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

Selective anticancer and antimicrobial metallodrugs based on Gold(III) dithiocarbamate complexes

New dithiocarbamate cycloaurated complexes have been synthesized and their physicochemical and in vitro antitumor properties have been evaluated. All the performed studies highlighted good transport through the blood and biodistribution, according to the balance between the properties of hydrophilicity/lipophilicity and the binding of moderate strength to the BSA protein. Furthermore, none of the complexes exhibited reduction or decomposition reactions, presenting excellent physiological stability. The in vitro cytotoxic effect was evaluated on human colon cancer cell line Caco-2/TC7, and the complexes showed great antiproliferative activity and excellent selectivity, as much less effect was detected on normal Caco-2/TC7 cells. Most of the complexes exhibit antiproliferative activity that was better than or similar to auranofin, and at least nine times better than that of cisplatin. Its action mechanism is still under discussion since no evidence of cell cycle arrest was found, but an antioxidant role was shown for some of the selective complexes. All complexes were also tested as antimicrobial drugs, exhibiting good activity towards S. aureus and E. coli. bacteria and C. albicans and C. neoformans fungi. © 2021 by the authors. Licensee MDPI, Basel, Switzerland. This article is an open access article distributed under the terms and conditions of the Creative Commons Attribution (CC BY) license (https:// creativecommons.org/licenses/by/ 4.0/)

Reconstruction and numerical modelling of the abdominal wall. Application to hernia surgery

Routine hernia repair surgery involves the implant of synthetic mesh. However, this proceduremay give rise to several problems causing considerable patient disability. With the aim ofimproving surgical procedures, the healthy and the herniated human abdomen are simulatedusing finite element (FE) simulations. For that purpose, a reconstruction of the physiologicalgeometry of a human abdomen was created using magnetic resonance images. Besides,following the anatomy of the abdomen, the different muscles and aponeurosis were defined.Furthermore, collagen fibres were included in each muscle layer and their passive anisotropicmechanical contribution was modelled within the framework of hyperelasticity. In the FEsimulation of the abdomen, the constraint imposed by the shoulder is applied and an internalpressure of 23 kPa was applied to the interior abdominal wall to reproduce the abdominal loadwhen standing jumping. After generating a hernia in the front of the abdomen, differentprostheses (Surgipro®, Optilene® and Infinit®), modelled using a membrane model, are placedin the defect to simulate the behaviour of the abdomen after the surgical procedure. In thehealthy abdomen, maximal principal stresses (MPS) and displacements (MD) appear in thefront of the belly. On the other hand, once the hernia is created and the mesh is placed, theMD and MPS are higher than those attained in the healthy abdomen. Thus, just after surgery,surgical repair procedure does not fully restore normal physiological conditions and the risk ofhernia recurrence by the suture is high due to the stress concentration

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)..

Toll-like receptors 2 and 4 modulate intestinal IL-10 differently in ileum and colon

Background: Inflammatory bowel diseases are consequence of an intestinal homeostasis breakdown in which innate immune dysregulation is implicated. Toll-like receptor (TLR)2 and TLR4 are immune recognition receptors expressed in the intestinal epithelium, the first physical-physiological barrier for microorganisms, to inform the host of the presence of Gram-positive and Gram-negative organisms. Interleukin (IL)-10 is an essential anti-inflammatory cytokine that contributes to maintenance of intestinal homeostasis. Aim: Our main aim was to investigate intestinal IL-10 synthesis and release, and whether TLR2 and TLR4 are determinants of IL-10 expression in the intestinal tract. Methods: We used Caco-2 cell line as an enterocyte-like cell model, and also ileum and colon from mice deficient in TLR2, TLR4 or TLR2/4 to test the involvement of TLR signaling. Results: Intestinal epithelial cells are able to synthesize and release IL-10 and their expression is increased after TLR2 or TLR4 activation. IL-10 regulation seems to be tissue specific, with IL-10 expression in the ileum regulated by a compensation between TLR2 and TLR4 expression, whereas in the colon, TLR2 and TLR4 affect IL-10 expression independently. Conclusions: Intestinal epithelial cells could release IL-10 in response to TLR activation, playing an intestinal tissue-dependent and critical intestinal immune role

Recursive internetwork architecture, investigating RINA as an alternative to TCP/IP (IRATI)

Driven by the requirements of the emerging applications and networks, the Internet has become an architectural patchwork of growing complexity which strains to cope with the changes. Moore’s law prevented us from recognising that the problem does not hide in the high demands of today’s applications but lies in the flaws of the Internet’s original design. The Internet needs to move beyond TCP/IP to prosper in the long term, TCP/IP has outlived its usefulness. The Recursive InterNetwork Architecture (RINA) is a new Internetwork architecture whose fundamental principle is that networking is only interprocess communication (IPC). RINA reconstructs the overall structure of the Internet, forming a model that comprises a single repeating layer, the DIF (Distributed IPC Facility), which is the minimal set of components required to allow distributed IPC between application processes. RINA supports inherently and without the need of extra mechanisms mobility, multi-homing and Quality of Service, provides a secure and configurable environment, motivates for a more competitive marketplace and allows for a seamless adoption. RINA is the best choice for the next generation networks due to its sound theory, simplicity and the features it enables. IRATI’s goal is to achieve further exploration of this new architecture. IRATI will advance the state of the art of RINA towards an architecture reference model and specifcations that are closer to enable implementations deployable in production scenarios. The design and implemention of a RINA prototype on top of Ethernet will permit the experimentation and evaluation of RINA in comparison to TCP/IP. IRATI will use the OFELIA testbed to carry on its experimental activities. Both projects will benefit from the collaboration. IRATI will gain access to a large-scale testbed with a controlled network while OFELIA will get a unique use-case to validate the facility: experimentation of a non-IP based Internet

Intestinal Serotonin Transporter Inhibition by Toll-Like Receptor 2 Activation. A Feedback Modulation

This is the final version of the article. Available from Public Library of Science via the DOI in this record.TLR2 is a microbiota recognition receptor that has been described to contribute to intestinal homeostasis and to ameliorate inflammatory intestinal injury. In this context, serotonin (5-HT) has shown to be an essential intestinal physiological neuromodulator that is also involved in intestinal inflammatory diseases. Since the interaction between TLR2 activation and the intestinal serotoninergic system remains non-investigated, our main aim was to analyze the effect of TLR2 on intestinal serotonin transporter (SERT) activity and expression and the intracellular pathways involved. Caco-2/TC7 cells were used to analyze SERT and TLR2 molecular expression and SERT activity by measuring 5-HT uptake. The results showed that apical TLR2 activation inhibits SERT activity in Caco-2/TC7 cells mainly by reducing SERT protein level either in the plasma membrane, after short-term TLR2 activation or in both the plasma membrane and cell lysate, after long-term activation. cAMP/PKA pathway appears to mediate short-term inhibitory effect of TLR2 on SERT; however, p38 MAPK pathway has been shown to be involved in both short- and long-term TLR2 effect. Reciprocally, 5-HT long-term treatment yielded TLR2 down regulation in Caco-2/TC7 cells. Finally, results from in vivo showed an augmented intestinal SERT expression in mice Tlr2-/-, thus confirming our inhibitory effect of TLR2 on intestinal SERT in vitro. The present work infers that TLR2 may act in intestinal pathophysiology, not only by its inherent innate immune role, but also by regulating the intestinal serotoninergic system.This work was funded by grants from the Spanish Ministry of Science and Innovation and the European Regional Development Fund (ERDF/FEDER) (BFU2010-18971), Zaragoza University (UZ2014-BIO-03), European Social Found (ESF) and the Aragon Regional Government (B61) and the Foundation for the Study of Inflammatory Bowel Diseases in Aragón (ARAINF 012/2008). ARAID Foundation supported J.P. (SAF2014-54763-C2-1-R) and E. Layunta is a PhD student fellow from Aragon Regional Government (B022/13). The funders had no role in study design, data collection and analysis, decision to publish, or preparation of the manuscript

Unreliable inter process communication in Ethernet: migrating to RINA with the shim DIF

There is often a requirement to interface a new model to a legacy implementation by creating a shim between them to make the legacy appear as close to the new model as possible. This is a common exercise, usually fraught with frustrations, but here we find the exercise reveals fundamental aspects about nature of layers that were previously not well understood. Here we will be primarily concerned with creating a shim between RINA and IEEE 802.1q (VLANs). The Recursive InterNet Architecture (RINA) proposes a network architecture derived from the fundamentals of InterProcess Communication (IPC). This yields a recursively layered architecture of Distributed IPC Facilities (DIFs)

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

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