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Dendritic cell (DC) activation by viral RNA sensors such as TLR3 and MDA-5 is critical for initiating antiviral immunity. Optimal DC activation is promoted by type I interferon (IFN) signaling which is believed to occur in either autocrine or paracrine fashion. Here, we show that neither autocrine nor paracrine type I IFN signaling can fully account for DC activation by poly(I:C) in vitro and in vivo. By controlling the density of type I IFN-producing cells in vivo, we establish that instead a quorum of type I IFN-producing cells is required for optimal DC activation and that this process proceeds at the level of an entire lymph node. This collective behavior, governed by type I IFN diffusion, is favored by the requirement for prolonged cytokine exposure to achieve DC activation. Furthermore, collective DC activation was found essential for the development of innate and adaptive immunity in lymph nodes. Our results establish how collective rather than cell-autonomous processes can govern the initiation of immune responses., (© 2021 The Authors.)

The inflammasome is activated in response to a variety of pathogens and has an important role in shaping adaptive immunity, yet the spatiotemporal orchestration of inflammasome activation in vivo and the mechanisms by which it promotes an effective immune response are not fully understood. Using an in vivo reporter to visualize inflammasome assembly, we establish the distribution, kinetics and propagation of the inflammasome response to a local viral infection. We show that modified vaccinia Ankara virus induces inflammasome activation in subcapsular sinus (SCS) macrophages, which is immediately followed by cell death and release of extracellular ASC specks. This transient inflammasome signaling in the lymph node generates a robust influx of inflammatory cells and mobilizes T cells from the circulation to increase the magnitude of T cell responses. We propose that after infection, SCS macrophages deliver a burst response of inflammasome activity and cell death that translates into the broadening of T cell responses, identifying an important aspect of inflammasome-driven vaccination strategies.

CD8(+) T cell responses generate effector cells endowed with distinct functional potentials but the contribution of early events in this process is unclear. Here, we have imaged T cells expressing a fluorescent reporter for the activation of the interferon-γ (IFN-γ) locus during priming in lymph nodes. We have demonstrated marked differences in the efficiency of gene activation during stable T cell-dentritic cell (DC) contacts, influenced in part by signal strength. Imaging the first cell division, we have demonstrated that heterogeneity in T cell functional potential was largely apparent as T cells initiated clonal expansion. Moreover, by analyzing the fate of single activated T cells ex vivo, we have provided evidence that these early differences resulted in clonal progenies with distinct functional properties. Thus, the early set of T cell-DC interactions in lymph nodes largely contribute to the heterogeneity of T cell responses through the generation of functionally divergent clonal progenies., (Copyright © 2010 Elsevier Inc. All rights reserved.)

In vitro studies have revealed that T cell activation occurs during the formation of either dynamic or stable interactions with antigen-presenting cells (APC), and the respective cell junctions have been referred to as immunological kinapses and synapses. However, the relevance and molecular dynamics of kinapses and synapses remain to be established in vivo. Using two-photon imaging, we tracked the distribution of LAT-EGFP molecules during antigen recognition by activated CD4(+) T cells in lymph nodes. At steady state, LAT-EGFP molecules were preferentially found at the uropod of rapidly migrating T cells. In contrast to naïve T cells that fully stopped upon systemic antigen delivery, recently activated T cells decelerated and formed kinapses, characterized by continuous extension of membrane protrusions and by the absence of persistent LAT-EGFP clustering. On the other hand, activated CD4(+) T cells formed stable immunological synapses with antigen-loaded B cells and displayed sustained accumulation of LAT-EGFP fluorescence at the contact zone. Our results show that the state of T cell activation and the type of APC largely influence T cell-APC contact dynamics in lymph nodes. Furthermore, we provide a dynamic look at immunological kinapses and synapses in lymph nodes and suggest the existence of distinct patterns of LAT redistribution during antigen recognition.

Summary: CD8+ T cell responses generate effector cells endowed with distinct functional potentials but the contribution of early events in this process is unclear. Here, we have imaged T cells expressing a fluorescent reporter for the activation of the interferon-γ (IFN-γ) locus during priming in lymph nodes. We have demonstrated marked differences in the efficiency of gene activation during stable T cell-dentritic cell (DC) contacts, influenced in part by signal strength. Imaging the first cell division, we have demonstrated that heterogeneity in T cell functional potential was largely apparent as T cells initiated clonal expansion. Moreover, by analyzing the fate of single activated T cells ex vivo, we have provided evidence that these early differences resulted in clonal progenies with distinct functional properties. Thus, the early set of T cell-DC interactions in lymph nodes largely contribute to the heterogeneity of T cell responses through the generation of functionally divergent clonal progenies. [ABSTRACT FROM AUTHOR]

The exact role of CD4+CD25high regulatory T cells (Treg) in adoptive immunotherapy of melanoma is still to be determined, and an association between an expansion of Treg cells and the expansion of therapeutic tumor-infiltrating lymphocytes (TIL) remains unelucidated. In this context, the aim of our study was to determine whether functional Treg cells were detectable and amplified among in vitro-expanded TIL from 10 metastatic melanoma lymph nodes (LNs). In this study, we investigated the expression of forkhead/winged helix transcription factor 3 (Foxp3) in melanoma-invaded LNs and determined proportion and functionality of Treg cells among TIL extracted from these 10 metastatic melanoma LNs at different steps of their in vitro expansion. We found that metastatic melanoma LNs expressed very heterogeneous levels of Foxp3 and that CD4+CD25high Treg cells extracted from these LNs were detectable at each step of the in vitro culture of TIL but decreasing during the culture. In addition, functional assays demonstrated that these CD4+CD25high T cells were capable of suppressing autologous CD8+ and CD4+CD25- T cell proliferation. These cells were indeed Treg cells as they expressed Foxp3. In conclusion, our work suggests that CD4+CD25high Foxp3 expressing T cells are not expanded during in vitro amplification of TIL obtained from melanoma-invaded LNs. [ABSTRACT FROM AUTHOR]