title: Naja nigricollis CMS-9 enhances the mitochondria-mediated death pathway in adaphostin-treated human leukaemia U937 cells.
abstract: 1. The aim of the present study was to explore the effect of the Naja nigricollis phospholipase A(2) CMS-9 on adaphostin-induced death of human leukaemia U937 cells. 2. Leukaemia U937 cells (Bcr/Abl-negative cells) were treated with adaphostin (0-10 μmol/L) and CMS-9 (0-1 μmol/L). The effects of CMS-9, adaphostin and their combination on cell viability, the generation reactive oxygen species (ROS), [Ca(2+) ](i) , p38 mitogen-activated protein kinase (MAPK) activation, Akt and extracellular signal-regulated kinase (ERK) inactivation, mitochondrial membrane potential (ΔΨ(m) ) and Bcl-2 family proteins were analysed. 3. Both adaphostin and CMS-9 induced U937 cell apoptosis, characterized by dissipation of ΔΨ(m) and ROS generation. Combined treatment further increased ΔΨ(m) loss and reduced the viability of adaphostin-treated cells. Unlike in CMS-9-treated cells, in adaphostin-treated cells ROS-induced increases in [Ca(2+) ](i) were observed. CMS-9-induced ROS generation resulted in p38 MAPK activation, whereas adaphostin treatment elicited ROS/Ca(2+) -mediated inactivation of Akt and ERK. Moreover, Akt was found to be involved in ERK phosphorylation. Suppression of p38 MAPK activation blocked CMS-9-induced ΔΨ(m) loss and Bcl-xL downregulation. Overexpression of constitutively active Akt and mitogen-activated protein kinase kinase (MEK) 1 rescued adaphostin-induced ΔΨ(m) loss and Bcl-2 downregulation. Similarly, CMS-9 augmented adaphostin toxicity in human leukaemia K562 cells via increased mitochondrial alterations. 4. The results suggest that two distinct pathways mediate adaphostin- and CMS-9-induced mitochondrial damage (i.e. the ROS-Ca(2+) -Akt-ERK and ROS-p38 MAPK pathways, respectively). These distinct pathway explain the augmentation by CMS-9 of ΔΨ(m) loss and apoptosis in adaphostin-treated U937 cells.
authors.affiliation: Institute of Biomedical Sciences, National Sun Yat-Sen University, Kaohsiung Medical University, Taiwan.
authors.forename: Ying-Jung
authors.initials: YJ
authors.lastname: Chen
authors.forename: Jeh-Jeng
authors.initials: JJ
authors.lastname: Wang
authors.forename: Long-Sen
authors.initials: LS
authors.lastname: Chang
mesh_terms: D000218:Adamantane / Q000031:analogs & derivatives* / Q000494:pharmacology; D000818:Animals; D017209:Apoptosis / Q000187:drug effects* / Q000235:genetics; D051017:Apoptosis Regulatory Proteins / Q000378:metabolism; D002118:Calcium / Q000378:metabolism; D016923:Cell Death / Q000187:drug effects / Q000235:genetics; D002470:Cell Survival / Q000187:drug effects / Q000235:genetics; D004546:Elapid Venoms / Q000494:pharmacology*; D048049:Extracellular Signal-Regulated MAP Kinases / Q000235:genetics / Q000378:metabolism; D006801:Humans; D006873:Hydroquinones / Q000494:pharmacology*; D020014:K562 Cells; D007938:Leukemia / Q000188:drug therapy / Q000378:metabolism* / Q000473:pathology; D053078:Membrane Potential, Mitochondrial / Q000187:drug effects / Q000235:genetics; D008928:Mitochondria / Q000187:drug effects* / Q000378:metabolism*; D054467:Phospholipases A2 / Q000494:pharmacology*; D010766:Phosphorylation / Q000187:drug effects / Q000235:genetics; D051057:Proto-Oncogene Proteins c-akt / Q000235:genetics / Q000378:metabolism; D019253:Proto-Oncogene Proteins c-bcl-2 / Q000378:metabolism; D017382:Reactive Oxygen Species / Q000378:metabolism; D014407:Tumor Cells, Cultured; D020298:U937 Cells; D048051:p38 Mitogen-Activated Protein Kinases / Q000378:metabolism
chemical_list: D051017:Apoptosis Regulatory Proteins; C550860:CMS-9 protein, Naja nigricollis; D004546:Elapid Venoms; D006873:Hydroquinones; C404397:NSC 680410; D019253:Proto-Oncogene Proteins c-bcl-2; D017382:Reactive Oxygen Species; D051057:Proto-Oncogene Proteins c-akt; D048049:Extracellular Signal-Regulated MAP Kinases; D048051:p38 Mitogen-Activated Protein Kinases; D054467:Phospholipases A2; D000218:Adamantane; D002118:Calcium