Ornithine decarboxylase prevents tumor necrosis factor alpha-induced apoptosis by decreasing intracellular reactive oxygen species

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Abstract
Ornithine decarboxylase (ODC) plays an essential role in various biological functions, including cell proliferation, differentiation and cell death. However, how it prevents the cell apoptotic mechanism is still unclear. Previous studies have demonstrated that decreasing the activity of ODC by difluoromethylornithine (DFMO), an irreversible inhibitor of ODC, causes the accumulation of intracellular reactive oxygen species (ROS) and cell arrest, thus inducing cell death. These findings might indicate how ODC exerts anti-oxidative and anti-apoptotic effects. In our study, tumor necrosis factor alpha (TNF-α) induced apoptosis in HL-60 and Jurkat T cells. The kinetic studies revealed that the TNF-α -induced apoptotic process included intracellular ROS generation (as early as 1 h after treatment), the activation of caspase 8 (3 h), the cleavage of Bid (3 h) and the disruption of mitochondrial membrane potential (Δ ψ m ) (6 h). Furthermore, ROS scavengers, such as glutathione (GSH) and catalase, maintained Δ ψ m and prevented apoptosis upon treatment. Putrescine and overexpression of ODC had similar effects as ROS scavengers in decreasing intracellular ROS and preventing the disruption of Δ ψ m and apoptosis. Inhibition of ODC by DFMO in HL-60 cells only could increase ROS generation, but did not disrupt Δ ψ m or induce apoptosis. However, DFMO enhanced the accumulation of ROS, disruption of Δ ψ m and apoptosis when cells were treated with TNF-α . ODC overexpression avoided the decline of Bcl-2, prevented cytochrome c release from mitochondria and inhibited the activation of caspase 8, 9 and 3. Overexpression of Bcl-2 maintained Δ ψ m and prevented apoptosis, but could not reduce ROS until four hours after TNF-α treatment. According to these data, we suggest that TNF-α induces apoptosis mainly by a ROS-dependent, mitochondria-mediated pathway. Furthermore, ODC prevents TNF-α -induced apoptosis by decreasing intracellular ROS to avoid Bcl-2 decline, maintain Δ ψ m , prevent cytochrome c release and deactivate the caspase cascade pathway.