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Caspase inhibition reveals functional cooperation between p55- and p75-TNF receptors in cell necrosis
M Pelagi1, F Curnis, B Colombo
1Department of Biological and Technological Research, San Raffaele H Scientific Institute, via Olgettina 58, 20132 Milan, Italy.
Abstract:
TNF-induced caspase activation is critically involved in both apoptosis and protection from cell necrosis. We have investigated the roles of the p55- and p75-TNF receptors (TNFR1 and TNFR2) in the induction of mouse L-M cell death in the presence of a caspase inhibitor (zVAD-fmk) and a transcription inhibitor (actinomycin D), i.e. under conditions in which protective pathways requiring caspase activation and protein synthesis were blocked. Cytometric analysis after TNF treatment showed that apoptosis was inhibited, while necrosis was highly activated. In contrast, apoptosis was observed in cells treated with TNF and actinomycin D alone. Stimulation of TNFR1 was sufficient to induce either cell necrosis or apoptosis, even when we blocked endogenous TNF with an anti-murine TNF antibody. Experiments based on the use of receptor-agonist and antagonist antibodies also showed that TNFR2 contributes to cell necrosis and apoptosis. Simultaneous stimulation of TNFR2 and TNFR1 with specific agonists indicated that TNFR2 functionally cooperates with TNFR1 to potentiate the response indirectly, by inducing endogenous TNF cytotoxicity. Caspase inhibitors enhanced the cytotoxic effect of endogenous TNF, suggesting that TNFR2 modulation can regulate the global necrotic response to TNF. TNFR2 modulation could play an important role in determining the response to TNF in pathophysiological conditions characterized by caspase down-regulation and local TNF production.
Insights
Tumor Necrosis Factor (TNF) receptor signaling dictates cell fate, influencing apoptosis or necrosis. TNFR2 modulation impacts cell death pathways, particularly when caspase activity is inhibited.
Area of Science:
- Cell Biology
- Immunology
- Molecular Biology
Background:
- Tumor Necrosis Factor (TNF) signaling is crucial for apoptosis and necrosis.
- The roles of p55-TNF receptor (TNFR1) and p75-TNF receptor (TNFR2) in cell death are complex.
- Understanding TNF receptor function is vital in conditions with altered cell death pathways.
Purpose of the Study:
- To investigate the distinct roles of TNFR1 and TNFR2 in TNF-induced cell death.
- To elucidate how TNFR1 and TNFR2 signaling pathways interact under conditions of blocked caspase activation and protein synthesis.
- To determine the potential of TNFR2 modulation in regulating cell death responses.
Main Methods:
- Utilized mouse L-M cells treated with TNF, caspase inhibitor (zVAD-fmk), and transcription inhibitor (actinomycin D).
- Employed flow cytometry for cell death analysis (apoptosis vs. necrosis).
- Used receptor-specific agonist and antagonist antibodies for TNFR1 and TNFR2 stimulation and blockade.
Main Results:
- TNF treatment with inhibitors blocked apoptosis and induced necrosis.
- TNFR1 stimulation alone induced either necrosis or apoptosis.
- TNFR2 contributed to both necrosis and apoptosis, cooperating with TNFR1 to enhance TNF cytotoxicity, especially when caspase activity was inhibited.
Conclusions:
- Both TNFR1 and TNFR2 are key mediators of TNF-induced cell death.
- TNFR2 plays a significant role in potentiating necrosis, particularly under conditions of caspase down-regulation.
- Modulation of TNFR2 offers a potential therapeutic target for controlling TNF-mediated cell death in specific pathophysiological contexts.