Conserved stromal-immune cell circuits secure B cell homeostasis and function.

B cell zone reticular cells (BRCs) form stable microenvironments that direct efficient humoral immunity with B cell priming and memory maintenance being orchestrated across lymphoid organs. However, a comprehensive understanding of systemic humoral immunity is hampered by the lack of knowledge of global BRC sustenance, function and major pathways controlling BRC-immune cell interactions. Here we dissected the BRC landscape and immune cell interactome in human and murine lymphoid organs. In addition to the major BRC subsets underpinning the follicle, including follicular dendritic cells, PI16+ RCs were present across organs and species. As well as BRC-produced niche factors, immune cell-driven BRC differentiation and activation programs governed the convergence of shared BRC subsets, overwriting tissue-specific gene signatures. Our data reveal that a canonical set of immune cell-provided cues enforce bidirectional signaling programs that sustain functional BRC niches across lymphoid organs and species, thereby securing efficient humoral immunity.

PI16+ reticular cells in human palatine tonsils govern T cell activity in distinct subepithelial niches.

Fibroblastic reticular cells (FRCs) direct the interaction and activation of immune cells in discrete microenvironments of lymphoid organs. Despite their important role in steering innate and adaptive immunity, the age- and inflammation-associated changes in the molecular identity and functional properties of human FRCs have remained largely unknown. Here, we show that human tonsillar FRCs undergo dynamic reprogramming during life and respond vigorously to inflammatory perturbation in comparison to other stromal cell types. The peptidase inhibitor 16 (PI16)-expressing reticular cell (PI16+ RC) subset of adult tonsils exhibited the strongest inflammation-associated structural remodeling. Interactome analysis combined with ex vivo and in vitro validation revealed that T cell activity within subepithelial niches is controlled by distinct molecular pathways during PI16+ RC-lymphocyte interaction. In sum, the topological and molecular definition of the human tonsillar stromal cell landscape reveals PI16+ RCs as a specialized FRC niche at the core of mucosal immune responses in the oropharynx.

Remodeling of light and dark zone follicular dendritic cells governs germinal center responses.

Efficient generation of germinal center (GC) responses requires directed movement of B cells between distinct microenvironments underpinned by specialized B cell-interacting reticular cells (BRCs). How BRCs are reprogrammed to cater to the developing GC remains unclear, and studying this process is largely hindered by incomplete resolution of the cellular composition of the B cell follicle. Here we used genetic targeting of Cxcl13-expressing cells to define the molecular identity of the BRC landscape. Single-cell transcriptomic analysis revealed that BRC subset specification was predetermined in the primary B cell follicle. Further topological remodeling of light and dark zone follicular dendritic cells required CXCL12-dependent crosstalk with B cells and dictated GC output by retaining B cells in the follicle and steering their interaction with follicular helper T cells. Together, our results reveal that poised BRC-defined microenvironments establish a feed-forward system that determines the efficacy of the GC reaction.

An Innate Checkpoint Determines Immune Dysregulation and Immunopathology during Pulmonary Murine Coronavirus Infection.

Hallmarks of life-threatening, coronavirus-induced disease include dysregulated antiviral immunity and immunopathological tissue injury. Nevertheless, the sampling of symptomatic patients overlooks the initial inflammatory sequela culminating in severe coronavirus-induced disease, leaving a fundamental gap in our understanding of the early mechanisms regulating anticoronavirus immunity and preservation of tissue integrity. In this study, we delineate the innate regulators controlling pulmonary infection using a natural mouse coronavirus. Within hours of infection, the cellular landscape of the lung was transcriptionally remodeled altering host metabolism, protein synthesis, and macrophage maturation. Genetic perturbation revealed that these transcriptional programs were type I IFN dependent and critically controlled both host cell survival and viral spread. Unrestricted viral replication overshooting protective IFN responses culminated in increased IL-1ß and alarmin production and triggered compensatory neutrophilia, interstitial inflammation, and vascular injury. Thus, type I IFNs critically regulate early viral burden, which serves as an innate checkpoint determining the trajectory of coronavirus dissemination and immunopathology.

Differentiation and activation of fibroblastic reticular cells.

Secondary lymphoid organs (SLO) are underpinned by fibroblastic reticular cells (FRC) that form dedicated microenvironmental niches to secure induction and regulation of innate and adaptive immunity. Distinct FRC subsets are strategically positioned in SLOs to provide niche factors and govern efficient immune cell interaction. In recent years, the use of specialized mouse models in combination with single-cell transcriptomics has facilitated the elaboration of the molecular FRC landscape at an unprecedented resolution. While single-cell RNA-sequencing has advanced the resolution of FRC subset characterization and function, the high dimensionality of the generated data necessitates careful analysis and validation. Here, we reviewed novel findings from high-resolution transcriptomic analyses that refine our understanding of FRC differentiation and activation processes in the context of infection and inflammation. We further discuss concepts, strategies, and limitations for the analysis of single-cell transcriptome data from FRCs and the wide-ranging implications for our understanding of stromal cell biology.

Integration of Th17- and Lymphotoxin-Derived Signals Initiates Meningeal-Resident Stromal Cell Remodeling to Propagate Neuroinflammation.

Tertiary lymphoid tissues (TLTs) have been observed in the meninges of multiple sclerosis (MS) patients, but the stromal cells and molecular signals that support TLTs remain unclear. Here, we show that T helper 17 (Th17) cells induced robust TLTs within the brain meninges that were associated with local demyelination during experimental autoimmune encephalitis (EAE). Th17-cell-induced TLTs were underpinned by a network of stromal cells producing extracellular matrix proteins and chemokines, enabling leukocytes to reside within, rather than simply transit through, the meninges. Within the CNS, interactions between lymphotoxin αß (LTαß) on Th17 cells and LTßR on meningeal radio-resistant cells were necessary for the propagation of de novo interleukin-17 responses, and activated T cells from MS patients expressed elevated levels of LTßR ligands. Therefore, input from both Th17 cells and the lymphotoxin pathway induce the formation of an immune-competent stromal cell niche in the meninges.

Stromal Cell Niches in the Inflamed Central Nervous System.

Inflammation in the CNS must be tightly regulated to respond efficiently to infection with neurotropic pathogens. Access of immune cells to the CNS and their positioning within the tissue are controlled by stromal cells that construct the barriers of the CNS. Although the role of the endothelium in regulating the passage of leukocytes and small molecules into the CNS has been studied extensively, the contribution of fibroblastic stromal cells as portals of entry into the CNS was only recently uncovered. We review the critical immune-stimulating role of meningeal fibroblasts in promoting recruitment and retention of lymphocytes during CNS inflammation. Activated meningeal fibroblastic stromal cells have the capacity to rapidly elaborate an immune-competent niche that sustains protective immune cells entering the CNS from the draining cervical lymph node. Such stromal cell niches can ultimately foster the establishment of tertiary lymphoid tissues during chronic neuroinflammatory conditions.

Substrain differences reveal novel disease-modifying gene candidates that alter the clinical course of a rodent model of multiple sclerosis.

Experimental autoimmune encephalomyelitis (EAE) is a rodent model of multiple sclerosis that is executed in animals by immunization with myelin Ag in adjuvant. The SJL/J autoimmune-prone strain of mouse has been used to model relapsing-remitting multiple sclerosis. However, significant variations in peak scores, timing of onset, and incidence are observed among laboratories, with the postacute (relapse) phase of the disease exhibiting significant inconsistency. We characterized two substrains of SJL/J mice that exhibit profoundly different EAE disease parameters. Induction of EAE in the first SJL/J substrain resulted in many cases of chronic EAE that was dominated by an aggressive B cell response to the immunizing Ag and to endogenous CNS Ags. In contrast, the other SJL/J substrain exhibited a relapsing-remitting form of EAE concomitant with an elevated number of cytokine-producing CD4(+) T cells in the CNS. Exploiting these interstrain differences, we performed a genome-wide copy number analysis on the two disparate SJL/J substrains and discovered numerous gene-dosage differences. In particular, one inflammation-associated gene, Naip1, was present at a higher copy number in the SJL/J substrain that exhibited relapsing-remitting EAE. These results demonstrate that substrain differences, perhaps at the level of genomic copy number, can account for variability in the postacute phase of EAE and may drive chronic versus relapsing disease.

B cell zone reticular cell microenvironments shape CXCL13 gradient formation.

Through the formation of concentration gradients, morphogens drive graded responses to extracellular signals, thereby fine-tuning cell behaviors in complex tissues. Here we show that the chemokine CXCL13 forms both soluble and immobilized gradients. Specifically, CXCL13+ follicular reticular cells form a small-world network of guidance structures, with computer simulations and optimization analysis predicting that immobilized gradients created by this network promote B cell trafficking. Consistent with this prediction, imaging analysis show that CXCL13 binds to extracellular matrix components in situ, constraining its diffusion. CXCL13 solubilization requires the protease cathepsin B that cleaves CXCL13 into a stable product. Mice lacking cathepsin B display aberrant follicular architecture, a phenotype associated with effective B cell homing to but not within lymph nodes. Our data thus suggest that reticular cells of the B cell zone generate microenvironments that shape both immobilized and soluble CXCL13 gradients.

Meningeal Tertiary Lymphoid Tissues and Multiple Sclerosis: A Gathering Place for Diverse Types of Immune Cells during CNS Autoimmunity.

Collections of leukocytes in the meningeal space have been documented in Multiple Sclerosis (MS). These meningeal aggregates, which in the context of other autoimmune diseases have often been termed tertiary lymphoid tissues (TLT), have been associated with sub-pial cortical damage and disease progression. However, the key molecular and cellular signals required for their formation and maintenance remain unclear. Herein, we review TLT structures in other disease states in order to provide a framework for understanding these structures in the MS meninges. We then assess the evidence that the meningeal compartment serves as an important nexus for immune cells as well as a location for drainage of antigen into cervical lymph nodes. Extrapolating what is known about the molecular and cellular cues that initiate the formation of leukocyte aggregates in non-lymphoid tissues, we speculate on what signals lead to the formation and maintenance of meningeal TLT structures. Referring to the animal model of MS [experimental autoimmune encephalomyelitis (EAE)], we also explore what is known about these structures in supporting B cell and T cell responses during neuroinflammation. Last, we examine the evidence that connects these structures to ongoing neuropathology. Collectively, our review points to the meningeal compartment as an important player in neuroinflammatory processes. Moreover, we hypothesize that in order to gain insights into pro- and anti-inflammatory properties of lymphocytes in MS, one must understand the cellular scaffolds that support lymphocyte retention within the meninges, thus highlighting the importance of non-immune cells (stromal cells) in the neuroinflammatory process.

B Cells in the Multiple Sclerosis Central Nervous System: Trafficking and Contribution to CNS-Compartmentalized Inflammation.

Clinical trial results of peripheral B cell depletion indicate abnormal proinflammatory B cell properties, and particularly antibody-independent functions, contribute to relapsing MS disease activity. However, potential roles of B cells in progressive forms of disease continue to be debated. Prior work indicates that presence of B cells is fostered within the inflamed MS central nervous system (CNS) environment, and that B cell-rich immune cell collections may be present within the meninges of patients. A potential association is reported between such meningeal immune cell collections and the subpial pattern of cortical injury that is now considered important in progressive disease. Elucidating the characteristics of B cells that populate the MS CNS, how they traffic into the CNS and how they may contribute to progressive forms of the disease has become of considerable interest. Here, we will review characteristics of human B cells identified within distinct CNS subcompartments of patients with MS, including the cerebrospinal fluid, parenchymal lesions, and meninges, as well as the relationship between B cell populations identified in these subcompartments and the periphery. We will further describe the different barriers of the CNS and the possible mechanisms of migration of B cells across these barriers. Finally, we will consider the range of human B cell responses (including potential for antibody production, cytokine secretion, and antigen presentation) that may contribute to propagating inflammation and injury cascades thought to underlie MS progression.