Involvement of APO2 ligand/TRAIL in activation-induced death of Jurkat and human peripheral blood T cells

Abstract

The interaction of Fas with Fas ligand (FasL) mediates activation-induced cell death (AICD) of T hybridomas and of mature T lymphocytes. The TNF/TNF receptor system also plays a significant role in AICD of mature T cells and in the maintenance of peripheral tolerance. We previously demonstrated that in human Jurkat leukemia cells, AICD is triggered mainly by the rapid release of preformed FasL upon TCR stimulation. In the present work, we show that the cytotoxic cytokine APO2 ligand (APO2L; also known as TRAIL) is constitutively expressed as an intracytoplasmic protein in Jurkat T cells and derived sublines. APO2L is also detected in fresh human peripheral blood mononuclear cells (PBMC) from a significant number of donors, and the amount of both FasL and APO2L substantially increases upon blast generation. A neutralizing anti-APO2L monoclonal antibody (mAb) partially suppresses the cytotoxicity induced by supernatants of phytohemagglutinin (PHA)-prestimulated Jurkat or human PBMC on non-activated Jurkat cells, indicating that APO2L is released by these cells and contributes to AICD. A combination of neutralizing anti-APO2L and anti-Fas mAb blocks around 60 % of the toxicity associated with supernatants from PHA-activated human PBMC. These results show that FasL and APO2L account for the majority of cytotoxic activity released during AICD, and suggest that additional uncharacterized factors may also contribute to this process.

Human CD8+ T cell blasts are more sensitive than CD4+ T cell blasts to regulation by APO2L/TRAIL

“…PHA or anti-CD3 mAb activation results in secretion of FasL and APO2L/TRAIL, while CD59 ligation results preferentially in APO2L/TRAIL secretion. These new results using separated CD4 + and CD8 + T cell blasts are compatible with secretion of the death ligands associated with microvesicles/exosomes into the supernatant, as described by our group in previous studies using mixed T cell blast populations [12,14,15]. We have also observed, using a bioassay on Jurkat cells, that although CD8 + T cell blasts are more sensitive to APO2L/TRAIL regulation, CD4 + T cell blasts are able to secrete higher amounts of bioactive APO2L/TRAIL upon PHA stimulation.…”

“…In fact, lpr and gld mice deficient in functional Fas or FasL expression, respectively [9,10], or humans with similar defects [11] have systemic autoimmune disease characterized by lymphoproliferation. Our group suggested for the first time the participation of APO2 ligand/TNF-related apoptosisinducing ligand (APO2L/TRAIL) and its receptors DR4 and DR5 in the down-regulation of T cell responses [12]. This participation has been confirmed recently in the APO2L/TRAIL knockout mice, which are more susceptible to autoimmune disease induction [13].…”

“…Other studies have shown that soluble HLA molecules induce apoptosis in polyclonal CD8 + human T cell blasts through release of bioactive FasL into the supernatant [32,33]. No data are available on a possible differential regulation of T cell subsets by APO2L/TRAIL or by CD59 ligation, which have been shown to be implicated in the down-modulation of mixed populations of human T cell blasts [12,14].In the present work, we have undertaken a systematic study using normal human T cell blasts from at least 20 healthy donors to characterize the mechanisms responsible for the down-modulation of normal human T cell activation, by separating CD4 + and CD8 + populations from the same donors. Results indicate that normal human CD8 + T cell blasts are more sensitive to APO2L/TRAIL immunoregulation than CD4 + T cell blasts.…”

“…The rApo2L preparation used in this study corresponds to Apo2L.0, in which the monomer form predominates [41] and which, in contrast to previous preparations, does not contain a polyhistidine tag promoting aggregation [42]. rApo2L was kindly provided by Dr. Avi Ashkenazi (Genentech, South San Francisco, CA).Supernatants from day 6 human CD4 + and CD8 + T cell blasts restimulated with a PHA pulse or by immobilized anti-CD3 or anti-CD59 mAb were obtained as indicated in our previous studies performed using mixed T cell blast populations [12,15]. Toxicity of supernatants from cells pulsestimulated with PHA, anti-CD3 or anti-CD59 mAb was tested on Jurkat cells, with supernatants from non-stimulated cells used as controls, in the presence or absence of the anti-Fas and/or anti-APO2L/TRAIL blocking mAb SM1/23 or 5C2, respectively, as described [12,15].…”

Induction of apoptosis and modulation of activation and effector function in T cells by immunosuppressive drugs

“…DEX, MPA and RAP did not increase AICD in Jurkat T cells or human peripheral T cells, suggesting that immunosuppressive therapy does not enhance autoregulatory apoptosis. Recently for TRAIL (Apo2L), a role has also been described in the induction of AICD in Jurkat T cells [47]. As more than 90% of cell death in CD3-triggered Jurkat cells in our system can be blocked with F(ab¢) 2 APO-1 fragments, by inhibition of the interaction of CD95 with its ligand, the CD95 system is the major mechanism of AICD in these cells (data not shown).…”