Anti-inflammatory potential of CB1-mediated cAMP elevation in mast cells.
Ontology highlight
ABSTRACT:
SUBMITTER: Small-Howard AL
PROVIDER: S-EPMC1138953 | biostudies-other | 2005 Jun
REPOSITORIES: biostudies-other
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Project description:BackgroundMast cells (MCs) are major contributors to an inflammatory milieu. One of the most potent drivers of inflammation is the cytokine IL-1?, which is produced in the cytoplasm in response to danger signals like LPS. Several controlling mechanisms have been reported which limit the release of IL-1?. Central to this regulation is the NLRP3 inflammasome, activation of which requires a second danger signal with the capacity to subvert the homeostasis of lysosomes and mitochondria. High concentrations of extracellular ATP have the capability to perturb the plasma membrane by activation of P2X7 channels and serve as such a danger signal. In this study we investigate the role of P2X7 channels and the ecto-5'-nucleotidase CD39 in ATP-triggered release of IL-1? from LPS-treated mast cells.ResultsWe report that in MCs CD39 sets an activation threshold for the P2X7-dependent inflammatory cell death and concomitant IL-1? release. Knock-out of CD39 or stimulation with non-hydrolysable ATP led to a lower activation threshold for P2X7-dependent responses. We found that stimulation of LPS-primed MCs with high doses of ATP readily induced inflammatory cell death. Yet, cell death-dependent release of IL-1? yielded only minute amounts of IL-1?. Intriguingly, stimulation with low ATP concentrations augmented the production of IL-1? in LPS-primed MCs in a P2X7-independent but caspase-1-dependent manner.ConclusionOur study demonstrates that the fine-tuned interplay between ATP and different surface molecules recognizing or modifying ATP can control inflammatory and cell death decisions.
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Project description:Allosteric modulation of G-protein-coupled receptors represents a key goal of current pharmacology. In particular, endogenous allosteric modulators might represent important targets of interventions aimed at maximizing therapeutic efficacy and reducing side effects of drugs. Here we show that the anti-inflammatory lipid lipoxin A(4) is an endogenous allosteric enhancer of the CB(1) cannabinoid receptor. Lipoxin A(4) was detected in brain tissues, did not compete for the orthosteric binding site of the CB(1) receptor (vs. (3)H-SR141716A), and did not alter endocannabinoid metabolism (as opposed to URB597 and MAFP), but it enhanced affinity of anandamide at the CB1 receptor, thereby potentiating the effects of this endocannabinoid both in vitro and in vivo. In addition, lipoxin A(4) displayed a CB(1) receptor-dependent protective effect against ?-amyloid (1-40)-induced spatial memory impairment in mice. The discovery of lipoxins as a class of endogenous allosteric modulators of CB(1) receptors may foster the therapeutic exploitation of the endocannabinoid system, in particular for the treatment of neurodegenerative disorders.
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Project description:The suppression of activated pro-inflammatory macrophages during immune response has a major impact on the outcome of many inflammatory diseases including sepsis and rheumatoid arthritis. The pro- and anti-inflammatory functions of macrophages have been widely studied, whereas their regulation under immunosuppressive treatments such as glucocorticoid (GC) therapy is less well-understood. GC-mediated glucocorticoid receptor (GR) activation is crucial to mediate anti-inflammatory effects. In addition, the anti-cancer drug roscovitine, that is currently being tested in clinical trials, was recently described to regulate inflammatory processes by inhibiting different Cdks such as cyclin-dependent kinase 5 (Cdk5). Cdk5 was identified as a modulator of inflammatory processes in different immune cells and furthermore described to influence GR gene expression in the brain. Whether roscovitine can enhance the immunosuppressive effects of GCs and if the inhibition of Cdk5 affects GR gene regulatory function in innate immune cells, such as macrophages, has not yet been investigated. Here, we report that roscovitine enhances the immunosuppressive Dexamethasone (Dex) effect on the inducible nitric oxide synthase (iNos) expression, which is essential for immune regulation. Cdk5 deletion in macrophages prevented iNos protein and nitric oxide (NO) generation after a combinatory treatment with inflammatory stimuli and Dex. Cdk5 deletion in macrophages attenuated the GR phosphorylation on serine 211 after Dex treatment alone and in combination with inflammatory stimuli, but interestingly increased the GR-dependent anti-inflammatory target gene dual-specificity phosphatase 1 (Dusp1, Mkp1). Mkp1 phosphatase activity decreases the activation of its direct target p38Mapk, reduced iNos expression and NO production upon inflammatory stimuli and Dex treatment in the absence of Cdk5. Taken together, we identified Cdk5 as a potential novel regulator of NO generation in inflammatory macrophages under GC treatment. Our data suggest that GC treatment in combination with specific Cdk5 inhibtior(s) provides a stronger suppression of inflammation and could thus replace high-dose GC therapy which has severe side effects in the treatment of inflammatory diseases.
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Project description:TRPM4 proteins form Ca(2+)-activated non selective cation (CAN) channels that affect transmembrane Ca(2+)-influx by determining the membrane potential. Tight control of the intracellular Ca(2+) concentration is essential for mast cell responses. In this study, we analyzed the expression of TRPM4 in peritoneal mast cells (PCMC) as a model for connective tissue type mast cells with respect to Fc?RI-evoked calcium changes and the subcellular localization of fluorescently labeled TRPM4 using two viral transduction systems before and following antigen stimulation. Our results show that TRPM4 is expressed in PCMCs, is an essential constituent of the endogenous CAN channels in PCMCs and regulates antigen-evoked increases in intracellular calcium that are significantly enhanced in TRPM4-deficient PCMCs. Compared to PCMCs analyzed before antigen stimulation, the cells depict a substantially increased localization of TRPM4 proteins towards the plasma membrane after Fc?RI stimulation. Thus, TRPM4 functions as a limiting factor for antigen evoked calcium rise in connective tissue type mast cells and concurrent translocation of TRPM4 into the plasma membrane is part of this mechanism.
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