Apoptosis of myelin-reactive T cells induced by reactive oxygen and nitrogen intermediates in vitro.
Apoptosis is a major mechanism of T cell elimination during ontogeny and tolerance induction as well as in autoimmunity. To assess the possible involvement of reactive oxygen and nitrogen intermediates (ROI and NO.) in T-cell apoptosis during autoimmune demyelination we investigated the effects of H2O2 and NO. in vitro on activated autoreactive CD4+ T cell lines capable of transferring experimental autoimmune encephalomyelitis (EAE) and experimental autoimmune neuritis (EAN). For detection and quantitation of apoptotic cells, DNA fragmentation was assessed by in situ tailing with fluorescein-ddUTP and subsequent flow cytometric analysis. H2O2 applied directly to the cell cultures for 6 to 18 hr at concentrations of 10 to 300 microM and ROI released by combination of hypoxanthine and xanthine oxidase (HX/XO) caused apoptosis in a dose-dependent manner in 13-33% of T cells of neuritogenic and encephalitogenic T cell lines. Apoptosis induction could be suppressed by the H2O2-neutralizing enzyme catalase. NO. released by the penicillamine derivative SNAP induced apoptosis to a similar extent as ROI. Maximum values were 38% in an encephalitogenic V beta 8.2-T cell receptor-bearing T cell line and 26% in a neuritogenic T cell line. T cell lines with specificity to ovalbumin revealed slightly lower susceptibility to apoptosis induction by all three kinds of trigger, which is, however, most probably not due to the different antigen specificity, but rather a result of fewer in vitro restimulation cycles of these cells. In neuritogenic cells high-dose (100 units/ml) exogenous interleukin-2 (IL-2) prevents H2O2-induced apoptosis. In conclusion, macrophage-derived reactive oxygen and nitrogen intermediates have the potency to limit inflammatory demyelination by elimination of autoreactive and bystander T cells via apoptotic cell death, and IL-2 is a rescue factor.
Zettl
U K
UK
Department of Neurology, University of Rostock, Germany.
Mix
E
E
Zielasek
J
J
Stangel
M
M
Hartung
H P
HP
Gold
R
R
D000818:Animals; D017209:Apoptosis / Q000187:drug effects*; D001327:Autoimmune Diseases / Q000276:immunology* / Q000473:pathology; D015496:CD4-Positive T-Lymphocytes / Q000187:drug effects* / Q000276:immunology / Q000473:pathology; D002478:Cells, Cultured; D053938:DNA Fragmentation; D006861:Hydrogen Peroxide / Q000494:pharmacology*; D019271:Hypoxanthine / Q000378:metabolism; D004676:Myelin Basic Protein / Q000276:immunology*; D009444:Neuritis, Autoimmune, Experimental / Q000276:immunology* / Q000473:pathology; D009569:Nitric Oxide / Q000494:pharmacology*; D010396:Penicillamine / Q000031:analogs & derivatives / Q000494:pharmacology; D051381:Rats; D011917:Rats, Inbred Lew; D017382:Reactive Oxygen Species / Q000378:metabolism*; D026423:S-Nitroso-N-Acetylpenicillamine; D014969:Xanthine Oxidase / Q000494:pharmacology
D004676:Myelin Basic Protein; D017382:Reactive Oxygen Species; D019271:Hypoxanthine; D009569:Nitric Oxide; D026423:S-Nitroso-N-Acetylpenicillamine; D006861:Hydrogen Peroxide; D014969:Xanthine Oxidase; D010396:Penicillamine