Search

The consequences of H[sub 2]O[sub 2]/Fe[sup 2+]-induced oxidative stress on translocation of ethanolamine phosphoglyceride (EPG) and serine phosphoglyceride (SPG) were studied in an oligodendroglia-like cell line (OLN 93) following 3 days of supplementation with 0.1 mm docosahexaenoic acid (DHA) and a series of polar head group precursors, including N-monomethyl- and N,N-dimethylethanolamine at millimolar concentrations. Added DHA was predominantly esterified in EPG species and those cells enriched in DHA showed enhanced sensitivity to oxidative stress and eventually died by apoptosis. Co-supplements with ethanolamine and DHA resulted in a rapid, but transient, EPG translocation with a maximum at 30 min following stress, as characterized by a trinitrobenzenesulfonic acid reagent. There was no significant translocation of SPG as evidenced by annexin V binding. Unlike SPG, which is usually irreversibly translocated to subserve as a tag for phagocytosis, EPG acted as a signaling molecule with biphasic kinetic characteristics. N-Monomethyl- and N,N-dimethylethanolamine supplements reduced EPG synthesis, prevented its externalization and rescued cells from apoptotic death. Following stress, the fatty acid profile of the externalized EPG showed marked losses in polyunsaturated fatty acids and aldehydes compared with the remaining intracellular EPG. Prevention of EPG species selective translocation to the outer membrane leaflet by altering phospholipid asymmetry may be important in the mechanism of rescue from cell death. [ABSTRACT FROM AUTHOR]

A major feature of Alzheimer's disease is the deposition of the amyloid beta peptide (Aβ) in the brain by mechanisms which remain unclear. One hypothesis suggests that oxidative stress and Aβ aggregation are interrelated processes. Protein kinase C, a major neuronal regulatory protein is activated after oxidative stress and is also altered in the Alzheimer's disease brain. Therefore, we examined the effects of Aβ[sub 1-40] peptide on the protein kinase C cascade and cell death in primary neuronal cultures following anoxic conditions. Treatment with Aβ[sub 1-40] for 48 h caused a significant increase in the content and activity of Ca[sup 2+]-dependent and Ca[sup 2+]-independent protein kinase C isoforms. By 72 h various protein kinase C isoforms were down-regulated. Following 90 min anoxia and 6 h normoxia, a decrease in protein kinase C isoforms was noticed, independent of Aβ[sub 1-40] treatment. A combination of Aβ[sub 1-40] and 30-min anoxia enhanced cytotoxicity as noticed by a marked loss in the mitochondrial ability to convert 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyl-tetrazolium bromide and by enhanced 4′,6-diamidino-2-phenylindole nuclear staining. Phosphorylation of two downstream protein kinase C substrates of apparent molecular mass 80 and 43 kDa, tentatively identified as the mirystoyl alanine-rich C-kinase substrate (MARCKS), were gradually elevated up to 72 h upon incubation with Aβ[sub 1-40]. Anoxia followed by 30 min normoxia enhanced MARCKS phosphorylation in the membrane but not in the cytosolic fraction. In the presence of Aβ[sub 1-40], phosphorylation of MARCKS was reduced. After 6 h normoxia, MARCKS phosphorylability was diminished possibly because of protein kinase C down-regulation. The data suggest that a biphasic modulation of protein kinase C and MARCKS by Aβ[sub 1-40] combined with anoxic stress may play a role in Alzheimer's disease pathology.[sup 1]. [ABSTRACT FROM AUTHOR]

The present work examines the effect of membrane lipid composition on activation of extracellular signal-regulated protein kinases (ERK) and cell death following oxidative stress. When subjected to 50 µm docosahexaenoic acid (DHA, 22 : 6 n-3), cellular phospholipids of OLN 93 cells, a clonal line of oligodendroglia origin low in DHA, were enriched with this polyunsaturated fatty acid. In the presence of 1 mm N,N-dimethylethanolamine (dEa) a new phospholipid species analog was formed in lieu of phosphatidylcholine. Exposure of DHA-enriched cells to 0.5 mm H[sub 2]O[sub 2], caused sustained activation of ERK up to 24 h. At this time massive apoptotic cell death was demonstrated by ladder and TUNEL techniques. H[sub 2]O[sub 2]-induced stress applied to dEa or DHA/dEa co-supplemented cells showed only a transient ERK activation and no cell death after 24 h. Moreover, while ERK was rapidly translocated into the nucleus in DHA-enriched cells, dEa supplements completely blocked ERK nuclear translocation. This study suggests that H[sub 2]O[sub 2]-induced apoptotic cell death is associated with prolonged ERK activation and nuclear translocation in DHA-enriched OLN 93 cells, while both phenomena are prevented by dEa supplements. Thus, the membrane lipid composition ultimately modulates ERK activation and translocation and therefore can promote or prevent apoptotic cell death. [ABSTRACT FROM AUTHOR]