We examined the role of the antiapoptotic molecule Bcl-2 in combating the proapoptotic molecule Bim in control of naive and memory T cell homeostasis using Bcl-2−/− mice that were additionally deficient in one or both alleles of Bim. Naive T cells were significantly decreased in Bim+/−Bcl-2−/− mice, but were largely restored in Bim−/−Bcl-2−/− mice. Similarly, a synthetic Bcl-2 inhibitor killed wild-type, but not Bim−/−, T cells. Further, T cells from Bim+/−Bcl-2−/− mice died rapidly ex vivo and were refractory to cytokine-driven survival in vitro. In vivo, naive CD8+ T cells required Bcl-2 to combat Bim to maintain peripheral survival, whereas naive CD4+ T cells did not. In contrast, Bim+/−Bcl-2−/− mice generated relatively normal numbers of memory T cells after lymphocytic choriomeningitis virus infection. Accumulation of memory T cells in Bim+/−Bcl-2−/− mice was likely caused by their increased proliferative renewal because of the lymphopenic environment of the mice. Collectively, these data demonstrate a critical role for a balance between Bim and Bcl-2 in controlling homeostasis of naive and memory T cells.

Reactive oxygen species (ROS) are known to regulate cell death in a variety of cell types. Differential effects of ROS on cell death are observed depending on the level of ROS within the cell (1). High levels of ROS can lead to lipid peroxidation, damage to cellular membranes, inactivation of caspase enzymes, and necrotic cell death (1). In contrast, low levels of ROS have been shown to activate protein kinases and phosphatases, mobilize Ca 2ϩ stores, activate or inactivate transcription factors, and lead to apoptotic cell death (1). Thus, high levels of ROS probably kill cells by means of direct damage, whereas low levels of ROS probably mediate apoptosis indirectly through their effects on protein kinases and͞or phosphatases, transcription factors, and the subsequent effects of these molecules on gene expression. However, little has been done to assess the extent to which ROS affect gene expression in cells undergoing apoptosis.It has been known for some time that Bcl-2 overexpression can protect cells from apoptosis mediated by ROS (2). However, the mechanism by which Bcl-2 prevents ROS-induced apoptosis is unknown. Bcl-2 itself does not possess antioxidant activity; rather, it may act indirectly to increase the levels and͞or activities of endogenous antioxidants (e.g., glutathione or superoxide dismutase) within cells (3-5). Thus, overexpression of Bcl-2 may allow cells to cope better with the effects of ROS, possibly by allowing increases in endogenous antioxidant enzymes.An alternative but not mutually exclusive hypothesis is that ROS act to down-regulate endogenous Bcl-2 levels within cells. Because levels of Bcl-2 within cells are critical to antiapoptotic activity, decreasing Bcl-2 could be a mechanism to sensitize cells to apoptosis. By detoxifying ROS, antioxidants may therefore reverse the ROS-induced decline in Bcl-2 and prevent apoptosis. Several studies suggest a reciprocal relationship between ROS and Bcl-2 levels within cells (6-8). Thus, in divergent cell types, decreases in ROS correlate with increases in Bcl-2 levels and vice versa.Recently we showed that, in vivo, activated T cells decrease their levels of Bcl-2 and become susceptible to the proapoptotic effects of Bim (9). We also showed that culture of these activated T cells with a synthetic catalytic antioxidant, Mn(III) tetrakis(5,10,15,20-benzoic acid)porphyrin (MnTBAP), lowered superoxide levels and prevented apoptosis (10). Here, we show that culture of T cells with MnTBAP reverses the decline in Bcl-2. The corollary of this finding is that ROS are responsible for Bcl-2 down-regulation within activated T cells. Taken together, these results suggest a ''twosignal'' model for activated T cell apoptosis. The first signal is driven by ROS down-regulation of Bcl-2, which is necessary, but not sufficient, to complete apoptosis. The second signal requires the expression of Bim. . RNA extraction, cDNA synthesis, and gene microarray analysis were performed as described (11,12) Mice. C57BL͞10 and C57BL͞6J (BL͞6) mice were purchased from The Ja...