Background and Purpose
Tobacco smoke contains many classes of carcinogens and although nicotine is unable to initiate tumourigenesis in humans and rodents, it promotes tumour growth and metastasis in ...lung tumours by acting on neuronal nicotinic ACh receptors (nAChRs). The aim of this study was to identify molecularly, biochemically and pharmacologically which nAChR subtypes are expressed and functionally activated by nicotine in lung cancer cell lines.
Experimental Approach
We used A549 and H1975 adenocarcinoma cell lines derived from lung tumours to test the in vitro effects of nicotine, and nAChR subtype‐specific peptides and compounds.
Key Results
The two adenocarcinoma cell lines express distinctive nAChR subtypes, and this affects their nicotine‐induced proliferation. In A549 cells, nAChRs containing the α7 or α9 subunits not only regulate nicotine‐induced cell proliferation but also the activation of the Akt and ERK pathways. Blocking these nAChRs by means of subtype‐specific peptides, or silencing their expression by means of subunit‐specific siRNAs, abolishes nicotine‐induced proliferation and signalling. Moreover, we found that the α7 antagonist MG624 also acts on α9–α10 nAChRs, blocks the effects of nicotine on A549 cells and has dose‐dependent cytotoxic activity.
Conclusions and Implications
These results highlight the pathophysiological role of α7‐ and α9‐containing receptors in promoting non‐small cell lung carcinoma cell growth and intracellular signalling and provide a framework for the development of new drugs that specifically target the receptors expressed in lung tumours.
Linked Articles
This article is part of a themed section on Nicotinic Acetylcholine Receptors. To view the other articles in this section visit http://onlinelibrary.wiley.com/doi/10.1111/bph.v175.11/issuetoc
Neuronal nicotinic acetylcholine receptors (nAChRs) are a family of ligand-gated ion channels present in the central and peripheral nervous systems, that are permeable to mono- and divalent cations. ...They share a common basic structure but their pharmacological and functional properties arise from the wide range of different subunit combinations making up distinctive subtypes.
nAChRs are involved in many physiological functions in the central and peripheral nervous systems, and are the targets of the widely used drug of abuse nicotine. In addition to tobacco dependence, changes in their number and/or function are associated with neuropsychiatric disorders, ranging from epilepsy to dementia.
Although some of the neural circuits involved in the acute and chronic effects of nicotine have been identified, much less is known about which native nAChR subtypes are involved in specific physiological functions and pathophysiological conditions.
We briefly review some recent findings concerning the structure and function of native nAChRs, focusing on the subtypes identified in the mesostriatal and habenulo-interpeduncular pathways, two systems involved in nicotine reinforcement and withdrawal. We also discuss recent findings concerning the effect of chronic nicotine on the expression of native subtypes.
Vagus nerve stimulation (VNS), most likely via enteric neurons, prevents postoperative ileus (POI) by reducing activation of alpha7 nicotinic receptor (α7nAChR) positive
macrophages (mMφ) and ...dampening surgery-induced intestinal inflammation. Here, we evaluated if 5-HT4 receptor (5-HT4R) agonist prucalopride can mimic this effect in mice and human.
Using Ca
imaging, the effect of electrical field stimulation (EFS) and prucalopride was evaluated in situ on mMφ activation evoked by ATP in jejunal
tissue. Next, preoperative and postoperative administration of prucalopride (1-5 mg/kg) was compared with that of preoperative VNS in a model of POI in wild-type and α7nAChR knockout mice. Finally, in a pilot study, patients undergoing a Whipple procedure were preoperatively treated with prucalopride (n=10), abdominal VNS (n=10) or sham/placebo (n=10) to evaluate the effect on intestinal inflammation and clinical recovery of POI.
EFS reduced the ATP-induced Ca
response of mMφ, an effect that was dampened by neurotoxins tetrodotoxin and ω-conotoxin and mimicked by prucalopride. In vivo, prucalopride administered before, but not after abdominal surgery reduced intestinal inflammation and prevented POI in wild-type, but not in α7nAChR knockout mice. In humans, preoperative administration of prucalopride, but not of VNS, decreased
and
expression in the
and improved clinical recovery.
Enteric neurons dampen mMφ activation, an effect mimicked by prucalopride. Preoperative, but not postoperative treatment with prucalopride prevents intestinal inflammation and shortens POI in both mice and human, indicating that preoperative administration of 5-HT4R agonists should be further evaluated as a treatment of POI.
NCT02425774.
The post-mortem diagnosis of acute myocardial ischemia remains a challenge for both clinical and forensic pathologists. We performed an experimental study (ligation of left anterior descending ...coronary artery in rats) in order to identify early markers of myocardial ischemia, to further apply to forensic and clinical pathology in cases of sudden cardiac death. Using immunohistochemistry, Western blots, and gene expression analyses, we investigated a number of markers, selected among those which are currently used in emergency departments to diagnose myocardial infarction and those which are under investigation in basic research and autopsy pathology studies on cardiovascular diseases. The study was performed on 44 adult male Lewis rats, assigned to three experimental groups: control, sham-operated, and operated. The durations of ischemia ranged between 5 min and 24 h. The investigated markers were troponins I and T, myoglobin, fibronectin, C5b-9, connexin 43 (dephosphorylated), JunB, cytochrome c, and TUNEL staining. The earliest expressions (≤30 min) were observed for connexin 43, JunB, and cytochrome c, followed by fibronectin (≤1 h), myoglobin (≤1 h), troponins I and T (≤1 h), TUNEL (≤1 h), and C5b-9 (≤2 h). By this investigation, we identified a panel of true early markers of myocardial ischemia and delineated their temporal evolution in expression by employing new technologies for gene expression analysis, in addition to traditional and routine methods (such as histology and immunohistochemistry). Moreover, for the first time in the autopsy pathology field, we identified, by immunohistochemistry, two very early markers of myocardial ischemia: dephosphorylated connexin 43 and JunB.
Control of synapse number and function in the developing central nervous system is critical to the formation of neural circuits. Astrocytes play a key role in this process by releasing factors that ...promote the formation of excitatory synapses. Astrocyte‐secreted thrombospondins (TSPs) induce the formation of structural synapses, which however remain post‐synaptically silent, suggesting that completion of early synaptogenesis may require a two‐step mechanism. Here, we show that the humoral innate immune molecule Pentraxin 3 (PTX3) is expressed in the developing rodent brain. PTX3 plays a key role in promoting functionally‐active CNS synapses, by increasing the surface levels and synaptic clustering of AMPA glutamate receptors. This process involves tumor necrosis factor‐induced protein 6 (TSG6), remodeling of the perineuronal network, and a β1‐integrin/ERK pathway. Furthermore, PTX3 activity is regulated by TSP1, which directly interacts with the N‐terminal region of PTX3. These data unveil a fundamental role of PTX3 in promoting the first wave of synaptogenesis, and show that interplay of TSP1 and PTX3 sets the proper balance between synaptic growth and synapse function in the developing brain.
Synopsis
Pentraxin 3 (PTX3) is a soluble innate immunity molecule that plays a key role in recognition and binding of microbial moieties and complement components. PTX3 also interacts with extracellular matrix components and contributes to the structural three‐dimensional organization of the extracellular matrix.
PTX3 is expressed and released by glial cells in the developing rodent brain.
PTX3 promotes formation and maturation of excitatory synapses by enhancing the synaptic recruitment of AMPARs.
PTX3 regulates synaptic AMPARs through the remodeling of ECM surrounding excitatory synapses.
Glial‐derived thrombospondin‐1 (TSP1) regulates PTX3 activity through direct interaction.
Murine astrocytes release innate immunity‐associated PTX3 at early postnatal stages, where it plays a key role in the functional maturation of excitatory synapses by promoting AMPAR recruitment at synapses.
Rationale
Prolinol aryl ethers and their rigidified analogues pyrrolidinyl benzodioxanes have a high affinity for mammalian α4β2 nicotinic acetylcholine receptors (nAChRs). Electrophysiological ...studies have shown that the former are full agonists and the latter partial agonists or antagonists of human α4β2 receptors, but their in vivo effects are unknown.
Objectives and methods
As α4β2 nAChRs play an important role in the cognition and the rewarding effects of nicotine, we tested the effects of two full agonists and one antagonist on spatial learning, memory and attention in zebrafish using a T-maze task and virtual object recognition test (VORT). The effect of a partial agonist in reducing nicotine-induced conditioned place preference (CPP) was also investigated.
Results
In comparison with the vehicle alone, the full agonists MCL-11 and MCL-28 induced a significant cognitive enhancement as measured by the reduced running time in the T-maze and increased attention as measured by the increased discrimination index in the VORT. MCL-11 was 882 times more potent than nicotine. The two compounds were characterised by an inverted U-shaped dose-response curve, and their effects were blocked by the co-administration of the antagonist MCL-117, which alone had no effect. The partial agonist MCL-54 induced CPP and had an inverted U-shaped dose-response curve similar to that of nicotine but blocked the reinforcing effect of co-administered nicotine. Binding studies showed that all of the compounds have a higher affinity for heteromeric
3
H-epibatidine receptors than
125
I-αBungarotoxin receptors. MCL-11 was the most selective of heteromeric receptors.
Conclusions
These behavioural studies indicate that full agonist prolinol aryl ethers are very active in increasing spatial learning, memory and attention in zebrafish. The benzodioxane partial agonist MCL-54 reduced nicotine-induced CPP, and the benzodioxane antagonist MCL-117 blocked all agonist-induced activities.
We examined α7β2-nicotinic acetylcholine receptor (α7β2-nAChR) expression in mammalian brain and compared pharmacological profiles of homomeric α7-nAChRs and α7β2-nAChRs. α-Bungarotoxin affinity ...purification or immunoprecipitation with anti-α7 subunit antibodies (Abs) was used to isolate nAChRs containing α7 subunits from mouse or human brain samples. α7β2-nAChRs were detected in forebrain, but not other tested regions, from both species, based on Western blot analysis of isolates using β2 subunit-specific Abs. Ab specificity was confirmed in control studies using subunit-null mutant mice or cell lines heterologously expressing specific human nAChR subtypes and subunits. Functional expression in Xenopus oocytes of concatenated pentameric (α7)5-, (α7)4(β2)1-, and (α7)3(β2)2-nAChRs was confirmed using two-electrode voltage clamp recording of responses to nicotinic ligands. Importantly, pharmacological profiles were indistinguishable for concatenated (α7)5-nAChRs or for homomeric α7-nAChRs constituted from unlinked α7 subunits. Pharmacological profiles were similar for (α7)5-, (α7)4(β2)1-, and (α7)3(β2)2-nAChRs except for diminished efficacy of nicotine (normalized to acetylcholine efficacy) at α7β2- versus α7-nAChRs. This study represents the first direct confirmation of α7β2-nAChR expression in human and mouse forebrain, supporting previous mouse studies that suggested relevance of α7β2-nAChRs in Alzheimer disease etiopathogenesis. These data also indicate that α7β2-nAChR subunit isoforms with different α7/β2 subunit ratios have similar pharmacological profiles to each other and to α7 homopentameric nAChRs. This supports the hypothesis that α7β2-nAChR agonist activation predominantly or entirely reflects binding to α7/α7 subunit interface sites.
We present the first high-quality catalog of early aftershocks of the three mainshocks of the 2016 central Italy Amatrice-Visso-Norcia normal faulting sequence. We located 10,574 manually picked ...aftershocks with a robust probabilistic, non-linear method achieving a significant improvement in the solution accuracy and magnitude completeness with respect to previous studies. Aftershock distribution and relocated mainshocks give insight into the complex architecture of major causative and subsidiary faults, thus providing crucial constraints on multi-segment rupture models. We document reactivation and kinematic inversion of a WNW-dipping listric structure, referable to the inherited Mts Sibillini Thrust (MST) that controlled segmentation of the causative normal faults. Spatial partitioning of aftershocks evidences that the MST lateral ramp had a dual control on rupture propagation, behaving as a barrier for the Amatrice and Visso mainshocks, and later as an asperity for the Norcia mainshock. We hypothesize that the Visso mainshock re-activated also the deep part of an optimally oriented preexisting thrust. Aftershock patterns reveal that the Amatrice Mw5.4 aftershock and the Norcia mainshock ruptured two distinct antithetic faults 3-4 km apart. Therefore, our results suggest to consider both the MST cross structure and the subsidiary antithetic fault in the finite-fault source modelling of the Norcia earthquake.
Starting from late May 2012, the Emilia region (Northern Italy) was severely shaken by an intense seismic sequence, originated from a ML 5.9 earthquake on May 20th, at a hypocentral depth of 6.3km, ...with thrust-type focal mechanism. In the following days, the seismic rate remained high, counting 50 ML≥2.0 earthquakes a day, on average. Seismicity spreads along a 30km east–west elongated area, in the Po river alluvial plain, in the nearby of the cities Ferrara and Modena. Nine days after the first shock, another destructive thrust-type earthquake (ML 5.8) hit the area to the west, causing further damage and fatalities. Aftershocks following this second destructive event extended along the same east-westerly trend for further 20km to the west, thus illuminating an area of about 50km in length, on the whole. After the first shock struck, on May 20th, a dense network of temporary seismic stations, in addition to the permanent ones, was deployed in the meizoseismal area, leading to a sensible improvement of the earthquake monitoring capability there. A combined dataset, including three-component seismic waveforms recorded by both permanent and temporary stations, has been analyzed in order to obtain an appropriate 1-D velocity model for earthquake location in the study area. Here we describe the main seismological characteristics of this seismic sequence and, relying on refined earthquakes location, we make inferences on the geometry of the thrust system responsible for the two strongest shocks.
•First complete analysis of the 2012 Emilia mainshocks–aftershocks seismic sequence.•New catalog of the seismic sequence including data from temporary stations•Sensible improvement in terms of the activated fault system geometry definition•The different dip of the activated fault segments is highlighted and discussed.•Basic information are provided for further specific studies and hazard scenarios.
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