Lymph Node Stroma Dynamics and Approaches for Their Visualization

doi:10.1016/j.it.2017.01.005

Trends in Immunology

Volume 38, Issue 4, April 2017, Pages 236-247

https://doi.org/10.1016/j.it.2017.01.005 Get rights and content

Trends

Lymphoid stromal cells are more than architectural cells; they are pivotal regulators of adaptive immunity.

While we begin to understand the functions of lymphoid stromal cells at the population level, we critically lack information on the heterogeneity of individual stromal cells.

Stromal cells are difficult to study; animal models are rare and flow cytometry approaches are limited.

Neuroscientists face similar technical limitations and have developed tools to decipher and manipulate the neuronal network; let them be an inspiration to us.

Lymphoid stromal cells are best known as the architectural cells of lymphoid organs. For decades, they have been considered as inert elements of the immune system but this view has changed dramatically in recent years, when it was discovered that they are endowed with critical immunoregulatory functions. It is now accepted that without them, the adaptive immune response would be compromised, if not abrogated entirely. Here, we review the function of the major lymphoid stromal cell types; the way they remodel upon inflammation; discuss the available tools to track their behavior; and introduce several methodological approaches that we believe will help improving our knowledge of these pivotal cell types.

Section snippets

Lymphoid Stromal Cells: Versatile and Reconfigurable 3D Immunological Networks

Secondary lymphoid organs such as lymph nodes (LNs) are organs in which adaptive immune responses develop. They are composed of 95% motile leukocytes and 5% sessile stromal cells [1]. In 1984, Nossal wrote: ‘A readership consisting of primarily anatomists has every right to question the favorite sport of research workers in cell immunology. This is to take a lymphoid tissue and totally destroy its beautiful and elaborately designed architecture to obtain simple cell suspension of lymphocytes,

Fibroblastic Reticular Cells (FRCs)

FRCs are contractile myofibroblasts that form the major mesenchymal stromal cell network of the T cell zone. Through secretion of chemokines such as CCL19 and CCL21, they delineate the boundaries of the T cell zone and physically support lymphocyte migration into that zone 4, 5. At steady state, FRCs secrete interleukin (IL)-7 and B cell activating factor (BAFF), the main survival factors for T and B cells, respectively 6, 7. Consistent with this prosurvival function, FRC network ablation using

Studying LN Stromal Cells: A Technical Challenge

Stromal cells regulate the immune system on several levels. Yet, our understanding of their biology is limited. This limitation of knowledge largely results from technical challenges inherent to the isolation and the culture of LN stromal cells, the inability to model the complexity of LN organization in vitro and the paucity of animal models dedicated to their study. Stromal cells are attached to each other, forming interconnected 3D networks. The 3D nature of all stromal cell networks is key

Stromal Cell Heterogeneity: Understanding the LN Stroma at the Single Cell Level

As summarized above, FDCs, FRCs, MRCs, LECs, and BECs are considered the major LN stromal cell populations. Nonetheless, additional subsets have been described, including CXCL12-producing follicular stromal cells, integrin α7-expressing pericytes and versatile stromal cells (VSCs) 29, 45, 46 (Tables 1 and 2). It is currently unknown if all these lymphoid stromal cell subsets represent true distinct populations or different anatomical aspects of a single mesenchymal stromal cell. Despite having

Inspiration from Neurosciences: Multicolor Lineage Tracing Models

Unlike most immunologists who work on cells that can be adoptively transferred or replaced by bone marrow grafting, neurobiologists face the same basic challenges as immunologists studying lymphoid stromal cells: neurons assemble in various, complex and intermingled 3D networks. It is impossible to graft, remove or replace these networks in vivo. Neuroscientists had thus no choice but to generate new models to study neurons in their natural, complex 3D environment in vivo. The recent

Multicolor Lineage Tracing Models to Study Stromal Cells

Far from being mere ‘fancy’ tools, Brainbow models have allowed seminal discoveries in the fields of developmental, cancer, and stem cell biology in several species (zebrafish, Drosophila and mice) 52, 53, 54, 55. How about immunology? Brainbow models are retrospective tools that allow the reconstruction of the proliferative and migratory history of an individual cell and its progeny by measuring the color, location, and size of clones at a given time. By definition, this system only applies to

Better Mouse Models

Stromal cell immunobiology is a young field that heavily relies on animal models. Paradoxically, however, suitable models are critically lacking. As an example, specific genetic deletion or fate mapping of lymphoid stromal cell subsets is currently impossible in the mouse (Table 2). How can we improve that? Precisely defining the roles of specific cell types is a challenging task within an entire organism. To this aim, biologists have engineered an important repository of genetically encoded

Concluding Remarks

Our knowledge of the origin and function of LN stromal cells has increased in recent years. However, we still lack a comprehensive view of the stromal cell dynamics that accompany and sustain LN growth during an immune response (see Outstanding Questions). A central and still unresolved question concerns the existence of an adult LN mesenchymal stromal cell progenitor. Similar to embryonic LTo during development, such a putative progenitor would extensively divide in the course of an immune

Acknowledgments

This work was supported by a grant from the European Research Council (ERC) under the European Union Horizon 2020 Research and Innovation Program grant agreement N° 647257STROMA.

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Lymphoid Tissue inducer (LTi) cell ontogeny and functioning in embryo and adult
2021, Biomedical Journal
Citation Excerpt :
This was later amended by the marginal reticular cells (MRC), positioned in the paracortex, adjacent to the B-cell follicles and just below the subcapsular sinus [75]. However, the use of single cell sequencing has allowed the identification of nine stromal cell clusters within the lymph node [76], each with specific expression profiles and functions (reviewed in Ref. [74]). These stromal cells differentiate from mesenchymal precursor cells, or LTo cells, in the first weeks after birth in the mouse [77–79].
Innate Lymphoid Cells (ILC) are involved in homeostasis and immunity. Their dynamic differentiation and characterization depend on their tissue of residency and is adapted to their role within these tissues. Lymphoid Tissue inducer (LTi) cells are an ILC member and essential for embryonic lymph node (LN) formation. LNs are formed at pre-defined and strategic positions throughout the body and how LTi cells are initially attracted towards these areas is under debate. Besides their role in LN formation, LTi-like and the closely related ILC type 3 (ILC3) cells have been observed within the embryonic gut. New studies have now shown more information on their origin and differentiation within the embryo.
This review will evaluate the embryonic LTi cell origin from a specific embryonic hemogenic wave, which has recently been described in mouse. Moreover, I will discuss their differentiation and similarities with the closely related ILC3 cells in embryo and adult.
High Endothelial Venules Accelerate Naive T Cell Recruitment by Tumor Necrosis Factor-Mediated R-Ras Upregulation
2021, American Journal of Pathology
Recruitment of naive T cells to lymph nodes is essential for the development of adaptive immunity. Upon pathogen infection, lymph nodes promptly increase the influx of naive T cells from the circulation in order to screen and prime the T cells. The precise contribution of the lymph node vasculature to the regulation of this process remains unclear. Here we show a role for the Ras GTPase, R-Ras, in the functional adaptation of high endothelial venules to increase naive T cell trafficking to the lymph nodes. R-Ras is transiently up-regulated in the endothelium of high endothelial venules by the inflammatory cytokine tumor necrosis factor (TNF) within 24 hours of pathogen inoculation. TNF induces R-Ras upregulation in endothelial cells via JNK and p38 mitogen-activated protein kinase but not NF-κB. Studies of T cell trafficking found that the loss of function of endothelial R-Ras impairs the rapid acceleration of naive T cell recruitment to the lymph nodes upon inflammation. This defect diminished the ability of naive OT-1 T cells to develop antitumor activity against ovalbumin-expressing melanoma. Proteomic analyses suggest that endothelial R-Ras facilitates TNF-dependent transendothelial migration (diapedesis) of naive T cells by modulating molecular assembly the at T cell–endothelial cell interface. These findings give new mechanistic insights into the functional adaptation of high endothelial venules to accelerate naive T cell recruitment to the lymph nodes.
Dietary lipids accumulate in macrophages and stromal cells and change the microarchitecture of mesenteric lymph nodes
2020, Journal of Advanced Research
Citation Excerpt :
Furthermore, capillary BECs are also detectable and provide nutrients and oxygen to the surrounding cells [28,29]. FDCs are located in the cortex, express CXCL13 for B cell migration and can function as antigen-presenting cells [30]. During immune reactions, FDCs remodel follicles into germinal centers, where B cells proliferate and undergo somatic hypermutations [30].
In obesity, increased dietary lipids are taken up and transported by the lymphatic systems into the circulatory system. Increased fat accumulation results in impairments in the lymph fluid and lymph node (LN) atrophy. LNs filter the lymph fluid for foreign antigens to induce and control immune responses, and the alteration of this function during obesity remains underexplored. Here, the changes within the microarchitecture of mesenteric LNs (mLNs) during high levels of lipid transport were investigated, and the role of stromal cells in mice fed a high-fat diet for 10 weeks was assessed. Microarray experiments revealed that gene probes involved in lipid metabolism are expressed by mLN stromal cells. Transmission electron microscopy enabled the identification of lipid droplets in lymphatic endothelial cells, different reticulum cells, and macrophages, and the lipid droplet sizes as well as their numbers and intercellular distances increased after 10 weeks of high-fat diet feeding. The results indicate that changes in the microarchitecture and increased accumulation of lipid droplets in stromal cells and macrophages influence the immunological function of mLNs.
Targeted deletion of RANKL in M cell inducer cells by the Col6a1-Cre driver
2017, Biochemical and Biophysical Research Communications
Citation Excerpt :
Mesenchymal cell-specific Cre drivers are indispensable for the functional analysis of lymphoid tissue-resident stromal cells [13]. The CCL19-Cre line is reported to have specificity for FRCs, the CD21-Cre line for FDCs and the Ve cadherin-Cre line for endothelial cells [13]. The Col6a1-Cre driver is utilized as a tool to target tissues, including the intestine, synovium, skin and secondary lymphoid organs [14–16].
The gut-associated lymphoid tissues (GALTs), including Peyer's patches (PPs), cryptopatches (CPs) and isolated lymphoid follicles (ILFs), establish a host-microbe symbiosis by the promotion of immune reactions against gut microbes. Microfold cell inducer (MCi) cells in GALTs are the recently identified mesenchymal cells that express the cytokine RANKL and initiate bacteria-specific immunoglobulin A (IgA) production via induction of microfold (M) cell differentiation. In the previous study, the Twist2-Cre driver was utilized for gene deletion in mesenchymal cells including MCi cells. In order to investigate MCi cells more extensively, it will be necessary to develop experimental tools in addition to the Twist2-Cre driver mice and characterize such drivers in specificity and efficiency. Here we show that M cell differentiation and IgA production are impaired in the targeted deletion of RANKL by the Col6a1-Cre driver. We compared Col6a1-Cre with Twist2-Cre in terms of the specificity for mesenchymal cells in GALTs. Col6a1-Cre CAG-CAT-EGFP mice exhibited EGFP expression in podoplanin⁺CD31^– cells including MCi cells, while Twist2-Cre mice were shown to target endothelial cells and podoplanin⁺CD31^– cells. Tnfsf11^fl/Δ Col6a1-Cre mice exhibited the absence of M cells and severe IgA reduction together with an alteration in gut microbial composition. Moreover, we analyzed germ free mice to test whether changes in the microbiota are the cause of M cell deficiency. M cell differentiation was normal in the CPs/ILFs of germ free mice, indicating that MCi cells induce M cells independently of microbial colonization. This study demonstrates that Col6a1-Cre driver mice are as useful as Twist2-Cre driver mice for functional analyses of GALT-resident mesenchymal cells, including MCi cells.
New tools for immunologists: models of lymph node function from cells to tissues
2023, Frontiers in Immunology
Postnatal expansion of mesenteric lymph node stromal cells towards reticular and CD34<sup>+</sup> stromal cell subsets
2022, Nature Communications

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Trends in Immunology

ReviewLymph Node Stroma Dynamics and Approaches for Their Visualization

Trends

Section snippets

Lymphoid Stromal Cells: Versatile and Reconfigurable 3D Immunological Networks

Fibroblastic Reticular Cells (FRCs)

Studying LN Stromal Cells: A Technical Challenge

Stromal Cell Heterogeneity: Understanding the LN Stroma at the Single Cell Level

Inspiration from Neurosciences: Multicolor Lineage Tracing Models

Multicolor Lineage Tracing Models to Study Stromal Cells

Better Mouse Models

Concluding Remarks

Acknowledgments

Trends Immunol.

Immunity

Immunity

Immunity

Semin. Immunol.

Immunity

Immunity

Cell

Immunol. Today

Trends Immunol.

Immunobiology

Semin. Immunol.

Immunity

Immunity

Cell

Immunity

Mol. Immunol.

Cell

Immunity

Methods Enzymol.

Neuron

Neuron

Cell

Blood

Dev. Biol.

Stromal cell contributions to the homeostasis and functionality of the immune system

Nat. Rev. Immunol.

Stromal cell-immune cell interactions

Annu. Rev. Immunol.

Lymph node fibroblastic reticular cells construct the stromal reticulum via contact with lymphocytes

J. Exp. Med.

B cell homeostasis and follicle confines are governed by fibroblastic reticular cells

Nat. Immunol.

Fibroblastic reticular cells in lymph nodes regulate the homeostasis of naive T cells

Nat. Immunol.

Topological small-world organization of the fibroblastic reticular cell network determines lymph node functionality

PLoS Biol.

Cords, channels, corridors and conduits: critical architectural elements facilitating cell interactions in the lymph node cortex

Immunol. Rev.

Lymph node fibroblastic reticular cells directly present peripheral tissue antigen under steady-state and inflammatory conditions

J. Exp. Med.

The CLEC-2-podoplanin axis controls the contractility of fibroblastic reticular cells and lymph node microarchitecture

Nat. Immunol.

Dendritic cells control fibroblastic reticular network tension and lymph node expansion

Nature

Multiple CD11c+ cells collaboratively express IL-1beta to modulate stromal vascular endothelial growth factor and lymph node vascular-stromal growth

J. Immunol.

Fibroblast-type reticular stromal cells regulate the lymph node vasculature

J. Immunol.

Fibroblastic reticular cells regulate intestinal inflammation via IL-15-mediated control of group 1 ILCs

Nat. Immunol.

Review
Lymph Node Stroma Dynamics and Approaches for Their Visualization