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Several diseases and conditions have been associated with an uncontrolled rise in bile acid (BA) concentrations. This is often the case when the tight feedback regulation of BA synthesis is compromised to the point that BAs become detrimental. BAs and their cognate receptors, farnesoid X receptor (FXR) and Takeda G-protein receptor 5 (TGR5), however, exert many beneficial roles as they enable tissues to adapt to environmental, nutritional, and physiological cues. Over the last two decades, BA mimetics targeting FXR, TGR5, or both, have been proven to be efficacious in alleviating chronic metabolic and inflammatory disorders, such as obesity, Type 2 diabetes (T2D), atherosclerosis and non-alcoholic steatohepatitis (NASH). While several aspects of BA signaling are still poorly understood, the first therapeutics targeting FXR are making their way into the clinic to treat liver diseases, such as primary biliary cholangitis (PBC) and NASH. Drugs targeting BA signaling may, hence, have a bright future and the continuing efforts on studying the impact of changing BA signaling pathways in humans will be beneficial to translate our emerging knowledge on BA physiology in model organisms into clinical benefits.

International audience; Bile acids (BAs) improve metabolism and exert anti-obesity effects through the activation of the Takeda G protein-coupled receptor 5 (TGR5) in peripheral tissues. TGR5 is also found in the brain hypothalamus, but whether hypothalamic BA signaling is implicated in body weight control and obesity pathophysiology remains unknown. Here we show that hypothalamic BA content is reduced in diet-induced obese mice. Central administration of BAs or a specific TGR5 agonist in these animals decreases body weight and fat mass by activating the sympathetic nervous system, thereby promoting negative energy balance. Conversely, genetic downregulation of hypothalamic TGR5 expression in the mediobasal hypothalamus favors the development of obesity and worsens established obesity by blunting sympathetic activity. Lastly, hypothalamic TGR5 signaling is required for the anti-obesity action of dietary BA supplementation. Together, these findings identify hypothalamic TGR5 signaling as a key mediator of a top-down neural mechanism that counteracts diet-induced obesity.

BACKGROUND & AIMS: Renewal and patterning of the intestinal epithelium is coordinated by intestinal stem cells (ISCs); dietary and metabolic factors provide signals to the niche that control ISC activity. Bile acids (BAs), metabolites in the gut, signal nutrient availability by activating the G protein-coupled bile acid receptor 1 (GPBAR1, also called TGR5). TGR5 is expressed in the intestinal epithelium, but it is not clear how its activation affects ISCs and regeneration of the intestinal epithelium. We studied the role of BAs and TGR5 in intestinal renewal, and regulation of ISC function in mice and intestinal organoids. METHODS: We derived intestinal organoids from wild-type mice and Tgr5(-/-) mice, incubated them with BAs or the TGR5 agonist INT-777, and monitored ISC function by morphologic analyses and colony-forming assays. We disrupted Tgr5 specifically in Lgr5-positive ISCs in mice (Tgr5ISC(-/-) mice) and analyzed ISC number, proliferation, and differentiation by flow cytometry, immunofluorescence, and organoid assays. Tgr5ISC(-/-) mice were given cholecystokinin; we measured the effects of BA release into the intestinal lumen and on cell renewal. We induced colitis in Tgr5ISC(-/-) mice by administration of dextran sulfate sodium; disease severity was determined based on body weight, colon length, and histopathology analysis of colon biopsies. RESULTS: BAs and TGR5 agonists promoted growth of intestinal organoids. Administration of cholecystokinin to mice resulted in acute release of BAs into the intestinal lumen and increased proliferation of the intestinal epithelium. BAs and Tgr5 expression in ISCs were required for homeostatic intestinal epithelial renewal and fatespecification, and for regeneration after colitis induction. Tgr5ISC(-/-) mice developed more severe colitis than mice without Tgr5 disruption in ISCs. ISCs incubated with INT-777 increased activation of yes-associated protein 1 (YAP1) and of its upstream regulator SRC. Inhibitors of YAP1 and SRC prevented organoid growth induced by TGR5 activation. CONCLUSIONS: BAs promote regeneration of the intestinal epithelium via activation of TGR5 in ISCs, resulting in activation of SRC and YAP and activation of their target genes. Release of endogenous BAs in the intestinal lumen is sufficient to promote ISC renewal and drives regeneration in response to injury.

In the past decade substantial progress has been made in understanding how the insurgence of chronic low-grade inflammation influences the physiology of several metabolic diseases. Tissue-resident immune cells have been identified as central players in these processes, linking inflammation to metabolism. The bile acid-responsive G-protein-coupled receptor TGR5 is expressed in monocytes and macrophages, and its activation mediates potent anti-inflammatory effects. Herein, we summarize recent advances in TGR5 research, focusing on the downstream effector pathways that are modulated by TGR5 activators, and on its therapeutic potential in inflammatory and metabolic diseases.

Bile acids and epithelial-derived human β-defensins (HβDs) are known to be important factors in the regulation of colonic mucosal barrier function and inflammation. We hypothesized that bile acids regulate colonic HβD expression and aimed to test this by investigating the effects of deoxycholic acid (DCA) and ursodeoxycholic acid on the expression and release of HβD1 and HβD2 from colonic epithelial cells and mucosal tissues. DCA (10-150 µM) stimulated the release of both HβD1 and HβD2 from epithelial cell monolayers and human colonic mucosal tissue in vitro In contrast, ursodeoxycholic acid (50-200 µM) inhibited both basal and DCA-induced defensin release. Effects of DCA were mimicked by the Takeda GPCR 5 agonist, INT-777 (50 μM), but not by the farnesoid X receptor agonist, GW4064 (10 μM). INT-777 also stimulated colonic HβD1 and HβD2 release from wild-type, but not Takeda GPCR 5-/-, mice. DCA stimulated phosphorylation of the p65 subunit of NF-κB, an effect that was attenuated by ursodeoxycholic acid, whereas an NF-κB inhibitor, BMS-345541 (25 μM), inhibited DCA-induced HβD2, but not HβD1, release. We conclude that bile acids can differentially regulate colonic epithelial HβD expression and secretion and discuss the implications of our findings for intestinal health and disease.-Lajczak, N. K., Saint-Criq, V., O'Dwyer, A. M., Perino, A., Adorini, L., Schoonjans, K., Keely, S. J. Bile acids deoxycholic acid and ursodeoxycholic acid differentially regulate human β-defensin-1 and -2 secretion by colonic epithelial cells.

Remodelling of energy storing white fat into energy expending beige fat could be a promising strategy to reduce adiposity. Here, we show that the bile acid-responsive membrane receptor TGR5 mediates beiging of the subcutaneous white adipose tissue (scWAT) under multiple environmental cues including cold exposure and prolonged high-fat diet feeding. Moreover, administration of TGR5-selective bile acid mimetics to thermoneutral housed mice leads to the appearance of beige adipocyte markers and increases mitochondrial content in the scWAT of Tgr5 +/+ mice but not in their Tgr5 −/− littermates. This phenotype is recapitulated in vitro in differentiated adipocytes, in which TGR5 activation increases free fatty acid availability through lipolysis, hence fuelling β-oxidation and thermogenic activity. TGR5 signalling also induces mitochondrial fission through the ERK/DRP1 pathway, further improving mitochondrial respiration. Taken together, these data identify TGR5 as a druggable target to promote beiging with potential applications in the management of metabolic disorders.

The vitamin D endocrine system has many extraskeletal targets, including adipose tissue. 1,25-Dihydroxyvitamin D-3, the active form of vitamin D, not only increases adipogenesis and the expression of typical adipocyte genes but also decreases the expression of uncoupling proteins. Mice with disrupted vitamin D action-owing to gene deletion of the nuclear receptor vitamin D receptor (Vdr) or the gene encoding 1 alpha-hydroxylase (Cyp27b1)-lose fat mass over time owing to an increase in energy expenditure, whereas mice with increased Vdr-mediated signalling in adipose tissue become obese. The resistance to diet-induced obesity in mice with disrupted Vdr signalling is caused at least partially by increased expression of uncoupling proteins in white adipose tissue. However, the bile acid pool is also increased in these animals, and bile acids are known to be potent inducers of energy expenditure through activation of several nuclear receptors, including Vdr, and G-protein-coupled receptors, such as GPBAR1 (also known as TGR5). By contrast, in humans, obesity is strongly associated with poor vitamin D status. A causal link has not been firmly proven, but most intervention studies have failed to demonstrate a beneficial effect of vitamin D supplementation on body weight. The reasons for the major discrepancy between mouse and human data are unclear, but understanding the link between vitamin D status and energy homeostasis could potentially be very important for the human epidemic of obesity and the metabolic syndrome.

PI3Ks are signaling enzymes engaged by different types of membrane receptors and activated in cardiovascular diseases such as hypertension, atherosclerosis, thrombosis and heart failure. Studies performed on genetically modified animals have provided proof-of-concept that general or isoform-specific blockade of these enzymes can modify disease development and progression. Hence, therapeutic inhibition of PI3Ks with novel pharmacological compounds constitutes a promising area of drug development. In particular, inhibitors of PI3Ks have the potential to reduce blood pressure, restrain the development of atherosclerosis and/or stabilize atherosclerotic plaques, blunt platelet aggregation, prevent left ventricular remodeling and preserve myocardial contractility in heart failure. This review summarizes the rationale of PI3K inhibition in the most prevalent cardiovascular diseases, and the available data on the therapeutic effects of PI3K inhibitors in their preclinical models. Implications for future drug development and human therapy are also discussed.

Phosphoinositide 3-kinases (PI3Ks) are protein and lipid kinases activated by different classes of membrane receptors, including G-protein coupled and tyrosine kinase receptors. Several lines of evidence have uncovered specific roles for distinct PI3K isoforms in the vascular system in both physiology and disease. The present review will summarize and discuss the most recent advances regarding PI3K-Akt signalling in endothelial cells, vascular smooth muscle cells, platelets, and inflammatory cells involved in the atherosclerotic process. Of interest, the development of novel isoform-selective PI3K inhibitor drugs offers a unique opportunity to selectively and differentially target PI3K-driven pathways in the vascular system and may give rise to new strategies for the treatment of cardiovascular diseases.

Bile acids are signalling molecules, which activate the transmembrane receptor TGR5 and the nuclear receptor FXR. BA sequestrants (BAS) complex bile acids 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 beta-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 glycaemia 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.

Reverse cholesterol transport (RCT) is an antiatherogenic process in which excessive cholesterol from peripheral tissues is transported to the liver and finally excreted from the body via the bile. The nuclear receptor liver receptor homolog 1 (LRH-1) drives expression of genes regulating RCT, and its activity can be modified by different posttranslational modifications. Here, we show that atherosclerosis-prone mice carrying a mutation that abolishes SUMOylation of LRH-1 on K289R develop less aortic plaques than control litter-mates when exposed to a high-cholesterol diet. The mechanism underlying this atheroprotection involves an increase in RCT and its associated hepatic genes and is secondary to a compromised interaction of LRH-1 K289R with the corepressor prospero homeobox protein 1 (PROX1). Our study reveals that the SUMOylation status of a single nuclear receptor lysine residue can impact the development of a complex metabolic disease such as atherosclerosis.

Phosphoinositides (PIs), a family of phosphorylated derivatives of the membrane lipid phosphatidylinositol, are established regulators of multiple cellular functions. An increasing amount of evidence has highlighted potential links between PI-mediated signaling pathways and the etiology of many human diseases, including cardiovascular pathologies. This chapter will provide a detailed overview of the peculiar functions of the major cardiovascular PIs in the pathogenesis of atherosclerosis, heart failure, and arrhythmias.

Significance: Oxidative stress is involved in the pathogenesis of heart failure but clinical antioxidant trials have been unsuccessful. This may be because effects of reactive oxygen species (ROS) depend upon their source, location, and concentration. Nicotinamide adenine dinucleotide phosphate oxidase (Nox) proteins generate ROS in a highly regulated fashion and modulate several components of the heart failure phenotype. Recent Advances: Two Nox isoforms, Nox2 and Nox4, are expressed in the heart. Studies using gene-modified mice deficient in Nox2 activity indicate that Nox2 activation contributes to angiotensin II–induced cardiomyocyte hypertrophy, atrial fibrillation, and the development of interstitial fibrosis but may also positively modulate physiological excitation-contraction coupling. Nox2 contributes to myocyte death under stress situations and plays important roles in postmyocardial infarction remodeling, in part by modulating matrix metalloprotease activity. In contrast to Nox2, Nox4 is constitutively active at a low level and induces protective effects in the heart under chronic stress, for example, by maintaining myocardial capillary density. However, high levels of Nox4 could have detrimental effects. Critical Issues: The effects of Nox proteins during the development of heart failure likely depend upon the isoform, activation level, and cellular distribution, and may include beneficial as well as detrimental effects. More needs to be learnt about the precise regulation of abundance and biochemical activity of these proteins in the heart as well as the downstream signaling pathways that they regulate. Future Directions: The development of specific approaches to target individual Nox isoforms and/or specific cell types may be important for the achievement of therapeutic efficacy in heart failure. Antioxid. Redox Signal. 18, 1024–1041.

Objective— Liver X receptors (LXRα, LXRβ) are master regulators of cholesterol homeostasis. In the endothelium, perturbations of cell cholesterol have an impact on fundamental processes. We, therefore, assessed the effects of LXR activation on endothelial functions related to angiogenesis in vitro and in vivo. Methods and Results— LXR agonists (T0901317, GW3965) blunted migration, tubulogenesis, and proliferation of human umbilical vein endothelial cells. By affecting endothelial cholesterol homeostasis, LXR activation impaired the compartmentation of vascular endothelial growth factor receptor-2 in lipid rafts/caveolae and led to defective phosphorylation and downstream signaling of vascular endothelial growth factor receptor-2 upon vascular endothelial growth factor-A stimulation. Consistently, the antiangiogenic actions of LXR agonists could be prevented by coadministration of exogenous cholesterol. LXR agonists reduced endothelial sprouting from wild-type but not from LXRα −/− /LXRβ −/− knockout aortas and blunted the vascularization of implanted angioreactors in vivo. Furthermore, T0901317 reduced the growth of Lewis lung carcinoma grafts in mice by impairing angiogenesis. Conclusion— Pharmacological activation of endothelial LXRs reduces angiogenesis by restraining cholesterol-dependent vascular endothelial growth factor receptor-2 compartmentation and signaling. Thus, administration of LXR agonists could exert therapeutic effects in pathological conditions characterized by uncontrolled angiogenesis.

Class IB phosphoinositide 3-kinase p110γ is a master regulator of both leukocyte and cardiomyocyte function. Whereas in the immune system p110γ is principally involved in the control of leukocyte chemotaxis and inflammatory reactions, in cardiomyocytes p110γ affects multiple aspects of β-adrenergic receptor signaling and cardiac function. Because inflammatory cell recruitment/activation and cardiac dysfunction are strictly connected, p110γ has recently been revealed as a promising target for drug design in the treatment of heart failure. This review discusses recent works that dissect the relative contribution of leukocyte p110γ and its cardiac counterpart in the onset and progression of pressure overload–induced cardiac remodeling and the ensuing therapeutic implications.

In the last decade, the availability of genetically modified animals has revealed interesting roles for phosphoinositide 3-kinases (PI3Ks) as signaling platforms orchestrating multiple cellular responses, both in health and pathology. By acting downstream distinct receptor types, PI3Ks nucleate complex signaling assemblies controlling several biological process, ranging from cell proliferation and survival to immunity, cancer, metabolism and cardiovascular control. While the involvement of these kinases in modulating immune reactions and neoplastic transformation has long been accepted, recent progress from our group and others has highlighted new and unforeseen roles of PI3Ks in controlling cardiovascular function. Hence, the view is emerging that pharmacological targeting of distinct PI3K isoforms could be successful in treating disorders such as myocardial infarction and heart failure, besides inflammatory diseases and cancer. Currently, PI3Ks represent attractive drug targets for companies interested in the development of novel and safe treatments for such diseases. Numerous hit and lead compounds are now becoming available and, for some of them, clinical trials can be envisaged in the near future. In the following sections, we will outline the impact of specific PI3K isoforms in regulating different cellular contexts, including immunity, metabolism, cancer and cardiovascular system, both in physiological and disease conditions.

Spatial and temporal organization of signal transduction is coordinated through the segregation of signaling enzymes in selected cellular compartments. This highly evolved regulatory mechanism ensures the activation of selected enzymes only in the vicinity of their target proteins. In this context, cAMP-responsive triggering of protein kinase A is modulated by a family of scaffold proteins referred to as A-kinase anchoring proteins. A-kinase anchoring proteins form the core of multiprotein complexes and enable simultaneous but segregated cAMP signaling events to occur in defined cellular compartments. In this review we will focus on the description of A-kinase anchoring protein function in the regulation of cardiac physiopathology.

Psoriasis is a chronic inflammatory skin disease triggered by interplay between immune mediators from both innate and adaptive immune systems and skin tissue, in which the IL-23/IL-17 axis is critical. PI3Kδ and PI3Kγ play important roles in various immune cell functions. We found that mice lacking functional PI3Kδ or PI3Kγ are largely protected from imiquimod (IMQ)-induced psoriasis-like dermatitis, correlating with reduced IL-17 levels in the lesions, serum, and the draining lymph nodes. TCRγδ T cells were the major IL-17–producing population in the draining lymph nodes and were significantly diminished in IMQ-treated PI3Kδ knockin and PI3Kγ knockout mice. We also show that PI3Kδ and PI3Kγ inhibitors reduced IFN-γ production by human TCRγδ T cells and IL-17 and IFN-γ production by PBMCs from psoriatic or healthy donors. In addition, inhibition of PI3Kγ, but not PI3Kδ, blocked chemotaxis of CCR6+IL-17–producing cells from IMQ-treated mice or healthy human donors. Taken together, these data indicate that PI3Kδ and/or PI3Kγ inhibitors should be considered for treating IL-17–driven diseases, such as psoriasis.

Adrenergic stimulation of the heart engages cAMP and phosphoinositide second messenger signaling cascades. Cardiac phosphoinositide 3-kinase p110 gamma participates in these processes by sustaining beta-adrenergic receptor internalization through its catalytic function and by controlling phosphodiesterase 3B (PDE3B) activity via an unknown kinase-independent mechanism. We have discovered that p110 gamma anchors protein kinase A (PKA) through a site in its N-terminal region. Anchored PKA activates PDE3B to enhance cAMP degradation and phosphorylates p110 gamma to inhibit PIP3 production. This provides local feedback control of PIP3 and cAMP signaling events. In congestive heart failure, p110 gamma is upregulated and escapes PKA-mediated inhibition, contributing to a reduction in beta-adrenergic receptor density. Pharmacological inhibition of p110 gamma normalizes beta-adrenergic receptor density and improves contractility in failing hearts.

Signal transduction events are key modulators of cellular function and, in the cardiovascular system, an emerging role is played by phosphoinositide 3-kinases (PI3Ks), a family of enzymes containing a 3-phosphorylated phosphoinositide that produce lipid second messengers. In the heart, multiple PI3K isoforms are expressed, but play potentially distinct roles. Among cardiac PI3Ks, PI3Kalpha is triggered by tyrosine kinase receptors and plays a role in adaptive hypertrophy, while PI3Kgamma is triggered by G protein-coupled receptors and is involved in maladaptive heart remodeling. This view has been recently complicated by the finding that PI3Ks can also be involved in protein-protein interactions and affect signaling independently of their kinase activity. This review will thus focus on the effects of these multiple signaling events, with particular emphasis on their involvement in cardiac hypertrophy and failure.

Class I phosphoinositide 3-kinases (PI3Ks) are heterodimeric enzymes involved in signal transduction triggered by growth factors and G-protein-coupled receptors. The catalytic function of PI3Ks is well known to promote a wide variety of biological processes, including proliferation, survival and migration, but a new layer of complexity in the function of PI3Ks has recently emerged, indicating that these proteins function not only as kinases but also as scaffold proteins. Knockout mice that lack PI3K protein expression show a different phenotype from knock-in mice expressing PI3K mutants that have lost their kinase activity, providing evidence for this novel role of PI3Ks. We will discuss such findings, highlighting the crucial scaffold function of PI3Kgamma in cAMP homeostasis and PI3Kbeta in receptor recycling.

PI3Kgamma is a phosphoinositide 3-kinase characterized by both lipid and protein kinase activity. It is activated by G-protein-coupled receptors and is predominantly expressed in leucocytes; in addition, recent work showed its presence in the heart and its involvement in regulating cardiac functions. In this tissue, PI3Kgamma acts as a negative modulator of contractility, by decreasing cAMP concentration through a kinase-independent mechanism. Indeed, whereas PI3Kgamma-deficient mice show an abnormal cAMP elevation, cAMP levels in knock-in mouse mutants, expressing a kinase-dead PI3Kgamma, are comparable with wild-type controls. PI3Kgamma regulates cardiac cAMP homoeostasis by forming a macromolecular complex containing PDE3B (phosphodiesterase 3B). In this complex, PI3Kgamma could regulate PDE3B activity through protein kinase A, a PDE activator.