Alessandra D'Alessandro

Enteroglial-derived S100B protein integrates bacteria-induced Toll-like receptor signalling in human enteric glial cells

Objective Enteric glial cells (EGC) have been suggested to participate in host–bacteria cross-talk, playing a protective role within the gut. The way EGC interact with microorganisms is still poorly understood. We aimed to evaluate whether: EGC participate in host–bacteria interaction; S100B and Toll-like receptor (TLR) signalling converge in a common pathway leading to nitric oxide (NO) production. Design Primary cultures of human EGC were exposed to pathogenic (enteroinvasive Escherichia coli; EIEC) and probiotic (Lactobacillus paracasei F19) bacteria. Cell activation was assessed by evaluating the expression of cFos and major histocompatibility complex (MHC) class II molecules. TLR expression in EGC was evaluated at both baseline and after exposure to bacteria by real-time PCR, fluorescence microscopy and western blot analysis. S100B expression and NO release from EGC, following exposure to bacteria, were measured in the presence or absence of specific TLR and S100B pathway inhibitors. Results EIEC activated EGC by inducing the expression of cFos and MHC II. EGC expressed TLR at baseline. Pathogens and probiotics differentially modulated TLR expression in EGC. Pathogens, but not probiotics, significantly induced S100B protein overexpression and NO release from EGC. Pretreatment with specific inhibitors of TLR and S100B pathways abolished bacterial-induced NO release from EGC. Conclusions Human EGC interact with bacteria and discriminate between pathogens and probiotics via a different TLR expression and NO production. In EGC, NO release is impaired in the presence of specific inhibitors of the TLR and S100B pathways, suggesting the presence of a novel common pathway involving both TLR stimulation and S100B protein upregulation.

Genetic contribution to motility disorders of the upper gastrointestinal tract.

Motility disorders of the upper gastrointestinal tract encompass a wide range of different diseases. Esophageal achalasia and functional dyspepsia are representative disorders of impaired motility of the esophagus and stomach, respectively. In spite of their variable prevalence, what both diseases have in common is poor knowledge of their etiology and pathophysiology. There is some evidence showing that there is a genetic predisposition towards these diseases, especially for achalasia. Many authors have investigated the possible genes involved, stressing the autoimmune or the neurological hypothesis, but there is very little data available. Similarly, studies supporting a post-infective etiology, based on an altered immune response in susceptible individuals, need to be validated. Further association studies can help to explain this complex picture and find new therapeutic targets. The aim of this review is to summarize current knowledge of genetics in motility disorders of the upper gastrointestinal tract, addressing how genetics contributes to the development of achalasia and functional dyspepsia respectively.

Palmitoylethanolamide Exerts Antiproliferative Effect and Downregulates VEGF Signaling in Caco‐2 Human Colon Carcinoma Cell Line Through a Selective PPAR‐α‐Dependent Inhibition of Akt/mTOR Pathway

Palmitoylethanolamide (PEA) is a nutraceutical compound that has been demonstrated to improve intestinal inflammation. We aimed at evaluating its antiproliferative and antiangiogenic effects in human colon adenocarcinoma Caco‐2 cell line. Caco‐2 cells were treated with increasing concentrations of PEA (0.001, 0.01 and 0.1 μM) in the presence of peroxisome proliferator‐activated receptor‐a (PPAR‐α) or PPAR‐γ antagonists. Cell proliferation was evaluated by performing a MTT assay. Vascular endothelial growth factor (VEGF) release was estimated by ELISA, while the expression of VEGF receptor and the activation of the Akt/mammalian target of rapamycin (mTOR) pathway were evaluated by western blot analysis. PEA caused a significant and concentration‐dependent decrease of Caco‐2 cell proliferation at 48 h. PEA administration significantly reduced in a concentration‐dependent manner VEGF secretion and VEGF receptor expression. Inhibition of Akt phosphorylation and a downstream decrease of phospho‐mTOR and of p‐p70S6K were observed as compared with untreated cells. PPAR‐α, but not PPAR‐γ antagonist, reverted all effects of PEA. PEA is able to decrease cell proliferation and angiogenesis. The antiangiogenic effect of PEA depends on the specific inhibition of the AkT/mTOR axis, through the activation of PPAR‐α pathway. If supported by in vivo models, our data pave the way to PEA co‐administration to the current chemotherapeutic regimens for colon carcinoma. Copyright

Enteric glia: A new player in inflammatory bowel diseases

In addition to the well-known involvement of macrophages and neutrophils, other cell types have been recently reported to substantially contribute to the onset and progression of inflammatory bowel diseases (IBD). Enteric glial cells (EGC) are the equivalent cell type of astrocyte in the central nervous system (CNS) and share with them many neurotrophic and neuro-immunomodulatory properties. This short review highlights the role of EGC in IBD, describing the role played by these cells in the maintenance of gut homeostasis, and their modulation of enteric neuronal activities. In pathological conditions, EGC have been reported to trigger and support bowel inflammation through the specific over-secretion of S100B protein, a pivotal neurotrophic factor able to induce chronic inflammatory changes in gut mucosa. New pharmacological tools that may improve the current therapeutic strategies for inflammatory bowel diseases (IBD), lowering side effects (i.e. corticosteroids) and costs (i.e. anti-TNFα monoclonal antibodies) represent a very important challenge for gastroenterologists and pharmacologists. Novel drugs capable to modulate enteric glia reactivity, limiting the pro-inflammatory release of S100B, may thus represent a significant innovation in the field of pharmacological interventions for inflammatory bowel diseases.

Endocannabinoid-related compounds in gastrointestinal diseases

The endocannabinoid system (ECS) is an endogenous signalling pathway involved in the control of several gastrointestinal (GI) functions at both peripheral and central levels. In recent years, it has become apparent that the ECS is pivotal in the regulation of GI motility, secretion and sensitivity, but endocannabinoids (ECs) are also involved in the regulation of intestinal inflammation and mucosal barrier permeability, suggesting their role in the pathophysiology of both functional and organic GI disorders. Genetic studies in patients with irritable bowel syndrome (IBS) or inflammatory bowel disease have indeed shown significant associations with polymorphisms or mutation in genes encoding for cannabinoid receptor or enzyme responsible for their catabolism, respectively. Furthermore, ongoing clinical trials are testing EC agonists/antagonists in the achievement of symptomatic relief from a number of GI symptoms. Despite this evidence, there is a lack of supportive RCTs and relevant data in human beings, and hence, the possible therapeutic application of these compounds is raising ethical, political and economic concerns. More recently, the identification of several EC‐like compounds able to modulate ECS function without the typical central side effects of cannabino‐mimetics has paved the way for emerging peripherally acting drugs. This review summarizes the possible mechanisms linking the ECS to GI disorders and describes the most recent advances in the manipulation of the ECS in the treatment of GI diseases.

Tu1452 Enteroglial-Derived S100B Protein Modulates Differentiation and Proliferation of Human Intestinal Epithelial Cells in a RAGE-Dependent Manner

Retrograde axonal transport was impaired in the gastric branch of the vagus nerve in animals with type 1 diabetes mellitus (DM). These studies evaluated active, retrograde axonal transport of the neural tracer fluorogold (FG) in gastric enteric neurons in KKAy mice 4 weeks prior to and at the onset of type 2 DM, and after 8, 16, and 24 weeks of uncontrolled DM. We hypothesized that the number of FG-labeled enteric neurons would progressively decrease with increasing durations of uncontrolled DM as compared to control KK mice. Under anesthesia, FG was injected into the ventral fundus or antrum. After 5-6 days, the stomach was removed and the numbers of FG-labeled enteric neurons with cell bodies in 2.8 mm2 of fundus, 1.7 mm2 of corpus, and 1.7 mm2 of antrum were counted. The total number of FG-labeled neurons in ascending pathways (cell bodies in corpus or antrum with nerve terminals in fundus), descending pathways (cell bodies in the fundus or corpus with nerve terminals in antrum), the local fundic pathway (cell body and nerve terminals in fundus), and the local antral pathway (cell body and nerve terminals in antrum) were compared in control and diabetic mice of different ages by 2-way ANOVA. Only main effects were significant; no interactions were significant. Data are mean±SEM with n representing the number of mice. In the ascending pathway, fewer FG-labeled neurons were observed in diabetic as compared to control mice [F(1,116)=10.668; control (n=68), 179±11; diabetic (n= 58), 145±11*; *p<0.05]. In contrast, similar numbers of FG-labeled neurons were observed in descending pathways [F(1,124)=0.489; p=0.486], local fundic pathways [F(1,53)=2.464; p=0.122], and local antral pathways [F(1,57)=0.728; p=0.397] of control and diabetic mice. Overall, the number of FG-labeled neurons decreased significantly with age in all pathways [ascending: F(4,116)=42.862; descending: F(4,124)=2.456; local fundic: F(4,53)=5.195; local antral: F(4,57)=8.920]. In the ascending pathway, the number of FG-labeled neurons was significantly less at 16 weeks of age [n=28, 146±12] as compared to 4 weeks [n=24, 246±8] or 8 weeks [n=28, 229±12] of age, reaching its lowest value at 24 weeks of age [n= 24, 80±10]. In conclusion, DM did not uniformly impair active tracer transport in all gastric enteric neurons suggesting specific axonal targets in ascending enteric neurons rendering them more susceptible to hyperglycemia and low effective insulin. Supported by ORSP at MWU.

RAGE-Dependent S100B Protein Modulation of Peripheral and Mucosal Immune Cells' Functions in Patients With Ulcerative Colitis

colitis (mortality; clinical scores, weight loss) was significantly greater in TPH2 KO than in WT animals. The enhanced severity was also evident in enteric histological scores, leukocyte infiltration, and cytokine expression. These data are compatible with the ideas that by promoting inflammation, mucosal 5-HT enhances the defense of the bowel against infection, while neuronal 5-HT makes such an effect less damaging by protecting the ENS from inflammation. Mucosal and neuronal 5-HT may thus exert antagonistic effects on inflammation that are functionally synergistic. Supported by a clinical and translational pilot grant from Columbia University (KGM) and NS12969 (MDG).

P.1.147: RAGE-DEPENDENT S100B PROTEIN MODULATION OF PERIPHERAL AND MUCOSAL IMMUNE CELLS' FUNCTIONS IN PATIENTS WITH ULCERATIVE COLITIS (UC)

colitis (mortality; clinical scores, weight loss) was significantly greater in TPH2 KO than in WT animals. The enhanced severity was also evident in enteric histological scores, leukocyte infiltration, and cytokine expression. These data are compatible with the ideas that by promoting inflammation, mucosal 5-HT enhances the defense of the bowel against infection, while neuronal 5-HT makes such an effect less damaging by protecting the ENS from inflammation. Mucosal and neuronal 5-HT may thus exert antagonistic effects on inflammation that are functionally synergistic. Supported by a clinical and translational pilot grant from Columbia University (KGM) and NS12969 (MDG).