Medizinische Universität Graz Austria/Österreich - Forschungsportal - Medical University of Graz
Gewählte Publikation:
Nusshold, C.
Uptake, Metabolism, and Cellular Activities of Native and Chlorinated Sphingolipids in Neurons
[ Dissertation ] Medical University of Graz; 2011. pp. 204
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- Autor*innen der Med Uni Graz:
- Nusshold Christoph
- Betreuer*innen:
- Heinemann Akos
- Sattler Wolfgang
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- Abstract:
- Normal brain function depends on a delicately balanced set of remarkably diverse lipids. Consequently, short- and long term alterations in brain lipid composition during neuroinflammatory conditions associated with oxidative stress are casually involved in central nervous system (CNS) disorders (e. g. Alzheimer¿s disease, Parkinson¿s disease, multiple sclerosis, stroke, or traumatic brain injury). Within the different cerebral lipid subclasses sphingolipids take a central role in the CNS and in particular neuronal function. Consequently dysfunctional brain sphingolipid homeostasis is associated with the development of neurodegenerative diseases. Besides mitochondria, Aß peptides, redox-active iron, and the NADPH-oxidase complex, recent studies point towards myeloperoxidase (MPO) as a source of reactive oxygen/radical species. After activation of phagocytes, MPO uses chloride ions and hydrogen peroxide generated from superoxide anion radicals to form hypochlorous acid (HOCl). This potent oxidant targets unsaturated lipids to form a battery of chlorinated lipotoxic compounds. Of note, due to the presence of an unsaturated said chain and amide group sphingomyelin (SM) could be subject to modification by MPO-derived HOCl. Therefore this study aimed at investigating SM uptake and metabolism in a neuronal cell line (CATH.a) as well as the impact of SM and HOCl-modified SM (HOCl-SM) on mitochondrial function, cell viability, and proteome alterations of PC12 neurons. In vitro experiments performed during the present study demonstrate that fluorescently-labeled (BODIPY and PYRENE derivatives) SM and ceramide (Cer) analogues are efficiently internalized by caveolae-mediated endocytosis and metabolized by differentiated and undifferentiated CATH.a cells. SMase-mediated BODIPY-SM hydrolysis is a process starting already at the plasma membrane. SM-derived BODIPY-Cer diffuses into intracellular compartments and accumulates in the Golgi apparatus and the endoplasmic reticulum. In contrast, the majority of BODIPY-SM remains at the extracellular leaflet of the plasma membrane. Intracellular Cer is partially used as SMSase substrate to re-synthesize SM. Another part of the intracellular Cer pool is metabolized to an as yet unidentified polar sphingolipid species that is secreted by the cells. - XIV - Using rat pheochromacytoma (PC12) cells I could demonstrate that HOCl-modified SM, generated by HOCl-treatment or enzymatically via the MPO/H2O2/Cl- system induces formation of reactive oxygen species in contrast to unmodified SM. Furtherore, HOCl-SM provokes mitochondrial dysfunction, decreases PC12 cell viability, induces the apoptotic machinery, and leads to pronounced proteomic alterations. In summary, the present study indicates that oxidation/chlorination of SM by MPO-derived HOCl might play a central role in neuronal cell death in CNS-related diseases. These findings may be valuable in understanding the role of MPO-derived oxidants on dysregulated sphingolipid homeostasis as observed in chronic neurodegenerative diseases.