Hypochlorite modification of sphingomyelin generates chlorinated lipid species that induce apoptosis and proteome alterations in dopaminergic PC12 neurons in vitro.
Recent observations link myeloperoxidase (MPO) activation to neurodegeneration. In multiple sclerosis MPO is present in areas of active demyelination where the potent oxidant hypochlorous acid (HOCl), formed by MPO from H(2)O(2) and chloride ions, could oxidatively damage myelin-associated lipids. The purpose of this study was (i) to characterize reaction products of sphingomyelin (SM) formed in response to modification by HOCl, (ii) to define the impact of exogenously added SM and HOCl-modified SM (HOCl-SM) on viability parameters of a neuronal cell line (PC12), and (iii) to study alterations in the PC12 cell proteome in response to SM and HOCl-SM. MALDI-TOF-MS analyses revealed that HOCl, added as reagent or generated enzymatically, transforms SM into chlorinated species. On the cellular level HOCl-SM but not SM induced the formation of reactive oxygen species. HOCl-SM induced severely impaired cell viability, dissipation of the mitochondrial membrane potential, and activation of caspase-3 and DNA damage. Proteome analyses identified differential expression of specific subsets of proteins in response to SM and HOCl-SM. Our results demonstrate that HOCl modification of SM results in the generation of chlorinated lipid species with potent neurotoxic properties. Given the emerging connections between the MPO-H(2)O(2)-chloride axis and neurodegeneration, this chlorinating pathway might be implicated in neuropathogenesis.
Institute of Molecular Biology and Biochemistry, Medical University of Graz, 8010 Graz, Austria.
Christoph
C
Nusshold
Manfred
M
Kollroser
Harald
H
Köfeler
Gerald
G
Rechberger
Helga
H
Reicher
Andreas
A
Ullen
Eva
E
Bernhart
Sabine
S
Waltl
Ingrid
I
Kratzer
Albin
A
Hermetter
Hubert
H
Hackl
Zlatko
Z
Trajanoski
Andelko
A
Hrzenjak
Ernst
E
Malle
Wolfgang
W
Sattler
D000818:Animals; D017209:Apoptosis / Q000187:drug effects* / Q000502:physiology; D015153:Blotting, Western; D001923:Brain Chemistry; D002460:Cell Line; D004298:Dopamine / Q000378:metabolism; D015180:Electrophoresis, Gel, Two-Dimensional; D005583:Fourier Analysis; D054879:Halogenation; D006997:Hypochlorous Acid / Q000494:pharmacology*; D007091:Image Processing, Computer-Assisted; D066298:In Vitro Techniques; D008055:Lipids / Q000737:chemistry; D053078:Membrane Potential, Mitochondrial; D051379:Mice; D008810:Mice, Inbred C57BL; D009474:Neurons / Q000378:metabolism*; D016877:Oxidants / Q000494:pharmacology*; D018384:Oxidative Stress / Q000502:physiology*; D020543:Proteome / Q000737:chemistry / Q000187:drug effects / Q000378:metabolism; D051381:Rats; D017382:Reactive Oxygen Species / Q000378:metabolism; D019032:Spectrometry, Mass, Matrix-Assisted Laser Desorption-Ionization; D013109:Sphingomyelins / Q000737:chemistry / Q000378:metabolism*
D008055:Lipids; D016877:Oxidants; D020543:Proteome; D017382:Reactive Oxygen Species; D013109:Sphingomyelins; D006997:Hypochlorous Acid; D004298:Dopamine