Claerwen M. Jones

The CD8alpha(+) dendritic cell is responsible for inducing peripheral self-tolerance to tissue-associated antigens.

We previously described a mechanism for the maintenance of peripheral self-tolerance. This involves the cross-presentation of tissue-associated antigens by a bone marrow–derived cell type that stimulates the proliferation and ultimate deletion of self-reactive CD8 T cells. This process has been referred to as cross-tolerance. Here, we characterize the elusive cell type responsible for inducing cross-tolerance as a CD8α+ dendritic cell (DC). To achieve this aim, transgenic mice were generated expressing yellow fluorescent protein (YFP) linked to CTL epitopes for ovalbumin and glycoprotein B (gB) of herpes simplex virus under the rat insulin promoter (RIP). Although tracking of YFP was inconclusive, the use of a highly sensitive gB-specific hybridoma that produced β-galactosidase on encounter with antigen, enabled detection of antigen presentation by cells isolated from the pancreatic lymph node. This showed that a CD11c+CD8α+ cell was responsible for cross-tolerance, the same DC subset as previously implicated in cross-priming. These data indicate that CD8α+ DCs play a critical role in both tolerance and immunity to cell-associated antigens, providing a potential mechanism by which cytotoxic T lymphocyte can be immunized to viral antigens while maintaining tolerance to self.

Long-lived epithelial immunity by tissue-resident memory T (TRM) cells in the absence of persisting local antigen presentation

Although circulating memory T cells provide enhanced protection against pathogen challenge, they often fail to do so if infection is localized to peripheral or extralymphoid compartments. In those cases, it is T cells already resident at the site of virus challenge that offer superior immune protection. These tissue-resident memory T (TRM) cells are identified by their expression of the α-chain from the integrin αE(CD103)β7, and can exist in disequilibrium with the blood, remaining in the local environment long after peripheral infections subside. In this study, we demonstrate that long-lived intraepithelial CD103+CD8+ TRM cells can be generated in the absence of in situ antigen recognition. Local inflammation in skin and mucosa alone resulted in enhanced recruitment of effector populations and their conversion to the TRM phenotype. The CD8+ TRM cells lodged in these barrier tissues provided long-lived protection against local challenge with herpes simplex virus in skin and vagina challenge models, and were clearly superior to the circulating memory T-cell cohort. The results demonstrate that peripheral TRM cells can be generated and survive in the absence of local antigen presentation and provide a powerful means of achieving immune protection against peripheral infection.

Rapid Cytotoxic T Lymphocyte Activation Occurs in the Draining Lymph Nodes After Cutaneous Herpes Simplex Virus Infection as a Result of Early Antigen Presentation and Not the Presence of Virus

Localized cutaneous herpes simplex virus type 1 (HSV-1) infection leads to arming and initial expansion of cytotoxic T lymphocytes (CTLs) in the draining popliteal lymph nodes (PLNs) followed by migration and further proliferation in the spleen. To accurately characterize the sequence of events involved in the activation and generation of anti-HSV CTLs, we used T cell receptor (TCR) transgenic mice specific for the immunodominant epitope from HSV glycoprotein B (gB498–505). We describe the detection of the initiation of antigen presentation in the draining lymph nodes by 4–6 h after infection with HSV-1. Analysis of CD69 up-regulation revealed activation of gB-specific CD8+ T cells by 6–8 h after infection. Furthermore, we show that T cell proliferation begins no sooner than 24 h after activation and is marked by the concurrent appearance of CTL activity in the PLNs. These events are not dependent on the presence of virus in the draining lymph nodes, and suggest a requirement for recruitment of professional antigen-presenting cells to the site of T cell activation. Consequently, we have defined the initiation of the CD8+ T cell–mediated response to cutaneous HSV-1 infection, demonstrating that the immune response to localized viral infection depends only on the appearance of cells presenting virus-derived antigen and commences with remarkable swiftness.

Characterization of two TCR transgenic mouse lines specific for herpes simplex virus

To better understand the T cell‐mediated processes involved in the immune response to herpes simplex virus type 1 (HSV‐1) infection, two HSV‐specific T cell receptor (TCR) transgenic mouse lines were produced. These mice (gBT‐I.1 and gBT‐I.3) are MHC class I‐restricted and specific for the immunodominant peptide from HSV glycoprotein B (gB), gB498–505. Although derived from the same clone, the mice differ in the chromosomal location of the TCR transgenes and show marked differences in TCR α/β expression on both CD4+ and CD8+ cells in the thymus. Despite this, peripheral CD8+ T cells from both mice express equally high levels of the transgenic TCR and bind the KbgB498–505 tetramer to the same degree. In concordance with this, both were shown to respond equally well in vitro upon stimulation with the gB498–505 peptide or HSV‐infected cells. These data show that selection of broadly equivalent peripheral T‐cell subsets can occur in the presence of distinctly different thymic T‐cell subsets.

CD8+ T Cells from a Novel T Cell Receptor Transgenic Mouse Induce Liver-Stage Immunity That Can Be Boosted by Blood-Stage Infection in Rodent Malaria

To follow the fate of CD8+ T cells responsive to Plasmodium berghei ANKA (PbA) infection, we generated an MHC I-restricted TCR transgenic mouse line against this pathogen. T cells from this line, termed PbT-I T cells, were able to respond to blood-stage infection by PbA and two other rodent malaria species, P. yoelii XNL and P. chabaudi AS. These PbT-I T cells were also able to respond to sporozoites and to protect mice from liver-stage infection. Examination of the requirements for priming after intravenous administration of irradiated sporozoites, an effective vaccination approach, showed that the spleen rather than the liver was the main site of priming and that responses depended on CD8α+ dendritic cells. Importantly, sequential exposure to irradiated sporozoites followed two days later by blood-stage infection led to augmented PbT-I T cell expansion. These findings indicate that PbT-I T cells are a highly versatile tool for studying multiple stages and species of rodent malaria and suggest that cross-stage reactive CD8+ T cells may be utilized in liver-stage vaccine design to enable boosting by blood-stage infections.

Long-Term Retention of Mature NK1.1+ NKT Cells in the Thymus

The NKT cell pool in the thymus contains immature (NK1.1−) and mature (NK1.1+) subsets that represent distinct linear stages of a linear developmental pathway. An unexplained paradox is why immature NK1.1− NKT cells are mainly exported to the periphery instead of the more mature and more abundant NK1.1+ NKT cells. In this study we have determined that mature NK1.1+ NKT cells are retained by the thymus to form an extremely long-lived resident population capable of rapid and prolonged production of IFN-γ and IL-4. The retention of mature NKT cells provides an explanation for why the periphery is mainly seeded by immature NK1.1− cells despite mature NK1.1+ NKT cells being more abundant in the thymus. This is the first study to identify a mature T cell subset retained within the thymus and is additional evidence of the distinct developmental pathways of mainstream T cells and NKT cells.

Maintenance of T Cell Function in the Face of Chronic Antigen Stimulation and Repeated Reactivation for a Latent Virus Infection

Persisting infections are often associated with chronic T cell activation. For certain pathogens, this can lead to T cell exhaustion and survival of what is otherwise a cleared infection. In contrast, for herpesviruses, T cells never eliminate infection once it is established. Instead, effective immunity appears to maintain these pathogens in a state of latency. We used infection with HSV to examine whether effector-type T cells undergoing chronic stimulation retained functional and proliferative capacity during latency and subsequent reactivation. We found that latency-associated T cells exhibited a polyfunctional phenotype and could secrete a range of effector cytokines. These T cells were also capable of mounting a recall proliferative response on HSV reactivation and could do so repeatedly. Thus, for this latent infection, T cells subjected to chronic Ag stimulation and periodic reactivation retain the ability to respond to local virus challenge.

CD4 + T Cells Can Protect APC from CTL-Mediated Elimination

Professional APC play a central role in generating antiviral CD8+ CTL immunity. However, the fate of such APC following interaction with these same CTL remains poorly understood. We have shown previously that prolonged Ag presentation persists in the presence of a strong CTL response following HSV infection. In this study, we examined the mechanism of survival of APC in vivo when presenting an immunodominant determinant from HSV. We show that transferred peptide-labeled dendritic cells were eliminated from draining lymph nodes in the presence of HSV-specific CTL. Maturation of dendritic cells with LPS or anti-CD40 before injection protected against CTL lysis in vivo. Furthermore, endogenous APC could be eliminated from draining lymph nodes early after HSV infection by adoptive transfer of HSV-specific CTL, yet the cotransfer of significant virus-specific CD4+ T cell help promoted prolonged Ag presentation. This suggests that Th cells may assist in prolonging class I-restricted Ag presentation, potentially enhancing CTL recruitment and allowing more efficient T cell priming.

CTL response compensation for the loss of an immunodominant class I-restricted HSV-1 determinant

The T‐cell response to even complex pathogens is often focused on only a handful of immunodominant determinants. Such narrow responses provoke a selective pressure that can drive the emergence of CTL escape variants, raising the question of whether a broader response, targeting multiple non‐dominant peptides may be more beneficial. To examine the ability of the T‐cell repertoire to respond to non‐dominant determinants, we have investigated how mutating the dominant peptide in HSV affects the magnitude of the CD8+ T‐cell response. We found that the CTL response to HSV lacking the dominant peptide was only modestly reduced compared with the wild‐type virus and, surprisingly, this compensation occurred without any enhancement in the response to an established minor epitope. These findings are supportive of a malleable T‐cell repertoire that can elicit strong responses to alternate, unknown determinants in the absence of the dominant response.

Herpes Simplex Virus Type 1-Specific Cytotoxic T-Lymphocyte Arming Occurs within Lymph Nodes Draining the Site of Cutaneous Infection

ABSTRACT Various studies have shown that major histocompatibility complex class I-restricted cytotoxic T lymphocytes (CTL) can be isolated from lymph nodes draining sites of cutaneous infection with herpes simplex virus type 1 (HSV-1). Invariably, detection of this cytolytic activity appeared to require some level of in vitro culture of the isolated lymph node cells, usually for 3 days, in the absence of exogenous viral antigen. This in vitro “resting” period was thought to represent the phase during which committed CD8+ T cells become “armed” killers after leaving the lymph nodes and prior to their entry into infected tissue as effector CTL. In this study we reexamined the issue of CTL appearance in the HSV-1 immune response and found that cytolytic activity can be isolated directly from draining lymph nodes, although at levels considerably below those found after in vitro culture. By using T-cell receptor elements that represent effective markers for class I-restricted T cells specific for an immunodominant glycoprotein B (gB) determinant from HSV-1, we show that the increase in cytotoxicity apparent after in vitro culture closely mirrors the expansion of gB-specific CTL during the same period. Taken together, our results suggest that HSV-1-specific CTL priming does not appear to require any level of cytolytic machinery arming outside the lymph node compartment despite the absence of any detectable infection within that site.