Defective proximal TCR signaling inhibits CD8+ tumor-infiltrating lymphocyte lytic function

CD8+ tumor-infiltrating lymphocytes (TIL) are severely deficient in cytolysis, a defect that may permit tumor escape from immune-mediated destruction. Because lytic function is dependent upon TCR signaling, we have tested the hypothesis that primary TIL have defective signaling by analysis of the localization and activation status of TIL proteins important in TCR-mediated signaling. Upon conjugate formation with cognate target cells in vitro, TIL do not recruit granzyme B+ granules, the microtubule-organizing center, F-actin, Wiskott-Aldrich syndrome protein, nor proline rich tyrosine kinase-2 to the target cell contact site. In addition, TIL do not flux calcium nor demonstrate proximal tyrosine kinase activity, deficiencies likely to underlie failure to fully activate the lytic machinery. Confocal microscopy and fluorescence resonance energy transfer analyses demonstrate that TIL are triggered by conjugate formation in that the TCR, p56lck, CD3zeta, LFA-1, lipid rafts, ZAP70, and linker for activation of T cells localize at the TIL:tumor cell contact site, and CD43 and CD45 are excluded. However, proximal TCR signaling is blocked upon conjugate formation because the inhibitory motif of p56lck is rapidly phosphorylated (Y505) and COOH-terminal Src kinase is recruited to the contact site, while Src homology 2 domain-containing protein phosphatase 2 is cytoplasmic. Our data support a novel mechanism explaining how tumor-induced inactivation of proximal TCR signaling regulates lytic function of antitumor T cells.     

Deficient anti-listerial immunity in the absence of perforin can be restored by increasing memory CD8+ T cell numbers

Compared with wild-type (WT) mice, Listeria monocytogenes (LM)-vaccinated perforin-deficient (PKO) mice have elevated levels of CD8(+) T cell memory, but exhibit reduced levels of protection against virulent LM. In this study, Ag-specific CD8(+) T cells from LM-vaccinated WT and PKO mice were used in adoptive transfer assays to determine the contribution of perforin-dependent cytolysis in protective immunity to LM. Perforin deficiency resulted in an approximately 5-fold reduction in the per-cell protective capacity of Ag-specific memory CD8(+) T cells that was not caused by differences in memory cell quality as measured by CD62L/CD27 expression, TCR repertoire use, functional avidity, differences in expansion of Ag-specific cells upon infection, or maintenance of memory levels over time. However, perforin-deficient CD8(+) T cells exhibited reduced in vivo cytotoxic function compared to WT CD8(+) T cells. Consistent with the existence of perforin-independent effector pathways, double-vaccinated PKO mice were as resistant to challenge with LM as single-vaccinated WT mice. Thus, increasing the number of memory CD8(+) T cells can overcome diminished per-cell protective immunity in the absence of perforin.     

Disparate primary and secondary allospecific CD8+ T cell cytolytic effector function in the presence or absence of host CD4+ T cells

The role of CD4+ T cells in promoting CD8+ T cell effector activity in response to transplant Ags in vivo has not been reported. We used a hepatocellular allograft model known to initiate both CD4-dependent and CD4-independent rejection responses to investigate the contribution of CD4+ T cells to the development, function, and persistence of allospecific CD8+ T cell effectors in vivo. Complete MHC-mismatched hepatocellular allografts were transplanted into C57BL/6 (CD4-sufficient) or CD4 knockout (CD4-deficient) hosts. The development of in vivo allospecific cytotoxicity was determined by clearance of CFSE-labeled target cells. CD8+ T cell cytotoxic effector activity was enhanced in response to allogeneic hepatocellular grafts with a greater magnitude of allocytotoxicity and a prolonged persistence of CTL effector activity in CD4-sufficient hosts compared with CD4-deficient hosts. Cytolytic activity was mediated by CD8+ T cells in both recipient groups. In response to a second hepatocyte transplant, rejection kinetics were enhanced in both CD4-sufficient and CD4-deficient hepatocyte recipients. However, only CD4-sufficient hosts developed recall CTL responses with an augmented magnitude and persistence of allocytotoxicity in comparison with primary CTL responses. These studies show important functional differences between alloreactive CD8+ T cell cytolytic effectors that mature in vivo in the presence or absence of CD4+ T cells.     

Somatostatin down-regulates LFA-1 activation by modulating Rap1 expression in CD4+ and CD8+ T cells

Leukocyte function-associated antigen-1 (LFA-1) is one of the integrins that are expressed on the leukocytes, and has been shown to play an important role in leukocyte trafficking. The adhesive activity of LFA-1 is governed partially by the Rap1. This study examined that the relationship between LFA-1 and Rap1 mRNA expressions by anti-CD3 and anti-CD3+SOM treatment in the CD4+ and CD8+ T cells. The LFA-1 mRNA expression levels following the anti-CD3 and anti-CD3+SOM treatment for 30 min was greater on the CD8+ T cells, and the LFA-1 expression of the CD8+ T cells with anti-CD+SOM treatment was affected more severely than that of the CD4+ T cells. The Rap1 mRNA expression patterns following anti-CD3 and anti-CD3+SOM stimulation in the CD4+ and CD8+ T cells were similar to the LFA-1 expression patterns, and the expression level following anti-CD3+SOM treatment was suppressed more significantly in the CD8+ T cells. These results suggest that the difference in the Rap1 expression level after stimulation might explain the differences in the LFA-1 expression level on the T cell subsets, and that the down-regulation of Rap1 expression following SOM treatment is closely related to the diminished LFA-1 expression.     

Anti-CD8 antibodies can trigger CD8+ T cell effector function in the absence of TCR engagement and improve peptide-MHCI tetramer staining

CD8(+) T cells recognize immunogenic peptides presented at the cell surface bound to MHCI molecules. Ag recognition involves the binding of both TCR and CD8 coreceptor to the same peptide-MHCI (pMHCI) ligand. Specificity is determined by the TCR, whereas CD8 mediates effects on Ag sensitivity. Anti-CD8 Abs have been used extensively to examine the role of CD8 in CD8(+) T cell activation. However, as previous studies have yielded conflicting results, it is unclear from the literature whether anti-CD8 Abs per se are capable of inducing effector function. In this article, we report on the ability of seven monoclonal anti-human CD8 Abs to activate six human CD8(+) T cell clones with a total of five different specificities. Six of seven anti-human CD8 Abs tested did not activate CD8(+) T cells. In contrast, one anti-human CD8 Ab, OKT8, induced effector function in all CD8(+) T cells examined. Moreover, OKT8 was found to enhance TCR/pMHCI on-rates and, as a consequence, could be used to improve pMHCI tetramer staining and the visualization of Ag-specific CD8(+) T cells. The anti-mouse CD8 Abs, CT-CD8a and CT-CD8b, also activated CD8(+) T cells despite opposing effects on pMHCI tetramer staining. The observed heterogeneity in the ability of anti-CD8 Abs to trigger T cell effector function provides an explanation for the apparent incongruity observed in previous studies and should be taken into consideration when interpreting results generated with these reagents. Furthermore, the ability of Ab-mediated CD8 engagement to deliver an activation signal underscores the importance of CD8 in CD8(+) T cell signaling.     

Defective alloreactive CD8 T cell function and memory response in allograft recipients in the absence of CD4 help

We have shown that alloreactive CD8 T cell activation may proceed via CD4-dependent and CD4-independent pathways, and that CD8 T cell activation in Ag-primed animals is independent of CD154 costimulation. In this report, we further analyzed the activation and function of alloreactive CD8 CTL effectors in CD4 knockout (KO) skin/cardiac allograft recipients. FACS analysis showed that alloreactive CD8 T cells were activated at a significantly reduced level in CD4 KO mice. Importantly, these helpless CD8 T cells failed to develop CD154 blockade resistance following reactivation by the same alloantigen, indicative of defective memory formation. Only transient CD4 help was required, as short-term CD4 blockade at the time of first skin graft challenge only delayed alloreactive CD8 activation, without affecting the CD8 T cell memory response to a second skin graft. Moreover, postoperative CD4 blockade had no effect on alloreactive CD8 activation. Alloreactive CD8 cells generated in the absence of CD4 help exhibited decreased effector responses. Interestingly, intragraft induction of T cell-targeted chemokines early after transplant was also dependent on CD4 help, as the induction kinetics of CXCL9 and CCL5 in CD4 KO recipients was significantly delayed, coupled with similarly delayed infiltration by CD3/CD8 cells. Remarkably, helpless CD8 cells ultimately entering the graft still displayed significantly diminished T cell effector molecules (IFN-gamma, granzyme B). Thus, CD4 help is critical for alloreactive CD8 activation, function, and memory formation.     

Targeting LFA-1 and cd154 suppresses the in vivo activation and development of cytolytic (cd4-Independent) CD8+ T cells

Short-term immunotherapy targeting both LFA-1 and CD40/CD154 costimulation produces synergistic effects such that long-term allograft survival is achieved in the majority of recipients. This immunotherapeutic strategy has been reported to induce the development of CD4+ regulatory T cells. In the current study, the mechanisms by which this immunotherapeutic strategy prevents CD8+ T cell-dependent hepatocyte rejection in CD4 knockout mice were examined. Combined blockade of LFA-1 and CD40/CD154 costimulation did not influence the overall number or composition of inflammatory cells infiltrating the liver where transplanted hepatocytes engraft. Expression of T cell activation markers CD43, CD69, and adhesion molecule CD103 by liver-infiltrating cells was suppressed in treated mice with long-term hepatocellular allograft survival compared to liver-infiltrating cells of untreated rejector mice. Short-term immunotherapy with anti-LFA-1 and anti-CD154 mAb also abrogated the in vivo development of alloreactive CD8+ cytotoxic T cell effectors. Treated mice with long-term hepatocyte allograft survival did not reject hepatocellular allografts despite adoptive transfer of naive CD8+ T cells. Unexpectedly, treated mice with long-term hepatocellular allograft survival demonstrated prominent donor-reactive delayed-type hypersensitivity responses, which were increased in comparison to untreated hepatocyte rejectors. Collectively, these findings support the conclusion that short-term immunotherapy with anti-LFA-1 and anti-CD154 mAbs induces long-term survival of hepatocellular allografts by interfering with CD8+ T cell activation and development of CTL effector function. In addition, these recipients with long-term hepatocellular allograft acceptance show evidence of immunoregulation which is not due to immune deletion or ignorance and is associated with early development of a novel CD8+CD25high cell population in the liver.     

TCR affinity promotes CD8+ T cell expansion by regulating survival

Ligation with high affinity ligands are known to induce T lymphocytes to become fully activated effector cells while ligation with low affinity ligands (or partial agonists) may result in a delayed or incomplete response. We have examined the quantitative features of CD8(+) T cell proliferation induced by peptides of different TCR affinities at a range of concentrations in the mouse OT-I model. Both the frequency of cells responding and the average time taken for cells to reach their first division are affected by peptide concentration and affinity. Consecutive division times, however, remained largely unaffected by these variables. Importantly, we identified affinity to be the sole regulator of cell death in subsequent division. These results suggest a mechanism whereby TCR affinity detection can modulate the subsequent rate of T cell growth and ensure the dominance of higher affinity clones over time.     

Disregulated influenza A virus-specific CD8+ T cell homeostasis in the absence of IFN-gamma signaling

Recent studies indicate that IFN-gamma may influence both the expansion and the trafficking of virus-specific CD8+ CTL, though the effects are not necessarily consistent for different models of viral and bacterial disease. Influenza A virus infection of mice deficient for IFN-gamma (IFN-gamma(-/-)) or deficient for the IFN-gamma receptor 1 (IFNGR1(-/-)) was, when compared with the wild-type (WT) B6 controls, associated with increased Ag-specific CD8+ T cell counts in the spleen and mediastinal lymph nodes. At the same time, fewer of these CTL effectors were found in the bronchoalveolar lavage population recovered from the IFN-gamma(-/-) mice. Comparable effects were observed for WT mice treated with a neutralizing IFN-gamma-specific mAb. Transfer of WT memory Thy1.1(+) CD8+ populations into Thy1.2+ B6 IFN-gamma(-/-) or IFNGR1(-/-) mice followed by intranasal virus challenge demonstrated both that IFN-gamma produced by the host was important for the regulation of Ag-specific CTL numbers and that IFN-gamma was likely to act directly on the T cells themselves. In addition, the prevalence of CTLs undergoing apoptosis in spleen was lower when measured directly ex vivo for IFN-gamma(-/-) vs WT B6 mice. The present analysis is the first comprehensive demonstration that IFN-gamma signaling can differentially regulate both Ag-specific CTL homeostasis in secondary lymphoid organs and trafficking to a site of virus-induced pathology.     

Defective CD8+ T cell peripheral tolerance in nonobese diabetic mice.

Nonobese diabetic (NOD) mice develop spontaneous autoimmune diabetes that involves participation of both CD4+ and CD8+ T cells. Previous studies have demonstrated spontaneous reactivity to self-Ags within the CD4+ T cell compartment in this strain. Whether CD8+ T cells in NOD mice achieve and maintain tolerance to self-Ags has not previously been evaluated. To investigate this issue, we have assessed the extent of tolerance to a model pancreatic Ag, the hemagglutinin (HA) molecule of influenza virus, that is transgenically expressed by pancreatic islet beta cells in InsHA mice. Previous studies have demonstrated that BALB/c and B10.D2 mice that express this transgene exhibit tolerance of HA and retain only low-avidity CD8+ T cells specific for the dominant peptide epitope of HA. In this study, we present data that demonstrate a deficiency in peripheral tolerance within the CD8+ T cell repertoire of NOD-InsHA mice. CD8+ T cells can be obtained from NOD-InsHA mice that exhibit high avidity for HA, as measured by tetramer (K(d)HA) binding and dose titration analysis. Significantly, these autoreactive CD8+ T cells can cause diabetes very rapidly upon adoptive transfer into NOD-InsHA recipient mice. The data presented demonstrate a retention in the repertoire of CD8+ T cells with high avidity for islet Ags that could contribute to autoimmune diabetes in NOD mice.     

Induction of polyclonal CD8+ T cell activation and effector function by Pertussis toxin

Pertussis toxin (PTX) has pronounced adjuvant activity and strongly enhances innate and adaptive immune responses, including increased antibody production and Th1/Th2 cytokine production. Adjuvant effects of PTX on Th1 and Th2 cells are primarily mediated via CD80/86 costimulation via enhanced expression of these molecules by APCs. However, it has remained unresolved whether PTX modulates the expression of costimulatory and inhibitory molecules on CD4+ and CD8+ T cells. To address this question, we determined the expression kinetics of CD28, CTLA-4, and CD40L on spleen CD4+ and CD8+ T cells after incubation with PTX. The results show that PTX upregulated the expression of CD28 by CD8+ T cells, but not by CD4+ T cells. In contrast, the expression of CTLA-4 and CD40L was not substantially altered on CD4+ or CD8+ T cells. CD28 upregulation by CD8+ T cells was paralleled by upregulation of CD69 and the induction of IFN-γ, Granzyme B (GrB), and IL-17. CD8+ T cell activation and cytokine production could be substantially blocked with anti-CD80 and CD86 antibodies, consistent with CD28 mediated signaling. Treatment of highly purified CD8+ T cells with PTX resulted in upregulation of CD28 and CD69, and production of IFN-γ. Incubation with CD28 mAb further enhanced this effect, suggesting that PTX has direct effects on CD8+ T cells which are enhanced by CD80/86-mediated costimulation provided by APCs.     

Potentiating CD8+ T cell antitumor activity by inhibiting PCSK9 to promote LDLR-mediated TCR recycling and signaling

Metabolic regulation has been proven to play a critical role in T cell antitumor immunity.However,cholesterol metabolism as a key component of this regulation remains largely unexplored.Herein,we found that the low-density lipoprotein receptor (LDLR),which has been previously identified as a transporter for cholesterol,plays a pivotal role in regulating CD8+ T cell antitumor activity.Besides the involvement of cholesterol uptake which is mediated by LDLR in T cell priming and clonal expansion,we also found a non-canonical function of LDLR in CD8+ T cells: LDLR interacts with the T-cell receptor (TCR) complex and regulates TCR recycling and signaling,thus facilitating the effector function of cytotoxic T-lymphocytes (CTLs).Furthermore,we found that the tumor microenvironment (TME) downregulates CD8+ T cell LDLR level and TCR signaling via tumor cell-derived proprotein convertase subtilisin/kexin type 9 (PCSK9) which binds to LDLR and prevents the recycling of LDLR and TCR to the plasma membrane thus inhibits the effector function of CTLs.Moreover,genetic deletion or pharmacological inhibition of PCSK9 in tumor cells can enhance the antitumor activity of CD8+ T cells by alleviating the suppressive effect on CD8+ T cells and consequently inhibit tumor progression.While previously established as a hypercholesterolemia target,this study highlights PCSK9/LDLR as a potential target for cancer immunotherapy as well.     

The role of the T cell receptor, CD8, and LFA-1 in different stages of the cytolytic reaction mediated by alloreactive T lymphocyte clones

The cytotoxic reaction mediated by cytotoxic T lymphocytes (CTL) consists of three phases: first, the CTL binds to the target cell; next, the CTL is triggered to lyse the target cell; and in the third phase, the CTL detaches from the target cell which is lysed in the absence of the CTL. Recently, we obtained evidence that human alloreactive CTL clones initially adhere to target cells without the involvement of the interaction between the T cell receptor (Tcr) and its specific target antigen. In the present study, we investigated the effect of monoclonal antibodies specific for the Tcr on the cytotoxic reaction of three CD8+ HLA-A2-specific CTL clones, using a single cell assay in which the binding event can be distinguished from the post-binding (lytic) phase of the cytolytic reaction. It was found that monoclonal antibodies directed at a variable part of the Tcr do not affect the binding phase but strongly block the lytic phase of the cytotoxic reaction. An anti-constant region Tcr antibody and an anti-CD3 reagent had a similar effect on the two phases of the reaction as the anti-variable part Tcr antibodies. In contrast, antibodies specific for LFA-1 strongly blocked the adhesion phase but did not affect the lytic phase. Antibodies specific for CD-8 had intermediate effects. They could block both the adhesion as well as the lytic phase. The effect of anti-CD8 appeared to be dependent on the CTL clone tested. One clone was found to be inhibited in the adhesion phase, but not in the lytic phase, whereas anti-CD8 hardly blocked the adhesion phase of two other CTL clones, but affected the lytic step of those clones. Our data indicate that LFA-1 is a major adhesion molecule in the CTL reaction, whereas the Tcr/CD3 complex is implicated in a phase after the initial formation of conjugates. CD8 is associated with both steps in the cytolytic reaction. In addition to its minor role in the adhesion phase, our data suggest strongly that CD-8 is involved in the triggering phase of the cytolytic reaction.     

In vivo CD86 blockade inhibits CD4+ T cell activation, whereas CD80 blockade potentiates CD8+ T cell activation and CTL effector function

To address whether a functional dichotomy exists between CD80 and CD86 in naive T cell activation in vivo, we administered anti-CD80 or CD86 blocking mAb alone or in combination to mice with parent-into-F(1) graft-vs-host disease (GVHD). In this model, the injection of naive parental T cells into unirradiated F(1) mice results in either a Th1 cytokine-driven, cell-mediated immune response (acute GVHD) or a Th2 cytokine-driven, Ab-mediated response (chronic GVHD) in the same F(1) recipient. Combined CD80/CD86 blockade beginning at the time of donor cell transfer mimicked previous results seen with CTLA4Ig and completely abrogated either acute or chronic GVHD by preventing the activation and maturation of donor CD4(+) T cells as measured by a block in acquisition of memory marker phenotype and cytokine production. Similar results were seen with selective CD86 blockade; however, the degree of CD4 inhibition was always less than that seen with combined CD80/CD86 blockade. A more striking effect was seen with selective CD80 blockade in that chronic GVHD was converted to acute GVHD. This effect was associated with the induction of Th1 cytokine production, donor CD8(+) T cell activation, and development of antihost CTL. The similarity of this effect to that reported for selective CTLA4 blockade suggests that CD80 is a critical ligand for CTLA4 in mediating the down-regulation of Th1 responses and CD8(+) T cell activation. In contrast, CD86 is critical for the activation of naive CD4(+) T cells in either a Th1 or a Th2 cytokine-mediated response.     

LFA-1-mediated costimulation of CD8+ T cell proliferation requires phosphatidylinositol 3-kinase activity

LFA-1 binding to ICAM-I provides a costimulatory signal for CD8(+) T cell activation that results in increased IL-2 mRNA levels and protein production to support proliferation. CD28 binding to its B7 ligands has the same effect, and the two costimulatory receptors activate some of the same intracellular signaling events, including up-regulation of phosphatidylinositol (PI) 3-kinase activity. However, costimulation by LFA-1 depends upon the activity of this enzyme, whereas costimulation by CD28 does not, as evidenced by differential effects of specific inhibitors of PI 3-kinase. When cells are costimulated with ICAM-1 in the presence of the inhibitors wortmannin or LY294002, proliferation is blocked, but increases in IL-2 mRNA levels and protein production are not. Costimulation also results in increased surface expression of CD25, which is essential for formation of an active IL-2R. This is blocked by the PI 3-kinase inhibitors when costimulation is via LFA-1 but not when it is via CD28. Finally, IL-2-driven proliferation is not blocked by the inhibitors once CD25 surface expression has increased. Thus, the PI 3-kinase-dependent step in CD8 T cell costimulation by LFA-1 is up-regulation of IL-2R expression. In contrast, CD28 engagement also increases IL-2R surface expression, but the up-regulation does not depend upon PI 3-kinase activity.     

PSGL-1 attenuates early TCR signaling to suppress CD8+ T cell progenitor differentiation and elicit terminal CD8+ T cell exhaustion

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Subpopulations of CD8+ cytotoxic T cell precursors collaborate in the absence of conventional CD4+ helper T cells.

Four different subpopulations (Ly6Cneg, Ly6Clow, Ly6Cint, and Ly6Chi) of CD8+ T cells were arbitrarily defined on the basis of differential expression of Ly6C Ag. By combining the processes of electronic cell sorting and automated cell deposition, small numbers of respective CD8+ T cell subpopulations were directly deposited into tissue culture wells in which mitogen-stimulated responses were studied. Anti-CD3-stimulated proliferation and IL-2 production were the strongest by Ly6Cneg/Ly6Clow T cells, moderate for Ly6Cint T cells, and highly deficient for Ly6Chi T cells. The level of IL-2 production for Ly6Cneg CD8+ T cells was comparable to that of conventional CD4+ Th cells. Allogeneic stimulator cells elicited a strong cytotoxic response by Ly6Cneg + low but not Ly6Chi CD8+ T cells in the absence of added lymphokines. When IL-2 was supplied in excess, anti-CD3 induced comparable levels of cell proliferation and cytotoxic activity in Ly6Cneg, Ly6Clow, Ly6Cint, and Ly6Chi CD8+ T cells whereas alloantigen stimulated an approximate fivefold higher cytotoxic response by Ly6Chi than Ly6Cneg + low CD8+ T cells. Stimulation of co-cultures of B10 (CD8b) Ly6Cneg + low and congenic B10.CD8a Ly6Chi CD8+ T cells in the absence of added lymphokines, followed by selective elimination of activated CD8.1+ (CD8.2+) T cells by anti-CD8.1 (anti-CD8.2) + C treatment, allowed the demonstration that help provided by Ly6Cneg + low T cells can be effectively used by both Ly6Cneg + low and Ly6Chi T cells in anti-CD3 and alloantigen induced proliferative and cytotoxic responses, respectively.     

Defective clonogenic potential of CD8+ T lymphocytes in patients with AIDS. Expansion in vivo of a nonclonogenic CD3+CD8+DR+CD25- T cell population

This study examines the potential mechanism(s) responsible for the defective clonability of CD8+ T lymphocytes in patients with AIDS. By the combined use of one- and two-color fluorescence cytofluorometry we have shown an increase in the number of circulating DR+ cells due to the expression of DR on a relatively large proportion of T lymphocytes (one-third of CD3+ cells), the majority of them belonging to the CD8+ subset. In addition, the majority of CD8+DR+ cells in AIDS patients did not express CD25 Ag (the receptor for IL-2), a surface marker generally expressed on normal activated T lymphocytes. Sorted CD8+DR+ and CD8+DR- cell populations were analyzed comparatively for their ability to proliferate in response to different stimuli, including anti-CD3, anti-CD2, alone or in combination with anti-CD28 mAb and mitogens such as PHA, alone or in combination with PMA. We have demonstrated that CD8+DR+ cells were severely defective in their proliferative response to triggering via these major pathways of T cell activation even when an exogenous source of IL-2 or IL-4 was added to the microcultures 24 h after initiating the cultures. In contrast, CD8+DR- cells showed a significant proliferation in response to the different stimuli and the proliferative response was strongly enhanced by the addition of IL-2 or IL-4. At the end of the stimulation period CD8+DR+ and CD8+DR- proliferating populations were analyzed for CD25 Ag expression. Only 1 to 10% of CD8+DR+ cells expressed CD25 antigen compared with 40 to 50% of CD8+DR- cells. The proliferative defect of CD8+DR+ cells was further confirmed in experiments performed at the clonal level. The analysis of the frequency of proliferating T lymphocyte-precursors in both CD8+DR+ and CD8+DR- subsets showed that the defective clonogenic potential of CD8+ cells in AIDS patients could be in large part ascribed to CD8+DR+ cells. Five percent of CD8+DR+ cells showed a clonogenic potential compared to the 25% of CD8+DR- cells. Finally, we analyzed the surface expression of VLA-2 Ag, a marker of a chronic state of T cell activation, on circulating T lymphocytes. We have shown that a large proportion of CD3+DR+CD25- cells (50 to 80% in the different patients with AIDS analyzed) expressed VLA-2 Ag.(ABSTRACT TRUNCATED AT 400 WORDS)     

The dependence for leukocyte function-associated antigen-1/ICAM-1 interactions in T cell activation cannot be overcome by expression of high density TCR ligand

The leukocyte-specific integrin, LFA-1, can enhance T cell activation. However, it is unclear whether the binding of LFA-1 to its ligand, ICAM-1, functions through intercellular adhesion alone, resulting in an augmentation of the TCR signal, or involves an additional LFA-1-mediated cellular signal transduction pathway. We have previously shown that naive CD4+ lymph node T cells, isolated from DO11.10 TCR transgenic mice, are activated by increasing doses of exogenous OVA peptide presented by transfectants expressing both class II and ICAM-1, but not by cells expressing class II alone. To determine whether LFA-1/ICAM-1 interactions were simply enhancing the presentation of low concentrations of specific MHC/peptide complexes generated from exogenously added peptide, we transfected cells with class II that is covalently coupled to peptide, alone or in combination with ICAM-1. These cells express 100-fold more specific class II/peptide complexes than can be loaded onto class II-positive cells at maximum concentrations of exogenous peptide. Despite this high density of TCR ligand, activation of naive CD4+ T cells still requires the coexpression of ICAM-1. LFA-1/ICAM-1 interactions are not required for effective conjugate formation and TCR engagement because presentation of class II/peptide complexes in the absence of ICAM-1 does induce up-regulation of CD25 and CD69. Thus, high numbers of engaged TCR cannot compensate for the lack of LFA-1/ICAM-1 interactions in the activation of naive CD4+ T cells.     

Direct interferon-gamma signaling dramatically enhances CD4+ and CD8+ T cell memory

Studies in IFN-gamma-deficient mice suggest that the delivery of IFN-gamma to CD8(+) T cells early in virus infection programs their eventual contraction, thereby reducing the abundance of CD8(+) memory T cells. In this study, we show that such mice fail to completely eliminate virus infection and that, when evaluated without the confounding factor of persisting Ag, both CD4(+) and CD8(+) T cells undergo profound contraction when they are unable to receive IFN-gamma signals. Furthermore, the abundance of CD4(+) and CD8(+) memory cells that express the IFN-gamma receptor is approximately 100-fold higher than cells lacking this molecule. Thus, direct IFN-gamma signaling is not required for T cell contraction during virus infection, and it enhances, rather than suppresses, the development of virus-specific CD4(+) and CD8(+) T cell memory.