Sabrina Cipriani

FXR activation reverses insulin resistance and lipid abnormalities and protects against liver steatosis in Zucker (fa/fa) obese rats.

The farnesoid X receptor (FXR) is a bile acid activated nuclear receptor. Zucker (fa/fa) rats, harboring a loss of function mutation of the leptin receptor, develop diabetes, insulin resistance, obesity, and liver steatosis. In this study, we investigated the effect of FXR activation by 6-ethyl-chenodeoxycholic acid, (6E-CDCA, 10 mg/kg) on insulin resistance and liver and muscle lipid metabolism in fa/fa rats and compared its activity with rosiglitazone (10 mg/kg) alone or in combination with 6E-CDCA (5 mg/kg each). In comparison to lean (fa/+), fa/fa rats on a normal diet developed insulin resistance and liver steatosis. FXR activation protected against body weight gain and liver and muscle fat deposition and reversed insulin resistance as assessed by insulin responsive substrate-1 phosphorylation on serine 312 in liver and muscles. Activation of FXR reduced liver expression of genes involved in fatty acid synthesis, lipogenesis, and gluconeogenesis. In the muscles, FXR treatment reduced free fatty acid synthesis. Rosiglitazone reduced blood insulin, glucose, triglyceride, free fatty acid, and cholesterol plasma levels but promoted body weight gain (20%) and liver fat deposition. FXR activation reduced high density lipoprotein plasma levels. In summary, FXR administration reversed insulin resistance and correct lipid metabolism abnormalities in an obesity animal model.

Bile-acid-activated receptors: targeting TGR5 and farnesoid-X-receptor in lipid and glucose disorders.

Bile acids are a family of steroid molecules generated in the liver by cholesterol oxidation. In addition to their role in nutrient absorption, bile acids are signaling molecules that exert genomic and non-genomic effects by activating TGR5 (M-BAR, GP-BAR1 or BG37) a G-protein-coupled receptor, and farnesoid X receptor (FXR), a member of the nuclear hormone receptor superfamily. Ligands for these receptors might be beneficial in treating disorders of lipid and glucose homeostasis. TGR5 ligands decrease blood glucose levels and increase energy expenditure by promoting intracellular thyroid hormone activation in thermogenically competent tissues. FXR agonists repress the synthesis of endogenous bile acids and reduce triglyceride, cholesterol and glucose plasma levels and are currently being tested in nonalcoholic steatohepatitis. FXR modulators are being developed to target selective gene clusters and avoid the negative impact of FXR on HDL biosynthesis. The development of dual FXR and TGR5 ligands could provide new opportunities for the treatment of lipid and glucose disorders.

The bile acid receptor GPBAR-1 (TGR5) modulates integrity of intestinal barrier and immune response to experimental colitis

Background GP-BAR1, a member G protein coupled receptor superfamily, is a cell surface bile acid-activated receptor highly expressed in the ileum and colon. In monocytes, ligation of GP-BAR1 by secondary bile acids results in a cAMP-dependent attenuation of cytokine generation. Aims To investigate the role GP-BAR1 in regulating intestinal homeostasis and inflammation-driven immune dysfunction in rodent models of colitis. Methods Colitis was induced in wild type and GP-BAR1−/− mice by DSS and TNBS administration. Potential GP-BAR1 agonists were identified by in silico screening and computational docking studies. Results GP-BAR1−/− mice develop an abnormal morphology of colonic mucous cells and an altered molecular architecture of epithelial tight junctions with increased expression and abnormal subcellular distribution of zonulin 1 resulting in increased intestinal permeability and susceptibility to develop severe colitis in response to DSS at early stage of life. By in silico screening and docking studies we identified ciprofloxacin as a GP-BAR1 ligand. In monocytes, ciprofloxacin increases cAMP concentrations and attenuates TNFα release induced by TLR4 ligation in a GP-BAR1 dependent manner. Treating mice rendered colitic by TNBS with ciprofloxacin and oleanolic acid, a well characterized GP-BAR1 ligand, abrogates signs and symptoms of colitis. Colonic expression of GP-BAR1 mRNA increases in rodent models of colitis and tissues from Crohns disease patients. Flow cytometry analysis demonstrates that ≈90% of CD14+ cells isolated from the lamina propria of TNBS-treated mice stained positively for GP-BAR1. Conclusions GP-BAR1 regulates intestinal barrier structure. Its expression increases in rodent models of colitis and Crohns disease. Ciprofloxacin is a GP-BAR1 ligand.

Bile acid-activated receptors in the treatment of dyslipidemia and related disorders.

Dyslipidemia is a metabolic disorder that constitutes a major risk factor for cardiovascular diseases and stroke and is often associated with diabetes mellitus and atherosclerosis. In recent years a number of ligand-activated receptors have been found to exert a role in integrating essential steps of lipid and glucose metabolism. Bile acid-activated receptors are a defined subset of nuclear and G-protein coupled receptors mainly expressed in entero-hepatic tissues for which bile acids function as signaling molecules. Primary bile acids (chenodeoxycholic acid and cholic acid) are physiological ligands/activators of farnesoid-X-receptor (FXR), pregnane-X-receptor (PXR) and constitutive androstane receptor (CAR), while litocholic acid is a ligand for the Vitamin D receptor (VDR) and the G-protein coupled receptor TGR5. Despite FXR demonstrates a high selectivity for bile acids, PXR and CAR are relatively promiscuous receptors integrating lipid homeostasis with xenobiotic metabolism. FXR, PXR, CAR and TGR exert synergistic activities in regulating lipid and glucose homeostasis and energy expenditure and liver and peripheral insulin sensitivity. Ligands for these receptors hold promise in the treatment of dyslipidemic conditions as revealed by results of a number of preclinical models but carry a defined risk for potential side effects.

Mesenchymal cells from human amniotic fluid survive and migrate after transplantation into adult rat brain

Amniotic fluid has been recently suggested as an alternative source of mesenchymal stem cells. However, the fate of amniotic fluid‐derived mesenchymal stem cells (AF‐MSCs) after in vivo transplantation has yet to be determined. In the present study we explored whether human AF‐MSCs could survive and migrate following transplantation into the striatum of normal and ischemic rat. We found that the grafted cells could survive and migrate towards multiple brain regions in the normal animals, while they moved towards the injured region in the ischemic rat. Double‐immunostaining analyses showed that the implanted human AF‐MSCs express markers for immature neurons (Doublecortin) at 10 days, and for astrocytes (GFAP) at 10, 30 and 90 after transplantation. This study provides the first evidence that human amniotic fluid contains cells having the potential to survive and integrate into adult rat brain tissue and, therefore, to function as effective stem cells for therapeutic strategies.

Modulation of intestinal microbiota by the probiotic VSL#3 resets brain gene expression and ameliorates the age-related deficit in LTP.

The intestinal microbiota is increasingly recognized as a complex signaling network that impacts on many systems beyond the enteric system modulating, among others, cognitive functions including learning, memory and decision-making processes. This has led to the concept of a microbiota-driven gut–brain axis, reflecting a bidirectional interaction between the central nervous system and the intestine. A deficit in synaptic plasticity is one of the many changes that occurs with age. Specifically, the archetypal model of plasticity, long-term potentiation (LTP), is reduced in hippocampus of middle-aged and aged rats. Because the intestinal microbiota might change with age, we have investigated whether the age-related deficit in LTP might be attenuated by changing the composition of intestinal microbiota with VSL#3, a probiotic mixture comprising 8 Gram-positive bacterial strains. Here, we report that treatment of aged rats with VSL#3 induced a robust change in the composition of intestinal microbiota with an increase in the abundance of Actinobacteria and Bacterioidetes, which was reduced in control-treated aged rats. VSL#3 administration modulated the expression of a large group of genes in brain tissue as assessed by whole gene expression, with evidence of a change in genes that impact on inflammatory and neuronal plasticity processes. The age-related deficit in LTP was attenuated in VSL#3-treated aged rats and this was accompanied by a modest decrease in markers of microglial activation and an increase in expression of BDNF and synapsin. The data support the notion that intestinal microbiota can be manipulated to positively impact on neuronal function.

'One night' sleep deprivation stimulates hippocampal neurogenesis.

Neurogenesis in the adult hippocampus can be up- or downregulated in response to a variety of physiological and pathological conditions. Among these, dysregulation of hippocampal neurogenesis has been recently implicated in the pathogenesis of depression. In addition, in animal models of depression, a variety of antidepressant treatments reverse that condition by increasing neurogenesis. As one night sleep deprivation is known to improve mood in depressed patients for at least 1 day, we investigated whether a comparable treatment may affect hippocampal neurogenesis in adult rats. Accordingly, rats were sleep-deprived by gentle handling for 12 h during their physiological period of rest, and were injected with bromodeoxyuridine 4 h and 2 h before the end of sleep deprivation. They were then perfused immediately thereafter, or after 15 days and 30 days. We found that 12 h sleep deprivation significantly increased cell proliferation and the total number of surviving cells in the hippocampal dentate gyrus soon after sleep deprivation, as well as 15 days and 30 days later, in comparison to control rats allowed to sleep. No changes were instead found in the subventricular zone of the lateral ventricles, indicating that 12 h sleep deprivation selectively triggers neurogenic signals to the hippocampus. The present data include acute sleep deprivation among the conditions which upregulate hippocampal neurogenesis and raise the possibility that such response could be implicated in the beneficial effects elicited in depressed patients by one night sleep deprivation. Thus, the findings could contribute to the understanding of the intriguing relationship between depression and neurogenesis in the adult brain.

The Bile Acid Sensor Farnesoid X Receptor Is a Modulator of Liver Immunity in a Rodent Model of Acute Hepatitis

Immune-mediated liver diseases including autoimmune and viral hepatitis are a major health problem worldwide. In this study, we report that activation of the farnesoid X receptor (FXR), a member of the ligand-activated nuclear receptor superfamily and bile sensor highly expressed in the liver, attenuates liver injury in a model of autoimmune hepatitis induced by Con A. We found that FXR gene ablation results in a time-dependent increase of liver expression (up to 20-fold in a 9-mo-old mouse) of osteopontin, a NKT cell-derived extracellular matrix protein and immunoregulatory cytokine. In comparison to wild-type, FXR−/− mice are more susceptible to Con A-induced hepatitis and react to Con A administration by an unregulated production of osteopontin. Administering wild-type mice with a synthetic FXR agonist attenuated Con A-induced liver damage and liver expression of the osteopontin gene. By in vitro studies, we found that FXR is expressed by primarily isolated NKT cells and its ablation favors ostepontin production in response to Con A. Chromatin immunoprecipitation assay and coimmunoprecipitation experiments demonstrate that the short heterodimer partner (SHP), a nuclear receptor and FXR target, was expressed by NKT cell hybridomas and increased in response to FXR activation. FXR activates SHP that interacts with and inhibits c-Jun binding to the osteopontin promoter. These data indicate that in NKT cells, FXR activation causes a SHP-mediated inhibition of osteopontin production. These data support the notion that the bile acid sensor FXR regulates the activation of liver NKT cells.

Pregnane-X-receptor mediates the anti-inflammatory activities of rifaximin on detoxification pathways in intestinal epithelial cells.

The pregnane-X-receptor (PXR) is master gene overseeing detoxification of wide number of xenobiotics and is critical for maintenance of intestinal integrity. The intestinal expression of genes involved in cellular detoxification is down-regulated in patients with inflammatory bowel diseases (IBD). Rifaximin is a non-absorbable antibiotic endowed with a PXR agonistic activity. In the present study we have investigated whether rifaximin activates PXR in primary human colon epithelial cells and human colon biopsies and assessed whether this antibiotic antagonizes the effect of tumor necrosis factor (TNF)-α on expression of PXR and PXR-related genes. Present results demonstrate that primary colon epithelial cells express PXR and that their exposure to rifaximin induces the expression of genes involved in cellular detoxification. Exposure to TNFα reduces the expression of PXR mRNA as well as expression of its target genes. This inhibitory effect was prevented by that co-treatment with rifaximin. Knocking down the expression of PXR in colon epithelial cells by an anti-PXR siRNA, abrogated the counter-regulatory effects exerted by rifaximin on cell exposed to TNFα. Finally, ex vivo exposure of colon biopsies obtained from ulcerative colitis patients to rifaximin increased the expression of genes involved in xenobiotics metabolism. In aggregate, these data illustrate that rifaximin increases the expression of PXR and PXR-regulated genes involved in the metabolism and excretion of xenobiotics and antagonizes the effects of TNFα in intestinal epithelial cells and colon biopsies. These non-antibiotic effects of rifaximin could contribute to the maintenance of the intestinal barrier integrity against xenobiotics and products generated by luminal bacteria.

Efficacy of the CCR5 Antagonist Maraviroc in Reducing Early, Ritonavir-Induced Atherogenesis and Advanced Plaque Progression in Mice

Background— CCR5 plays an important role in atherosclerosis and ischemic cardiovascular diseases, as well as in HIV replication and diffusion. HIV infection is characterized by a high burden of cardiovascular diseases, particularly in subjects exposed to ritonavir-boosted protease inhibitors. Maraviroc, a CCR5 antagonist antiretroviral drug, might provide benefit for patients with M-tropic HIV infections at high risk for cardiovascular diseases. Methods and Results— Exposure to maraviroc limits the evolution and associated systemic inflammation of ritonavir-induced atherosclerotic in ApoE−/− mice and inhibits plaques development in a late model of atherosclerosis in which dyslipidemia plays the main pathogenic role. In ritonavir-treated mice, maraviroc reduced plaque areas and macrophage infiltration; downregulated the local expression of vascular cell adhesion molecule-1, intercellular adhesion molecule-1, monocyte chemoattractant protein-1, and interleukin-17A; and reduced tumor necrosis factor-&agr; and RANTES (regulated on activation, normal T cell expressed, and secreted). Moreover, maraviroc counterregulated ritonavir-induced lipoatrophy and interlelukin-6 gene expression in epididymal fat, along with the splenic proinflammatory profile and expression of CD36 on blood monocytes. In the late model, maraviroc inhibited atherosclerotic progression by reducing macrophage infiltration and lowering the expression of adhesion molecules and RANTES inside the plaques. However, limited systemic inflammation was observed. Conclusions— In a mouse model of genetic dyslipidemia, maraviroc reduced the atherosclerotic progression by interfering with inflammatory cell recruitment into plaques. Moreover, in mice characterized by a general ritonavir-induced inflammation, maraviroc reversed the proinflammatory profile. Therefore, maraviroc could benefit HIV-positive patients with residual chronic inflammation who are at a high risk of acute coronary disease despite a suppressive antiretroviral therapy. To determine these benefits, large clinical studies are needed.