptor and a key regulator of liver cholesterol and triglyceride homeostasis. Scavenger receptor class B type I (SR-BI) is critical for reverse cholesterol transport (RCT) by transporting high-density lipoprotein (HDL) into liver. FXR induces SR-BI, however, the underlying molecular mechanism of this induction is not known. The current study confirmed induction of SR-BI mRNA by activated FXR in mouse livers, a human hepatoma cell line, and primary human hepatocytes. Genome-wide FXR binding analysis in mouse livers identified 4 putative FXR response elements in the form of inverse repeat separated by one nucleotide (IR1) at the first intron and 1 IR1 at the downstream of the mouse Sr-bi gene. ChIP-qPCR analysis revealed FXR binding to only the intronic IR1s, but not the downstream one. Luciferase assays and site-directed mutagenesis further showed that 3 out of 4 IR1s were able to activate gene transcription. A 16-week high-fat diet (HFD) feeding in mice increased hepatic Sr-bi gene expression in a FXR-dependent manner. In addition, FXR bound to the 3 bona fide IR1s in vivo, which was increased following HFD feeding. Serum total and HDL cholesterol levels were increased in FXR knockout mice fed the HFD, compared to wild-type mice. In conclusion, the Sr-bi/SR-BI gene is confirmed as a FXR target gene in both mice and humans, and at least in mice, induction of Sr-bi by FXR is via binding to intronic IR1s. This study suggests that FXR may serve as a promising molecular target for increasing reverse cholesterol transport.

Farnesoid X receptor (FXR) is a ligand-activated nuclear receptor and serves as a key regulator to maintain health of the liver and intestine. Bile acids are endogenous ligands of FXR, and there are increasing efforts to identify FXR modulators to serve as biological probes and/or pharmaceutical agents. Natural FXR ligands isolated from plants may serve as models to synthesize novel FXR modulators. In this study, we demonstrated that epigallocatechin-3-gallate (EGCG), a major tea catechin, specifically and dose-dependently activates FXR. In addition, EGCG induced FXR target gene expression in vitro. Surprisingly, in a co-activator (SRC2) recruitment assay, we found that EGCG does not recruit SRC2 to FXR, but it dose-dependently inhibits recruitment of SRC2 to FXR (IC(50), 1μM) by GW6064, which is a potent FXR synthetic ligand. In addition, EGCG suppressed FXR target gene expression induced by either GW4064 or chenodeoxycholic acid in vitro. Furthermore, wild-type and FXR knockout mice treated with an acute dose of EGCG had induced mRNA expression in a subset of FXR target genes in the intestine but not in the liver. In conclusion, EGCG is a unique modulator of FXR in the intestine and may serve as an important model for future development of FXR modulators.