Nitric oxide induces apoptosis in mouse thymocytes.
Nitric oxide (NO) produced at high concentrations by the inducible NO synthase is an important effector molecule involved in immune regulation and defense. We have examined whether NO represents a signal for triggering apoptosis in thymocytes. Freshly isolated thymocytes were incubated with different chemical NO donors for various intervals. Apoptosis was determined by detection of DNA strand breaks with in situ nick translation. All NO donors induced thymocyte apoptosis with 30% positive thymocytes vs 10% in controls after 8 h. Apoptosis was prevented by addition of ZnSO4. Short-term pre-exposure to NO resulted in protection from apoptosis induced by glucocorticoids comparable with the protective effect of heat shock. Flow cytometry revealed that NO treatment as well as heat shock or dexamethasone incubation is accompanied by reduction in the CD4+ CD8+ thymocyte subpopulation. Apoptosis induction was accompanied by increased expression of p53, as detected by PCR analysis 2 h after NO donor addition. In vivo treatment of mice with endotoxin results in increased thymic apoptosis. Focal apoptosis was found to occur in close proximity to blood vessels 18 h after LPS treatment. Capillary endothelium and dendritic cells adjacent to apoptotic foci were found to stain strongly for inducible NO synthase expression. Furthermore, in an in vitro experiment using cocultures of thymocytes with LPS/cytokine-activated endothelial cells expressing inducible NO synthase, a significantly increased rate of thymocyte apoptosis was found, and this could be prevented completely by inhibiting NO production. Addition of dexamethasone to these cocultures did not lead to a further increase in the percentage of apoptotic thymocytes, underlining the protective effect of NO on dexamethasone-induced apoptosis.
Fehsel
K
K
Institute of Immunobiology, Heinrich Heine University, Düsseldorf, Germany.
Kröncke
K D
KD
Meyer
K L
KL
Huber
H
H
Wahn
V
V
Kolb-Bachofen
V
V
D000818:Animals; D017209:Apoptosis / Q000187:drug effects* / Q000276:immunology; D016516:CD4-CD8 Ratio / Q000187:drug effects; D002478:Cells, Cultured; D004249:DNA Damage / Q000187:drug effects; D003713:Dendritic Cells / Q000473:pathology; D004730:Endothelium, Vascular / Q000473:pathology; D008070:Lipopolysaccharides / Q000008:administration & dosage; D051379:Mice; D009569:Nitric Oxide / Q000494:pharmacology*; D013601:T-Lymphocytes / Q000276:immunology / Q000473:pathology*; D016159:Tumor Suppressor Protein p53 / Q000096:biosynthesis
D008070:Lipopolysaccharides; D016159:Tumor Suppressor Protein p53; D009569:Nitric Oxide