Organized Lymphoid Tissue Research Articles

Interleukin 9 mediates T follicular helper cell activation to promote antibody responses.

Antigen-specific humoral responses are orchestrated through complex interactions among immune cells in lymphoid tissues, including the collaboration between B cells and T follicular helper (Tfh) cells. Accumulating evidence indicates a crucial role for interleukin-9 (IL-9) in the formation of germinal centers (GCs), enhancing the generation of class-switched high-affinity antibodies. However, the exact function of IL-9 in Tfh cell regulation remains unclear. In this study, we examined the humoral immune responses of CD4Cre/+Il9rafl/fl mice, which lack an IL-9-specific receptor in Tfh cells. Upon intraperitoneal immunization with sheep red blood cells (SRBCs), CD4Cre/+Il9rafl/fl mice displayed diminished levels of SRBC-specific IgG antibodies in their sera, along with reduced levels of GC B cells and plasma cells. Notably, Il9ra-deficient Tfh cells in the spleen exhibited decreased expression of their signature molecules such as B-cell lymphoma 6, C-X-C chemokine receptor 5, IL-4, and IL-21 compared to control mice. In models of allergic asthma induced by house dust mite (HDM) inhalation, CD4Cre/+Il9rafl/fl mice failed to elevate serum levels of HDM-specific IgE and IgG. This was accompanied by reductions in Tfh cells, GC B cells, and plasma cells in mediastinal lymph nodes. Furthermore, group 2 innate lymphoid cells (ILC2s) were identified as producers of IL-9 under immunizing conditions, possibly induced by leukotrienes released by activated IgD+ B cells around the T-B border. These observations may indicate the critical role of IL-9 receptor signaling in the activation of Tfh cells, with ILC2s potentially capable of supplying IL-9 in organized lymphoid tissues.

CGAS-activated endothelial cell-T cell cross-talk initiates tertiary lymphoid structure formation.

Aberrant activation of the cyclic guanosine monophosphate-adenosine monophosphate synthase-stimulator of interferon genes (cGAS-STING) pathway causes autoimmunity in humans and mice; however, the exact mechanism by which the cGAS-STING pathway initiates adaptive immunity and tissue pathology is still not fully understood. Here, we used a cGAS knockin (KI) mouse model that develops systemic autoimmunity. In the lungs of cGAS-KI mice, blood vessels were enclosed by organized lymphoid tissues that resemble tertiary lymphoid structures (TLSs). Cell-intrinsic cGAS induction promoted up-regulation of CCR5 in CD8+ T cells and led to CCL5 production in vascular endothelial cells. Peripheral CD8+ T cells were recruited to the lungs and produced CXCL13 and interferon-γ. The latter triggered endothelial cell death, potentiated CCL5 production, and was essential for TLS establishment. Blocking CCL5 or CCR5, or depleting CD8+ T cells, impaired TLS formation. cGAS-mediated TLS formation also enhanced humoral and antitumor responses. These data demonstrate that cGAS signaling drives a specialized lymphoid structure that underlies autoimmune tissue pathology.

Abstract 3988: Deep spatial immunophenotyping of lymphoid aggregates in pancreatic cancer using multi-omic integration of ultra high-plex proteomics and transcriptomics

Abstract The most well-described mechanism of immune evasion in PDAC is exclusion of immune cells by a desmoplastic stroma, but such immune exclusion is not uniform. Through single-nucleus RNA-seq and whole-transcriptome digital spatial profiling of patient tumors, we previously reported marked heterogeneity in the immune composition of PDAC. Here, we developed a novel multi-omics workflow integrating single cell spatial proteomics (96-plex, Akoya Phenocycler®-Fusion) and transcriptomics (990-plex, Nanostring CosMx) in primary patient PDAC tumors to systematically study spatial relationships and functional states of malignant, stromal, and immune cells. Consecutive sections from four independent tumors enabled whole slide proteomic assessment of >3.5M cells, and transcriptomic assessment of >200K cells. Strikingly, we observed an abundance of immune cell aggregates in all tissue sections, up to 57 within a 1 cm2 tumor section. Aggregates were diverse in composition with representation of myeloid cells, CD8+ T cells, and stromal cells, including fibroblasts and nerves. B/T lymphoid aggregates were well vascularized with co-localized CD31+CD34+ endothelial cells and appeared at various phases of maturation towards putative tertiary lymphoid structures (TLSs) containing CD38+ B cells, CD4+ T cells, and CD21+ follicular DCs. These tissues were enriched for interferon stimulated genes in T cells and type 1 interferon response signatures in malignant cells and cancer associated fibroblasts (CAFs) forming inflammatory multicellular hubs that have been associated with improved response to immune checkpoint blockade. Surprisingly, B cells, in addition to macrophages and CD8+ T cells, were major producers of interferon gamma (B cells: 13.7%, macrophages: 36.6%, CD8+ T cells: 12.1% of IFNG+ cells). Finally, to gain insight into the necessary factors for TLS formation in PDAC, we quantified the expression of chemokines and adhesion molecules associated with TLS initiation. Activated dendritic cells expressed CCL19 and CCL21, endothelial cells expressed CCL21, and in concert with inflammatory CAFs (iCAFs), CXCL12. Malignant cells expressed CXCL13 and CCL20 and displayed high levels of cell adhesion molecules ICAM1, VCAM1, and ICAM2. Together, our data suggests that the functions of lymphoid tissue organizer cells can be substituted by endothelial cells, dendritic cells, iCAFs, and, surprisingly, malignant cells. Our spatial immunophenotyping study of pancreatic cancer thus highlights the marked immune heterogeneity of the disease and suggests a central role for malignant and stromal cells in mediating diverse mechanisms of immune evasion and engagement. Functional studies will be conducted to investigate these mechanisms of tumor-immune crosstalk and identify novel immunotherapeutic vulnerabilities for this deadly disease. Citation Format: Dennis Gong, Jingyi Cao, Peter Wang, Niyati Jhaveri, Yue Hou, Bassem B. Cheikh, HaYeun Ji, Mark Gregory, Jason Reeves, Michael Patrick, Carina Shiau, Jennifer Su, Jimmy A. Guo, Joseph M. Beechem, Martin Hemberg, William L. Hwang, Ryan Park. Deep spatial immunophenotyping of lymphoid aggregates in pancreatic cancer using multi-omic integration of ultra high-plex proteomics and transcriptomics [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 3988.

Crosstalk between CD8+ T cells and mesenchymal stromal cells in intestine homeostasis and immunity.

The intestine represents the most complex cellular network in the whole body. It is constantly faced with multiple types of immunostimulatory agents encompassing from food antigen, gut microbiome, metabolic waste products, and dead cell debris. Within the intestine, most T cells are found in three primary compartments: the organized gut-associated lymphoid tissue, the lamina propria, and the epithelium. The well-orchestrated epithelial-immune-microbial interaction is critically important for the precise immune response. The main role of intestinal mesenchymal stromal cells is to support a structural framework within the gut wall. However, recent evidence from stromal cell studies indicates that they also possess significant immunomodulatory functions, such as maintaining intestinal tolerance via the expression of PDL1/2 and MHC-II molecules, and promoting the development of CD103+ dendritic cells, and IgA+ plasma cells, thereby enhancing intestinal homeostasis. In this review, we will summarize the current understanding of CD8+ T cells and stromal cells alongside the intestinal tract and discuss the reciprocal interactions between T subsets and mesenchymal stromal cell populations. We will focus on how the tissue residency, migration, and function of CD8+ T cells could be potentially regulated by mesenchymal stromal cell populations and explore the molecular mediators, such as TGF-β, IL-33, and MHC-II molecules that might influence these processes. Finally, we discuss the potential pathophysiological impact of such interaction in intestine hemostasis as well as diseases of inflammation, infection, and malignancies.

Recurrence of follicular dendritic cell sarcoma in tonsils after 7 years: a rare case report

Follicular dendritic cell sarcoma (FDCS) is a rare mesenchymal neoplasm. It arises from not only from lymph nodes but also from extra nodal tissues, either as acquired lymphoid tissue or as part of the organized constitutive lymphoid tissue. We here report this rare entity which developed again after 7 years post tonsillectomy in a 5-year old male patient. Patient underwent radical tonsillectomy and adjuvant treatment is awaited. Differential diagnosis includes large cell lymphoma, peripheral nerve sheath tumor, extracranial meningioma, malignant melanoma, metastatic carcinoma, ectopic thymoma, malignant fibrous histiocytoma, and interstitial reticulum cell sarcoma. Currently, the management of FDCS includes the therapeutic guidelines similar to that of high-grade soft tissue sarcomas that is complete surgical resection of the lesion with possibility of adjuvant radiotherapy and/or chemotherapy. The ideal combination of management of FDCS has yet to be defined.

CXCL13 Serum Level as a Potential Biomarker in Vitiligo Patients: Correlation with Disease Activity and Severity

Background: Vitiligo is a prevalent autoimmune pigmentary skin disorder, with 0.5 - 2% of the global population prevalence. Human follicular T helper cells, dendritic follicular cells, and mesenchymal lymphoid tissue organizer cells all generate the B-cell chemokine (C-X-C motif) ligand 13 (CXCL13). Chemokine (C-X-C motif) ligand 13 and its receptor CXCR5 have been linked to the etiopathogenesis of various autoimmune and inflammatory diseases in the latest studies. Objectives: This study aimed to assess the serum CXCL13 level in vitiligo patients and its relation to the activity and severity of the disease. Methods: This case-control study involved 21 vitiligo patients and 21 age and gender-matched healthy controls. Full history taking, clinical evaluation, and assessment of the disease activity and severity were performed for all the patients. The CXCL13 serum levels were measured by enzyme-linked immunosorbent assay (ELISA). Results: The serum CXCL13 levels were significantly higher in vitiligo patients than in healthy controls (227.35 ± 125.70 and 130.58 ± 26.52 ng/L, respectively; P = 0.002). There was a significant relationship between CXCL13 and the clinical pattern of activity among patients with vitiligo; however, there was no correlation with disease activity or severity index scores. Conclusions: Although CXCL13 was greater in vitiligo patients than in healthy controls, it was unrelated to the activity or severity of the illness. This finding suggests that CXCL13 might have a role in the immunological pathogenesis of vitiligo.

Histological and immunohistochemical characterization of mucosa-associated lymphoid tissue and antigen-presenting cells in trachea and lung of cattle.

The presence of bronchus-associated lymphoid tissue (BALT) and its structural components has been described in different healthy animal species and in animals with diseases of the respiratory tract. In contrast to normal mammals, BALT is absent in healthy human adult lungs, but has been found in the lungs of children. The histological characteristics of organized mucosa-associated lymphoid tissue (MALT), its subsets of immune cells and their in situ distribution in the lung of healthy subadult and adult cattle shows close similarities with BALT in humans and other animal species such as sheep, horses and pigs. This study clearly demonstrates that organized MALT also occurs in the tracheal mucosa of cattle. The absence of tracheal MALT and BALT in calves suggest that these structures are not constitutive. In the mucosa of bovine trachea, bronchi and bronchioli, MHC II+ and CD11c+ dendritic cells (DCs) are located in the epithelium and in the lamina propria mucosae. These DCs are already present in calves soon after birth. Examination of tangential epithelial sheets shows that in the bovine tracheal epithelium, like in man and rat, a dense network of MHC II+ and CD11c+ DCs exists and that their number is considerably higher than in conventional transverse sections. In the bovine tracheal and bronchial epithelium, MHC II+ DCs are extending their dendrites towards the lumen indicating that these DCs possibly are involved in sampling of luminal antigens. The presence of significantly higher numbers of MHC II+ DCs in the tracheal and bronchial/bronchiolar mucosa of older cattle in than in calves possibly results from local stimulation with exogenous antigens during postnatal life. Detection of DCs expressing the costimulatory molecules CD80 and CD86 in calves and cattle suggests maturation of DCs, which is most likely induced by stimulation with exogenous antigens.

Abstract B030: Form and function in intratumoral immune organization: Understanding the cellular composition of TCF1+ CD8+ T cell niches in human cancer

Abstract Tumor infiltrating T cells are a positive prognosticator in many tumor types, and our prior work demonstrated that in renal cell carcinoma, patients with higher CD8 T cell infiltration have improved survival (Jansen Nature 2019). We described that this T cell response is supported by TCF1+ stem-like CD8 T cells, which reside within dense regions of closely clustered antigen presenting cells within the tumor. Interestingly, aggregations of immune cells have also been described in other settings and termed ‘tertiary lymphoid structures’ (TLS), raising an important question of whether the described immune niches are similar to these TLS or if they represent a distinct type of peripheral immune organization. In the work presented here, we use quantitative immunofluorescence image analysis to examine the cellular composition and organization of intratumoral immune niches, TLS, and secondary lymphoid tissue. The novel data presented here describes how immune niches are distinct from TLS and are more similar to the organization of T cell zones in secondary lymphoid tissue. Immune niches are T cell dominant structures, with T cell and antigen presenting cell compartments similar to T cell zones, whereas these niches profoundly lack the B cell presence seen in TLS and B cell zones of secondary lymphoid tissue. Immune niches are comparatively small in size (50-200um), while TLS are much larger (often >1mm), and, importantly, immune niches are found inside the tumor border, while TLS are located peripherally in tumor tissue. Notably, we find that immune niches have a marked presence of aSMA+ stromal cells, which are similar to those found in T cell zones but absent in B cell zones and TLS. When probed by flow cytometry and RNAseq, we find that aSMA+ stroma in immune niches are myofibroblast-like, similar those known to maintain the structure of secondary lymphoid tissues, and are distinct from those found in healthy kidney tissue. These data suggest these aSMA+ cells may play an important role in the organization of immune niches in tumor tissue. As we have shown that a robust T cell response is beneficial for patient outcomes, it is crucial to understand the mechanisms which support that response. Recently, our group defined that CD8 T cell activation in cancer is comprised of two phases, with an initial priming phase in the lymph node and a second phase of effector program acquisition in the tumor tissue (Prokhnevska Immunity 2023). We suggest that these immune niches represent an important hub for this development of T cell function, which is required for the robust, effective anti-tumor T cell response that supports improved patient outcomes and is the basis for the response to immunotherapy. Advancing our understanding of intratumoral immune organization will allow for enhanced biomarker discovery, improvement in existing therapies, and innovation in developing novel therapeutic strategies that will convert immunologically “cold” tumors to be immunologically “hot” and enhance anti-tumor immunity. Citation Format: Caroline S. Jansen, BaoHan Vo, Ewelina Sobierajska, Rachel Greenwald, Patrick Mullane, Nataliya Prokhnevska, Maria Cardenas, Mehmet Asim Bilen, Adeboye O. Osunkoya, Viraj A. Master, Haydn T. Kissick. Form and function in intratumoral immune organization: Understanding the cellular composition of TCF1+ CD8+ T cell niches in human cancer [abstract]. In: Proceedings of the AACR Special Conference: Advances in Kidney Cancer Research; 2023 Jun 24-27; Austin, Texas. Philadelphia (PA): AACR; Cancer Res 2023;83(16 Suppl):Abstract nr B030.

A local subset of mesenchymal cells expressing the transcription factor Osr1 orchestrates lymph node initiation

During development, lymph node (LN) initiation is coordinated by lymphoid tissue organizer (LTo) cells that attract lymphoid tissue inducer (LTi) cells at strategic positions within the embryo. The identity and function of LTo cells during the initial attraction of LTi cells remain poorly understood. Using lineage tracing, we demonstrated that a subset of Osr1-expressing cells was mesenchymal LTo progenitors. By investigating the heterogeneity of Osr1+ cells, we uncovered distinct mesenchymal LTo signatures at diverse anatomical locations, identifying a common progenitor of mesenchymal LTos and LN-associated adipose tissue. Osr1 was essential for LN initiation, driving the commitment of mesenchymal LTo cells independent of neural retinoic acid, and for LN-associated lymphatic vasculature assembly. The combined action of chemokines CXCL13 and CCL21 was required for LN initiation. Our results redefine the role and identity of mesenchymal organizer cells and unify current views by proposing a model of cooperative cell function in LN initiation.

Pseudotemporal and spatial organization of B cell activation and differentiation in Peyer's patches

Abstract Organized lymphoid tissues are important sites of immune induction where B cells activate and differentiate. High-resolution transcriptomic technology has revolutionized current understandings of B cell activation and differentiation in organized lymphoid tissues such as lymph nodes but has not been used to dissect B cell activation and differentiation in Peyer’s patches, important sites of intestinal immune induction. We utilized single-cell RNA sequencing and spatial transcriptomics to define pseudotemporal and spatial organization, respectively, of B cell activation and differentiation in porcine jejunal and ileal Peyer’s patches (JPPs and IPPs). IPPs had greater follicular mass and more defined follicular borders than JPPs, resulting in greater follicular B cell abundance in IPPs and non-follicular B cell abundance in JPPs. Despite follicular morphology and B cell abundances varying between JPPs and IPPs, pseudotemporal trajectories and predicted spatial locations of B cell activation and differentiation were similar between intestinal locations. Transition of resting into follicular B cells was demarcated by increased expression of follicle- and cell cycle-associated genes and movement of cells from interfollicular zones (IFZs) into follicles. Another trajectory indicated transition of resting B cells located in IFZs directly into antibody-secreting cells located in IFZs and crypts, suggesting a route of extrafollicular B cell activation and differentiation. Altogether, results indicate conserved patterns of both follicular and extrafollicular B cell activation and differentiation in Peyer’s patches across intestinal locations, despite differences in cellular abundances and follicular morphology. Research was funded by (1) USDA-ARS CRIS project #5030-3200- 225-00D, (2) the SCINet project of the USDA ARS project number 0500-00093-001- 00-D, and (3) an appointment to the Agricultural Research Service (ARS) Research Participation Program administered by the Oak Ridge Institute for Science and Education (ORISE) through an interagency agreement between the U.S. Department of Energy (DOE) and the U.S. Department of Agriculture (USDA). ORISE is managed by ORAU under DOE contract number DE-SC0014664. All opinions expressed in this paper are the authors’ and do not necessarily reflect the policies and views of USDA, ARS, DOE, or ORAU/ORISE.

Murine neonatal dermal fibroblast acquires a lymphoid tissue organizer cell-like activity upon synergistic activation of TNF-α receptor and LTβ receptor

Tertiary lymphoid organs (TLOs) are ectopic aggregates of immune cells. As accumulating studies demonstrate TLOs as a predictor of better prognosis in certain cancers, targeting TLO formation, which is tightly regulated by the lymphoid tissue organizer cells (LTOs), has become intriguing in cancer treatment. However, the clinical outcome of these attempts is limited, because the approaches for activating tumor adjacent LTO is lack and little is known about what type of self-cell can be used as LTO to initiate TLO formation. Here we demonstrate that co-stimulation with membrane-bound ligand LTα1β2 and soluble TNF-α could induced an LTO-like activity in murine neonatal dermal fibroblast, featured by high expression of cell migration-associated chemokines and adhesion molecules that resemble typical LTO gene signature. Furthermore, the LTO-phenotypic dermal fibroblast could enhance the attachment and survival of T and B cell and proliferation of T cell. These findings suggest dermal fibroblast as a promising target for TLO induction to improve cancer immunotherapy.

Lead suppresses interferon γ to induce splenomegaly via modification on splenic endothelial cells and lymphoid tissue organizer cells in mice.

Splenomegaly is a symptom characterized by the presence of an enlarged spleen. The impact of environmental factors on splenomegaly is largely unknown. In this study, C57BL/6 mice were treated with 125ppm or 1250ppm lead (Pb) via drinking water for 8 wk, and the process of splenomegaly was evaluated. Treatment with 1250ppm Pb, but not 125ppm Pb, caused splenomegaly, which was associated with increased capacity for erythrocyte clearance. Intriguingly, Pb-caused splenomegaly was independent of lymphoid tissue inducer (LTi) cells, which produce lymphotoxins α and β (LTα/β) to activate endothelial cells and LT organizer (LTo) cells and drive the development of spleen physiologically. A direct action of Pb on endothelial cells and LTo cells did not impact their proliferation. On the other hand, during steady state, a tonic level of interferon (IFN)γ acted on endothelial cells and LTo cells to suppress splenomegaly, as IFNγ receptor (IFNγR)-deficient mice had enlarged spleens relative to wild-type mice; during Pb exposure, splenic IFNγ production was suppressed, thus leading to a loss of the inhibitory effect of IFNγ on splenomegaly. Mechanically, Pb acted on splenic CD4+ T cells to suppress IFNγ production, which impaired the Janus kinase (Jak)1/ signal transducer and activator of transcription (STAT)1 signaling in endothelial cells and LTo cells; the weakened Jak1/STAT1 signaling resulted in the enhanced nuclear factor-κB (NF-κB) signaling in endothelial cells and LTo cells, which drove their proliferation and caused splenomegaly. The present study reveals a previously unrecognized mechanism for the immunotoxicity of Pb, which may extend our current understanding for Pb toxicology.

Lymphatic Drainage Conduit and Macro Anatomy of Lymphoid System in Large Intestine of Post Weaned Pigs

Background: The gut mucosa is effectively protected by a local organization of lymphoid tissue. In large intestine, isolated lymphoid follicles give rise to solitary lymphoid nodules and lympho glandular complexes, which are responsible for sampling of gut antigen and production of local immunity. Large intestine requires higher level of mucosal protection as because it is the site where food staff remains for a longer period and also for ascending rectal infection. Present study was undertaken highlighting the lymphatic drainage from large intestine and solitary lymphoid nodules of pig. Methods: 1% solution of methylene blue up to 100 micro litters in the one place of submucosa of large intestine as per the standard technique advocated by Smedley et al. (2014) and then stained nearest mesenteric lymph nodes were recorded and re-injected to map the drainage to the next draining lymph node to trace the lymphatic tract. Result: The study was designed on 12 (Twelve) apparently healthy weaned crossbred piglets of 3 to 4 months age irrespective of sex. Gross study revealed the presence of two different types of solitary lymphatic nodules (SLN) viz. scrotiform and faviform. The scrotiform SLN consisted of central small pore was appreciated in colon and rectum, whereas the faviform SLN although present in colon and rectum was devoid of a central pore. The average solitary lymphoid nodules in the large intestine was 427, of which 403 in colon and 24 in rectum and in terminal part of rectum patches of solitary lymphoid nodules of more than 100 numbers were observed. The different lymph nodes involved in the lymphatic drainage of large intestine of pig were colic caecal, anorectal , lumbar aortic, renal , medial lumbar and lateral lumbar lymph nodes. Again, except the right and left lumbar duct, one minor lymphatic trunk was observed parallel to the rectum, draining the lymph from anorecral lymph node to caudal mesenteric lymph node and then to renal lymph node. The celiac trunk and the jejunal trunk joined and formed a common trunk which was again joined by the colic trunk that finally opened into the caudal aspect of cysternachyli.

Discovery of an organized nasopharynx-associated lymphoid tissue in the nasal cavity of rainbow trout and its role in secondary adaptive immune responses to nasal vaccines

Abstract The olfactory organ of all vertebrates is constantly exposed to pathogens. The nasal mucosa is therefore equipped with a local immune system that protects the host from infection. In teleost fish, we previously described the presence of a diffuse nasopharynx-associated lymphoid tissue (NALT) found in the olfactory lamellae. This diffuse NALT (d-NALT) consists of myeloid and lymphoid cells but lacks an organized structure. Rainbow trout d-NALT mounts quick innate immune responses as well as adaptive immune responses to nasal vaccines. Here we report the discovery of a novel lymphoid structure located in the inner epithelial lining of the nasal cavity, opposite to the tips of the olfactory lamellae, in rainbow trout. This lymphoid structure represents the first description of an organized NALT (O-NALT) in teleost fish. Rainbow trout O-NALT is rich in B and T lymphocytes at the steady state with a proportion of approximately 8:3:1 IgM+ B cells, CD8+ T cells and IgT+ cells, respectively. Using laser capture microdissection we show that compared to d-NALT, O-NALT is highly enriched in expression of CD4, CD8 and IgM but not IgT. Strikingly, O-NALT expresses 8-fold higher levels of activation-induced cytidine deaminase aidc mRNA compared to d-NALT. Following primary and secondary intranasal vaccination with a live attenuated viral vaccine against infectious hematopoietic necrosis (IHN), trout O-NALT was expanded with increased numbers of B and T cell lymphocytes. Our results indicate that teleost fish posses organized lymphoid structures in the nasal cavity that participate in the maturation of the adaptive immune response and that need to be considered as important players in the response to mucosal vaccines. This work was supported by USDA NIFA award # 2020-67015-31457

Role of the CXCL13/CXCR5 Axis in Autoimmune Diseases.

CXCL13 is a B-cell chemokine produced mainly by mesenchymal lymphoid tissue organizer cells, follicular dendritic cells, and human T follicular helper cells. By binding to its receptor, CXCR5, CXCL13 plays an important role in lymphoid neogenesis, lymphoid organization, and immune responses. Recent studies have found that CXCL13 and its receptor CXCR5 are implicated in the pathogenesis of several autoimmune diseases, such as rheumatoid arthritis, multiple sclerosis, systemic lupus erythematosus, primary Sjögren’s syndrome, myasthenia gravis, and inflammatory bowel disease. In this review, we discuss the biological features of CXCL13 and CXCR5 and the recent findings on the pathogenic roles of the CXCL13/CXCR5 axis in autoimmune diseases. Furthermore, we discuss the potential role of CXCL13 as a disease biomarker and therapeutic target in autoimmune diseases.

RNA-Seq of Single Fish Cells - Seeking Out the Leukocytes Mediating Immunity in Teleost Fishes.

The immune system is a complex and sophisticated biological system, spanning multiple levels of complexity, from the molecular level to that of tissue. Our current understanding of its function and complexity, of the heterogeneity of leukocytes, is a result of decades of concentrated efforts to delineate cellular markers using conventional methods of antibody screening and antigen identification. In mammalian models, this led to in-depth understanding of individual leukocyte subsets, their phenotypes, and their roles in health and disease. The field was further propelled forward by the development of single-cell (sc) RNA-seq technologies, offering an even broader and more integrated view of how cells work together to generate a particular response. Consequently, the adoption of scRNA-seq revealed the unexpected plasticity and heterogeneity of leukocyte populations and shifted several long-standing paradigms of immunology. This review article highlights the unprecedented opportunities offered by scRNA-seq technology to unveil the individual contributions of leukocyte subsets and their crosstalk in generating the overall immune responses in bony fishes. Single-cell transcriptomics allow identifying unseen relationships, and formulating novel hypotheses tailored for teleost species, without the need to rely on the limited number of fish-specific antibodies and pre-selected markers. Several recent studies on single-cell transcriptomes of fish have already identified previously unnoticed expression signatures and provided astonishing insights into the diversity of teleost leukocytes and the evolution of vertebrate immunity. Without a doubt, scRNA-seq in tandem with bioinformatics tools and state-of-the-art methods, will facilitate studying the teleost immune system by not only defining key markers, but also teaching us about lymphoid tissue organization, development/differentiation, cell-cell interactions, antigen receptor repertoires, states of health and disease, all across time and space in fishes. These advances will invite more researchers to develop the tools necessary to explore the immunology of fishes, which remain non-conventional animal models from which we have much to learn.

Mucosa-Associated Lymphoid Tissue (MALT) Lymphoma in the Gastrointestinal Tract in the Modern Era.

Simple SummaryExtranodal marginal zone lymphoma of mucosa-associated lymphoid tissue (MALT lymphoma), which is frequently linked to chronic antigenic stimulation, is clinically an indolent B-cell lymphoma. The most common site is the stomach, accounting for 35% of MALT lymphomas, with a strong association with Helicobacter pylori (H. pylori) infection. Although antibiotic eradication is first-line therapy for gastric MALT lymphoma, a recent trend showing increasing H. pylori-negative cases may require different treatment strategies. Intestinal MALT lymphoma, including immunoproliferative small intestinal disease, is relatively rare, and the pathogenesis and therapeutic approach have not been fully elucidated. In addition, the gastrointestinal tract was recently considered a preferable site of Epstein–Barr-virus-negative marginal zone lymphoma in the post-transplant setting. We review the updated clinicopathological features, treatment, and evolving concepts of MALT lymphoma of the gastrointestinal tract.Extranodal marginal zone lymphoma of mucosa-associated lymphoid tissue (MALT lymphoma) typically arises from sites such as the stomach, where there is no organized lymphoid tissue. Close associations between Helicobacter pylori and gastric MALT lymphoma or Campylobacter jejuni and immunoproliferative small intestinal disease (IPSID) have been established. A subset of tumors is associated with chromosomal rearrangement and/or genetic alterations. This disease often presents as localized disease, requiring diverse treatment approaches, from antibiotic therapy to radiotherapy and immunochemotherapy. Eradication therapy for H. pylori effectively cures gastric MALT lymphoma in most patients. However, treatment strategies for H. pylori-negative gastric MALT lymphoma are still challenging. In addition, the effectiveness of antibiotic therapy has been controversial in intestinal MALT lymphoma, except for IPSID. Endoscopic treatment has been noted to usually achieve complete remission in endoscopically resectable colorectal MALT lymphoma with localized disease. MALT lymphoma has been excluded from post-transplant lymphoproliferative disorders with the exception of Epstein–Barr virus (EBV)-positive marginal zone lymphoma (MZL). We also describe the expanding spectrum of EBV-negative MZL and a close association of the disease with the gastrointestinal tract.

High-Resolution, 3D Imaging of the Zebrafish Gill-Associated Lymphoid Tissue (GIALT) Reveals a Novel Lymphoid Structure, the Amphibranchial Lymphoid Tissue.

The zebrafish is extensively used as an animal model for human and fish diseases. However, our understanding of the structural organization of its immune system remains incomplete, especially the mucosa-associated lymphoid tissues (MALTs). Teleost MALTs are commonly perceived as diffuse and scattered populations of immune cells throughout the mucosa. Yet, structured MALTs have been recently discovered in Atlantic salmon (Salmo salar L.), including the interbranchial lymphoid tissue (ILT) in the gills. The existence of the ILT was only recently identified in zebrafish and other fish species, highlighting the need for in-depth characterizations of the gill-associated lymphoid tissue (GIALT) in teleosts. Here, using 3-D high-resolution microscopy, we analyze the GIALT of adult zebrafish with an immuno-histology approach that reveals the organization of lymphoid tissues via the labeling of T/NK cells with an antibody directed to a highly conserved epitope on the kinase ZAP70. We show that the GIALT in zebrafish is distributed over at least five distinct sub-regions, an organization found in all pairs of gill arches. The GIALT is diffuse in the pharyngeal part of the gill arch, the interbranchial septum and the filaments/lamellae, and structured in two sub-regions: the ILT, and a newly discovered lymphoid structure located along each side of the gill arch, which we named the Amphibranchial Lymphoid Tissue (ALT). Based on RAG2 expression, neither the ILT nor the ALT constitute additional thymi. The ALT shares several features with the ILT such as presence of abundant lymphoid cells and myeloid cells embedded in a network of reticulated epithelial cells. Further, the ILT and the ALT are also a site for T/NK cell proliferation. Both ILT and ALT show structural changes after infection with Spring Viraemia of Carp Virus (SVCV). Together, these data suggest that ALT and ILT play an active role in immune responses. Comparative studies show that whereas the ILT seems absent in most neoteleosts (“Percomorphs”), the ALT is widely present in cyprinids, salmonids and neoteleosts, suggesting that it constitutes a conserved tissue involved in the protection of teleosts via the gills.

Immune mechanisms orchestrate tertiary lymphoid structures in tumors via cancer-associated fibroblasts.

SUMMARYTumor-associated tertiary lymphoid structures (TA-TLS) are associated with enhanced patient survival and responsiveness to cancer therapies, but the mechanisms underlying their development are unknown. We show here that TA-TLS development in murine melanoma is orchestrated by cancer-associated fibroblasts (CAF) with characteristics of lymphoid tissue organizer cells that are induced by tumor necrosis factor receptor signaling. CAF organization into reticular networks is mediated by CD8 T cells, while CAF accumulation and TA-TLS expansion depend on CXCL13-mediated recruitment of B cells expressing lymphotoxin-α1β2. Some of these elements are also overrepresented in human TA-TLS. Additionally, we demonstrate that immunotherapy induces more and larger TA-TLS that are more often organized with discrete T and B cell zones, and that TA-TLS presence, number, and size are correlated with reduced tumor size and overall response to checkpoint immunotherapy. This work provides a platform for manipulating TA-TLS development as a cancer immunotherapy strategy.

Vasculature-Associated Lymphoid Tissue: A Unique Tertiary Lymphoid Tissue Correlates With Renal Lesions in Lupus Nephritis Mouse Model.

Lupus nephritis (LN) is a common complication in young patients and the most predominant cause of glomerulonephritis. Infiltrating immune cells and presence of immunocomplexes in the kidney are hallmarks of LN, which is closely associated with renal lesions (RLs). However, their regulatory mechanism in the kidney remains unclear, which is valuable for prevention of RL development. Here, we show the development of vasculature-associated lymphoid tissue (VALT) in LN, which is related to renal inflammatory cytokines, indicating that VALT is a unique tertiary lymphoid tissue. Transcriptomic analysis revealed different chemokines and costimulatory molecules for VALT induction and organization. Vascular and perivascular structures showed lymphoid tissue organization through lymphorganogenic chemokine production. Transcriptional profile and intracellular interaction also demonstrated antigen presentation, lymphocyte activity, clonal expansion, follicular, and germinal center activity in VALT. Importantly, VALT size was correlated with infiltrating immune cells in kidney and RLs, indicating its direct correlation with the development of RLs. In addition, dexamethasone administration reduced VALT size. Therefore, inhibition of VALT formation would be a novel therapeutic strategy against LN.