Lymphoid Tissue Inducer Research Articles

Multiparameter flow cytometric analysis of RORγt expression in mouse models of T cell development and differentiation (163.25)

Abstract Retinoic acid-related Orphan Receptor gamma t (RORγt) is a DNA-binding transcription factor that is selectively expressed by lymphoid cells including CD4+CD8+ thymocytes, CD4+ Th17 and CD8+ Tc17 cells, NKT cells and lymphoid tissue inducer (LTi) cells. RORγt plays essential roles in thymopoiesis, T cell homeostasis, effector T cell differentiation and the development of secondary lymphoid tissues, eg, lymph nodes and Peyer’s patches. A new mouse mAb, Q31-378, that recognizes mouse RORγt was developed to provide a sensitive probe for analyzing the cellular and molecular mechanisms that underlie the development and differentiation of T cells and lymphoid tissues. With a newly developed transcription factor buffer, Q31-378 was successfully used in multicolor flow cytometric analyses to analyze RORγt expression in thymocyte subsets defined by CD4 and CD8 expression. Among all thymocytes, CD4+CD8+ cells comprised the main cell subset that expressed RORγt. Analyses of splenocytes cultured under Th17-inducing conditions revealed that most activated CD4+IL-17A+IFN-γ± T cells expressed RORγt when compared with other T cell subsets. These new reagents, along with additional fluorescent antibodies to other cytokines, transcription factors and phenotypic markers are being applied to analyze RORγt expression patterns in the development and differentiation of T cells and LTi cells.

RANK-ligand activates stroma in postnatal lymph node growth (69.4)

Abstract The lymph node (LN) is a well-organized immune organ, favoring immune cell interactions in a spacio-temporally-controlled manner. Studies of embryonic LN formation have identified TNFSF11 (RANKL), the lymphoid tissue inducer cells and the fibroblastic stromal cells as key players in this process. Yet the cellular and molecular mechanisms that underlie postnatal LN growth remain unclear. We have studied a mouse mutant overexpressing RANKL in the skin and found massive postnatal growth of skin-draining LNs. T cells and dendritic cells are not activated and respond well to immunization by foreign antigen and adjuvant. The overall organ architecture is preserved with the exception of an increase in the numbers of small B cell follicles. Reticular fibroblastic and vascular stromal cells, important for secondary lymphoid organ formation and organization, express RANK and undergo hyperproliferation, which is abrogated by RANKL neutralization. In addition, they express higher levels of CXCL13 and CCL19 chemokines as well as MAdCAM-1 and VCAM-1 cell adhesion molecules. As for hematopoietic cells, they appear to receive anti-apoptotic signals as T cells from hyperplasic LNs survive better in lymphopenic hosts than T cells from control mice. These findings identify RANKL as a key molecule for controlling homeostasis of the immune system through stroma.

Interleukin-22 drives endogenous thymic regeneration in mice (44.6)

Abstract Thymic regeneration is a crucial function that allows for renewal of immune competance after stress, infection or immunodepletion, however the mechanisms governing this rejuvenation remain poorly understood. Here we detail a framework of endogenous thymic regeneration centred on IL-22, a recently identified cytokine primarily implicated in maintenance of epithelial barrier function. Although IL-22 was redundant for steady-state thymopoiesis, thymic recovery was impaired in IL-22-deficient mice and intrathymic levels of IL-22 were significantly increased in WT mice following thymic damage. IL-22, which signalled through thymic epithelial cells and promoted their proliferation and survival, was upregulated by radio-resistant RORγ(t)+NKp46-CCR6+ lymphoid tissue-inducer cells after thymic injury under the control of dendritic cell-derived IL-23 and triggered by the depletion of CD4+CD8+ double positive (DP) thymocytes. Importantly, administration of IL-22 enhanced thymic recovery following irradiation damage. These studies reveal a network of endogenous thymus regeneration where 1) the depletion of DP thymocytes triggers 2) upregulation of IL-23 by DCs that induces 3) the production of IL-22 by thymic LTi. This cascade of events leads to regeneration of the epithelial microenvironment and, ultimately, to rejuvenation of thymopoiesis; presenting an innovative strategy for improving immune competence in patients whose thymus has been damaged from infection or cytoreductive therapy.

Type 2 innate lymphoid cells–new members of the “type 2 franchise” that mediate allergic airway inflammation

Type 2 innate lymphoid cells (ILC2s) are members of an ILC family, which contains NK cells and Rorγt(+) ILCs, the latter including lymphoid tissue inducer (LTi) cells and ILCs producing IL-17 and IL-22. ILC2s are dedicated to the production of IL-5 and IL-13 and, as such, ILC2s provide an early and important source of type 2 cytokines critical for helminth expulsion in the gut. Several studies have also demonstrated a role for ILC2s in airway inflammation. In this issue of the European Journal of Immunology, Klein Wolterink et al. [Eur. J. Immunol. 2012. 42: 1106-1116] show that ILC2s are instrumental in several models of experimental asthma where they significantly contribute to production of IL-5 and IL-13, key cytokines in airway inflammation. This study sheds light over the relative contribution of ILC2s versus T helper type 2 cells (Th2) in type 2 mediated allergen-specific inflammation in the airways as discussed in this commentary.

Interleukin-7: a key Mediator in T Cell-driven Autoimmunity, Inflammation, and Tissue Destruction

IL-7, expressed by stromal cells in primary lymphoid organs, is known for its critical role in the development and homeostatic expansion of T cells in humans and mice. IL-7 is equally important for B cell development in human and mice, but only in mice seems critical for B cell development and expansion. Recent studies demonstrate that this potent immunostimulatory cytokine is overexpressed in inflamed tissues of patients with (rheumatic) autoimmune diseases and that expression levels correlate with clinical parameters of disease. In inflamed tissues several cell types, including macrophages, dendritic cells, and fibroblasts produce IL-7. IL-7 primarily acts on T cells that abundantly express the IL-7 receptor and that are increased at the inflammatory sites, and predominantly induces Th1 and Th17-associated cytokine secretion. IL-7-mediated T cell-dependent activation of macrophages, dendritic cells and B cells is accompanied by up regulation of T cell differentiating factors, chemokines, adhesion/co-stimulatory molecules and catabolic cytokines and enzymes. Moreover, overexpression of IL-7 is associated with ectopic lymphoid aggregate formation, corresponding with the capacity of IL-7 to induce LTβ and TNFα and to activate innate lymphoid tissue inducer cells. Additionally, IL-7 promotes T cell-driven osteoclastogenesis and fibroblast activation, processes involved in tissue destruction in chronic inflammation. Altogether this suggests that IL-7 is an important proinflammatory mediator in several chronic (rheumatic) inflammatory autoimmune diseases. The substantial amelioration of inflammation and immunopathology in experimental animal models for these diseases by blocking IL-7(receptor) supports this role of IL-7 and demonstrates that IL-7 and its receptor represent novel targets for immunotherapy.

Notch, Id2, and RORγt sequentially orchestrate the fetal development of lymphoid tissue inducer cells

Lymphoid tissue development is initiated during embryogenesis by the migration of lymphoid tissue inducer (LTi) cells from the fetal liver to the periphery, where they induce the formation of lymph nodes and Peyer's patches. In the fetal liver, a subset of common lymphoid progenitors (CLPs) that expresses the integrin α4β7 gives rise to LTi cells, a process strictly dependent on the expression of the transcriptional repressor Id2 and the nuclear hormone receptor retinoic acid-related orphan receptor γ t (RORγt). In this study, we show that Id2 and RORγt are sequentially up-regulated during LTi cell development, matching two waves of differentiation with opposite requirements for Notch signaling. Both the expression of Id2 and Notch are required for the generation of α4β7(+) RORγt(-) fetal progenitors, but Notch subsequently blocks progression to the RORγt(+) stage and final maturation of LTi cells. Notch is therefore a necessary switch to engage the LTi developmental pathway, but needs to be turned off later to avoid diversion to the T cell fate.

Rank Signaling Links the Development of Invariant γδ T Cell Progenitors and Aire+ Medullary Epithelium

SummaryThe thymic medulla provides a specialized microenvironment for the negative selection of T cells, with the presence of autoimmune regulator (Aire)-expressing medullary thymic epithelial cells (mTECs) during the embryonic-neonatal period being both necessary and sufficient to establish long-lasting tolerance. Here we showed that emergence of the first cohorts of Aire+ mTECs at this key developmental stage, prior to αβ T cell repertoire selection, was jointly directed by Rankl+ lymphoid tissue inducer cells and invariant Vγ5+ dendritic epidermal T cell (DETC) progenitors that are the first thymocytes to express the products of gene rearrangement. In turn, generation of Aire+ mTECs then fostered Skint-1-dependent, but Aire-independent, DETC progenitor maturation and the emergence of an invariant DETC repertoire. Hence, our data attributed a functional importance to the temporal development of Vγ5+ γδ T cells during thymus medulla formation for αβ T cell tolerance induction and demonstrated a Rank-mediated reciprocal link between DETC and Aire+ mTEC maturation.

IL17A (interleukin 17A)

Interleukin-17A (IL17A), a characteristic cytokine produced by the T helper 17 cells (Th17 cells), can form either a homodimer or a heterodimer with IL17F.It is produced not only by Th17 cells, but also by cytotoxic CD8+ T cells (Tc17 cells), ?d T cells, invariant natural killer T cells (iNKT cells), lymphoid tissue inducer cells (LTi cells), and other hematopoietic and non-hematopoietic cells. During development, these cells exhibit flexible or plastic features distinct from those of Th1 and Th2 cells. IL17A plays important roles in the pathogenesis of autoimmune diseases and in the host defenses against bacterial and fungal infections.Expression of IL17A and its related factors, as well as the infiltration of IL17A-producing cells into the tumor microenvironment, has been implicated in anti-tumor or pro-tumor effects in various cancers.

Expression pattern changes and function of RANKL during mouse lymph node microarchitecture development

Receptor activator of nuclear factor kappa-B ligand (RANKL) expression was examined during the development of mouse fetal peripheral lymphoid organs. A shift in the expression pattern was detected during the transition from lymphoid tissue inducer (LTi) cells to lymphoid tissue organizer (LTo) cells in the lymph node (LN) anlagen but not in the Peyer's patch anlagen. In order to understand the functional impact of these changes in the fetal expression of RANKL, the RANKL function was blocked by a blocking antibody. Excess anti-RANKL antibody was administered to pregnant mice between 13.5 and 16.5 dpc and was found to completely block LN anlagen development, suggesting that RANKL function during this period is critical for LN development. In addition, small amounts of anti-RANKL antibodies were injected directly into the amniotic space at 13.5 dpc, resulting in perturbed B-cell follicle formation and high endothelial venule differentiation after birth. These results suggest that RANKL expression on LTi cells during the early phase of LN development is critical for the development LN microarchitecture.

Pilot gene expression analysis of inflamed synovium

Background and objectivesWithin the synovial membrane of an rheumatoid arthritis (RA) joint, tissue architecture varies depending on organisation of the infiltrating inflammatory cells (diffuse, aggregates and germinal centres). It has...

Cooperation of innate and acquired immune system derived signals induces stromal cell activation in chronic inflammation and ectopic lymphoneogenesis

BackgroundAcquisition of lymphoid-like features is a hallmark of ectopic lymphoneogenesis. In lymph nodes, gp8+ fibroblastic reticular cells (FRC) are responsible for lymphoid chemokines/cytokine production and lymphocyte compartmentalisation. gp38 up-regulation on...

Stroma cell priming in enteric lymphoid organ morphogenesis

The lymphoid system is equipped with a network of specialized platforms located at strategic sites, which grant strict immune-surveillance and efficient immune responses. The development of these peripheral secondary lymphoid organs (SLO) occurs mainly in utero, while tertiary lymphoid structures can form in adulthood generally in response to persistent infection and inflammation. Regardless of the lymphoid tissue and intrinsic cellular and molecular differences, it is now well established that the recruitment of fully functional lymphoid tissue inducer (LTi) cells to presumptive lymphoid organ sites, and their consequent close and reciprocal interaction with resident stroma cells, are central to SLO formation. In contrast, the nature of events that initially prime resident sessile stroma cells to recruit and retain LTi cells remains poorly understood. Recently, new findings revealed early phases of SLO development putting emphasis on mesenchymal and lymphoid tissue initiator cells. Herein we discuss the main tenets of enteric lymphoid organs genesis and focus in the most recent findings that open new perspectives to the understanding of the early phases of lymphoid morphogenesis.

The Aryl Hydrocarbon Receptor: A Sentinel Safeguarding the Survival of Immune Cells in the Gut

Innate lymphoid cells (ILCs) regulate the epithelial barrier function and immunity in the gut. How ILC numbers are maintained is unknown. In this issue of Immunity, Qiu et al. (2012) report that the transcription factor aryl hydrocarbon receptor controls survival and function of gut-residing ILCs.

Sterile inflammation – do innate lymphoid cell subsets play a role?

The recent identification of several novel innate lymphoid cell (iLC) subsets has increased our understanding of the mechanisms which link the innate and adaptive immune systems. While the contribution of these subsets toward the pathogenesis of human disease remains largely to be determined, it seems likely that they will play a particularly important role in sterile inflammatory settings where the innate response is seen as a critical mediator of inflammation. Several recent studies have highlighted the role of endogenous damage-associated molecular patterns such as IL-33, IL-1α, and IL-1β in promoting lymphoid cell responses. This review discusses the influence of such endogenous danger signals on novel iLCs such as lymphoid tissue-inducer cells, innate type 2 helper cells, and γδ T cells and explores how these responses may contribute to the development of an inflammatory response in a sterile setting.

Pairing experimentation and computational modeling to understand the role of tissue inducer cells in the development of lymphoid organs

The use of genetic tools, imaging technologies and ex vivo culture systems has provided significant insights into the role of tissue inducer cells and associated signaling pathways in the formation and function of lymphoid organs. Despite advances in experimental technologies, the molecular and cellular process orchestrating the formation of a complex three-dimensional tissue is difficult to dissect using current approaches. Therefore, a robust set of simulation tools have been developed to model the processes involved in lymphoid tissue development. Specifically, the role of different tissue inducer cell populations in the dynamic formation of Peyer’s patches has been examined. Utilizing approaches from systems engineering, an unbiased model of lymphoid tissue inducer cell function has been developed that permits the development of emerging behaviors that are statistically not different from that observed in vivo. These results provide the confidence to utilize statistical methods to explore how the simulator predicts cellular behavior and outcomes under different physiological conditions. Such methods, known as sensitivity analysis techniques, can provide insight into when a component part of the system (such as a particular cell type, adhesion molecule, or chemokine) begins to have an influence on observed behavior, and quantifies the effect a component part has on the end result: the formation of lymphoid tissue. Through use of such a principled approach in the design, calibration, and analysis of a computer simulation, a robust in silico tool can be developed which can both further the understanding of a biological system being explored, and act as a tool for the generation of hypotheses which can be tested utilizing experimental approaches.

Lymphoid tissue inducer cells: pivotal cells in the evolution of CD4 immunity and tolerance?

Phylogeny suggests that the evolution of placentation in mammals was accompanied by substantial changes in the mammalian immune system: in particular lymph nodes and CD4 high affinity memory antibody responses co-evolved during the same period. Lymphoid tissue inducer cells (LTi) are members of an emerging family of innate lymphoid cells (ILCs) that are crucial for lymph node development, but our studies have indicated that they also play a pivotal role in the long-term maintenance of memory CD4 T cells in adult mammals through their expression of the tumor necrosis family members, OX40- and CD30-ligands. Additionally, our studies have shown that these two molecules are also key operators in CD4 effector function, as their absence obviates the need for the FoxP3 dependent regulatory T (Tregs) cells that prevent CD4 driven autoimmune responses. In this perspective article, we summarize findings from our group over the last 10 years, and focus specifically on the role of LTi in thymus. We suggest that like memory CD4 T cells, LTi also play a role in the selection and maintenance of the Tregs that under normal circumstances are absolutely required to regulate CD4 effector cells.

Mesenchymal cell differentiation during lymph node organogenesis

Secondary lymphoid tissues such as lymph nodes are essential for the interactions between antigen presenting cells and lymphocytes that result in adaptive immune responses that protect the host against invading pathogens. The specialized architecture of these organs facilitates the cognate interactions between antigen-loaded dendritic cells and lymphocytes expressing their specific receptor as well as B–T cell interactions that are at the core of long lasting adaptive immune responses. Lymph nodes develop during embryogenesis as a result of a series of cross-talk interactions between a hematopoietically derived cell lineage called lymphoid tissue inducer cells and stromal cells of mesenchymal origin to form the anlagen of these organs. This review will present an overview of the different signaling pathways and maturation steps that mesenchymal cells undergo during the process of lymph node formation such as cell specification, priming, and maturation to become lymphoid tissue stromal organizer cells.

Lymphoid tissue inducer cells: innate cells critical for CD4+ T cell memory responses?

Lymphoid tissue inducer cells (LTi) are a relatively new arrival on the immunological cellular landscape, having first been characterized properly only 15 years ago. They are members of an emerging family of innate lymphoid cells (ILCs). Elucidation of their function reveals links not only with the ancient innate immune system, but also with adaptive immune responses, in particular the development of lymph nodes and CD4(+) T cell memory immune responses, which on one hand underpin the success of vaccination strategies, and on the other hand drive many human immunologically mediated diseases. This perspective article is not an exhaustive account of the role of LTi in the development of lymphoid tissues, as there have been many excellent reviews published already. Instead, we combine current knowledge of genetic phylogeny and comparative immunology, together with classical mouse genetics, to suggest how LTi might have evolved from a primitive lymphocytic innate cell in the ancestral 500-million-year-old vertebrate immune system into a cell critical for adaptive CD4(+) T cell immune responses in mammals.

Functional Differences between Human NKp44− and NKp44+ RORC+ Innate Lymphoid Cells

Human RORC+ lymphoid tissue inducer cells are part of a rapidly expanding family of innate lymphoid cells (ILC) that participate in innate and adaptive immune responses as well as in lymphoid tissue (re) modeling. The assessment of a potential role for innate lymphocyte-derived cytokines in human homeostasis and disease is hampered by a poor characterization of RORC+ innate cell subsets and a lack of knowledge on the distribution of these cells in adults. Here we show that functionally distinct subsets of human RORC+ innate lymphoid cells are enriched for secretion of IL-17a or IL-22. Both subsets have an activated phenotype and can be distinguished based on the presence or absence of the natural cytotoxicity receptor NKp44. NKp44+ IL-22 producing cells are present in tonsils while NKp44− IL-17a producing cells are present in fetal developing lymph nodes. Development of human intestinal NKp44+ ILC is a programmed event that is independent of bacterial colonization and these cells colonize the fetal intestine during the first trimester. In the adult intestine, NKp44+ ILC are the main ILC subset producing IL-22. NKp44− ILC remain present throughout adulthood in peripheral non-inflamed lymph nodes as resting, non-cytokine producing cells. However, upon stimulation lymph node ILC can swiftly initiate cytokine transcription suggesting that secondary human lymphoid organs may function as a reservoir for innate lymphoid cells capable of participating in inflammatory responses.

RANKL-RANK interaction in immune regulatory systems

The interaction between the receptor activator of NF-κB ligand (RANKL) and its receptor RANK plays a critical role in the development and function of diverse tissues. This review summarizes the studies regarding the functions of RANKL signaling in immune regulatory systems. Previous in vitro and in vivo studies have indicated that the RANKL signal promotes the survival of dendritic cells (DCs), thereby activating the immune response. In addition, RANKL signaling to DCs in the body surface barriers controls self-tolerance and oral-tolerance through regulatory T cell functions. In addition to regulating DC functions, the RANKL and RANK interaction is critical for the development and organization of several lymphoid organs. The RANKL signal initiates the formation of clusters of lymphoid tissue inducer cells, which is crucial for lymph node organogenesis. Moreover, the RANKL-RANK interaction controls the differentiation of M cells, specialized epithelial cells in mucosal tissues, that take up and transcytose antigen particles to control the immune response to pathogens or commensal bacterium. The development of epithelial cells localized in the thymic medulla (mTECs) is also regulated by the RANKL-RANK signal. Given that the unique property of mTECs to express a wide variety of tissue-specific self-antigens is critical for the elimination of self-antigen reactive T cells in the thymus, the RANKL-RANK interaction contributes to the suppression of autoimmunity. Future studies on the roles of the RANKL-RANK system in immune regulatory functions would be informative for the development and application of inhibitors of RANKL signaling for disease treatment.