Early primed KLRG1- CMV-specific T cells determine the size of the inflationary T cell pool.

Memory T cell inflation is a process in which a subset of cytomegalovirus (CMV) specific CD8 T cells continuously expands mainly during latent infection and establishes a large and stable population of effector memory cells in peripheral tissues. Here we set out to identify in vivo parameters that promote and limit CD8 T cell inflation in the context of MCMV infection. We found that the inflationary T cell pool comprised mainly high avidity CD8 T cells, outcompeting lower avidity CD8 T cells. Furthermore, the size of the inflationary T cell pool was not restricted by the availability of specific tissue niches, but it was directly related to the number of virus-specific CD8 T cells that were activated during priming. In particular, the amount of early-primed KLRG1- cells and the number of inflationary cells with a central memory phenotype were a critical determinant for the overall magnitude of the inflationary T cell pool. Inflationary memory CD8 T cells provided protection from a Vaccinia virus challenge and this protection directly correlated with the size of the inflationary memory T cell pool in peripheral tissues. These results highlight the remarkable protective potential of inflationary CD8 T cells that can be harnessed for CMV-based T cell vaccine approaches.

Tissue maintenance of CMV-specific inflationary memory T cells by IL-15.

Cytomegalovirus (CMV) infection induces an atypical CD8 T cell response, termed inflationary, that is characterised by accumulation and maintenance of high numbers of effector memory like cells in circulation and peripheral tissues-a feature being successfully harnessed for vaccine purposes. Although stability of this population depends on recurrent antigen encounter, the requirements for prolonged survival in peripheral tissues remain unknown. Here, we reveal that murine CMV-specific inflationary CD8 T cells are maintained in an antigen-independent manner and have a half-life of 12 weeks in the lung tissue. This half-life is drastically longer than the one of phenotypically comparable inflationary effector cells. IL-15 alone, and none of other common γ-cytokines, was crucial for survival of inflationary cells in peripheral organs. IL-15, mainly produced by non-hematopoietic cells in lung tissue and being trans-presented, promoted inflationary T cell survival by increasing expression of Bcl-2. These results indicate that inflationary CD8 T cells are not just simply effector-like cells, rather they share properties of both effector and memory CD8 T cells and they appear to be long-lived cells compared to the effector cells from acute virus infections.

The Salivary Gland Acts as a Sink for Tissue-Resident Memory CD8(+) T Cells, Facilitating Protection from Local Cytomegalovirus Infection.

Tissue-resident memory T cells (TRM) reside in barrier tissues and provide local immediate protective immunity. Here, we show that the salivary gland (SG) most-effectively induces CD8(+) and CD4(+) TRM cells against murine cytomegalovirus (MCMV), which persists in and spreads from this organ. TRM generation depended on local antigen for CD4(+), but not CD8(+), TRM cells, highlighting major differences in T cell subset-specific demands for TRM development. CMV-specific CD8(+) T cells fail to control virus replication upon primary infection in the SG due to CMV-induced MHC I downregulation in glandular epithelial cells. Using intraglandular infection, we challenge this notion and demonstrate that memory CD8(+) T cells confer immediate protection against locally introduced MCMV despite active viral immune evasion, owing to early viral tropism to cells that largely withstand MHC I downregulation. Thus, we unravel a yet-unappreciated role for memory CD8(+) T cells in protecting mucosal tissues against CMV infection.

Salivary gland resident APCs are Flt3L- and CCR2-independent macrophage-like cells incapable of cross-presentation.

Cytomegaloviruses (CMVs) disseminate within the human population via mucosal excretions, for example, from the salivary glands (SGs), which represent a privileged site of viral immune evasion and persistence. The murine CMV (MCMV) model has served to identify factors that maintain a unique virus-host relationship in this organ. In contrast to all other organs, the SG is resistant to CD8(+) T-cell mediated control of MCMV replication due to virally induced MHC class I downregulation, which is exceptionally efficient in acinar glandular epithelial cells. Uniquely to the SG, IFN-γ producing CD4(+) T cells are required for virus control. While T-cell responses have been extensively characterized in the SG, the ontogeny and function of APCs in this organ remain to be assessed. Here, we show that macrophage-like cells constitute the population of SG-resident APCs in steady state and during MCMV-induced inflammation in mice. Inflammatory monocytes, monocyte-derived DCs as well as conventional, Flt3L-dependent DCs do not contribute to this population. Despite supporting contact formation to CD4(+) and CD8(+) T cells in principle, SG-resident APCs fail to activate the latter due to their inability to cross-present MCMV-derived antigen.

Adoptive transfer of cytomegalovirus-specific effector CD4+ T cells provides antiviral protection from murine CMV infection.

Cytomegalovirus (CMV) infects a majority of the human population and establishes a life-long persistence. CMV infection is usually asymptomatic but the virus carries pathogenic potential and causes severe disease in immunocompromised individuals. T-cell-mediated immunity plays an essential role in control of CMV infection and adoptive transfer of CMV-specific CD8(+) T cells restores viral immunity in immunosuppressed patients but a role for CD4(+) T cells remains elusive. Here, we analyzed in adoptive transfer studies the features and antiviral functions of virus-specific CD4(+) T cells during primary murine CMV (MCMV) infection. MCMV-specific CD4(+) T cells expanded upon MCMV infection and displayed an effector phenotype and function. Adoptive transfer of in vivo activated MCMV-specific CD4(+) T cells to immune-compromised mice was protective during pathogenic MCMV infection and IFN-γ was a crucial mediator of this protective capacity. Moreover, co-transfer of low doses of both MCMV-specific CD4(+) T cells and CD8(+) T cells synergized in control of lytic viral replication in immune-compromised mice. Our data reveal a pivotal antiviral role for virus-specific CD4(+) T cells in protection from pathogenic CMV infection and provide evidence for their antiviral therapeutic potential.

T cell memory in the context of persistent herpes viral infections.

The generation of a functional memory T cell pool upon primary encounter with an infectious pathogen is, in combination with humoral immunity, an essential process to confer protective immunity against reencounters with the same pathogen. A prerequisite for the generation and maintenance of long-lived memory T cells is the clearance of antigen after infection, which is fulfilled upon resolution of acute viral infections. Memory T cells play also a fundamental role during persistent viral infections by contributing to relative control and immuosurveillance of active replication or viral reactivation, respectively. However, the dynamics, the phenotype, the mechanisms of maintenance and the functionality of memory T cells which develop upon acute/resolved infection as opposed to chronic/latent infection differ substantially. In this review we summarize current knowledge about memory CD8 T cell responses elicited during α-, ß-, and γ-herpes viral infections with major emphasis on the induction, maintenance and function of virus-specific memory CD8 T cells during viral latency and we discuss how the peculiar features of these memory CD8 T cell responses are related to the biology of these persistently infecting viruses.

Non-hematopoietic cells in lymph nodes drive memory CD8 T cell inflation during murine cytomegalovirus infection.

During human and murine cytomegalovirus (MCMV) infection an exceptionally large virus-specific CD8 T cell pool is maintained in the periphery lifelong. This anomalous response is only seen for specific subsets of MCMV-specific CD8 T cells which are referred to as 'inflationary T cells'. How memory CD8 T cell inflation is induced and maintained is unclear, though their activated phenotype strongly suggests an involvement of persistent antigen encounter during MCMV latency. To dissect the cellular and molecular requirements for memory CD8 T cell inflation, we have generated a transgenic mouse expressing an MHC class I-restricted T cell receptor specific for an immunodominant inflationary epitope of MCMV. Through a series of adoptive transfer experiments we found that memory inflation was completely dependent on antigen presentation by non-hematopoietic cells, which are also the predominant site of MCMV latency. In particular, non-hematopoietic cells selectively induced robust proliferation of inflationary CD8 T cells in lymph nodes, where a majority of the inflationary CD8 T cells exhibit a central-memory phenotype, but not in peripheral tissues, where terminally differentiated inflationary T cells accumulate. These results indicate that continuous restimulation of central memory CD8 T cells in the lymph nodes by infected non-hematopoietic cells ensures the maintenance of a functional effector CD8 T pool in the periphery, providing protection against viral reactivation events.

Absence of cross-presenting cells in the salivary gland and viral immune evasion confine cytomegalovirus immune control to effector CD4 T cells.

Horizontal transmission of cytomegaloviruses (CMV) occurs via prolonged excretion from mucosal surfaces. We used murine CMV (MCMV) infection to investigate the mechanisms of immune control in secretory organs. CD4 T cells were crucial to cease MCMV replication in the salivary gland (SG) via direct secretion of IFNγ that initiated antiviral signaling on non-hematopoietic cells. In contrast, CD4 T cell helper functions for CD8 T cells or B cells were dispensable. Despite SG-resident MCMV-specific CD8 T cells being able to produce IFNγ, the absence of MHC class I molecules on infected acinar glandular epithelial cells due to viral immune evasion, and the paucity of cross-presenting antigen presenting cells (APCs) prevented their local activation. Thus, local activation of MCMV-specific T cells is confined to the CD4 subset due to exclusive presentation of MCMV-derived antigens by MHC class II molecules on bystander APCs, resulting in IFNγ secretion interfering with viral replication in cells of non-hematopoietic origin.

Batf3 transcription factor-dependent DC subsets in murine CMV infection: differential impact on T-cell priming and memory inflation.

Priming of CD8(+) T cells specific for viruses that interfere with the MHC class I presentation pathway is a challenge for the immune system and is believed to rely on cross-presentation. Cytomegalovirus (CMV) infection induces vigorous CD8(+) T-cell responses despite its potent immune evasion strategies. Furthermore, CD8(+) T cells specific for a subset of viral epitopes accumulate and are maintained at high levels exhibiting an activated phenotype - referred to as "inflationary T cells". Taking advantage Batf3(-/-) mice in which the development of cross-presenting CD8α(+) and CD103(+) DCs is severely compromised, we analyzed their role in the induction and inflation of murine (M)CMV-specific CD8(+) T-cell responses. We found that priming of MCMV-specific CD8(+) T cells was severely impaired in the absence of cross-presenting DCs. However, inflation of two immuno-dominant MCMV-specific CD8(+) T-cell populations was largely normal in the absence of cross-presenting DCs, indicating that inflation during latency was mainly dependent on direct antigen presentation. These results highlight differential antigen presentation requirements during acute and latent MCMV infection.

T-cell help permits memory CD8(+) T-cell inflation during cytomegalovirus latency.

CD4(+) T cells are implied to sustain CD8(+) T-cell responses during persistent infections. As CD4(+) T cells are often themselves antiviral effectors, they might shape CD8(+) T-cell responses via help or via controlling antigen load. We used persistent murine CMV (MCMV) infection to dissect the impact of CD4(+) T cells on virus-specific CD8(+) T cells, distinguishing between increased viral load in the absence of CD4(+) T cells and CD4(+) T-cell-mediated helper mechanisms. Absence of T-helper cells was associated with sustained lytic MCMV replication and led to a slow and gradual reduction of the size and function of the MCMV-specific CD8(+) T-cell pool. However, when virus replication was controlled in the absence of CD4(+) T cells, CD8(+) T-cell function was comparably impaired, but in addition CD8(+) T-cell inflation, a hallmark of CMV infection, was completely abolished. Thus, CD8(+) T-cell inflation during latent CMV infection is strongly dependent on CD4(+) T-cell helper functions, which can partially be compensated by ongoing lytic viral replication in the absence of CD4(+) T cells.