title: ATP-induced apoptosis involves a Ca2+-independent phospholipase A2 and 5-lipoxygenase in macrophages.
abstract: Macrophages express P2X(7) and other nucleotide (P2) receptors, and display the phenomena of extracellular ATP (ATP(e))-induced P2X(7)-dependent membrane permeabilization and cell death by apoptosis and necrosis. P2X(7) receptors also cooperate with toll-like receptors (TLRs) to induce inflammasome activation and IL-1beta secretion. We investigated signaling pathways involved in the induction of cell death by ATP(e) in intraperitoneal murine macrophages. Apoptosis (hypodiploid nuclei) and necrosis (LDH release) were detected 6h after an induction period of 20 min in the presence of ATP. Apoptosis was blocked by caspase 3 and caspase 9 inhibitors and by cyclosporin A. The MAPK inhibitors PD-98059, SB-203580 and SB-202190 provoked no significant effect on apoptosis, but SB-203580 blocked LDH release. Neither apoptosis nor necrosis was inhibited when both intra- and extracellular Ca(2+) were chelated during the induction period. Mepacrine, a generic PLA(2) inhibitor and BEL, an inhibitor of Ca(2+)-independent PLA(2) (iPLA(2)) blocked apoptosis, while pBPB and AACOOPF(3), inhibitors of secretory and Ca(2+)-dependent PLA(2) respectively, had no significant effect. Cycloxygenase inhibitors had no effect on apoptosis, while the inhibitors of lipoxygenase (LOX) and leukotriene biosynthesis nordihydroguaiaretic acid (NDGA), zileuton, AA-861, and MK-886 significantly decreased apoptosis. Neither NDGA nor MK-886 blocked apoptosis of 5-LOX(-/-) macrophages. CP-105696 and MK-571, antagonists of leukotriene receptors, had no significant effect on apoptosis. None of the inhibitors of PLA(2) and LOX/leukotriene pathway had a significant inhibitory effect on LDH release. Our results indicate that a Ca(2+)-independent step involving an iPLA(2) and 5-LOX are involved in the triggering of apoptosis but not necrosis by P2X(7) in macrophages.
authors.affiliation: Laboratório de Imunobiofísica, Instituto de Biofísica Carlos Chagas Filho, Universidade Federal do Rio de Janeiro, 21941-902 Rio de Janeiro, RJ, Brazil.
authors.forename: Helio Miranda
authors.initials: HM
authors.lastname: Costa-Junior
authors.forename: Anderson Nogueira
authors.initials: AN
authors.lastname: Mendes
authors.forename: Gustavo Henrique Nolasco Grimmer
authors.initials: GH
authors.lastname: Davis
authors.forename: Cristiane Monteiro
authors.initials: CM
authors.lastname: da Cruz
authors.forename: Ana Lúcia Marques
authors.initials: AL
authors.lastname: Ventura
authors.forename: Carlos Henrique
authors.initials: CH
authors.lastname: Serezani
authors.forename: Lucia Helena
authors.initials: LH
authors.lastname: Faccioli
authors.forename: Auro
authors.initials: A
authors.lastname: Nomizo
authors.forename: Célio G
authors.initials: CG
authors.lastname: Freire-de-Lima
authors.forename: Rodrigo da Cunha
authors.initials: Rda C
authors.lastname: Bisaggio
authors.forename: Pedro Muanis
authors.initials: PM
authors.lastname: Persechini
mesh_terms: D000255:Adenosine Triphosphate / Q000378:metabolism / Q000494:pharmacology*; D000818:Animals; D017209:Apoptosis; D001094:Arachidonate 5-Lipoxygenase / Q000378:metabolism*; D002118:Calcium / Q000378:metabolism; D016923:Cell Death; D008264:Macrophages / Q000201:enzymology*; D051379:Mice; D008810:Mice, Inbred C57BL; D020928:Mitogen-Activated Protein Kinases / Q000378:metabolism; D054512:Phospholipases A2, Calcium-Independent / Q000378:metabolism*
chemical_list: D000255:Adenosine Triphosphate; D001094:Arachidonate 5-Lipoxygenase; D020928:Mitogen-Activated Protein Kinases; D054512:Phospholipases A2, Calcium-Independent; D002118:Calcium