Evidence for nucleotide receptor modulation of cross talk between MAP kinase and NF-κB signaling pathways in murine RAW 264.7 macrophages

Abstract

. Evidence for nucleotide receptor modulation of cross talk between MAP kinase and NF-B signaling pathways in murine RAW 264.7 macrophages. Am J Physiol Cell Physiol 286: C923-C930, 2004. First published December 18, 2003; 10.1152/ajpcell.00417.2003.-Extracellular nucleotides such as ATP are present in abundance at sites of inflammation and tissue damage, and these agents exert a potent modulatory effect on macrophage/monocyte function via the nucleotide receptor P2X 7. In this regard, after exposure to bacterial LPS, P2X 7 activation augments expression of the inducible nitric oxide (NO) synthase and production of NO in macrophages. Because P2X 7 has been reported to stimulate certain members of the MAP kinase family (ERK1/2) and can enhance the DNA-binding activity of NF-B, we tested the hypothesis that LPS and nucleotides regulate NF-B-dependent inflammatory events via cross talk with MAPK-associated pathways. In this regard, the present studies revealed that cotreatment of macrophages with LPS and the P2X 7-selective ligand 2Ј-3Ј-O-(4-benzoylbenzoyl)adenosine 5Ј-triphosphate (BzATP) results in the cooperative activation of NF-B DNA-binding activity and a sustained attenuation of levels of the NF-B inhibitory protein IB␣. Interestingly, a persistent reduction in IB␣ levels is also observed when the MEK1/2 inhibitor U0126 is coadministered with LPS, suggesting that components of the MEK/ERK pathway are involved in regulating IB␣ protein expression and/or turnover. The observation that U0126 and BzATP exhibit overlapping actions with respect to LPS-induced changes in IB␣ levels is supported by the finding that Ras activation, which is upstream of MEK/ERK activation, is reduced upon macrophage cotreatment with BzATP and LPS compared with the effects of BzATP treatment alone. These data are consistent with the concept that the Ras/MEK/ERK pathways are involved in regulating NF-B/ IB-dependent inflammatory mediator production and suggest a previously unidentified mechanism by which nucleotides can modulate LPS-induced action via cross talk between NF-B and Ras/MEK/ MAPK-associated pathways. nucleotide receptors; mitogen-activated protein kinases; nuclear factor-B; monocytes/macrophages; cytokines ALTHOUGH ATP IS KNOWN to be an important source of intracellular energy, numerous studies have revealed that high concentrations of ATP (5 mM) in the extracellular milieu following tissue damage or platelet degranulation can play a key role in the regulation of biological responses such as inflammation, platelet aggregation, and smooth muscle contraction (13,20,25,38). Nucleotides have been shown to greatly enhance the effects of bacterial LPS on macrophage and monocyte activation by augmenting the production of mediators such as nitric oxide (NO) and other free radicals, in addition to numerous cytokines, such as interleukin (IL)-1␤ and tumor necrosis factor (TNF)-␣ (26, 27, 39, 51, 55). The overproduction of these mediators, although important for the activation of the immune system and for bactericidal effects, can als...

P2X receptors regulate adenosine diphosphate release from hepatic cells

“…Extracellular nucleotides are known to activate nuclear factor kappa B [3,4] and trigger the release of proinflammatory cytokines [5,6]. Elevated circulating nucleotide levels and sustained purinergic signaling may consequently promote inflammatory diseases [7,8].…”

An indomethacin analogue, N-(4-chlorobenzoyl)-melatonin, is a selective inhibitor of aldo-keto reductase 1C3 (type 2 3α-HSD, type 5 17β-HSD, and prostaglandin F synthase), a potential target for the treatment of hormone dependent and hormone independent malignancies

“…Of its endogenously relevant substrates, AKR1C3 has the highest catalytic activity for the reduction of PGD 2 [9;10]. The PGF 2 isomers will activate the Gq-coupled F-prostanoid receptor and initiate protein kinase C and MAPK signaling cascades that stimulate proliferation through mechanisms that include inhibition of the peroxisome proliferator-activated receptor γ (PPARγ) and activation of NF-κB [11][12][13][14]. The AKR1C3-mediated depletion of PGD 2 will also prevent the formation of anti-proliferative PGJ 2 isomers, including 15-deoxy-Δ12,14-PGJ 2 , which are natural PPARγ ligands and inhibitors of NF-κB signaling [15][16][17][18].…”

Cell signaling via the P2X7 nucleotide receptor: linkage to ROS production, gene transcription, and receptor trafficking

“…The transactivation of c-Jun is augmented by the phosphorylation of Ser-63 and -73 by JNK1/2. Studies from our lab and others have noted JNK1/ 2 activation following stimulation of monocytic cells with P2X 7 agonists [21,68,95]. Furthermore, co-stimulation of RAW 264.7 cells with LPS and BzATP results in enhanced JNK activation compared to either stimulus alone [68].…”

“…Studies from our lab and others have noted JNK1/ 2 activation following stimulation of monocytic cells with P2X 7 agonists [21,68,95]. Furthermore, co-stimulation of RAW 264.7 cells with LPS and BzATP results in enhanced JNK activation compared to either stimulus alone [68]. Of note, P2X 7 agonist-induced JNK activation in RAW 264.7 cells is attenuated by N-acetylcysteine and ascorbic acid, implicating a role for ROS in P2X 7 -dependent AP-1 activation [21].…”

“…Past studies have shown that P2X 7 agonists are able to modulate macrophage activation in response to LPS, resulting in increased pro-inflammatory mediator production [17,18,23,56,[67][68][69][70][71][72][73][74][75][76]. Because LPS is known to activate ROS production in leukocytes [77,78], and because P2X 7 activation is able to augment several LPSinduced signaling events, we tested the hypothesis that P2X 7 -stimulated ROS production is enhanced in LPSprimed macrophages.…”