Regulation of the ileal bile acid-binding protein gene: An approach to determine its physiological function(s)

Ileal bile acid-binding protein (I-BABP) is a soluble bile acids (BA) carrier protein which belongs to the fatty acid-binding protein (FABP) family. In the gut, its expression is strictly restricted to the ileum, where it is thought to be involved in the active BA reabsorption. Therefore, I-BABP gene expression levels might be rate limiting for the BA enterohepatic circulation, and hence, might be crucial for cholesterol (CS) homeostasis. Indeed, BA not reclaimed by intestinal absorption constitute the main way to eliminate a CS excess. However, such a function is not yet established. Because generally rate limiting genes are tightly controlled, we have undertaken the study of the I-BABP gene regulation. It was found that both BA and CS, probably via oxysterols, are able to up-regulate the trancription rate of I-BABP gene. The fact that intracellular sterol sensors (FXR, LXR and SREBP1c) are involved in the control of I-BABP gene expression strongly suggest a crucial role for I-BABP in the ileum.     

Liver X receptor alpha interferes with SREBP1c-mediated Abcd2 expression. Novel cross-talk in gene regulation

The peroxisomal ATP binding cassette (ABC) transporter adrenoleukodystrophy-related protein, encoded by ABCD2, displays functional redundancy with the X-linked adrenoleukodystrophy-associated protein, making ABCD2 up-regulation of therapeutic value. Cholesterol lowering activates human ABCD2 in cultured cells. To investigate in vivo regulation by sterols, we first characterized a sterol regulatory element (SRE) in the murine Abcd2 promoter that is directly bound by SRE-binding proteins (SREBPs). Intriguingly, this element overlaps with a direct repeat 4, which serves as binding site for liver X receptor (LXR)/retinoid X receptor heterodimers, suggesting novel cross-talk between SREBP and LXR/retinoid X receptor in gene regulation. Using fasting-refeeding and cholesterol loading, SREBP accessibility to the SRE/direct repeat 4 was tested. Results suggest that adipose Abcd2 is induced by SREBP1c, whereas hepatic Abcd2 expression is down-regulated by concurrent activation of LXRalpha and SREBP1c. In cell culture, SREBP1c-mediated Abcd2 induction is counteracted by ligand-activated LXRalpha. Finally, hepatic Abcd2 expression in LXRalpha,beta-deficient mice is inducible to levels vastly exceeding wild type. Together, we identify LXRalpha as negative modulator of Abcd2, acting through a novel regulatory mechanism involving overlapping SREBP and LXRalpha binding sites.     

LXR-alpha/SREBP-1c通路参与HCV NS5A诱导的肝细胞脂质代谢紊乱

肝细胞脂肪变性是丙型肝炎患者的突出病理特征,但丙肝病毒（HCV）诱导脂肪变性的分子机制尚不清楚.为探究HCV非结构蛋白5A（NS5A）参与诱导脂肪变性的可能分子机制,本研究以HCV NS5A转基因小鼠为研究对象,采集3～16月龄NS5A转基因小鼠和同窝非转基因小鼠的肝组织,进行病理学检测,并用气相色谱 质谱（GC-MS）法分析脂质主要成分胆固醇酯的含量.采用RT-PCR法检测肝细胞中与脂质代谢密切相关基因肝X受体（LXR-alpha）、固醇调节元件结合蛋白（SREBP-1c）、脂肪酸合成酶（FAS）、硬脂酰辅酶A去饱和酶1（SCD1）、过氧化物酶体增殖物受体alpha（PPAR-alpha）的mRNA表达水平.结果表明,与同窝非转基因对照小鼠相比,3～5月龄NS5A转基因小鼠的肝组织没有发生显著的病理性变化,但6～16月龄的NS5A转基因小鼠的肝脏发生了显著的脂肪变性（47.1% vs 130%;P=0.003）.与此相一致,胆固醇酯的含量在NS5A转基因小鼠的肝脏中显著升高（P < 0.01）.RT PCR检测结果表明,与对照小鼠相比,14月龄NS5A转基因小鼠肝组织中与脂质代谢相关的基因LXR.alpha、SREBP.1c、FAS、SCD1的mRNA表达水平显著增高（P < 0.05）,而PPAR alpha的表达则没有显著变化（P > 0.05）. 以上结果提示,NS5A在小鼠肝细胞中可能通过调高LXR.alpha/SREBP.1c信号通路相关基因的表达,进而促进脂质重新合成,诱导脂肪变性.     

Synthetic LXR agonist attenuates plaque formation in apoE-/- mice without inducing liver steatosis and hypertriglyceridemia

Liver X receptors (LXRs) are important regulators of cholesterol and lipid metabolism. LXR agonists have been shown to limit the cellular cholesterol content by inducing reverse cholesterol transport, increasing bile acid production, and inhibiting intestinal cholesterol absorption. Most of them, however, also increase lipogenesis via sterol regulatory element-binding protein-1c (SREBP1c) and carbohydrate response element-binding protein activation resulting in hypertriglyceridemia and liver steatosis. We report on the antiatherogenic properties of the steroidal liver X receptor agonist N,N-dimethyl-3beta-hydroxy-cholenamide (DMHCA) in apolipoprotein E (apoE)-deficient mice. Long-term administration of DMHCA (11 weeks) significantly reduced lesion formation in male and female apoE-null mice. Notably, DMHCA neither increased hepatic triglyceride (TG) levels in male nor female apoE-deficient mice. ATP binding cassette transporter A1 and G1 and cholesterol 7alpha-hydroxylase mRNA abundances were increased, whereas SREBP1c mRNA expression was unchanged in liver, and even decreased in macrophages and intestine. Short-term treatment revealed even higher changes on mRNA regulation. Our data provide evidence that DMHCA is a strong candidate as therapeutic agent for the treatment or prevention of atherosclerosis, circumventing the negative side effects of other LXR agonists.     

The X-ray structure of zebrafish (Danio rerio) ileal bile acid-binding protein reveals the presence of binding sites on the surface of the protein molecule

The ileal bile acid-binding proteins (I-BABPs), also called ileal lipid-binding proteins or gastrotropins, belong to the family of the fatty acid-binding proteins and play an important role in the solubilization and transport of bile acids in the enterocyte. This article describes the expression, purification, crystallization, and three-dimensional structure determination of zebrafish (Danio rerio) I-BABP both in its apo form and bound to cholic acid. This is the first X-ray structure of an I-BABP. The structure of the apoprotein was determined to a resolution of 1.6 Å, and two different monoclinic crystal forms of the holoprotein were solved and refined to 2.2 Å resolution. Three protein molecules are present in the asymmetric unit of one of the co-crystal forms and two in the other, and therefore, the results of this study refer to observations made on five different protein molecules in the crystalline state. In every case, two cholate ligands were found bound in approximately the same position in the internal cavity of the protein molecules, but an unexpected result is the presence of clear and unambiguous electron density for several cholate molecules bound on hydrophobic patches on the surface of all the five independent protein molecules examined. Isothermal titration calorimetry was used for the thermodynamic characterization of the binding mechanism and has yielded results that are consistent with the X-ray data. Ligand binding is described in detail, and the conformational changes undergone by the protein molecule in the apo-to-holo transition are examined by superposition of the apo- and holoprotein models. The structure of the holoprotein is also compared with that of the liver BABP from the same species and those of other I-BABPs determined by NMR.     

The LXR ligand T0901317 induces severe lipogenesis in the db/db diabetic mouse

Liver X receptor (LXR) ligands are currently being evaluated as potential therapeutic agents for the treatment of low HDL. The LXR ligand T0901317 elevates ATP binding cassette transporter A1 (ABCA1) and HDL levels in animal models and induces moderate lipogenesis through upregulation of sterol regulatory element binding protein 1c (SREBP1c). Because insulin may also regulate lipogenesis through SREBP1c and fatty acid synthase (FAS), we investigated the effect of an LXR ligand in hyperinsulinemic mice. Administration of T0901317 to male db/db mice for 12 days resulted in a more severe hypertriacylglycerolemia and hepatic triacylglycerol accumulation than observed in nondiabetic mice. The LXR target genes ABCA1, SREBP1c, FAS, and stearoyl-CoA desaturase 1 were upregulated by T0901317 treatment in both diabetic db/db and nondiabetic C57BLKS mice. Changes in lipogenic gene expression were independent of mouse strain, indicating that the severe lipogenesis observed in LXR ligand-treated db/db mice was not due to additive effects of insulin on lipogenic gene expression. Phosphoenolpyruvate carboxykinase expression was suppressed, suggesting that a shift from gluconeogenesis toward lipogenesis could partially explain our observations in db/db mice. Our data suggest that LXR ligands that have effects on both fatty acid and carbohydrate metabolism should be carefully evaluated in obesity, insulin, and leptin resistance.     

Nimbolide attenuate the lipid accumulation,oxidative stress and antioxidant in primary hepatocytes

Nimbolide is a bioactive compound found in Azadirachta indica. This work was devised to investigate the potential effects of nimbolide on intracellular lipid deposition and its associated redox modulation in primary hepatocytes (Heps). Lipid accumulation was induced in Heps by supplementing 1 mM oleic acid for 24 h which was marked by significant accumulation of lipids. The results demonstrated that nimbolide can decrease intracellular cholesterol, free fatty acids and triglycerides. Nimbolide may also improve hepatocytes function through its antioxidant effects by inhibiting oxidative DNA damage and lipid peroxidation by curtailing the reactive oxygen species levels. Further it also restore the mitochondrial potential, improving the endogenous antioxidant levels such as GSH and antioxidant enzyme activities. Nimbolide increased (P?<?0.05) liver X receptor-α (LXRα), peroxisome proliferator-activated receptor-γ (PPARγ) and sterol regulatory element-binding protein-1c (SREBP1c) gene expression in Heps. The biological significance of nimbolide may involve hypolipidemic effect, lipid peroxidation inhibition, DNA damage inhibition, ROS inhibition, restoring mitochondrial function, increases in GSH and SOD & CAT activities, and direct regulation of LXRα, PPARγ and SREBP1c gene expression. Nimbolide may be used as effective lipid lowering compound and lipid deposition-induced Heps changes.