title: Pharmacological characterization of ATP- and LPS-induced IL-1beta release in human monocytes.
abstract: 1. We have utilized the human monocytic cell line, THP-1, and freshly isolated adherent human monocytes with the compounds pyridoxalphosphate-6-azophenyl-2',4'-disuphonic acid (PPADS), oxidized ATP, and 1-(N, O-bis[5-isoquinolinesufonyll]-N-methyl-L-tyrosyl)-4-phenylpiper  azi ne (KN-62) to pharmacologically characterize the P2 receptor involved in ATP-induced release of interleukin 1beta (IL-1beta). We have also investigated the involvement of P2 receptors in lipopolysaccharide (LPS)-induced IL-1beta release from both cell types. 2. ATP caused release of IL-1beta from LPS primed THP-1 cells in both a time- and concentration-dependent manner, with a minimal effective ATP concentration of 1 mM. Stimulation of cells with 5 mM ATP resulted in detectable concentrations of IL-1beta in cell supernatants within 30 min. 3. The ATP analogue benzoylbenzoyl ATP (DBATP), a P2X7 receptor agonist, was approximately 10 fold more potent than ATP at eliciting IL-1beta release. 4. KN-62 (1 micro M), PPADS (100 microM) or oxidized ATP (100 uM) significantly inhibited 5 mM ATP-induced IL-1beta release by 81, 90 and 66% respectively, but failed to significantly inhibit LPS-induced IL-1beta release in both THP-1 cells and in freshly isolated human monocytes. 5. In both THP-1 cells and freshly isolated human monocytes, addition of the ATP degrading enzyme apyrase (0.4 U ml(-1)) to cell supernatants prior to LPS activation failed to significantly inhibit the LPS-induced IL-1beta release. In addition there was no correlation between extracellular ATP concentrations and IL-1beta release in THP-1 cells when studied over a 6 h time period. 6. In conclusion our data confirm the involvement of P2X7 receptors in ATP-induced IL-1beta release in human monocytes. However no evidence was obtained which would support the involvement of either endogenous ATP release or P2X7 receptor activation as the mechanism by which LPS-induces IL-1beta release in either the THP-1 cell line or in freshly isolated human monocytes.
authors.affiliation: Glaxo Institute of Applied Pharmacology, Department of Pharmacology, University of Cambridge, Tennis Court Rd, Cambridge CB2 1QJ.
authors.forename: C B
authors.initials: CB
authors.lastname: Grahames
authors.forename: A D
authors.initials: AD
authors.lastname: Michel
authors.forename: I P
authors.initials: IP
authors.lastname: Chessell
authors.forename: P P
authors.initials: PP
authors.lastname: Humphrey
mesh_terms: D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine / Q000031:analogs & derivatives / Q000494:pharmacology; D000255:Adenosine Triphosphate / Q000031:analogs & derivatives / Q000378:metabolism / Q000494:pharmacology*; D002460:Cell Line; D004791:Enzyme Inhibitors / Q000494:pharmacology; D006801:Humans; D007375:Interleukin-1 / Q000378:metabolism*; D007770:L-Lactate Dehydrogenase / Q000187:drug effects / Q000378:metabolism; D008070:Lipopolysaccharides / Q000494:pharmacology*; D009000:Monocytes / Q000187:drug effects* / Q000378:metabolism; D058919:Purinergic P2 Receptor Antagonists; D011732:Pyridoxal Phosphate / Q000031:analogs & derivatives / Q000494:pharmacology; D018048:Receptors, Purinergic P2 / Q000187:drug effects*; D058486:Receptors, Purinergic P2X7
chemical_list: D004791:Enzyme Inhibitors; D007375:Interleukin-1; D008070:Lipopolysaccharides; C546468:P2RX7 protein, human; D058919:Purinergic P2 Receptor Antagonists; D018048:Receptors, Purinergic P2; D058486:Receptors, Purinergic P2X7; C077792:pyridoxal phosphate-6-azophenyl-2',4'-disulfonic acid; C017199:2',3'-dialdehyde ATP; D011732:Pyridoxal Phosphate; C063302:KN 62; D019307:1-(5-Isoquinolinesulfonyl)-2-Methylpiperazine; D000255:Adenosine Triphosphate; D007770:L-Lactate Dehydrogenase