Inhibition of Caspases Increases the Sensitivity of L929 Cells to Necrosis Mediated by Tumor Necrosis Factor

doi:10.1084/jem.187.9.1477

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J. Exp. Med., Volume 187, Number 9, May 4, 1998 1477-1485

Inhibition of Caspases Increases the Sensitivity of L929 Cells to Necrosis Mediated by Tumor Necrosis Factor

By Dominique Vercammen, Rudi Beyaert, Geertrui Denecker, Vera Goossens, Geert Van Loo, Wim Declercq, Johan Grooten, Walter Fiers, and Peter Vandenabeele

From the Laboratory of Molecular Biology, Flanders Interuniversity Institute for Biotechnology and University of Ghent, B-9000 Ghent, Belgium

Murine L929 fibrosarcoma cells treated with tumor necrosis factor (TNF) rapidly die in a necrotic way, due to excessive formationof reactive oxygen intermediates. We investigated the role ofcaspases in the necrotic cell death pathway. When the cytokineresponse modifier A (CrmA), a serpin-like caspase inhibitor ofviral origin, was stably overexpressed in L929 cells, the latterbecame 1,000-fold more sensitive to TNF-mediated cell death. Inaddition, TNF sensitization was also observed when the cells werepretreated with Ac-YVAD-cmk or zDEVD-fmk, which inhibits caspase-1-and caspase-3-like proteases, respectively. zVAD-fmk and zD-fmk,two broad-spectrum inhibitors of caspases, also rendered the cellsmore sensitive, since the half-maximal dose for TNF-mediated necrosisdecreased by a factor of 1,000. The presence of zVAD-fmk alsoresulted in a more rapid increase of TNF-mediated production ofoxygen radicals. zVAD-fmk-dependent sensitization of TNF cytotoxicitycould be completely inhibited by the oxygen radical scavengerbutylated hydroxyanisole. These results indicate an involvementof caspases in protection against TNF-induced formation of oxygenradicals and necrosis.

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