Imaging of the cross-presenting dendritic cell subsets in the skin-draining lymph node

Abstract

Dendritic cells (DCs) are antigen-presenting cells specialized for activating T cells to elicit effector T-cell functions. Cross-presenting DCs are a DC subset capable of presenting antigens to CD8+ T cells and play critical roles in cytotoxic T-cell-mediated immune responses to microorganisms and cancer. Although their importance is known, the spatiotemporal dynamics of cross-presenting DCs in vivo are incompletely understood. Here, we study the T-cell zone in skin-draining lymph nodes (SDLNs) and find it is compartmentalized into regions for CD8 + T-cell activation by cross-presenting DCs that express the chemokine (C motif) receptor 1 gene, Xcr1 and for CD4 + T-cell activation by CD11b + DCs. Xcr1-expressing DCs in the SDLNs are composed of two different populations: migratory (CD103 hi ) DCs, which immigrate from the skin, and resident (CD8α hi ) DCs, which develop in the nodes. To characterize the dynamic interactions of these distinct DC populations with CD8 + T cells during their activation in vivo, we developed a photoconvertible reporter mouse strain, which permits us to distinctively visualize the migratory and resident subsets of Xcr1-expressing DCs. After leaving the skin, migratory DCs infiltrated to the deep T-cell zone of the SDLNs over 3 d, which corresponded to their half-life in the SDLNs. Intravital two-photon imaging showed that after soluble antigen immunization, the newly arriving migratory DCs more efficiently form sustained conjugates with antigen-specific CD8 + T cells than other Xcr1-expressing DCs in the SDLNs. These results offer in vivo evidence for differential contributions of migratory and resident cross-presenting DCs to CD8 + T-cell activation.dendritic cell | CD8 + T cell | cross-presentation | intravital two-photon imaging | photoconversion

CD8+ T Cells Orchestrate pDC-XCR1+ Dendritic Cell Spatial and Functional Cooperativity to Optimize Priming

“…In the steady state, XCR1 + DC were dispersed throughout the paracortex and the interfollicular area of the LN, with some XCR1 + DC located within the subcapsular sinus (Kitano et al, 2016) (Figure 7A). Importantly, after infection we found that XCR1 + DC accumulated around OT-I T cell clusters in a pattern similar to that of recruited pDC, creating a new microenvironment around the initial site of CD8 + T cell engagement with antigen-presenting cDC (Figure 7B, 7C, 7F, 7G and Movie S7).…”

In vivo multiphoton imaging of immune cell dynamics

“…Generally, helper T cells are developed from naïve CD4 + T cells, and cytotoxic T cells from naïve CD8 + T cells after these naïve T cells are activated by dendritic cells. Several recent imaging studies showed that in the early phase of immune responses, naïve CD4 + T cells and naïve CD8 + T cells are activated by interactions with different subsets of dendritic cells in different locations [12, 18, 29, 36]. Interestingly, by comparing the results of these studies, it is suggested that subsets of dendritic cells involved in the early activation of CD4 + T cells and CD8 + T cells vary depending on the form of antigens and types of pathogens.…”

“…However, mice that express the photochromic proteins in specific subsets of immune cells have been also generated [36]. The abovementioned XCR1 + dendritic cells in the lymph node are actually a mixture of a population that develops in the lymph node and a population that develops in other organs including the skin and then migrates to the lymph node.…”

“…The abovementioned XCR1 + dendritic cells in the lymph node are actually a mixture of a population that develops in the lymph node and a population that develops in other organs including the skin and then migrates to the lymph node. By irradiating the skin of a knock-in mouse strain harboring the KikGR gene inserted in the Xcr1 locus, these two populations of XCR1 + dendritic cells in the lymph node can be distinguished by in vivo imaging to analyze their interactions with T cells [36]. …”

“…2Multiphoton photoconversion of KikGR-expressing cells in the ex vivo lymph node. An inguinal lymph node was excised from an Xcr1 KikGR/+ mouse [36], and imaged and irradiated by multiphoton microscopy. a Single xy -slice images of the lymph node.…”