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Resistance to therapeutic drugs is a major challenge in the treatment of cancers, including breast cancer. Long noncoding RNAs (lncRNA) are known to have diverse physiologic and pathophysiologic functions, including in cancer. In searching for lncRNA responsible for cancer drug resistance, we identified an intergenic lncRNA ERINA (estrogen inducible lncRNA) as a novel lncRNA highly expressed in multiple cancer types, especially in estrogen receptor–positive (ER+) breast cancers. Expression of ERINA was inversely correlated with survival of patients with ER+ breast cancer and sensitivity to CDK inhibitor in breast cancer cell lines. Functional characterization established ERINA as an oncogenic lncRNA, as knockdown of ERINA in breast cancer cells inhibited cell-cycle progression and tumor cell proliferation in vitro and xenograft tumor growth in vivo. In contrast, overexpression of ERINA promoted cell growth and cell-cycle progression. ERINA promoted cell-cycle progression by interacting with the E2F transcription factor 1 (E2F1), which prevents the binding of E2F1 to the tumor suppressor retinoblastoma protein 1 (RB1). ERINA also functioned as an estrogen and ER-responsive gene, and an intronic ER-binding site was identified as an enhancer that mediates the transactivation of ERINA. In summary, ERINA is an estrogen-responsive oncogenic lncRNA that may serve as a novel biomarker and potential therapeutic target in breast cancer.Significance:These findings identify ERINA as an estrogen-responsive, oncogenic lncRNA, whose elevated expression may contribute to drug resistance and poor survival of patients with ER+ breast cancer.

Supplementary Figures 1-5. Supplementary Figure 1. ERINA is a nuclear lncRNA that affects the survival and cell proliferation marker of ER-positive breast cancer patients. Supplementary Figure 2. ERINA knockdown and overexpression efficiency and the morphology change of knockdown cells. Supplementary Figure 3. RNA sequencing analysis showed the number of up-regulated genes and downregulated genes in T47D cells. Supplementary Figure 4. GSEA analysis revealed ERINA played a crucial role in RB1-involved pathway. Supplementary Figure 5. ChIP-seq analysis reveals the binding of RNA polymerase II within the promoter region of ERINA. (A) Alignment of RNA polymerase II ChIP-seq in MCF7 cells. The binding peak and genomic locus are shown.

Crosstalk between xenobiotic metabolism and energy metabolism in the liver has provided a potential opportunity to target xenobiotic receptors to treat metabolic diseases. Activation of constitutive androstane receptor (CAR), a xenobiotic-sensing nuclear receptor, has been shown to inhibit obesity, suppress hepatic gluconeogenesis, and ameliorate hyperglycemia in rodent models of obesity and type 2 diabetes. However, the underlying molecular mechanism remains to be defined. The growth arrest and DNA damage-inducible gene 45b (Gadd45b), a well-known anti-apoptotic factor, has been shown to be an inducible coactivator of CAR in promoting rapid liver growth. It is unknown whether the effect of CAR on energy metabolism depends on GADD45B. In the present study and by using a high fat diet (HFD)-induced obesity model, we show that reduced body weight gain and improved insulin sensitivity by the CAR agonist 1,4-bis[2-(3,5-dichloropyridyloxy)] benzene (TCPOBOP) were markedly blunted in Gadd45b knockout mice. Mechanistically, the TCPOBOP-responsive inhibition of hepatic lipogenesis, gluconeogenesis, and adipose inflammation observed in wild type mice were largely abolished in Gadd45b knockout mice. We conclude that Gadd45b is required in part for the metabolic benefits of CAR activation., Graphical abstract Activation of xenobiotic receptor constitutive androstane receptor (CAR) by its agonist such as TCPOBOP conveys benefits for obesity and diabetes. In this study, it showes that anti-apoptotic factor GADD45B, a coactivator of CAR, is in part required for the anti-obesity effects of CAR activation. Inhibition of hepatic lipogenesis and gluconeogenesis and adipose inflammation by TCPOBOP was largely abolished in mice in the absence of GADD45B.Image 1

Autoimmune hepatitis (AIH) is an inflammatory disease of the liver. Liver X receptors (LXRs), including the α and β isoforms, are previously known for their anti‐inflammatory activities. The goal of this study is to determine whether and how LXR plays a role in AIH. LXRα gain‐of‐function and loss‐of‐function mouse models were used, in conjunction with the concanavalin A (ConA) model of T‐cell mediated hepatitis. We first showed that the hepatic expression of LXRα was decreased in the ConA model of hepatitis and in human patients with AIH. In the ConA model, we were surprised to find that activation of LXRα in the constitutively activated VP‐LXRα whole‐body knock‐in (LXRα‐KI) mice exacerbated ConA‐induced AIH, whereas the LXRα−/− mice showed attenuated ConA‐induced AIH. Interestingly, hepatocyte‐specific activation of LXRα in the fatty acid binding protein–VP‐LXRα transgenic mice did not exacerbate ConA‐induced hepatitis. Mechanistically, the sensitizing effect of the LXRα‐KI allele was invariant natural killer T (iNKT)–cell dependent, because the sensitizing effect was abolished when the LXRα‐KI allele was bred into the NKT‐deficient CD1d−/− background. In addition, LXRα‐enhanced ConA‐induced hepatitis was dependent on interferon gamma. In contrast, adoptive transfer of hepatic iNKT cells isolated from LXRα‐KI mice was sufficient to sensitize CD1d−/− mice to ConA‐induced AIH. Conclusion: Activation of LXRα sensitizes mice to ConA‐induced AIH in iNKT and interferon gamma–dependent manner. Our results suggest that LXRα plays an important role in the development of AIH., Our results showed that activation of LXRα sensitizes mice to ConA‐induced AIH in an iNKT‐dependent and IFN‐γ‐dependent manner. Our results suggest that LXRα plays an important role in the development of AIH.

Diabetes and related metabolic syndrome are common metabolic disorders. Gestational diabetes mellitus (GDM) is rather prevalent in the clinic. Although most GDM resolves after therapeutic intervention and/or after delivery, the long-term health effect of GDM remains to be better understood. The constitutive androstane receptor (CAR), initially characterized as a xenobiotic receptor, was more recently proposed to be a therapeutic target for obesity and type 2 diabetes mellitus (T2DM). In this study, high-fat diet (HFD) feeding was used to induce GDM. Upon delivery, GDM mice were returned to chow diet until the metabolic parameters were normalized. Parous non-GDM control females or metabolically normalized GDM females were then subjected to HFD feeding to induce nongestational obesity and T2DM. Our results showed that GDM sensitized mice to metabolic abnormalities induced by a second hit of HFD. Treatment with the CAR agonist 1,4-bis [2-(3,5 dichloropyridyloxy)] benzene efficiently attenuated GDM-sensitized and HFD-induced obesity and T2DM, including decreased body weight, improved insulin sensitivity, inhibition of hyperglycemia and hepatic steatosis, increased oxygen consumption, and decreased adipocyte hypertrophy. In conclusion, our results have established GDM as a key risk factor for the future development of metabolic disease. We also propose that CAR is a therapeutic target for the management of metabolic disease sensitized by GDM.

Phenobarbital (PB), a widely used antiepileptic drug, is known to upregulate the expression of numerous drug-metabolizing enzymes and transporters in the liver primarily via activation of the constitutive androstane receptor (CAR, NR1I3). The solute carrier family 13 member 5 (SLC13A5), a sodium-coupled citrate transporter, plays an important role in intracellular citrate homeostasis that is associated with a number of metabolic syndromes and neurological disorders. Here, we show that PB markedly elevates the expression of SLC13A5 through a pregnane X receptor (PXR)-dependent but CAR-independent signaling pathway. In human primary hepatocytes, the mRNA and protein expression of SLC13A5 was robustly induced by PB treatment, while genetic knockdown or pharmacological inhibition of PXR significantly attenuated this induction. Utilizing genetically modified HepaRG cells, we found that PB induces SLC13A5 expression in both wild type and CAR-knockout HepaRG cells, whereas such induction was fully abolished in the PXR-knockout HepaRG cells. Mechanistically, we identified and functionally characterized three enhancer modules located upstream from the transcription start site or introns of the SLC13A5 gene that are associated with the regulation of PXR-mediated SLC13A5 induction. Moreover, metformin, a deactivator of PXR, dramatically suppressed PB-mediated induction of hepatic SLC13A5 as well as its activation of the SLC13A5 luciferase reporter activity via PXR. Collectively, these data reveal PB as a potent inducer of SLC13A5 through the activation of PXR but not CAR in human primary hepatocytes.

Activation of the hepatic stellate cells (HSCs) is a key pathogenic event in liver fibrosis. Protein S-glutathionylation (PSSG) of cysteine residues is a distinct form of oxidative response that mo...

Conjugated equine estrogens (CEEs), whose brand name is Premarin, are widely used as a hormone-replacement therapy (HRT) drug to manage postmenopausal symptoms in women. Extracted from pregnant mare urine, CEEs are composed of nearly a dozen estrogens existing in an inactive sulfated form. To determine whether the hepatic steroid sulfatase (STS) is a key contributor to the efficacy of CEEs in HRT, we performed estrogen-responsive element (ERE) reporter gene assay, real-time PCR, and UPLC-MS/MS to assess the STS-dependent and inflammation-responsive estrogenic activity of CEEs in HepG2 cells and human primary hepatocytes. Using liver-specific STS-expressing transgenic mice, we also evaluated the effect of STS on the estrogenic activity of CEEs in vivo. We observed that CEEs induce activity of the ERE reporter gene in an STS-dependent manner and that genetic or pharmacological inhibition of STS attenuates CEE estrogenic activity. In hepatocytes, inflammation enhanced CEE estrogenic activity by inducing STS gene expression. The inflammation-responsive estrogenic activity of CEEs, in turn, attenuated inflammation through the anti-inflammatory activity of the active estrogens. In vivo, transgenic mice with liver-specific STS expression exhibited markedly increased sensitivity to CEE-induced estrogenic activity in the uterus resulting from increased levels of liver-derived and circulating estrogens. Our results reveal a critical role of hepatic STS in mediating the hormone-replacing activity of CEEs. We propose that caution needs to be applied when Premarin is used in patients with chronic inflammatory liver diseases because such patients may have heightened sensitivity to CEEs due to the inflammatory induction of STS activity.

Overdose of acetaminophen (APAP) is the leading cause of acute liver failure (ALF) in the United States. The sulfotransferase-mediated sulfation of APAP is widely believed to be a protective mechanism to attenuate the hepatotoxicity of APAP. The cholesterol sulfotransferase SULT2B1b is best known for its activity in catalyzing the sulfoconjugation of cholesterol to synthesize cholesterol sulfate. SULT2B1b can be transcriptionally and positively regulated by the hepatic nuclear factor 4α (HNF4α). In this study, we uncovered an unexpected role for SULT2B1b in APAP toxicity. Hepatic overexpression of SULT2B1b sensitized mice to APAP-induced liver injury, whereas ablation of the Sult2B1b gene in mice conferred resistance to the APAP hepatotoxicity. Consistent with the notion that Sult2B1b is a transcriptional target of HNF4α, overexpression of HNF4α sensitized mice or primary hepatocytes to APAP-induced hepatotoxicity in a Sult2B1b-dependent manner. We conclude that the HNF4α-SULT2B1b axis has a unique role in APAP-induced acute liver injury, and SULT2B1b induction might be a risk factor for APAP hepatotoxicity.

Sulfoconjugation of small molecules or protein peptides is a key mechanism to ensure biochemical and functional homeostasis in mammals. The PAPS synthase 2 (PAPSS2) is the primary enzyme to synthesize the universal sulfonate donor 3'-phosphoadenosine 5'-phosphosulfate (PAPS). Acetaminophen (APAP) overdose is the leading cause of acute liver failure (ALF), in which oxidative stress is a key pathogenic event, whereas sulfation of APAP contributes to its detoxification. The goal of this study was to determine whether and how PAPSS2 plays a role in APAP-induced ALF.Gene expression was analyzed in APAP-induced ALF in patients and mice. Liver-specific Papss2-knockout mice using Alb-Cre (Papss2The hepatic expression of PAPSS2 was decreased in APAP-induced ALF in patients and mice. Surprisingly, Papss2We have uncovered a previously unrecognized and p53-mediated role of PAPSS2 in controlling oxidative response. Inhibition of p53 sulfation may be explored for the clinical management of APAP overdose.

Resistance to therapeutic drugs is a major challenge in the treatment of cancers, including breast cancer. Long noncoding RNAs (lncRNA) are known to have diverse physiologic and pathophysiologic functions, including in cancer. In searching for lncRNA responsible for cancer drug resistance, we identified an intergenic lncRNA ERINA (estrogen inducible lncRNA) as a novel lncRNA highly expressed in multiple cancer types, especially in estrogen receptor–positive (ER+) breast cancers. Expression of ERINA was inversely correlated with survival of patients with ER+ breast cancer and sensitivity to CDK inhibitor in breast cancer cell lines. Functional characterization established ERINA as an oncogenic lncRNA, as knockdown of ERINA in breast cancer cells inhibited cell-cycle progression and tumor cell proliferation in vitro and xenograft tumor growth in vivo. In contrast, overexpression of ERINA promoted cell growth and cell-cycle progression. ERINA promoted cell-cycle progression by interacting with the E2F transcription factor 1 (E2F1), which prevents the binding of E2F1 to the tumor suppressor retinoblastoma protein 1 (RB1). ERINA also functioned as an estrogen and ER-responsive gene, and an intronic ER-binding site was identified as an enhancer that mediates the transactivation of ERINA. In summary, ERINA is an estrogen-responsive oncogenic lncRNA that may serve as a novel biomarker and potential therapeutic target in breast cancer. Significance: These findings identify ERINA as an estrogen-responsive, oncogenic lncRNA, whose elevated expression may contribute to drug resistance and poor survival of patients with ER+ breast cancer.

Steroid sulfatase (STS), a desulfating enzyme that converts steroid sulfates to hormonally active steroids, plays an important role in the homeostasis of sex hormones. STS is expressed in the adipose tissue of both male and female mice, but the role of STS in the development and function of adipose tissue remains largely unknown. In this report, we show that the adipose expression of Sts was induced in the high-fat diet (HFD) and ob/ob models of obesity and type 2 diabetes. Transgenic overexpression of the human STS in the adipose tissue of male mice exacerbated the HFD-induced metabolic phenotypes, including increased body weight gain and fat mass, and worsened insulin sensitivity, glucose tolerance, and energy expenditure, which were accounted for by adipocyte hypertrophy, increased adipose inflammation, and dysregulation of adipogenesis. The metabolic harm of the STS transgene appeared to have resulted from increased androgen activity in the adipose tissue, and castration abolished most of the phenotypes. Interestingly, the transgenic effects were sex specific, because the HFD-fed female STS transgenic mice exhibited improved metabolic functions, which were associated with attenuated adipose inflammation. The metabolic benefit of the STS transgene in female mice was accounted for by increased estrogenic activity in the adipose tissue, whereas such benefit was abolished upon ovariectomy. Our results revealed an essential role of the adipose STS in energy homeostasis in sex- and sex hormone-dependent manner. The adipose STS may represent a therapeutic target for the management of obesity and type 2 diabetes.

Cholesterol is essential for numerous biologic functions and processes, but an excess of intracellular cholesterol can be toxic. Intestinal cholesterol absorption is a major determinant of plasma cholesterol level. The liver X receptor (LXR) is a nuclear receptor known for its activity in cholesterol efflux and reverse cholesterol transport. In this study, we uncovered a surprising function of LXR in intestinal cholesterol absorption and toxicity. Genetic or pharmacologic activation of LXRα-sensitized mice to a high-cholesterol diet (HCD) induced intestinal toxicity and tissue damage, including the disruption of enterocyte tight junctions, whereas the same HCD caused little toxicity in the absence of LXR activation. The gut toxicity in HCD-fed LXR-KI mice may have been accounted for by the increased intestinal cholesterol absorption and elevation of enterocyte and systemic levels of free cholesterol. The increased intestinal cholesterol absorption preceded the gut toxicity, suggesting that the increased absorption was not secondary to tissue damage. The heightened sensitivity to HCD in the HCD-fed LXRα-activated mice appeared to be intestine-specific because the liver was not affected despite activation of the same receptor in this tissue. Moreover, heightened sensitivity to HCD cannot be reversed by ezetimibe, a Niemann-Pick C1-like 1 inhibitor that inhibits intestinal cholesterol absorption, suggesting that the increased cholesterol absorption in LXR-activated intestine is mediated by a mechanism that has yet to be defined.

Estrogen sulfotransferase catalyzes the sulfoconjugation and deactivation of estrogens. Previously, we showed that loss of Est in male ob/ob mice, but not in female ob/ob mice, exacerbated the diabetic phenotype, but the underlying mechanism was unclear. In this study, we show that transgenic reconstitution of Est in the adipose tissue, but not in the liver, attenuated diabetic phenotype in Est-deficient ob/ob mice (obe mice). Mechanistically, adipose reconstitution of Est in obe mice (oae mice) resulted in reduced local and systemic inflammation, improved insulin sensitivity, and increased energy expenditure. At the molecular level, adipose induction of lipocalin-2 (Lcn2) in oae males may have contributed to the inhibition of inflammation because the level of Lcn2 was negatively associated with tumor necrosis factor (Tnf) α expression, and treatment of differentiated adipocytes with Lcn2 antagonized Tnfα-responsive inhibition of insulin signaling. The metabolic benefit of adipose reconstitution of Est was sex specific, because adipose reconstitution of Est in obe females had little effect. Interestingly, despite their improved metabolic functions, obe male mice with reconstituted Est in their adipose tissue failed to ameliorate the impairment of the structure and function of the pancreatic islets. In summary, our study uncovers a crucial adipose- and male-specific role of Est in maintaining the whole-body energy homeostasis.

Hemorrhagic shock (HS) is a potential life-threatening condition that may lead to injury to multiple organs, including the lung. The estrogen sulfotransferase (EST, or SULT1E1) is a conjugating enzyme that sulfonates and deactivates estrogens. In this report, we showed that the expression of Est was markedly induced in the liver but not in the lung of female mice subject to HS and resuscitation. Genetic ablation or pharmacological inhibition of Est effectively protected female mice from HS-induced acute lung injury (ALI), including interstitial edema, neutrophil mobilization and infiltration, and inflammation. The pulmonoprotective effect of Est ablation or inhibition was sex-specific, because the HS-induced ALI was not affected in male Est-/- mice. Mechanistically, the pulmonoprotective phenotype in female Est-/- mice was accompanied by increased lung and circulating levels of estrogens, attenuated pulmonary inflammation, and inhibition of neutrophil mobilization from the bone marrow and neutrophil infiltration to the lung, whereas the pulmonoprotective effect was abolished upon ovariectomy, suggesting that the protection was estrogen dependent. The pulmonoprotective effect of Est ablation was also tissue specific, as loss of Est had little effect on HS-induced liver injury. Moreover, transgenic reconstitution of human EST in the liver of global Est-/- mice abolished the pulmonoprotective effect, suggesting that it is the EST in the liver that sensitizes mice to HS-induced ALI. Taken together, our results revealed a sex- and tissue-specific role of EST in HS-induced ALI. Pharmacological inhibition of EST may represent an effective approach to manage HS-induced ALI.

Background & Aims The role of aryl hydrocarbon receptor (AhR) in liver fibrosis is controversial because loss and gain of AhR activity both lead to liver fibrosis. The goal of this study was to investigate how the expression of AhR by different liver cell types, hepatic stellate cells (HSCs) in particular, affects liver fibrosis in mice. Methods We studied the effects of AhR on primary mouse and human HSCs, measuring their activation and stimulation of fibrogenesis using RNA-sequencing analysis. C57BL/6J mice were given the AhR agonists 2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD) or 2-(1′H-indole-3′-carbonyl)-thiazole-4-carboxylic acid methyl ester (ITE); were given carbon tetrachloride (CCl4); or underwent bile duct ligation. We also performed studies in mice with disruption of Ahr specifically in HSCs, hepatocytes, or Kupffer cells. Liver tissues were collected from mice and analyzed by histology, immunohistochemistry, and immunoblotting. Results AhR was expressed at high levels in quiescent HSCs, but the expression decreased with HSC activation. Activation of HSCs from AhR-knockout mice was accelerated compared with HSCs from wild-type mice. In contrast, TCDD or ITE inhibited spontaneous and transforming growth factor β–induced activation of HSCs. Mice with disruption of Ahr in HSCs, but not hepatocytes or Kupffer cells, developed more severe fibrosis after administration of CCl4 or bile duct ligation. C57BL/6J mice given ITE did not develop CCl4-induced liver fibrosis, whereas mice without HSC AhR given ITE did develop CCl4-induced liver fibrosis. In studies of mouse and human HSCs, we found that AhR prevents transforming growth factor β–induced fibrogenesis by disrupting the interaction of Smad3 with β-catenin, which prevents the expression of genes that mediate fibrogenesis. Conclusions In studies of human and mouse HSCs, we found that AhR prevents HSC activation and expression of genes required for liver fibrogenesis. Development of nontoxic AhR agonists or strategies to activate AhR signaling in HSCs might be developed to prevent or treat liver fibrosis.

Conjugated equine estrogen (CEE) is widely used in hormone replacement therapy in postmenopausal women. Extracted from pregnant mare urine, CEE is composed of nearly a dozen estrogens existing in their inactively sulfated forms. The goal of this study is to determine whether the steroid sulfatase (STS), the enzyme responsible for steroid desulfation and re-activation, is the major contributor to the hormone replacement therapy effect of CEE. To study the role of STS in CEE’s effect, we performed estrogen responsive element (ERE) reporter gene assay, real-time PCR and UPLC/MS-MS to measure the STS dependent estrogenic activity of CEE in HepG2 cells and human primary hepatocytes (HPH). The effect of STS on the estrogenic activity of CEE was also valuated in vivo by using the STS transgenic mice. Our results showed that the activity of ERE reporter was induced by CEE in a STS-dependent manner, because the estrogenic activity of CEE was attenuated by genetic or pharmacological inhibition of STS. In HepG2 cells and HPH, the estrogenic activity of CEE was enhanced by inflammation via the induction of endogenous STS gene expression. The inflammatory responsive estrogenic activity of CEE in return can attenuate inflammation due to the anti-inflammatory activity of estrogens. In vivo, STS transgenic mice exhibited markedly increased sensitivity to CEE-induced estrogenic activity in the uterus. In conclusion, our results revealed an essential role of STS in mediating the hormone replacement activity of CEE. Cautions need to apply when CEE is used in patients of chronic inflammatory disease, because these patients may have heightened sensitivity to CEE due to the inflammatory induction of STS. Unless otherwise noted, all abstracts presented at ENDO are embargoed until the date and time of presentation. For oral presentations, the abstracts are embargoed until the session begins. Abstracts presented at a news conference are embargoed until the date and time of the news conference. The Endocrine Society reserves the right to lift the embargo on specific abstracts that are selected for promotion prior to or during ENDO.

Hemorrhagic shock (HS) is a life-threatening condition associated with tissue hypoperfusion, and often leads to injury of multiple organs including the liver. Pregnane X receptor (PXR) is a species-specific xenobiotic receptor that regulates the expression of drug-metabolizing enzymes (DMEs) such as the cytochrome P450 3A (CYP3A). Many clinical drugs, including those often prescribed to trauma patients, are known to activate PXR and induce CYP3A. The goal of this study is to determine whether PXR plays a role in the regulation of DMEs in the setting of HS, and whether activation of PXR is beneficial or detrimental to HS-induced hepatic injury. PXR transgenic, knockout, and humanized mice were subject to HS, and the liver injury was assessed histologically and biochemically. The expression and/or activity of PXR and CYP3A were manipulated genetically or pharmacologically in order to determine their effects on HS-induced liver injury. Our results showed that genetic or pharmacological activation of PXR sensitized wild-type and hPXR/CYP3A4 humanized mice to HS-induced hepatic injury, whereas knockout of PXR protected mice from HS-induced liver injury. Mechanistically, the sensitizing effect of PXR activation was accounted for by PXR-responsive induction of CYP3A and increased oxidative stress in the liver. The sensitizing effect of PXR was attenuated by ablation or pharmacological inhibition of CYP3A, treatment with the antioxidant N-acetylcysteine amide (NACA), or treatment with a PXR antagonist. CONCLUSIONS: We have uncovered a novel function of PXR in HS-induced hepatic injury. Our results suggest that the unavoidable use of PXR-activating drugs in trauma patients has the potential to exacerbate HS-induced hepatic injury, which can be mitigated by the co-administration of anti-oxidative agents, CYP3A inhibitors, or PXR antagonists.

The common complications in obesity and type 2 diabetes include hepatic steatosis and disruption of glucose-glycogen homeostasis, leading to hyperglycemia. Fatty acid translocase (FAT/CD36), whose expression is inducible in obesity, is known for its function in fatty acid uptake. Previous work by us and others suggested that CD36 plays an important role in hepatic lipid homeostasis, but the results have been conflicting and the mechanisms were not well understood. In this study, by using CD36-overexpressing transgenic (CD36Tg) mice, we uncovered a surprising function of CD36 in regulating glycogen homeostasis. Overexpression of CD36 promoted glycogen synthesis, and as a result, CD36Tg mice were protected from fasting hypoglycemia. When challenged with a high-fat diet (HFD), CD36Tg mice showed unexpected attenuation of hepatic steatosis, increased very low-density lipoprotein (VLDL) secretion, and improved glucose tolerance and insulin sensitivity. The HFD-fed CD36Tg mice also showed decreased levels of proinflammatory hepatic prostaglandins and 20-hydroxyeicosatetraenoic acid (20-HETE), a potent vasoconstrictive and proinflammatory arachidonic acid metabolite. We propose that CD36 functions as a protective metabolic sensor in the liver under lipid overload and metabolic stress. CD36 may be explored as a valuable therapeutic target for the management of metabolic syndrome.

Liver X receptors (LXRs) were identified as receptors that sense oxidized cholesterol derivatives. LXRs are best known for their hepatic functions in regulating cholesterol metabolism and triglyceride synthesis, but whether and how LXRs play a role in the lung diseases is less understood. To study the function of LXRs in acute respiratory distress syndrome (ARDS), we applied the oleic acid (OA) model of ARDS to mice whose LXR was genetically or pharmacologically activated. The VP-LXRα knock-in (LXR-KI) mice, in which a constitutively activated LXRα (VP-LXRα) was inserted into the mouse LXRα locus, were used as the genetic gain-of-function model. We showed that the OA-induced lung damages, including the cytokine levels and total cell numbers and neutrophil numbers in the bronchoalveolar lavage fluid, the wet/dry weight ratio, and morphological abnormalities were reduced in the LXR-KI mice and wild-type mice treated with the LXR agonist GW3965. The pulmonoprotective effect of GW3965 was abolished in the LXR-null mice. Consistent with the pulmonoprotective effect of LXR and the induction of antioxidant enzymes by LXR, the OA-induced suppression of superoxide dismutase and catalase was attenuated in LXR-KI mice and GW3965-treated wild-type mice. Taken together, our results demonstrate that activation of LXRs can alleviate OA-induced ARDS by attenuating the inflammatory response and enhancing antioxidant capacity.

2,3,7,8-tetrachlorodibenzo-p-dioxin (TCDD), or dioxin, is a potent liver cancer promoter through its sustained activation of the aryl hydrocarbon receptor (Ahr) in rodents. However, the carcinogenic effect of TCDD and AHR in humans has been controversial. It has been suggested that the inter-species difference in the carcinogenic activity of AhR is largely due to different ligand affinity in that TCDD has a 10-fold lower affinity for the human AHR compared with the mouse Ahr. It remains unclear whether the activation of human AHR is sufficient to promote hepatocellular carcinogenesis. The goal of this study is to clarify whether activation of human AHR can promote hepatocarcinogenesis. Here we reported the oncogenic activity of human AHR in promoting hepatocellular carcinogenesis. Constitutive activation of the human AHR in transgenic mice was as efficient as its mouse counterpart in promoting diethylnitrosamine (DEN)-initiated hepatocellular carcinogenesis. The growth arrest and DNA damage-inducible gene 45 β (Gadd45b), a signaling molecule inducible by external stress and UV irradiation, is highly induced upon AHR activation. Further analysis revealed that Gadd45b is a novel AHR target gene and a transcriptional coactivator of AHR. Interestingly, ablation of Gadd45b in mice did not abolish the tumor promoting effects of the human AHR. Collectively, our findings suggested that constitutive activation of human AHR was sufficient to promote hepatocarcinogenesis.

The constitutive androstane receptor (CAR) and peroxisome proliferator-activated receptor gamma coactivator-1α (PGC1α) are master regulators of drug metabolism and gluconeogenesis, respectively. In supporting the cross talk between drug metabolism and energy metabolism, activation of CAR has been shown to suppress hepatic gluconeogenesis and ameliorate hyperglycemia in vivo, but the underlying molecular mechanism remains elusive. In this study, we demonstrated that CAR suppressed hepatic gluconeogenic gene expression through posttranslational regulation of the subcellular localization and degradation of PGC1α. Activated CAR translocated into the nucleus and served as an adaptor protein to recruit PGC1α to the Cullin1 E3 ligase complex for ubiquitination. The interaction between CAR and PGC1α also led to their sequestration within the promyelocytic leukemia protein-nuclear bodies, where PGC1α and CAR subsequently underwent proteasomal degradation. Taken together, our findings revealed an unexpected function of CAR in recruiting an E3 ligase and targeting the gluconeogenic activity of PGC1α. Both drug metabolism and gluconeogenesis are energy-demanding processes. The negative regulation of PGC1α by CAR may represent a cellular adaptive mechanism to accommodate energy-restricted conditions.

Estrogen sulfotransferase (EST) regulates estrogen homeostasis by sulfonating and deactivating estrogens. Liver ischemia and reperfusion (I/R) involves both hypoxia during the ischemic phase and oxidative damage during the reperfusion phase. In this report, we showed that the expression of EST was markedly induced by I/R. Mechanistically, oxidative stress-induced activation of Nrf2 was responsible for the EST induction, which was abolished in Nrf2−/− mice. EST is a direct transcriptional target of Nrf2. In female mice, the I/R-responsive induction of EST compromised estrogen activity. EST ablation attenuated I/R injury as a result of decreased estrogen deprivation, whereas this benefit was abolished upon ovariectomy. The effect of EST ablation was sex-specific because the EST−/− males showed heightened I/R injury. Reciprocally, both estrogens and EST regulate the expression and activity of Nrf2. Estrogen deprivation by ovariectomy abolished the I/R-responsive Nrf2 accumulation, whereas the compromised estrogen deprivation in EST−/− mice was associated with increased Nrf2 accumulation. Our results suggested a novel I/R-responsive feedback mechanism to limit the activity of Nrf2 in which Nrf2 induces the expression of EST, which subsequently increases estrogen deactivation and limits the estrogen-responsive activation of Nrf2. Inhibition of EST, at least in females, may represent an effective approach to manage hepatic I/R injury. Background: EST regulates estrogen homeostasis by sulfonating and deactivating estrogens. Results: Liver ischemia and reperfusion (I/R) induced the expression of EST and comprised estrogen activity in an Nrf2-dependent manner. EST ablation gender-specifically affected I/R sensitivity. Conclusion: EST is an oxidative stress responsive gene that affects liver I/R injury. Significance: Inhibition of EST, at least in females, represents an effective approach to manage hepatic I/R injury.

The nuclear receptor farnesoid X receptor (FXR) (nuclear receptor subfamily 1, group H, member 4, or NR1H4) is highly expressed in the liver and intestine. Previous reports have suggested beneficial functions of FXR in the homeostasis of bile acids, lipids, and glucose, as well as in promoting liver regeneration and inhibiting carcinogenesis. To investigate the effect of chronic FXR activation in vivo, we generated transgenic mice that conditionally and tissue specifically express the activated form of FXR in the liver and intestine. Unexpectedly, the transgenic mice showed several intriguing phenotypes, including partial neonatal lethality, growth retardation, and spontaneous liver toxicity. The transgenic mice also displayed heightened sensitivity to a high-cholesterol diet-induced hepatotoxicity but resistance to the gallstone formation. The phenotypes were transgene specific, because they were abolished upon treatment with doxycycline to silence the transgene expression. The perinatal toxicity, which can be rescued by a maternal vitamin supplement, may have resulted from vitamin deficiency due to low biliary bile acid output as a consequence of inhibition of bile acid formation. Our results also suggested that the fibroblast growth factor-inducible immediate-early response protein 14 (Fn14), a member of the proinflammatory TNF family, is a FXR-responsive gene. However, the contribution of Fn14 induction in the perinatal toxic phenotype of the transgenic mice remains to be defined. Because FXR is being explored as a therapeutic target, our results suggested that a chronic activation of this nuclear receptor may have an unintended side effect especially during the perinatal stage.

The aryl hydrocarbon receptor (AHR), also known as the dioxin receptor, was originally characterized as a xenobiotic receptor that senses xenotoxicants. Here we investigated the endobiotic and hepatic role of AHR in fatty liver and energy metabolism, and identified the endocrine factor that mediates the metabolic function of AHR. Wild type and liver-specific constitutively activated human AHR transgenic (TG) mice were used to investigate the role of AHR in fatty liver and energy homeostasis. Adenovirus expressing short hairpin RNA targeting the fibroblast growth factor 21 (FGF21) were used to determine the involvement of FGF21 in the metabolic effect of AHR. We showed that despite their severe fatty liver, the TG mice were protected from diet-induced obesity and type 2 diabetes. We identified the endocrine hormone FGF21 as a mediator for the metabolic benefit of AHR and established FGF21 as a direct transcriptional target of AHR. Interestingly, the transactivation of FGF21 by AHR contributed to both hepatic steatosis and systemic insulin hypersensitivity, both of which were largely abolished upon FGF21 knockdown. Conclusions The AHR-FGF21 endocrine signaling pathway establishes AHR as a pivotal environmental modifier that integrates signals from chemical exposure in the regulation of lipid and energy metabolism.

The cholesterol sulfotransferase SULT2B1b converts cholesterol to cholesterol sulfate (CS). We previously reported that SULT2B1b inhibits hepatic gluconeogenesis by antagonizing the gluconeogenic activity of hepatocyte nuclear factor 4α (HNF4α). In this study, we showed that the SULT2B1b gene is a transcriptional target of HNF4α, which led to our hypothesis that the induction of SULT2B1b by HNF4α represents a negative feedback to limit the gluconeogenic activity of HNF4α. Indeed, downregulation of Sult2B1b enhanced the gluconeogenic activity of HNF4α, which may have been accounted for by the increased acetylation of HNF4α as a result of decreased expression of the HNF4α deacetylase sirtuin 1 (Sirt1). The expression of Sult2B1b was also induced by HNF4α upon fasting, and the Sult2B1b null (Sult2B1b-/-) mice showed increased gluconeogenic gene expression and an elevated fasting glucose level, suggesting that SULT2B1b also plays a restrictive role in HNF4α-mediated fasting-responsive gluconeogenesis. We also developed thiocholesterol, a hydrolysis-resistant derivative of CS, which showed superior activity to that of the native CS in inhibiting gluconeogenesis and improving insulin sensitivity in high-fat-diet-induced diabetic mice. We conclude that the HNF4α-SULT2B1b-CS axis represents a key endogenous mechanism to prevent uncontrolled gluconeogenesis. Thiocholesterol may be used as a therapeutic agent to manage hyperglycemia.

Background and Aim: Sulfotransferase (SULT)-mediated sulfation and steroid sulfatase (STS)-mediated desulfation represent two critical mechanisms that regulate the chemical and functional homeostasis of endogenous and exogenous molecules. STS catalyzes the hydrolysis of steroid sulfates to form hydroxysteroids. Oxygenated cholesterol derivative oxysterols are known to be endogenous ligands of the liver X receptor (LXR), a nuclear receptor with anti-cholestasis activity, whereas the sulfated oxysterols antagonize LXR signaling. The conversion of sulfated oxysterols to their non-sulfated counterparts is catalyzed by STS. The aim of this study is to determine whether STS can alleviate cholestasis by increasing the activity of LXR. Methods: Liver-specific STS transgenic mice were created and subject to the lithocholic acid (LCA)-induced model of cholestasis. Results: Transgenic overexpression of STS in the liver promoted bile acid elimination and alleviated LCA-induced cholestasis. The protective effect of the STS transgene was associated with the activation of LXR and induction of LXR target genes, likely because of the increased conversion of the antagonistic oxysterol sulfates to the agonistic oxysterols. Conclusions: STS has a novel function in controlling the homeostasis of bile acids by regulating endogenous LXR ligands. Keywords: Steroid sulfatase, Bile acid, Toxicity, Cholestasis, Liver X receptor

Acute kidney injury (AKI) is associate with high mortality. Despite evidence of AKI-induced distant organ injury, a relationship between AKI and liver injury has not been clearly established. The goal of this study is to investigate whether renal ischemia-reperfusion (IR) can affect liver pathophysiology. We showed that renal IR in mice induced fatty liver and compromised liver function through the downregulation of constitutive androstane receptor (CAR; −90.4%) and inhibition of hepatic very-low-density lipoprotein triglyceride (VLDL-TG) secretion (−28.4%). Treatment of mice with the CAR agonist 1,4-bis[2-(3,5 dichloropyridyloxy)] benzene (TCPOBOP) prevented the development of AKI-induced fatty liver and liver injury, which was associated with the attenuation of AKI-induced inhibition of VLDL-TG secretion. The hepatoprotective effect of TCPOBOP was abolished in CAR(−/−) mice. Interestingly, alleviation of fatty liver by TCPOBOP also improved the kidney function, whereas CAR ablation sensitized mice to AKI-induced kidney injury and lethality. The serum concentrations of interleukin-6 (IL-6) were elevated by 27-fold after renal IR, but were normalized in TCPOBOP-treated AKI mice, suggesting that the increased release of IL-6 from the kidney may have mediated the AKI responsive liver injury. Taken together, our results revealed an interesting kidney-liver organ cross-talk in response to AKI. Given the importance of CAR in the pathogenesis of renal IR–induced fatty liver and impaired kidney function, fatty liver can be considered as an important risk factor for kidney injury, and a timely management of hepatic steatosis by CAR activation may help to restore kidney function in patients with AKI or kidney transplant.

Nonalcoholic fatty liver disease (NAFLD) is a prevalent chronic liver disease. The incidence of NAFLD has increased steadily due to its close association with the global epidemic of obesity and type 2 diabetes. However, there is no effective pharmacological therapy approved for NAFLD. Farnesoid X receptor (FXR), a member of the nuclear receptor subfamily, plays important roles in maintaining the homeostasis of bile acids, glucose, and lipids. FXR agonists have shown promise for the treatment of NAFLD. In this study, we report altenusin (2076A), a natural nonsteroidal fungal metabolite, as a novel selective agonist of FXR with an EC50 value of 3.2 ± 0.2 μM. Administration of 2076A protected mice from high-fat diet (HFD)-induced obesity by reducing the body weight and fat mass by 22.9% and 50.0%, respectively. Administration of 2076A also decreased the blood glucose level from 178.3 ± 12.4 mg/dl to 116.2 ± 4.1 mg/dl and the serum insulin level from 1.4 ± 0.6 ng/dl to 0.4 ± 0.1 ng/dl. Moreover, 2076A treatment nearly reversed HFD-induced hepatic lipid droplet accumulation and macrovesicular steatosis. These metabolic effects were abolished in FXR knockout mice. Mechanistically, the metabolic benefits of 2076A might have been accounted for by the increased insulin sensitivity and suppression of genes that are involved in hepatic gluconeogenesis and lipogenesis. In summary, we have uncovered a new class of nonsteroidal FXR agonist that shows promise in treating NAFLD and the associated metabolic syndrome.

The steroid sulfatase (STS)-mediated desulfation is a critical metabolic mechanism that regulates the chemical and functional homeostasis of endogenous and exogenous molecules. In this report, we first showed that the liver expression of Sts was induced in both the high fat diet (HFD) and ob/ob models of obesity and type 2 diabetes and during the fed to fasting transition. In defining the functional relevance of STS induction in metabolic disease, we showed that overexpression of STS in the liver of transgenic mice alleviated HFD and ob/ob models of obesity and type 2 diabetes, including reduced body weight, improved insulin sensitivity, and decreased hepatic steatosis and inflammation. Interestingly, STS exerted its metabolic benefit through sex-specific mechanisms. In female mice, STS may have increased hepatic estrogen activity by converting biologically inactive estrogen sulfates to active estrogens and consequently improved the metabolic functions, whereas ovariectomy abolished this protective effect. In contrast, the metabolic benefit of STS in males may have been accounted for by the male-specific decrease of inflammation in white adipose tissue and skeletal muscle as well as a pattern of skeletal muscle gene expression that favors energy expenditure. The metabolic benefit in male STS transgenic mice was retained after castration. Treatment with the STS substrate estrone sulfate also improved metabolic functions in both the HFD and ob/ob models. Our results have uncovered a novel function of STS in energy metabolism and type 2 diabetes. Liver-specific STS induction or estrogen/estrogen sulfate delivery may represent a novel approach to manage metabolic syndrome.

To investigate the effects of three natural product compounds, carapin, santonin and isokobusone, on the activity of pregnane X receptor (PXR) and constitutive androstane receptor (CAR) in induction of drug-metabolizing enzymes and inhibition of inflammation.The monkey kidney-derived fibroblast (CV-1) cells and human embryonic kidney HEK293 cells were used for transient transfection and luciferase reporter gene assays. Human primary hepatocytes and primary hepatocytes from wild type, PXR-/-, and hPXR transgenic mice were used to study the induction of drug-metabolizing enzymes and the implication of these compounds in inflammation.Carapin, santonin and isokobusone activated both PXR and CAR in transient transfection and luciferase reporter gene assays. Mutagenesis studies showed that two amino acid residues, Phe305 of the rodent PXR and Leu308 of the human PXR, are critical for the recognition of these compounds by PXR. Importantly, the activation of PXR and CAR by these compounds induced the expression of drug-metabolizing enzymes in primary human and mouse hepatocytes. Furthermore, activation of PXR by these compounds inhibited the expression of inflammatory mediators in response to lipopolysaccharide (LPS). The effects of these natural compounds on drug metabolism and inflammation were abolished in PXR-/- hepatocytes.Our results show that carapin, santonin and isokobusone activate PXR and CAR and induce drug-metabolizing enzymes. In addition, these compounds inhibited the expression of inflammatory mediators in response to LPS through the activation of PXR.

Chronic hepatitis B virus (HBV) infection is a major risk factor for hepatocellular carcinoma (HCC). Hepatitis B virus X protein (HBx) is a HBV protein that has multiple cellular functions, but its role in HCC pathogenesis has been controversial. The farnesoid X receptor (FXR) is a nuclear receptor known to have activities in anti-inflammation and inhibition of hepatocarcinogenesis. However, whether or how FXR can impact HBV/HBx-induced hepatocarcinogenesis remains unclear. In this study, we showed that HBx can interact with FXR and function as a co-activator of FXR. Expression of HBx in vivo enhanced FXR responsive gene regulation. HBx also increased the transcriptional activity of FXR in luciferase reporter gene assay. The HBx-FXR interaction was confirmed by co-immunoprecipitation and GST-pull down assays, and the FXR AF-1 domain was mapped to bind to the third α helix in the C-terminal of HBx. We also found that the C-terminal truncated variants of HBx, which were found in clinical HCC, were not effective in transactivating FXR. Interestingly, recruitment of the full-length HBx, but not the C-terminal truncated HBx, enhanced the binding of FXR to its response element. In vivo, FXR ablation markedly sensitized mice to HBx-induced hepatocarcinogenesis. Conclusions: We propose that transactivation of FXR by the full-length HBx may represent a protective mechanism to inhibit HCC, and this inhibition may have been compromised upon the appearance of the C-terminal truncated HBx, or when the expression and/or activity of FXR is decreased. This article is protected by copyright. All rights reserved.

Estrogen sulfotransferase (EST), the enzyme responsible for the sulfonation and inactivation of estrogens, plays an important role in estrogen homeostasis. In this study, we showed that induction of hepatic Est is a common feature of type 2 diabetes. Loss of Est in female mice improved metabolic function in ob/ob, dexamethasone-, and high-fat diet–induced mouse models of type 2 diabetes. The metabolic benefit of Est ablation included improved body composition, increased energy expenditure and insulin sensitivity, and decreased hepatic gluconeogenesis and lipogenesis. This metabolic benefit appeared to have resulted from decreased estrogen deprivation and increased estrogenic activity in the liver, whereas such benefit was abolished in ovariectomized mice. Interestingly, the effect of Est was sex-specific, as Est ablation in ob/ob males exacerbated the diabetic phenotype, which was accounted for by the decreased islet β-cell mass and failure of glucose-stimulated insulin secretion in vivo. The loss of β-cell mass in ob/ob males deficient in Est was associated with increased macrophage infiltration and inflammation in white adipose tissue. Our results revealed an essential role of EST in energy metabolism and the pathogenesis of type 2 diabetes. Inhibition of EST, at least in females, may represent a novel approach to manage type 2 diabetes.

Background & Aims Cholesterol gallstone disease (CGD) results from a biochemical imbalance of lipids and bile salts in the gallbladder bile. We investigated whether the xenobiotic receptor pregnane X receptor (PXR) has a role in pathogenesis of CGD. Methods Wild-type, PXR-null ( PXR −/− ), and CGD-sensitive C57L mice were placed on a lithogenic diet and then analyzed for CGD at the biochemical, histological, and gene-regulation levels. Results Loss of PXR sensitized mice to lithogenic diet-induced CGD, characterized by decreases in biliary concentrations of bile salts and phospholipids and an increases in the cholesterol saturation index and formation of cholesterol crystals. The decreased bile acid pool size in PXR −/− mice that received lithogenic diets was associated with reduced expression of cholesterol 7α-hydroxylase, the rate-limiting enzyme of cholesterol catabolism and bile acid formation. The reduced expression of cholesterol 7α-hydroxylase most likely resulted from activation of farnesoid X receptor and induction of fibroblast growth factor 15 in the intestine. In C57L mice given the PXR agonist, pregnenolone-16α-carbonitrile, or the herbal medicine, St John's wort, cholesterol precipitation was prevented by increases in concentrations of biliary bile salt and a reduced cholesterol saturation index. PXR prevented CGD via its coordinate regulation of the biosynthesis and transport of bile salts in the liver and intestine. Conclusions PXR maintains biliary bile acid homeostasis and may be developed as a therapeutic target for CGD.

Sepsis is the host's deleterious systemic inflammatory response to microbial infections. Here we report an essential role for the oestrogen sulfotransferase (EST or SULT1E1), a conjugating enzyme that sulfonates and deactivates estrogens, in sepsis response. Both the caecal ligation and puncture (CLP) and lipopolysaccharide models of sepsis induce the expression of EST and compromise the activity of oestrogen, an anti-inflammatory hormone. Surprisingly, EST ablation sensitizes mice to sepsis-induced death. Mechanistically, EST ablation attenuates sepsis-induced inflammatory responses due to compromised oestrogen deactivation, leading to increased sepsis lethality. In contrast, transgenic overexpression of EST promotes oestrogen deactivation and sensitizes mice to CLP-induced inflammatory response. The induction of EST by sepsis is NF-κB dependent and EST is a NF-κB-target gene. The reciprocal regulation of inflammation and EST may represent a yet-to-be-explored mechanism of endocrine regulation of inflammation, which has an impact on the clinical outcome of sepsis.

Background & Aims Fatty acid binding protein 4 (FABP4) has been known as a mediator of inflammatory response in the macrophages and adipose tissue, but its hepatic function is poorly understood. The goal of this study is to investigate the role of FABP4 in liver ischemia/reperfusion (I/R), a clinical condition that involves both hypoxia and inflammation. Methods To examine the I/R regulation of FABP4, mice were subjected to I/R surgery before being measured for FABP4 gene expression. Both loss-of-function (by using a pharmacological FABP4 inhibitor) and gain-of-function (by adenoviral overexpression of FABP4) were used to determine the functional relevance of FABP4 expression and its regulation during I/R. To determine the hypoxia responsive regulation of FABP4, primary mouse hepatocytes were exposed to hypoxia. The FABP4 gene promoter was cloned and its regulation by hypoxia inducible factor 1α (HIF-1α) was characterized by luciferase reporter gene, electrophoretic mobility shift, and chromatin immunoprecipitation assays. Results We found that the hepatic expression of FABP4 was markedly induced by I/R. At the functional level, pharmacological inhibition of FABP4 alleviated the I/R injury, whereas adenoviral overexpression of FABP4 sensitized mice to I/R injury. We also showed that exposure of primary hepatocytes to hypoxia or transgenic overexpression of HIF-1α in the mouse liver was sufficient to induce the expression of FABP4. Our promoter analysis established FABP4 as a novel transcriptional target of HIF-1α. Conclusions FABP4 is a hypoxia inducible gene that sensitizes mice to liver I/R injury. FABP4 may represent a novel therapeutic target, and FABP4 inhibitors may be used as therapeutic agents to manage hepatic I/R injury.

The nuclear receptor liver X receptor (LXR) plays an important role in the metabolism and homeostasis of cholesterol, lipids, bile acids, and steroid hormones. In this study, we uncovered a function of LXRα (NR1H3) in regulating the human hydroxysteroid sulfotransferase SULT2A1, a phase II conjugating enzyme known to sulfonate bile acids, hydroxysteroid dehydroepiandrosterone, and related androgens. We showed that activation of LXR induced the expression of SULT2A1 at mRNA, protein, and enzymatic levels. A combination of promoter reporter gene and chromatin immunoprecipitation assays showed that LXRα transactivated the SULT2A1 gene promoter through its specific binding to the −500- to −258-base pair region of the SULT2A1 gene promoter. LXR small interfering RNA knockdown experiments suggested that LXRα, but not LXRβ, played a dominant role in regulating SULT2A1. In primary human hepatocytes, we found a positive correlation between the expression of SULT2A1 and LXRα, which further supported the regulation of SULT2A1 by LXRα. In summary, our results established human SULT2A1 as a novel LXRα target gene. The expression of LXRα is a potential predictor for the expression of SULT2A1 in human liver.

Sulfotransferase (SULT)-mediated sulfation represents a critical mechanism in regulating the chemical and functional homeostasis of endogenous and exogenous molecules. The cholesterol sulfotransferase SULT2B1b catalyzes the sulfoconjugation of cholesterol to synthesize cholesterol sulfate (CS). In this study, we showed that the expression of SULT2B1b in the liver was induced in obese mice and during the transition from the fasted to the fed state, suggesting that the regulation of SULT2B1b is physiologically relevant. CS and SULT2B1b inhibited gluconeogenesis by targeting the gluconeogenic factor hepatocyte nuclear factor 4α (HNF4α) in both cell cultures and transgenic mice. Treatment of mice with CS or transgenic overexpression of the CS-generating enzyme SULT2B1b in the liver inhibited hepatic gluconeogenesis and alleviated metabolic abnormalities both in mice with diet-induced obesity (DIO) and in leptin-deficient (ob/ob) mice. Mechanistically, CS and SULT2B1b inhibited gluconeogenesis by suppressing the expression of acetyl coenzyme A (acetyl-CoA) synthetase (Acss), leading to decreased acetylation and nuclear exclusion of HNF4α. Our results also suggested that leptin is a potential effector of SULT2B1b in improving metabolic function. We conclude that SULT2B1b and its enzymatic by-product CS are important metabolic regulators that control glucose metabolism, suggesting CS as a potential therapeutic agent and SULT2B1b as a potential therapeutic target for metabolic disorders.