Transplantation of embryonic spleen tissue reveals a role for adult non-lymphoid cells in initiating lymphoid tissue organization

In this report we describe a transplantation system where embryonic spleens are grafted into adult hosts. This model can be used to analyze the cellular and molecular requirements for the development and organization of splenic microenvironments.Whole embryonic day 15 (ED15) spleens, grafted under the kidney capsule of adult mice, were colonized by host-derived lymphocytes and DC and developed normal splenic architecture. Grafts were also able to form germinal centers in response to T-dependent antigen. Using this system we demonstrated that adult host-derived lymphotoxin (LT) a was sufficient for the development of ED15 LTa/ grafts. Grafting of ED15 LTa/ spleens into RAG/ hosts followed by transfer of LT a/ splenocytes revealed no requirement for lymphocyte-derived LT a in the induction of CCL21 or the development of T-zone stroma. These data suggest that interactions between adult lymphoid-tissue inducer-like cells and embryonic stromal cells initiated T-zone development. Furthermore,adult lymphoid tissue inducer-like cells were shown to develop from bone marrow-derived progenitors. The model described here demonstrates a method of transferring whole splenic microenvironments and dissecting the stromal and hematopoietic signals involved in spleen development and organization

Functionally distinct resident macrophage subsets differentially shape responses to infection in the bladder

International audienceResident macrophages are abundant in the bladder, playing key roles in immunity to uropathogens. Yet, whether they are heterogeneous, where they come from, and how they respond to infection remain largely unknown. We identified two macrophage subsets in mouse bladders, MacM in muscle and MacL in the lamina propria, each with distinct protein expression and transcriptomes. Using a urinary tract infection model, we validated our transcriptomic analyses, finding that MacM macrophages phagocytosed more bacteria and polarized to an anti-inflammatory profile, whereas MacL macrophages died rapidly during infection. During resolution, monocyte-derived cells contributed to tissue-resident macrophage pools and both subsets acquired transcriptional profiles distinct from naïve macrophages. Macrophage depletion resulted in the induction of a type 1-biased immune response to a second urinary tract infection, improving bacterial clearance. Our study uncovers the biology of resident macrophages and their responses to an exceedingly common infection in a largely overlooked organ, the bladder

Visualizing early splenic memory CD8+ T cells reactivation against intracellular bacteria in the mouse

International audienceMemory CD8(+) T cells represent an important effector arm of the immune response in maintaining long-lived protective immunity against viruses and some intracellular bacteria such as Listeria monocytogenes (L.m). Memory CD8(+) T cells are endowed with enhanced antimicrobial effector functions that perfectly tail them to rapidly eradicate invading pathogens. It is largely accepted that these functions are sufficient to explain how memory CD8(+) T cells can mediate rapid protection. However, it is important to point out that such improved functional features would be useless if memory cells were unable to rapidly find the pathogen loaded/infected cells within the infected organ. Growing evidences suggest that the anatomy of secondary lymphoid organs (SLOs) fosters the cellular interactions required to initiate naive adaptive immune responses. However, very little is known on how the SLOs structures regulate memory immune responses. Using Listeria monocytogenes (L.m) as a murine infection model and imaging techniques, we have investigated if and how the architecture of the spleen plays a role in the reactivation of memory CD8(+) T cells and the subsequent control of L.m growth. We observed that in the mouse, memory CD8(+) T cells start to control L.m burden 6 hours after the challenge infection. At this very early time point, L.m-specific and non-specific memory CD8(+) T cells localize in the splenic red pulp and form clusters around L.m infected cells while naïve CD8(+) T cells remain in the white pulp. Within these clusters that only last few hours, memory CD8(+) T produce inflammatory cytokines such as IFN-gamma and CCL3 nearby infected myeloid cells known to be crucial for L.m killing. Altogether, we describe how memory CD8(+) T cells trafficking properties and the splenic micro-anatomy conjugate to create a spatio-temporal window during which memory CD8(+) T cells provide a local response by secreting effector molecules around infected cells

Characterizing natural hydrogel for reconstruction of three‐dimensional lymphoid stromal network to model T‐cell interactions

Hydrogels have been used in regenerative medicine because they provide a three‐dimensional environment similar to soft tissues, allow diffusion of nutrients, present critical biological signals, and degrade via endogenous enzymatic mechanisms. Herein, we developed in vitro system mimicking cell–cell and cell–matrix interactions in secondary lymphoid organs (SLOs). Existing in vitro culture systems cannot accurately represent the complex interactions happening between T‐cells and stromal cells in immune response. To model T‐cell interaction in SLOs in vitro, we encapsulated stromal cells in fibrin, collagen, or fibrin–collagen hydrogels and studied how different mechanical and biological properties affect stromal network formation. Overall, fibrin supplemented with aprotinin was superior to collagen and fibrin–collagen in terms of network formation and promotion of T‐cell penetration. After 8 days of culture, stromal networks formed through branching and joining with other adjacent cell populations. T‐cells added to the newly formed stromal networks migrated and attached to stromal cells, similar to the T‐cell zones of the lymph nodes in vivo. Our results suggest that the constructed three‐dimensional lymphoid stromal network can mimic the in vivo environment and allow the modeling of T‐cell interaction in SLOs. © 2015 Wiley Periodicals, Inc. J Biomed Mater Res Part A: 103A: 2701–2710, 2015.Peer Reviewedhttp://deepblue.lib.umich.edu/bitstream/2027.42/112007/1/jbma35409.pd

Association of T-Zone Reticular Networks and Conduits with Ectopic Lymphoid Tissues in Mice and Humans

Ectopic or tertiary lymphoid tissues (TLTs) are often induced at sites of chronic inflammation. They typically contain various hematopoietic cell types, high endothelial venules, and follicular dendritic cells; and are organized in lymph node–like structures. Although fibroblastic stromal cells may play a role in TLT induction and persistence, they have remained poorly defined. Herein, we report that TLTs arising during inflammation in mice and humans in a variety of tissues (eg, pancreas, kidney, liver, and salivary gland) contain stromal cell networks consisting of podoplanin+ T-zone fibroblastic reticular cells (TRCs), distinct from follicular dendritic cells. Similar to lymph nodes, TRCs were present throughout T-cell–rich areas and had dendritic cells associated with them. They expressed lymphotoxin (LT) β receptor (LTβR), produced CCL21, and formed a functional conduit system. In rat insulin promoter–CXCL13–transgenic pancreas, the maintenance of TRC networks and conduits was partially dependent on LTβR and on lymphoid tissue inducer cells expressing LTβR ligands. In conclusion, TRCs and conduits are hallmarks of secondary lymphoid organs and of well-developed TLTs, in both mice and humans, and are likely to act as important scaffold and organizer cells of the T-cell–rich zone

Dynamic Imaging of CD8+ T Cells and Dendritic Cells during Infection with Toxoplasma gondii

To better understand the initiation of CD8+ T cell responses during infection, the primary response to the intracellular parasite Toxoplasma gondii was characterized using 2-photon microscopy combined with an experimental system that allowed visualization of dendritic cells (DCs) and parasite specific CD8+ T cells. Infection with T. gondii induced localization of both these populations to the sub-capsular/interfollicular region of the draining lymph node and DCs were required for the expansion of the T cells. Consistent with current models, in the presence of cognate antigen, the average velocity of CD8+ T cells decreased. Unexpectedly, infection also resulted in modulation of the behavior of non-parasite specific T cells. This TCR-independent process correlated with the re-modeling of the lymph node micro-architecture and changes in expression of CCL21 and CCL3. Infection also resulted in sustained interactions between the DCs and CD8+ T cells that were visualized only in the presence of cognate antigen and were limited to an early phase in the response. Infected DCs were rare within the lymph node during this time frame; however, DCs presenting the cognate antigen were detected. Together, these data provide novel insights into the earliest interaction between DCs and CD8+ T cells and suggest that cross presentation by bystander DCs rather than infected DCs is an important route of antigen presentation during toxoplasmosis

Frequency and phenotype of natural killer cells and natural killer cell subsets in bovine lymphoid compartments and blood

Natural killer (NK) cells are widely distributed in lymphoid and non‐lymphoid tissues, but little is known about the recirculation of NK cells between blood and tissues. This is relevant to understanding recirculation in the steady‐state and also for determining the roles for NK cells in vaccine‐induced immunity and responses to infection. Therefore, the percentage of NK cells and their phenotype across peripheral blood, afferent lymph and lymph nodes in steady‐state conditions was investigated in cattle using the pseudo‐afferent lymphatic cannulation model. CD2(+) CD25(lo) NK cells were the predominant subset of NK cells within the blood. In contrast, CD2(−) CD25(hi) NK cells were the main subset present within the skin‐draining afferent lymphatic vessels and lymph nodes, indicating that CD2(−) NK cells are the principal NK cell subset trafficking to lymph nodes via the afferent lymphatic vessel. Furthermore, a low percentage of NK cells were present in efferent lymph, which were predominantly of the CD2(−) subset, indicating that NK cells can egress from lymph nodes and return to circulation in steady‐state conditions. These compartmentalization data indicate that NK cells represent a population of recirculating lymphocytes in steady‐state conditions and therefore may be important during immune responses to vaccination or infection

Tunable Chemokine Production by Antigen Presenting Dendritic Cells in Response to Changes in Regulatory T Cell Frequency in Mouse Reactive Lymph Nodes

BACKGROUND: Although evidence exists that regulatory T cells (Tregs) can suppress the effector phase of immune responses, it is clear that their major role is in suppressing T cell priming in secondary lymphoid organs. Recent experiments using two photon laser microscopy indicate that dendritic cells (DCs) are central to Treg cell function and that the in vivo mechanisms of T cell regulation are more complex than those described in vitro. PRINCIPAL FINDINGS: Here we have sought to determine whether and how modulation of Treg numbers modifies the lymph node (LN) microenvironment. We found that pro-inflammatory chemokines -- CCL2 (MCP-1) and CCL3 (MIP-la) -- are secreted in the LN early (24 h) after T cell activation, that this secretion is dependent on antigen-specific DC-T cell interactions, and that it was inversely related to the frequency of Tregs specific for the same antigen. Furthermore, we demonstrate that Tregs modify the chemoattractant properties of antigen-presenting DCs, which, as the frequency of Tregs increases, fail to produce CCL2 and CCL3 and to attract antigen-specific T cells. CONCLUSIONS: These results substantiate a major role of Tregs in LN patterning during antigen-specific immune responses