Bile acids (BA) are signalling molecules which activate the transmembrane receptor TGR5 and the nuclear receptor FXR. BA sequestrants (BAS) complex BA in the intestinal lumen and decrease intestinal FXR activity. The BAS-BA complex also induces Glucagon-Like Peptide-1 (GLP-1) production by L-cells which potentiates β-cell glucose-induced insulin secretion. Whether FXR is expressed in L-cells and controls GLP-1 production is unknown. Here we show that FXR activation in L-cells decreases proglucagon expression by interfering with the glucose-responsive factor Carbohydrate-Responsive Element Binding Protein (ChREBP) and GLP-1 secretion by inhibiting glycolysis. In vivo, FXR-deficiency increases GLP-1 gene expression and secretion in response to glucose hence improving glucose metabolism. Moreover, treatment of ob/ob mice with the BAS colesevelam increases intestinal proglucagon gene expression and improves glycemia in a FXR-dependent manner. These findings identify the FXR/GLP-1 pathway as a new mechanism of BA control of glucose metabolism and a pharmacological target for type 2 diabetes.

OBJECTIVEBile acids (BA) participate in the maintenance of metabolic homeostasis acting through different signaling pathways. The nuclear BA receptor farnesoid X receptor (FXR) regulates pathways in BA, lipid, glucose, and energy metabolism, which become dysregulated in obesity. However, the role of FXR in obesity and associated complications, such as dyslipidemia and insulin resistance, has not been directly assessed.RESEARCH DESIGN AND METHODSHere, we evaluate the consequences of FXR deficiency on body weight development, lipid metabolism, and insulin resistance in murine models of genetic and diet-induced obesity.RESULTSFXR deficiency attenuated body weight gain and reduced adipose tissue mass in both models. Surprisingly, glucose homeostasis improved as a result of an enhanced glucose clearance and adipose tissue insulin sensitivity. In contrast, hepatic insulin sensitivity did not change, and liver steatosis aggravated as a result of the repression of β-oxidation genes. In agreement, liver-specific FXR deficiency did not protect from diet-induced obesity and insulin resistance, indicating a role for nonhepatic FXR in the control of glucose homeostasis in obesity. Decreasing elevated plasma BA concentrations in obese FXR-deficient mice by administration of the BA sequestrant colesevelam improved glucose homeostasis in a FXR-dependent manner, indicating that the observed improvements by FXR deficiency are not a result of indirect effects of altered BA metabolism.CONCLUSIONSOverall, FXR deficiency in obesity beneficially affects body weight development and glucose homeostasis.

. We report here that peripheral blood mononuclear cells (PBMC) from individuals with the CX3CR1-IM haplotype adhered more potently to membrane-bound CX3CL1 than did PBMC from homozygous CX3CR1-VT donors. Similar excess adhesion was observed with CX3CR1-IMtransfected human embryonic kidney (HEK) cell lines tested with two different methods: the parallel plate laminar flow chamber and the dual pipette aspiration technique. Suppression of the extra adhesion in the presence of pertussis toxin indicates that G-protein mediated the underlying transduction pathway, in contrast to the G-protein-independent adhesion previously described for CX3CR1-VT. Surprisingly, HEK and PBMC that expressed CX3CR1-IM and -VT were indistinguishable when tested with the soluble form of CX3CL1 for chemotaxis, calcium release, and binding capacity. In conclusion, only the membrane-anchored form of CX3CL1 functionally discriminated between these two allelic isoforms of CX3CR1. These results suggest that each form of this ligand may lead to a different signaling pathway. The extra adhesion of CX3CR1-IM may be related to immune defenses and to atherogenesis, both of which depend substantially on adhesive intercellular events.Adhesion, a critical stage in cell trafficking and migration (1, 2), requires the presence of numerous adhesion molecules, such as integrins, that need divalent ions to function. Recent studies show that two chemokines, namely, CX3CL1 and CXCL16, are not only chemoattractant as soluble molecules, but also function as adhesion molecules since they are membrane-anchored, regardless of the presence of divalent ions (3-7). The best known of these is CX3CL1, also called fractalkine. It is expressed on the surface of many types of cells, in particular interleukin-1-and tumor necrosis factor-activated endothelial (4) and dendritic cells (8), as a membrane molecule containing the classic chemokine domain, a mucin-like stalk, and a transmembrane domain tethering it to the cell membrane. CX3CL1 may be cleaved by TACE and released from cells (9, 10). In this soluble form, it behaves like a chemoattractant molecule, just as other chemokines do. The transmembrane feature of the native CX3CL1 protein, combining it with its receptor, CX3CR1, produces a strongly adhesive pair (4, 5, 11) that mediates the rapid capture and firm adhesion of leukocytes. Because this activity persists in the absence of divalent cations, it is thought to be independent of integrins (5,11,12). This adhesive feature is also independent of the G i pathway, since it is still present after pertussis toxin (PTX) 1 treatment (4, 5, 11). The CX3CR1 molecule is expressed on leukocytes, especially monocytes (4) and cytotoxic cells (13,14), on dendritic cells (15), and on neurons and microglial cells (16,17). Recently, we identified two common polymorphisms in strong linkage disequilibrium in the CX3CR1 gene: V249I and T280M (18). We also found that these mutations are associated with more rapid progression to AIDS (18,19), although two studies have failed to confirm t...

Gastro-intestinal exclusion by Roux-en-Y gastric bypass (RYGB) improves glucose metabolism, independent of weight loss. Although changes in intestinal bile trafficking have been shown to play a role, the underlying mechanisms are unclear. We performed RYGB in minipigs and showed that the intestinal uptake of ingested glucose is blunted in the bile-deprived alimentary limb (AL). Glucose uptake in the AL was restored by the addition of bile, and this effect was abolished when active glucose intestinal transport was blocked with phlorizin. Sodium-glucose cotransporter 1 remained expressed in the AL, while intraluminal sodium content was markedly decreased. Adding sodium to the AL had the same effect as bile on glucose uptake. It also increased postprandial blood glucose response in conscious minipigs following RYGB. The decrease in intestinal uptake of glucose after RYGB was confirmed in humans. Our results demonstrate that bile diversion affects postprandial glucose metabolism by modulating sodium-glucose intestinal cotransport.

Rationale: In atherosclerotic plaques, iron preferentially accumulates in macrophages where it can exert pro-oxidant activities. Objective: The objective of this study was, first, to better characterize the iron distribution and metabolism in macrophage subpopulations in human atherosclerotic plaques and, second, to determine whether iron homeostasis is under the control of nuclear receptors, such as the liver X receptors (LXRs). Methods and Results: Here we report that iron depots accumulate in human atherosclerotic plaque areas enriched in CD68 and mannose receptor (MR)-positive (CD68 + MR + ) alternative M2 macrophages. In vitro IL-4 polarization of human monocytes into M2 macrophages also resulted in a gene expression profile and phenotype favoring iron accumulation. However, M2 macrophages on iron exposure acquire a phenotype favoring iron release, through a strong increase in ferroportin expression, illustrated by a more avid oxidation of extracellular low-density lipoprotein by iron-loaded M2 macrophages. In line, in human atherosclerotic plaques, CD68 + MR + macrophages accumulate oxidized lipids, which activate LXRα and LXRβ, resulting in the induction of ABCA1, ABCG1, and apolipoprotein E expression. Moreover, in iron-loaded M2 macrophages, LXR activation induces nuclear factor erythroid 2-like 2 expression, thereby increasing ferroportin expression, which, together with a decrease of hepcidin mRNA levels, promotes iron export. Conclusions: These data identify a role for M2 macrophages in iron handling, a process regulated by LXR activation.

The tested animal models share similar quantitative RYGBP-induced increases in peripheral blood BAs as humans, which render them useful for mechanistic studies. However, they also present qualitative differences in BA profiles, which may result in different signaling responses. Such differences need to be taken into account when translating results to humans.

Rationale: Vascular calcification is a process similar to bone formation leading to an inappropriate deposition of calcium phosphate minerals in advanced atherosclerotic plaques. Monocyte-derived macrophages, located in atherosclerotic lesions and presenting heterogeneous phenotypes, from classical proinflammatory M1 to alternative anti-inflammatory M2 macrophages, could potentially display osteoclast-like functions.Objective: To characterize the phenotype of macrophages located in areas surrounding the calcium deposits in human atherosclerotic plaques. Methods and Results: Macrophages near calcium deposits display an alternative phenotype being both CD68and mannose receptor-positive, expressing carbonic anhydrase type II, but relatively low levels of cathepsin K. In vitro interleukin-4-polarization of human primary monocytes into macrophages results in lower expression and activity of cathepsin K compared with resting unpolarized macrophages. Moreover, interleukin-4 polarization lowers expression levels of the osteoclast transcriptional activator nuclear factor of activated T cells type c-1, associated with increased gene promoter levels of the transcriptional repression mark H3K27me3 (histone 3 lysine 27 trimethylation). Despite higher expression of the receptor activator of nuclear factor κB receptor, receptor activator of nuclear factor κB ligand/macrophage colony-stimulating factor induction of nuclear factor of activated T cells type c-1 and cathepsin K expression is defective in these macrophages because of reduced Erk/cfos-mediated downstream signaling resulting in impaired bone resorption capacity. Conclusions:

In its native form, the chemokine CX3CL1 is a firmly adhesive molecule promoting leukocyte adhesion and migration and hence involved, along with its unique receptor CX3CR1, in various inflammatory processes. Here we investigated the role of molecular aggregation in the CX3CL1 adhesiveness. Assays of bioluminescence resonance energy transfer (BRET) and homogeneous time-resolved fluorescence (HTRF) in transfected cell lines and in primary cells showed specific signals indicative of CX3CL1 clustering. Truncation experiments showed that the transmembrane domain played a central role in this aggregation. A chimera with mutations of the 12 central transmembrane domain residues had significantly reduced BRET signals and characteristics of a non-clustering molecule. This mutant was weakly adhesive according to flow and dual pipette adhesion assays and was less glycosylated than CX3CL1, although, as we demonstrated, loss of glycosylation did not affect the CX3CL1 adhesive potency. We postulate that cell surfaces express CX3CL1 as a constitutive oligomer and that this oligomerization is essential for its adhesive potency. Inhibition of CX3CL1 self-assembly could limit the recruitment of CX3CR1-positive cells and may be a new pathway for anti-inflammatory therapies.Migration of circulating leukocytes to injury sites is the first step of the inflammation process, which involves a sequence of coordinated interactions between leukocytes and endothelial cells (1). Central to this physiological and pathological event are chemokines, a family of low molecular weight soluble proteins, that function to attract leukocytes bearing the appropriate receptors (2). Chemokines trigger activation of leukocytes and their firm adhesion to inflamed endothelium, mainly through the mediation of integrins and their cognate ligands (3).Among chemokines, there are two exceptions: CXCL16 and CX3CL1 are type-I membranous proteins. In addition to their chemokine domain (CD), 4 they are composed of a long mucinlike stalk, a transmembrane domain, and a cytoplasmic tail (4, 5) (Fig. 1). The CX3CL1 molecule, with its unique CX3CR1 receptor (6), has been shown to be central in defenses against neurodegenerative disorders (7,8) and against several cancers in murine models (9, 10). The CX3CL1-CX3CR1 axis is also involved in various inflammatory diseases (2), including renal inflammation (11) and atherosclerosis (12, 13). Understanding the structure of this pair of molecules is necessary for exploring pharmacological methods to regulate their activity. The structure of CX3CR1, a receptor of the G protein-coupled receptor (GPCR) family, is relatively well known, but that of CX3CL1 much less so.CX3CL1-CD (76 residues), a globular protein domain 3 nm in diameter (14) and maintained by two disulfide bridges, is structurally similar to other chemokines. It is composed of a disordered N terminus up to the first cysteine (Cys-8), a long loop followed by a three-stranded antiparallel ␤-sheet (residues 24 -51) and a C-terminal ␣-helix (residues 56 -67) packed agains...