title: The NFĸB pathway inhibitors Bay 11-7082 and parthenolide induce programmed cell death in anucleated Erythrocytes.
abstract: The preclinical compounds Bay 11-7082 and parthenolide trigger apoptosis, an effect contributing to their antiinflammatory action. The substances interfere with the activation and nuclear translocation of nuclear factor NFκB, by inhibiting NFκB directly (parthenolide) or by interfering with the inactivation of the NFκB inhibitory protein IκB-α (Bay 11-7082). Beyond that, the substances may be effective in part by nongenomic effects. Similar to apoptosis of nucleated cells, erythrocytes may undergo apoptosis-like cell death (eryptosis) characterized by cell membrane scrambling with phosphatidylserine exposure, and cell shrinkage. Thus, erythrocytes allow the study of nongenomic mechanisms contributing to suicidal cell death, e.g. Ca(2+) leakage or glutathione depletion. The present study utilized Western blotting to search for NFκB and IκB-α expression in erythrocytes, FACS analysis to determine cytosolic Ca(2+) (Fluo3 fluorescence), phosphatidylserine exposure (annexin V binding), and cell volume (forward scatter), as well as an enzymatic method to determine glutathione levels. As a result, both NFκB and IκB-α are expressed in erythrocytes. Targeting the NFκB pathway by Bay 11-7082 (IC(50) ≈ 10 μM) and parthenolide (IC(50) ≈ 30 μM) triggered suicidal erythrocyte death as shown by annexin V binding and decrease of forward scatter. Bay 11-7082 treatment further increased intracellular Ca(2+) and led to depletion of reduced glutathione. The effects of Bay 11-7082 and parthenolide on annexin V binding could be fully reversed by the antioxidant N-acetylcysteine. In conclusion, the pharmacological inhibitors of NFκB, Bay 11-7082 and parthenolide, interfere with the survival of erythrocytes involving mechanisms other than disruption of NFκB-dependent gene expression.
authors.affiliation: Department of Physiology, University of Tübingen, Tübingen, Germany.
authors.forename: Mehrdad
authors.initials: M
authors.lastname: Ghashghaeinia
authors.forename: Mahmoud
authors.initials: M
authors.lastname: Toulany
authors.forename: Mohammad
authors.initials: M
authors.lastname: Saki
authors.forename: Diwakar
authors.initials: D
authors.lastname: Bobbala
authors.forename: Birgit
authors.initials: B
authors.lastname: Fehrenbacher
authors.forename: Rudolf
authors.initials: R
authors.lastname: Rupec
authors.forename: H Peter
authors.initials: HP
authors.lastname: Rodemann
authors.forename: Kamran
authors.initials: K
authors.lastname: Ghoreschi
authors.forename: Martin
authors.initials: M
authors.lastname: Röcken
authors.forename: Martin
authors.initials: M
authors.lastname: Schaller
authors.forename: Florian
authors.initials: F
authors.lastname: Lang
authors.forename: Thomas
authors.initials: T
authors.lastname: Wieder
mesh_terms: D000814:Aniline Compounds / Q000737:chemistry; D017304:Annexin A5 / Q000378:metabolism; D000894:Anti-Inflammatory Agents, Non-Steroidal / Q000494:pharmacology*; D017209:Apoptosis; D002118:Calcium / Q000378:metabolism; D048429:Cell Size; D004912:Erythrocytes / Q000187:drug effects / Q000378:metabolism*; D005978:Glutathione / Q000378:metabolism; D006801:Humans; D051550:I-kappa B Kinase / Q000378:metabolism; D016328:NF-kappa B / Q000037:antagonists & inhibitors / Q000378:metabolism*; D009570:Nitriles / Q000494:pharmacology*; D010718:Phosphatidylserines / Q000378:metabolism; D011485:Protein Binding; D012717:Sesquiterpenes / Q000494:pharmacology*; D015398:Signal Transduction; D013450:Sulfones / Q000494:pharmacology*; D014966:Xanthenes / Q000737:chemistry
chemical_list: C434003:3-(4-methylphenylsulfonyl)-2-propenenitrile; D000814:Aniline Compounds; D017304:Annexin A5; D000894:Anti-Inflammatory Agents, Non-Steroidal; D016328:NF-kappa B; D009570:Nitriles; D010718:Phosphatidylserines; D012717:Sesquiterpenes; D013450:Sulfones; D014966:Xanthenes; C059715:Fluo-3; C002669:parthenolide; D051550:I-kappa B Kinase; D005978:Glutathione; D002118:Calcium