title: Prostacyclin prevents nitric oxide-induced megakaryocyte apoptosis.
abstract: 1 We have previously demonstrated that nitric oxide (NO) triggers CD34(+)-derived megakaryocyte apoptosis. We here show that prostacyclin (PGI(2)) inhibits PAPA/NO-induced megakaryocyte death detected by fluorescent microscopy and flow cytometry. 2 The cAMP-specific phosphodiesterase inhibitor, Ro 20-1724, and the permeable analog dibutyryl-cAMP also delayed apoptosis. PGI(2) effect was fully prevented when adenylyl cyclase activity was suppressed by SQ 22536, and partially reversed by the permeable protein kinase A inhibitor PKI 14-22 amide. ELISA showed that while both PGI(2) and NO alone or synergistically raised cAMP, only NO was able to increase intracellular cGMP levels. 3 Treatment of megakaryocytes with PGI(2) abolished both basal and NO-raised cGMP levels. Addition of 8-pCPT-cGMP or activation of soluble guanylyl cyclase by BAY 41-2272 induced cell death in a concentration-dependent manner, and ODQ, an inhibitor of guanylyl cyclase, prevented both PAPA/NO- or BAY 41-2272-induced apoptosis. Specific cGMP phosphodiesterase inhibition by Zaprinast or suppression of adenylyl cyclase by SQ 22536 enhanced the PAPA/NO proapoptotic effect. 4 PGI(2) completely inhibited NO-mediated generation and the increased activity of the cleaved form of caspase-3. 5 In conclusion, our results demonstrate that contrary to their well-known direct and synergistic inhibitory effects on platelets, PGI(2) and NO regulate opposite megakaryocyte survival responses through a delicate balance between intracellular cyclic nucleotide levels and caspase-3 activity control.
authors.affiliation: Department of Thrombosis and Haemostasis, Hematological Research Institute, National Academy of Medicine, National Research Council (CONICET), Pacheco de Melo 3081 (1425), Buenos Aires, Argentina.
authors.forename: Roberto Gabriel
authors.initials: RG
authors.lastname: Pozner
authors.forename: Soledad
authors.initials: S
authors.lastname: Negrotto
authors.forename: Lina Paola
authors.initials: LP
authors.lastname: D'Atri
authors.forename: Mónica Lidia
authors.initials: ML
authors.lastname: Kotler
authors.forename: María Angela
authors.initials: MA
authors.lastname: Lazzari
authors.forename: Ricardo Martin
authors.initials: RM
authors.lastname: Gomez
authors.forename: Mirta
authors.initials: M
authors.lastname: Schattner
mesh_terms: D017209:Apoptosis / Q000187:drug effects* / Q000502:physiology; D002478:Cells, Cultured; D004305:Dose-Response Relationship, Drug; D004730:Endothelium, Vascular / Q000187:drug effects / Q000378:metabolism; D011464:Epoprostenol / Q000494:pharmacology*; D006801:Humans; D008533:Megakaryocytes / Q000187:drug effects* / Q000378:metabolism; D009569:Nitric Oxide / Q000037:antagonists & inhibitors* / Q000494:pharmacology*
chemical_list: D009569:Nitric Oxide; D011464:Epoprostenol