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Inhibition of TNFalpha in vivo prevents hyperoxia-mediated activation of caspase 3 in type II cells.

AbstractBACKGROUND:
The mechanisms during the initial phase of oxygen toxicity leading to pulmonary tissue damage are incompletely known. Increase of tumour necrosis factor alpha (TNFalpha) represents one of the first pulmonary responses to hyperoxia. We hypothesised that, in the initial phase of hyperoxia, TNFalpha activates the caspase cascade in type II pneumocytes (TIIcells).
METHODS:
Lung sections or freshly isolated TIIcells of control and hyperoxic treated rats (48 hrs) were used for the determination of TNFalpha (ELISA), TNF-receptor 1 (Western blot) and activity of caspases 8, 3, and 9 (colorimetrically). NF-kappaB activation was determined by EMSA, by increase of the p65 subunit in the nuclear fraction, and by immunocytochemistry using a monoclonal anti-NF-kappaB-antibody which selectively stained the activated, nuclear form of NF-kappa B. Apoptotic markers in lung tissue sections (TUNEL) and in TIIcells (cell death detection ELISA, Bax, Bcl-2, mitochondrial membrane potential, and late and early apoptotic cells) were measured using commercially available kits.
RESULTS:
In vivo, hyperoxia activated NF-kappaB and increased the expression of TNFalpha, TNF-receptor 1 and the activity of caspase 8 and 3 in freshly isolated TIIcells. Intratracheal application of anti-TNFalpha antibodies prevented the increase of TNFRI and of caspase 3 activity. Under hyperoxia, there was neither a significant change of cytosolic cytochrome C or of caspase 9 activity, nor an increase in apoptosis of TIIcells. Hyperoxia-induced activation of caspase 3 gradually decreased over two days of normoxia without increasing apoptosis. Therefore, activation of caspase 3 is a temporary effect in sublethal hyperoxia and did not mark the "point of no return" in TIIcells.
CONCLUSION:
In the initiation phase of pulmonary oxygen toxicity, an increase of TNFalpha and its receptor TNFR1 leads to the activation of caspase 8 and 3 in TIIcells. Together with the hyperoxic induced increase of Bax and the decrease of the mitochondrial membrane potential, activation of caspase 3 can be seen as sensitisation for apoptosis. Eliminating the TNFalpha effect in vivo by anti-TNFalpha antibodies prevents the pro-apoptotic sensitisation of TIIcells.
AuthorsFlorian Guthmann, Heide Wissel, Christian Schachtrup, Angelika Tölle, Mario Rüdiger, Friedrich Spener, Bernd Rüstow
JournalRespiratory research (Respir Res) Vol. 6 Pg. 10 (Jan 21 2005) ISSN: 1465-993X [Electronic] England
PMID15663790 (Publication Type: Journal Article)
Chemical References
  • Cytokines
  • Tumor Necrosis Factor-alpha
  • Casp3 protein, rat
  • Casp8 protein, rat
  • Caspase 3
  • Caspase 8
  • Caspases
Topics
  • Animals
  • Caspase 3
  • Caspase 8
  • Caspases (metabolism)
  • Cells, Cultured
  • Cytokines (metabolism)
  • Enzyme Activation
  • Hyperoxia (metabolism)
  • Lung (metabolism)
  • Oxidative Stress
  • Rats
  • Rats, Wistar
  • Tumor Necrosis Factor-alpha (metabolism)

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