TLR2 and TLR4 Modulate Mouse Ileal Motility by the Interaction with Muscarinic and Nicotinic Receptors; 35681486

Irritable bowel syndrome (IBS) is a chronic functional bowel disorder characterized by intestinal dysmotility. Changes in intestinal microbiota (dysbiosis) can lead to alterations in neuro-muscular functions in the gut. Toll-like receptors (TLRs) 2 and 4 recognize intestinal bacteria and are involved in the motor response induced by gastrointestinal (GI) neurotransmitters. Acetylcholine (ACh) is a well-known neurotransmitter involved in the regulation of GI motility. This study aimed to evaluate the role of TLR2 and TLR4 in the intestinal motor-response induced by ACh in the mouse ileum, as well as the expression and function of the muscarinic and nicotinic ACh receptors. Muscle contractility studies showed that the contractions induced by ACh were significantly lower in TLR2-/- and TLR4-/- with respect to WT mice. In WT mice, the contractions induced by ACh were reduced in the presence of AF-DX AF-DX 116 (a muscarinic ACh receptor (mAChR) M2 antagonist), 4-DAMP (a mAChR M3 antagonist), mecamylamine (a nicotinic AChR receptor (nAChR) a3ß4 antagonist) and a-bungarotoxin (a nAChR a7 antagonist). In TLR2-/- mice, the contractions induced by ACh were increased by AF-DX 116 and mecamylamine. In TLR4-/- mice, the contractions induced by ACh were reduced by a-bungarotoxin and 4-DAMP. The mRNA and protein expressions of M3 and a3 receptors were diminished in the ileum from TLR2-/- and TLR4-/- with respect to WT mice. However, the levels of mRNA and protein of ß4 were diminished only in TLR4-/- but not in TLR2-/- mice. In conclusion, our results show that TLR2 and TLR4 modulates the motor responses to ACh in the mouse ileum. TLR2 acts on muscarinic M2 and M3 and nicotinic a3ß4 ACh receptors, while TLR4 acts on muscarinic M3 and nicotinic a3ß4 and a7 ACh receptors

Development of Synthetic Ca-based CO2 Sorbents for Sorption Enhanced Reforming Coupled to Ca/Cu Chemical Loop

Sorption Enhanced Reforming (SER) is a very promising option that allows H2 production coupled with capturing CO2 by sorption. The present research work focus in the sorbent point of view and several synthetic materials with a high CO2 absorption capacity and chemically and mechanically stable during multi-cyclic operation (at least 40 calcination-carbonation cycles and 100 cycles in some cases) under the typical SER process conditions have been developed. Two different synthesis routes (co-precipitation and mechanical mixing) have been followed trying to evaluate the effect of two different inert supports (magnesium oxide and mayenite) on the evolution of the CO2 carrying capacity of the materials. Also, sorbents have been synthesized with different morphologies (powder and particles) and sizes (from 100 microns up to 2 mm) in order to test their reactivity and mechanical suitability for large scale reactor system operation. Synthetic dolomite with a molar CaO/MgO ratio of 2:1 synthesized by co-precipitation method exhibited very stable performance for 100 consecutive carbonation/calcination cycles and a residual CO2 carrying capacity of 0.29 grCO2/grcalcined sorbent. Also a very promising result has been obtained for this sorbent mixed with a commercial catalyst in a preliminary SER experiment that yield a H2 production of 92% vol. (d.b.)

Toll-like receptor 2 modulates the inhibitory motor response induced by hydrogen sulphide in mouse colon

Introduction: The recognition of intestinal microbiota is in part carried out by toll-like receptors (TLR), which are responsible for initiating the innate immune response. Alterations in the intestinal microbiota and its recognition may contribute to the development of intestinal inflammatory pathologies. Otherwise, hydrogen sulphide (H2S) is an endogenous gaseous signalling molecule and it potentially plays a relevant role in the intestinal motility. In mammals, two pyridoxalphosphate-dependent enzymes are responsible for H2S synthesis: cystathionine b-synthase (CBS) and cystathionine -lyase (CSE)..

Development of Suitable CuO-Based Materials Supported on Al2O3, MgAl2O4, and ZrO2 for Ca/Cu H2 Production Process

Functional materials for the sorption enhanced reforming process for H2 production coupled to a Cu/CuO chemical loop have been synthesized. The performance of CuO-based materials supported on Al2O3, MgAl2O4, and ZrO2 and synthesized by different routes has been analyzed. Highly stable materials supported on Al2O3 or MgAl2O4 synthesized by coprecipitation and mechanical mixing with sufficient Cu loads (around 65% wt) have been successfully developed. However, it has been found that coprecipitation under these conditions is not a suitable route for ZrO2. Spray-drying and deposition precipitation did not provide the best chemical features to the materials. As the Ca/Cu process is operated in fixed bed reactors, the best candidates were pelletized and their stability was again assessed. Pellets with high chemical and mechanical stability, high oxygen transport capacity, and good mechanical properties have been finally obtained by coprecipitation. The good homogeneity that provides this route would allow an easy scaling up

NOD2 Modulates Serotonin Transporter and Interacts with TLR2 and TLR4 in Intestinal Epithelial Cells

Background/Aims: Serotonin (5-HT) is a chief modulator of intestinal activity. The effects of 5-HT depend on its extracellular availability, which is mainly controlled by serotonin transporter (SERT), expressed in enterocytes. On the other hand, innate immunity, mediated by Toll-like receptors (TLRs) and nucleotide oligomerization domain (NOD)-like receptors (NLRs), is known to control intestinal microbiota and maintain intestinal homeostasis. The dysregulation of the intestinal serotonergic system and innate immunity has been observed in in ammatory bowel diseases (IBD), the incidence of which has severely increased all over the world. The aim of the present study, therefore, was to analyze the effect of NOD2 on intestinal SERT activity and expression, as well as to study the crosstalk of NOD2 with TLR2 and TLR4. Methods: Intestinal epithelial cell line Caco-2/TC7 was used to analyze SERT activity and SERT, NOD2, TLR2 and TLR4 molecular expression by real-time PCR and western blotting. Moreover, intestinal tract (ileum and colon) from mice de cient in TLR2, TLR4 or TLR2/4 receptors was used to test the interdependence of NOD2 with these TLR receptors. Results: NOD2 activation inhibits SERT activity in Caco-2/TC7 cells, mainly due to the decrement of SERT molecular expression, with RIP2/RICK being the intracellular pathway involved in this effect. This inhibitory effect on SERT would yield an increment of extracellular 5-HT availability. In this sense, 5-HT strongly inhibits NOD2 expression. In addition, NOD2 showed greater interdependence with TLR2 than with TLR4. Indeed, NOD2 expression signi cantly increased in both cells treated with TLR2 agonists and the intestinal tract of Tlr2-/- mice. Conclusions: It may be inferred from our data that NOD2 could play a role in intestinal pathophysiology not only through its inherent innate immune role but also due to its interaction with other receptors as TLR2 and the modulation of the intestinal serotonergic system decreasing SERT activity and expression

Toll-like receptor 9 modifies intestinal serotonergic system.

Introduction: Toll-like receptor 9 (TLR9) is expressed in intestinal epithelial cells, which recognize microbiota developing different responses 1. Several studies have shown that TLR9 seems to be involved in Inflammatory Bowel Diseases (IBD) due to an inappropriate defensive response against microorganisms 2. Moreover, intestinal serotonergic system is also altered in IBD, where extracellular serotonin (5–HT) levels are increased 3. 5-HT bioavailability is mainly regulated by the serotonin transporter (SERT), expressed in enterocytes 4. Aims & Methods: The aim of the present study was to analyse whether TLR9 activation affects SERT expression and activity, and expression of other elements from the serotonergic system (TPH1, TPH2 and 5-HT receptors). Human enterocyte-like Caco-2 cells, and ileum and colon from TLR9-/- mice and Dextran Sulphate Sodium (DSS) mouse colitis model were used as experimental models. mRNA expression was determined by RT-qPCR, and protein expression by western blot..

NOD1 downregulates intestinal serotonin transporter and interacts with other pattern recognition receptors

Serotonin (5-HT) is an essential gastrointestinal modulator whose effects regulate the intestinal physiology. 5-HT effects depend on extracellular 5-HT bioavailability, which is controlled by the serotonin transporter (SERT) expressed in both the apical and basolateral membranes of enterocytes. SERT is a critical target for regulating 5-HT levels and consequently, modulating the intestinal physiology. The deregulation of innate immune receptors has been extensively studied in inflammatory bowel diseases (IBD), where an exacerbated defense response to commensal microbiota is observed. Interestingly, many innate immune receptors seem to affect the serotonergic system, demonstrating a new way in which microbiota could modulate the intestinal physiology. Therefore, our aim was to analyze the effects of NOD1 activation on SERT function, as well as NOD1's interaction with other immune receptors such as TLR2 and TLR4. Our results showed that NOD1 activation inhibits SERT activity and expression in Caco-2/TC7 cells through the extracellular signal-regulated kinase (ERK) signaling pathway. A negative feedback between 5-HT and NOD1 expression was also described. The results showed that TLR2 and TLR4 activation seems to regulate NOD1 expression in Caco-2/TC7 cells. To assess the extend of cross-talk between NOD1 and TLRs, NOD1 expression was measured in the intestinal tract (ileum and colon) of wild type mice and mice with individual knockouts of TLR2, and TLR4 as well as double knockout TLR2/TLR4 mice. Hence, we demonstrate that NOD1 acts on the serotonergic system decreasing SERT activity and molecular expression. Additionally, NOD1 expression seems to be modulated by 5-HT and other immune receptors as TLR2 and TLR4. This study could clarify the relation between both the intestinal serotonergic system and innate immune system, and their implications in intestinal inflammation

Treatment of rats with a self-selected hyperlipidic diet, increases the lipid content of the main adipose tissue sites in a proportion similar to that of the lipids in the rest of organs and tissues

Adipose tissue (AT) is distributed as large differentiated masses, and smaller depots covering vessels, and organs, as well as interspersed within them. The differences between types and size of cells makes AT one of the most disperse and complex organs. Lipid storage is partly shared by other tissues such as muscle and liver. We intended to obtain an approximate estimation of the size of lipid reserves stored outside the main fat depots. Both male and female rats were made overweight by 4-weeks feeding of a cafeteria diet. Total lipid content was analyzed in brain, liver, gastrocnemius muscle, four white AT sites: subcutaneous, perigonadal, retroperitoneal and mesenteric, two brown AT sites (interscapular and perirenal) and in a pool of the rest of organs and tissues (after discarding gut contents). Organ lipid content was estimated and tabulated for each individual rat. Food intake was measured daily. There was a surprisingly high proportion of lipid not accounted for by the main macroscopic AT sites, even when brain, liver and BAT main sites were discounted. Muscle contained about 8% of body lipids, liver 1-1.4%, four white AT sites lipid 28-63% of body lipid, and the rest of the body (including muscle) 38-44%. There was a good correlation between AT lipid and body lipid, but lipid in"other organs" was highly correlated too with body lipid. Brain lipid was not. Irrespective of dietary intake, accumulation of body fat was uniform both for the main lipid storage and handling organs: large masses of AT (but also liver, muscle), as well as in the"rest" of tissues. These storage sites, in specialized (adipose) or not-specialized (liver, muscle) tissues reacted in parallel against a hyperlipidic diet challenge. We postulate that body lipid stores are handled and regulated coordinately, with a more centralized and overall mechanisms than usually assumed

Sperm survival and heterogeneity are correlated with fertility after intrauterine insemination in superovulated ewes

Abstract Efficient animal production involves accurate estimations of fertilizing ability. One key factor is the plasma membrane of the sperm cell, which is actively involved in the cascade of events before oocyte fusion. Many methods are used to analyze the characteristics of this membrane, including partition in aqueous two-phase systems which is an efficient method to analyze sperm surface changes accounting for loss of viability and different functional states. Centrifugal countercurrent distribution (CCCD) analysis can also be used in an aqueous two-phase system to determine the relationship between sperm parameters and in vivo fertility in ewes. In a previous work, we found a significant correlation between two post-CCCD parameters (heterogeneity and recovered viability) and field fertility when the same sample was used after cervical AI. The present study was intended to find out whether the control of several external factors that affect reproductive efficiency is able to increase the correlation coefficient between post-CCCD parameters and fertility. Thus, 90 Rasa aragonesa ewes were controlled on the same farm and received intrauterine inseminations using the same technical equipment. The fertilizing ability of the raw semen and sperm samples selected by a dextran/swim-up process was compared using a low number of spermatozoa per insemination (7 Â 10 7 ) to enhance possible fertility differences. A new post-CCCD parameter was considered; the loss of viability (LV) occurred during the CCCD process. This variable denotes the sperm surviving ability and corresponds to the difference between the total number of viable cells loaded and recovered after the CCCD run. The mean fertility of eight sperm control samples was 60% (range: 25-76%), and there was no significant correlation between standard parameters and in vivo fertility. LV ranged from 2 to 69% (average 27%) and was negatively correlated with fertility (r ¼ À0.914, P < 0.01). Ejaculate heterogeneity (H) ranged from 20 to 47% and was positively, but not significantly, correlated with fertility (r ¼ 0.391). A predictive equation for fertility was deduced by multiple analysis with a very high correlation coefficient (r ¼ 0.967), and level of significance (P < 0.005): predictive fertility PF ¼ 52.546 À 0.594 LV þ 0.665 H. The mean fertility of 13 swim-up selected samples was 63% (range: 25-86%). Again, only parameters derived from the CCCD analysis were highly correlated with fertility, especially LV and H (P < 0.05)