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

Respiratory Research

Bernd Rüstow¹  Friedrich Spener²  Mario Rüdiger¹  Angelika Tölle¹  Christian Schachtrup²  Heide Wissel¹  Florian Guthmann¹ 

[1] Humboldt-Universität zu Berlin, Klinik für Neonatologie, Charité Campus Mitte, D-10098 Berlin, Germany;Westfälische Wilhelms-Universität Münster, Institut für Biochemie, Wilhelm-Klemm-Str. 2, D-48149 Münster, Germany

关键词: apoptosis;    caspase;    tumour necrosis factor receptor;    TNFα;    alveolar type II cells;    lung;    Hyperoxia;   

Others  :  1227337
DOI  :  10.1186/1465-9921-6-10

 received in 2004-08-19, accepted in 2005-01-21,  发布年份 2005

【 摘 要 】

Background

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.

【 授权许可】

   
2005 Guthmann et al; licensee BioMed Central Ltd.

【 图 表 】

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