Epitope Specificity and Relative Clonal Abundance Do Not Affect CD8 Differentiation Patterns during Lymphocytic Choriomeningitis Virus Infection (original) (raw)
Related papers
International Immunology, 2007
CD8 T cell responses to vaccinia virus (VV) and a virus-encoded ovalbumin peptide (OVAP) epitope were examined using adoptively transferred OT-I T cells. The results demonstrate that upon intraperitoneal challenge with ovalbumin-expressing VV (VV-OVAP), OT-I T cell proliferation occurs initially in lymph nodes and spleens followed by migration of the divided cells to the peritoneal cavity. Massive clonal expansion occurs in response to both the virus and the virus-encoded ovalbumin (OVA) epitope, as demonstrated using low numbers of adoptively transferred cells, and the responding OT-I cells display marked site-dependent functional heterogeneity with respect to IFN-g and tumor necrosis factor-a (TNF-a) production and granzyme B expression. OT-I cells responding to VV-OVAP develop the capacity to produce IFN-g in response to antigen as they proliferate and differentiate. In marked contrast, naive OT-I cells rapidly produce TNF-a upon antigen recognition, and this capacity declines as the cells proliferate in response to the virus, suggesting that this potent inflammatory cytokine may be important primarily during initiation of the response. At the peak of clonal expansion, a large fraction (30-60%) of the OT-I cells responding to the virus express high IL-7Ra levels, and the majority of these cells is subsequently lost. While high IL-7Ra expression may be necessary for a CD8 T cell to transition to memory, it is clearly not sufficient. Thus, OT-I cells responding to VV infection exhibit a high degree of heterogeneity within the responding population that differs depending on their anatomical location, despite the specificity and affinity of the TCR being identical on all of the cells.
absence of TCR -activation maintain an intact immune competence
2003
Gene transfer into T lymphocytes is currently tested for the treatment of lymphohematologic disorders. We previously showed that suicide gene transfer into donor lymphocytes infused to treat leukemic relapse after allogeneic hematopoietic stem cell transplantation allowed control of graft-versus-host disease. However, the T-cell receptor activation and sustained proliferation required for retroviral vector transduction may impair the half-life and immune competence of transduced cells and reduce graft-versus-leukemia activity. Thus, we tested lentiviral vectors (LV) and stimulation with cytokines involved in antigen-independent T-cell homeostasis, such as IL-7, IL-2 and IL-15. Late-generation LV transduced efficiently nonproliferating T cells which had progressed from G 0 to the G 1 phase of the cell cycle upon cytokine treatment. Importantly, IL-2 and IL-7, but not IL-15 stimulation preserved physiologic CD4/CD8 and naïve/memory ratios in transduced cells with only minor induction of some activation markers. Functional analysis of immune response to cytomegalovirus (CMV) showed that, although CMV-specific T cells were preserved by all conditions of transduction, proliferation and specific killing of autologous cells presenting a CMV epitope were higher for IL-2 and IL-7 than IL-15. Thus, LV transduction of IL-2 or IL-7 pre-stimulated cells overcome the limitations of retroviral vectors and may significantly improve the efficacy of T cell-based gene therapy. (Word count 202) For personal use only. by guest on June 10, 2013. bloodjournal.hematologylibrary.org From recently shown that cytokines, such as IL-7, IL-2 and IL-15, able to promote longterm survival in vitro of memory and naïve T lymphocytes (24-25), can render T cells susceptible to LV transduction in the absence of TCR-activation (22-23) (26). In this study, we confirmed and extended these findings using late-generation LV with advanced performance and safety profiles and showing efficient transduction of T cells which have progressed from G 0 into the G 1 phase of the cell cycle upon cytokine treatment without becoming committed to proliferation. Most importantly, we showed that culture conditions allowing for LV transduction of T cells in the absence of TCR-triggering result in preservation of full immune competence, thus allowing to overcome the intrinsic functional limitations of retroviral-mediated transduction. Methods Cell culture HeLa and 293T human embryonic kidney cell lines were cultured in Iscove's Modified Dulbecco's Medium (IMDM, Sigma Chemical Co, Milan, Italy) supplemented with 10% Fetal Bovine Serum (FBS, Gibco BRL, Inchinnam, Scotland UK) and a combination of Penicillin (100IU/ml), Streptomycin (100µg/ml) and L-Glutamine (2mM). Peripheral blood mononuclear cells (PBMC) were isolated from healthy donors by leukapheresis and Ficoll-Hypaque gradient separation (Pharmacia, Uppsala, Sweden). Cells were cultured in IMDM supplemented with 10% Fetal Calf Serum (FCS, Gibco BRL) at a density of 1x10 6 cells/ml in the presence of different cytokines: human recombinant IL-2 10U/ml (EuroCetus Italia S.r.l., Milan, Italy); human recombinant IL-7 5ng/ml (Boehringer Mannheim-Roche GmbH, Mannheim, Germany); human recombinant IL-15 10ng/ml (R&D system or PeproTech Inc,Rocky Hill, NH). TCR
Journal of Virology, 2007
Measuring the magnitudes and specificities of antiviral CD8 T-cell responses is critical for understanding the dynamics and regulation of adaptive immunity. Despite many excellent studies, the accurate measurement of the total CD8 T-cell response directed against a particular infection has been hampered by an incomplete knowledge of all CD8 T-cell epitopes and also by potential contributions of bystander expansion among CD8 T cells of irrelevant specificities. Here, we use several techniques to provide a more complete accounting of the CD8 T-cell response generated upon infection of C57BL/6 mice with lymphocytic choriomeningitis virus (LCMV). Eight days following infection, we found that 85 to 95% of CD8 T cells exhibit an effector phenotype as indicated by granzyme B, 1B11, CD62L, CD11a, and CD127 expression. We demonstrate that CD8 T-cell expansion is due to cells that divide >7 times, whereas heterologous viral infections only elicited <3 divisions among bystander memory CD8 T cells. Furthermore, we found that approximately 80% of CD8 T cells in spleen were specific for ten different LCMV-derived epitopes at the peak of primary infection. These data suggest that following a single LCMV infection, effector CD8 T cells divide >15 times and account for at least 80%, and possibly as much as 95%, of the CD8 T-cell pool. Moreover, the response targeted a very broad array of peptide major histocompatibility complexes (MHCs), even though we examined epitopes derived from only two of the four proteins encoded by the LCMV genome and C57BL/6 mice only have two MHC class I alleles. These data illustrate the potential enormity, specificity, and breadth of CD8 T-cell responses to viral infection and demonstrate that bystander activation does not contribute to CD8 T-cell expansion.
The Influenza Virus–Specific CTL Immunodominance Hierarchy in Mice Is Determined by the Relative Frequency of High-Avidity T Cells, 2014
Virus-specific CTL responses typically fall into reproducible hierarchies with particular epitopes eliciting either immunodominant or subdominant responses after viral challenge. The recently acquired capacity to directly enumerate naive CTL precursors (CTLps) in both mice and humans has implicated CTLp frequency as a key predictor of immune response magnitude after Ag challenge. However, recent studies have indicated that naive CTLp frequencies do not necessarily predict the size of the Ag driven response, indicating an important role for differential CTLp recruitment and/or expansion. This study characterizes the early emergence of various influenza epitope-specific CTL responses at multiple sites in C57BL/6 mice, and probes the role of Ag dose and TCR avidity in dictating immune response hierarchies. Despite large naive CTLp numbers, subdominance was found to arise largely as a consequence of the abrupt and premature cessation of CTL proliferation, at least for one epitope specificity. Investigation into the possible drivers of the poor proliferation observed for subdominant specificities showed that the immunodominance hierarchy endured irrespective of epitope abundance, and correlated with the prevalence of high-avidity T cells in both the naive and immune compartments. Our study strongly indicates that the quality, and not simply the quantity, of antiviral CTLs dictate response magnitude.
European Journal of Immunology, 1993
The question of functional differentiation within the CD8 subset has been addressed in a model of TcR-transgenic (TcR-tg) mice expressing a TcR specific for H-2Kb (Ti). CD8+ Ti+ T cells present in the periphery of these mice have no cytotoxic T lymphocyte (CTL) activity unless they are stimulated with H-2Kb-expressing cells. In contrast to T cells from normal H-2k littermates, alloantigen induction of CTL fromTcR-tg mice is independent of CD4+ T helper (Th) cells and is accompanied by high level secretion of interleukin-(11)-2 by Ti+ CD8+ T cells. Precursor frequency analysis performed on CD8+ cells fromTcR-tg mice revealed a high frequency of T h as compared to CTL precursors. This raised the possibility of the existence of distinct subpopulations within CD8+ precursors with different requirements for differentiation to functional CTL. FACS analyses (performed on resting and on in vitro stimulated Tcells from normal and TcR-tg mice) demonstrated a heterogeneous expression of Ly-6C on CD8+ cells with a large enrichment of Ly-6C-cells among the Ti+ cells which persisted after stimulation with H-2b cells in conditions that led to a homogeneous expression of the activation markers pgp-1 and CD69. The possibility that Ly-6C expression could mark functionally different subpopulations in CD8+ T cells was investigated. Stimulation of sorted populations of Ly-6C-and Ly-6C+ cells allowed detection of CTL precursors in both these subsets and the majority of limiting dilution wells containing one pCTL also scored positive for IL-2 secretion. Thus, for CD8+ Tcells expressing the same TcR, differentiation led to acquisition of both IL-2 secretion and CTL function and there was no evidence for the existence of a distinct population of helper-dependent CTL precursors.
Journal of General Virology, 2004
The ability of virus-specific CD8 + T cells to produce cytokines was studied in mice infected with lymphocytic choriomeningitis virus and vesicular stomatitis virus. Intracellular staining was used to visualize cytokine-producing CD8 + and CD4 + T cells. Overall, virus-specific CD8 + T cells produce a similar range of cytokines (IFN-c, TNF-a, IL-2, GM-CSF, RANTES, MIP-1a and MIP-1b) as CD4 + T cells, but the relative distribution of cytokine-producing subsets is different. Moreover, cytokine-producing CD8 + T cells were found to dominate numerically at all time-points tested. Co-staining for more than one cytokine revealed that while all cytokine-producing CD8 + T cells synthesized IFN-c, additional cytokines were produced by partly overlapping subsets of this population. The frequency of cells producing more than one cytokine was higher in a tertiary site (peritoneum) and generally increased with transition into the memory phase; however, GM-CSF producing cells were only present transiently. Concerning factors predicted to influence the distribution of cytokine-producing subsets, IFN-c and IL-12 did not play a role, nor was extensive virus replication essential. Notably, regarding the heterogeneity in cytokine production by individual cells with similar epitope specificity, variation in TCR avidity was not the cause, since in vivo-activated TCR transgene-expressing cells were as heterogeneous in cytokine expression as polyclonal cells specific for the same epitope.
The Journal of Immunology, 2010
Lymphocytic choriomeningitis virus (LCMV)-specific CD8 + T cell responses are considered to be independent of CD28-B7 costimulation. However, the LCMV-specific response has never been evaluated in B7.1/B7.2 2/2 mice. For this reason, we decided to study the T cell response in B7.1/B7.2 2/2 mice infected with two different strains of LCMV, one (Traub strain) typically causing low-grade chronic infection, and another (Armstrong clone 53b) displaying very limited capacity for establishing chronic infection. Using Traub virus we found that most B7.1/B7.2 2/2 mice were unable to rid themselves of the infection. Chronic infection was associated with a perturbed CD8 + T cell epitope hierarchy, as well as with the accumulation of cells expressing markers of terminal differentiation and being unable to respond optimally to Ag restimulation. Examination of matched CD28 2/2 mice revealed a similar albeit less pronounced pattern of CD8 + T cell dysfunction despite lack of virus persistence. Finally, analysis of B7.1/B7.2 2/2 mice infected with Armstrong virus revealed a scenario quite similar to that in Traub infected CD28 2/2 mice; that is, the mice displayed evidence of T cell dysfunction, but no chronic infection. Taken together, these results indicate that B7 costimulation is required for induction and maintenance of LCMV-specific CD8 + T cell memory, irrespective of the LCMV strain used for priming. However, the erosion of CD8 + T cell memory in B7.1/B7.2 2/2 mice was more pronounced in association with chronic infection. Finally, virus-specific T cell memory was more impaired in the absence of B7 molecules than in the absence of the CD28 receptor, supporting earlier data suggesting the existence of additional stimulatory receptors for B7.
Private specificities of CD8 T cell responses control patterns of heterologous immunity
Journal of Experimental Medicine, 2005
CD8 T cell cross-reactivity between viruses can play roles in protective heterologous immunity and damaging immunopathology. This cross-reactivity is sometimes predictable, such as between lymphocytic choriomeningitis virus (LCMV) and Pichinde virus, where crossreactive epitopes share six out of eight amino acids. Here, however, we demonstrate more subtle and less predictable cross-reactivity between LCMV and the unrelated vaccinia virus (VV). Epitope-specific T cell receptor usage differed between individual LCMV-infected C57BL/6 mice, even though the mice had similar epitope-specific T cell hierarchies.