The loss of telomerase activity in highly differentiated CD8+CD28-CD27- T cells is associated with decreased Akt (Ser473) phosphorylation.

The enzyme telomerase is essential for maintaining the replicative capacity of memory T cells. Although CD28 costimulatory signals can up-regulate telomerase activity, human CD8(+) T cells lose CD28 expression after repeated activation. Nevertheless, telomerase is still inducible in CD8(+)CD28(-) T cells. To identify alternative costimulatory pathways that may be involved, we introduced chimeric receptors containing the signaling domains of CD28, CD27, CD137, CD134, and ICOS in series with the CD3 zeta (zeta) chain into primary human CD8(+) T cells. Although CD3 zeta-chain signals alone were ineffective, triggering of all the other constructs induced proliferation and telomerase activity. However, not all CD8(+)CD28(-) T cells could up-regulate this enzyme. The further fractionation of CD8(+)CD28(-) T cells into CD8(+)CD28(-) CD27(+) and CD8(+)CD28(-)CD27(-) subsets showed that the latter had significantly shorter telomeres and extremely poor telomerase activity. The restoration of CD28 signaling in CD8(+)CD28(-)CD27(-) T cells could not reverse the low telomerase activity that was not due to decreased expression of human telomerase reverse transcriptase, the enzyme catalytic subunit. Instead, the defect was associated with decreased phosphorylation of the kinase Akt, that phosphorylates human telomerase reverse transcriptase to induce telomerase activity. Furthermore, the defective Akt phosphorylation in these cells was specific for the Ser(473) but not the Thr(308) phosphorylation site of this molecule. Telomerase down-regulation in highly differentiated CD8(+)CD28(-)CD27(-) T cells marks their inexorable progress toward a replicative end stage after activation. This limits the ability of memory CD8(+) T cells to be maintained by continuous proliferation in vivo.

The impact of telomere erosion on memory CD8+ T cells in patients with X-linked lymphoproliferative syndrome.

Patients with X-linked lymphoproliferative syndrome (XLP) experience excessive T cell proliferation after primary Epstein-Barr virus (EBV) infection, due to mutations in the signalling lymphocyte activation molecule (SLAM) associated protein (SAP) molecule. We examined the impact of dysfunctional proliferative control on the extent of CD8+ T cell differentiation in XLP patients who recovered from primary EBV infection. Although these young patients have normal numbers of lytic and latent EBV-epitope-specific CD8+ T cells, they were extremely differentiated as defined by loss of CCR7 and CD27, low telomerase activity and very short telomeres. This was not a direct effect arising from the loss of SAP, but was due to excessive T cell stimulation due to this defect. Thus, transduction of XLP CD8+ T cells with the catalytic component of telomerase (hTERT), but not SAP, prevented telomere loss and considerably extended proliferative lifespan in vitro. These results indicate that excessive proliferation in CD8+ T cells in XLP patients may lead to end-stage differentiation and loss of functional EBV-specific CD8+ T cells through replicative senescence. This may contribute to the defective immunity found in XLP patients who survive acute EBV infection who develop EBV-related B cell lymphomas before the fourth decade of life.

Quiescence and functional reprogramming of Epstein-Barr virus (EBV)-specific CD8+ T cells during persistent infection.

After acute infection Epstein-Barr virus (EBV)-specific memory CD8+ T cells exit cell cycle, and a proportion of these antigen-experienced cells re-express CD45RA (CD45 which predominantly express exon A). However, the signals involved are not known. We investigated the roles of interleukin 15 (IL-15) and interferon-alpha/beta (IFN-I) in these processes, since these mediators have a crucial but undefined role in the maintenance of CD8+ T-cell memory. We show that IFN-I (but not IL-15) allows activated EBV-specific CD8+ T cells to leave cell cycle without entering apoptosis. This was associated with up-regulation of the cyclin inhibitor p27, but not of CD45RA. In contrast, IL-15 (but not IFN-I) induced "homeostatic" proliferation and CD45RA re-expression by these cells in vitro. Different signals, therefore, induce quiescence and CD45RA re-expression in activated EBV-specific CD8+ T cells. After T-cell receptor (TCR) activation freshly isolated CD45RA+ antigen-experienced CD8+ T cells show poor proliferative activity but are highly cytotoxic and secrete IFN-gamma efficiently. This suggests functional reprogramming toward effector function but away from proliferation. The induction of quiescence and the generation of proliferation-independent effector CD8+ T cells that re-express CD45RA may minimize the impact of replicative senescence in virus-specific populations that would otherwise occur during decades of persistent infection.

Integration of apoptosis and telomere erosion in virus-specific CD8+ T cells from blood and tonsils during primary infection.

Human-virus-specific CD8+ T cells that are found during primary infection have been studied almost exclusively in the peripheral blood, and it is unclear whether these cells are regulated in the same way as those in secondary lymphoid tissue. We investigated, therefore, the control of apoptosis and telomere erosion of Epstein-Barr virus (EBV)-specific CD8+ T cells found in the blood and tonsils of the same patients during acute infectious mononucleosis (AIM). Although the clonal composition of CD8+ T cells as determined by heteroduplex analysis was similar in both compartments, there was greater CD28 expression in the tonsil population, indicating that they were less differentiated. EBV-specific CD8+ T cells in both tissue types were extremely susceptible to apoptosis related to low Bcl-2 expression and were dependent on exogenous cytokines such as interleukin-2 (IL-2), IL-15, and interferon-alpha/beta (IFN-alpha/beta) for survival. In both compartments, however, these cells maintained their telomere lengths through telomerase induction. Thus, apoptosis-prone EBV-specific CD8+ T cells found during acute infection have to be rescued from death to persist as a memory population. However, signals that induce telomerase ensure that the rescued cells retain their replicative capacity. Significantly, these processes operate identically in cells found in blood and secondary lymphoid tissue.

Anergic T cells exert antigen-independent inhibition of cell-cell interactions via chemokine metabolism.

Due to their ability to inhibit antigen-induced T-cell activation in vitro and in vivo, anergic T cells can be considered part of the spectrum of immunoregulatory T lymphocytes. Here we report that both murine and human anergic T cells can impair the ability of parenchymal cells (including endothelial and epithelial cells) to establish cell-cell interactions necessary to sustain leukocyte migration in vitro and tissue infiltration in vivo. The inhibition is reversible and cell-contact dependent but does not require cognate recognition of the parenchymal cells to occur. Instrumental to this effect is the increased cell surface expression and enzymatic activity of molecules such as CD26 (dipeptidyl-peptidase IV), which may act by metabolizing chemoattractants bound to the endothelial/epithelial cell surface. These results describe a previously unknown antigen-independent anti-inflammatory activity by locally generated anergic T cells and define a novel mechanism for the long-known immunoregulatory properties of these cells.