Expression of CD137 (4-1BB) on human follicular dendritic cells

Follicular dendritic cells (FDCs) are the antigen (Ag)-trapping accessory cells of the germinal centres (GCs), essential for the development of humoral immune responses and memory. FDCs reside in the microenvironment of secondary lymphoid tissue where Ag-activated B cells expand, and undergo isotype switching and affinity maturation prior to becoming memory B cells. In addition to delivering Ag, FDCs also provide potent nonspecific accessory signals to the B cells, which are important for the GC reaction. In this report, we show that human tonsilar FDCs express the costimulatory molecule CD137. Surface expression of CD137 on FDCs was confirmed by immunofluorescent labelling and fluorescence-activated cell sorter analysis. CD137 was also highly expressed by the human cell line HK, which displays many characteristics of in vivo FDCs. The interaction between B cells and FDCs is essential for the GC reactions, and our finding suggests that CD137 plays a role in FDC-regulated B-cell responses.     

The microstructure of secondary lymphoid organs that support immune cell trafficking

Immune cell trafficking in the secondary lymphoid organs is crucial for an effective immune response. Recirculating T cells constantly patrol not only secondary lymphoid organs but also the whole peripheral organs. Thoracic duct lymphocytes represent an ideal cell source for analyzing T cell trafficking: high endothelial venules (HEVs) allow recirculating lymphocytes to transmigrate from the blood directly, and recirculating T cells form a cluster with dendritic cells (DCs) to survey antigen invasions even in a steady state. This cluster becomes an actual site for the antigen presentation when DCs have captured antigens. On activation, effector and memory T cells differentiate into several subsets that have different trafficking molecules and patterns. DCs also migrate actively in a manner depending upon their maturational stages. Danger signals induce the recruitment of several DC precursor subsets with different trafficking patterns and functions. In this review, we describe general and specialized structures of the secondary lymphoid organs for the trafficking of T cells and DCs by a multicolor immunoenzyme staining technique. The lymph nodes, spleen, and Peyer's patches of rats were selected as the major representatives. In vivo trafficking of subsets of T cells and DCs within these organs under steady or emergency states are shown and discussed, and unsolved questions and future prospects are also considered.     

滤泡树突状细胞与生发中心反应研究的新进展

生发中心是在T细胞依赖性抗体应答过程中于外周淋巴组织内形成的一个特殊的结构。在GC内,受抗原刺激而活化的B细胞进行克隆扩增、IgV区基因的体细胞高度突变、亲和力成熟以及同类型转换,最终形成记忆性B细胞或是产生Ig的浆细胞。在GC内B细胞增殖的同时,也启动了凋亡机制,以确保最终形成的记忆B细胞或浆细胞对抗原的高度特异性。FDCs是参与再次免疫应答的重要细胞,它主要是通过表面的FcR和CR将免疫复合物结合在细胞膜上,并选择性的将抗原递呈给表达高亲和力BCR的B细胞,使之激活并产生抗体或形成记忆B细胞。因此,FDCs在生发中心反应、免疫记忆的维持、B细胞的分化、成熟以及记忆B细胞的形成具有极其重要的作用。但最近的研究对FDCs及其结合的免疫复合物的重要性提出了质疑,认为FDCs在生发中心反应、B细胞的分化、成熟以及记忆B细胞的形成中的作用很可能是非特异性的,并对驻留在FDCs表面的免疫复合物的其它潜在功能进行了讨论。     

Plasticity and heterogeneity of lymphoid organs. What are the criteria to call a lymphoid organ primary, secondary or tertiary?

Lymphoid organs are generally classified in a hierarchy with primary lymphoid organs such as the thymus and bone marrow for the production of receptor specific T and B lymphocytes, respectively, independent of antigens. In secondary lymphoid organs such as lymph nodes, spleen, and tonsils, the lymphocytes are expanded due to antigen exposure, producing memory T cells and effector B cells, resulting in plasma cells. Tertiary lymphoid tissues are often defined as aggregations of lymphoid cells in autoimmune diseases. It will be outlined that all these organs have a high plasticity and also the thymic medulla is included in the route of migrating mature T cells and the bone marrow, not only in the traffic of CD4+ but also of CD8+ lymphocytes. The mucosa-associated lymphoid organs depend to a much larger extent on microbial antigen and are much more diverse than often described. The role of structural elements as well as blood and lymphatic vessels as an entry and exit site of lymphocytes will be outlined. Using a precise terminology, taking account of the plasticity of these organs at different ages and considering species differences will reduce misunderstandings among immunologists.     

Mulberry cells in the thyroid: warthin–finkeldey‐like cells in hashimoto thyroiditis‐associated lymphoma

Warthin‐Finkeldey type giant cells were first described in autopsies performed on young children who died during the highly lethal measles epidemic in Palermo during the winter of 1908. The cells had 8‐15 nuclei without identifiable cytoplasm within the germinal centers of lymphoid organs resembling megakaryocytes. We describe a case of Hashimoto thyroiditis with an enlarging substernal throid mass. The resection specimen contained many Warthin‐Finkeldey‐Like Cells (WFLC) in an extranodal marginal zone lymphoma (MALT type) with focal transformation to diffuse large B‐cell lymphoma. The WFLC showed nuclear features similar to those of neighboring follicular dendritic cells (FDCs), favoring the hypothesis that these cells might be the product of fusion of FDCs. This is supported by immunostaining results and the occurrence of similar cells in follicular dendritic cell sarcomas and in “dysplastic” FDCs in hyaline vascular type Castleman disease, a possible precursor of follicular dendritic cell tumors. Diagn. Cytopathol. 2017;45:212–216. © 2016 Wiley Periodicals, Inc.     

B lymphocytes as antigen-presenting cell-based genetic vaccines

Summary:  Inoculation of plasmid DNA is a simple way to immunize, but it is characterized by low immunogenicity, which has hampered the development of effective DNA vaccines for human use. Here, we discuss how poor immunogenicity can be solved and present our proposal: genetically programmed B lymphocytes as antigen-presenting cell (APC) vaccines. First, we demonstrate that mature B lymphocytes take up plasmid DNA spontaneously, i.e., in the absence of any facilitating molecule or event, spontaneous lymphocyte transgenesis. Second, we demonstrate that transgenic B lymphocytes are easily and reproducibly turned into functional APCs with dual characteristics: upregulation of costimulatory molecules and endogenous synthesis of antigen. Used as immunogens in mice, transgenic B lymphocytes induce robust and long-lasting T-cell immunity after single intravenous injection. Surprisingly, immunity and protection against lethal virus challenge can be obtained with a single intravenous injection of 3 × 10² transgenic lymphocytes. The new approach is discussed relative to the advantage of targeting secondary lymphoid organs with genetically programmed B lymphocytes and the advantage offered with respect to low antigen dose. We suggest that these properties reflect on simple characteristics, such as time synchronization and initial localization to secondary lymphoid organs of APCs endowed with protracted synthesis and presentation of antigen to T cells.     

Lymphocyte trafficking and immunesurveillance]

The homeostasis of the immune system is maintained by the recirculation of naive lymphocytes through the secondary lymphoid tissues, such as the lymph nodes, Peyer's patches and spleen. Upon antigen encounter in the secondary lymphoid tissues, lymphocytes become activated and undergo a reprogramming of their trafficking properties. Most antigen-experienced lymphocytes traffic through the secondary lymphoid organs, but they can also migrate to extralymphoid tissues, where they exert effector functions. Dendritic cells in the secondary lymphoid tissues are crucial for the reprogramming of trafficking properties of activated T-lymphocytes. The exquisite specificity of such lymphocyte trafficking is determined by tissue-specific guidance signals expressed by the vascular endothelial cells, combined with counter receptors expressed by circulating lymphocytes. The high endothelial venules can selectively recruit naive lymphocytes into the lymph nodes and Peyer's patches by expressing a unique combination of vascular addressins and chemoattractants. The inflamed postcapillary venules in extralymphoid tissues also use a distinct array of endothelial adhesion molecules and tissue selective chemokines to support the recruitment of effector and memory lymphocytes that express appropriate trafficking receptors. Exit of lymphocytes from lymphoid and extralymphoid tissues into circulation is actively regulated by signals through specific receptors for sphingosine-1-phosphate and a certain chemokine(s), respectively. This review summarizes the present understandings of the mechanisms regulating homeostatic recirculation of naive lymphocytes through the secondary lymphoid tissues and tissue-specific trafficking of antigen-experienced lymphocytes.     

The adhesion molecules, l-selectin and sialyl lewis x, relate to the formation of the follicular dendritic cell-lymphocyte cluster in the mantle zone

The follicular dendritic cell (FDC)-lymphocyte cluster is rich in the follicular light zone of the secondary lymphoid follicles (LFs). Although, the mantle zone (MZ) also has FDC-lymphocyte cluster, it has not known about what kind of adhesion molecules relates to cluster formation. In the present study, we investigated whether the adhesion molecules, L-selectin (CD62L) and sialyl Lewis x (CD15s) can mediate the formation of the cluster in human tonsillar LFs. The MZ only expressed both the adhesion molecules in the secondary LF. Isolated FDC-lymphocyte clusters were composed of CD62L(+) lymphocytes and CD15s(+) FDCs. Stamper-Woodruff binding assay revealed that the binding of IgD(+) lymphocytes was significantly inhibited by pretreatment with anti-CD62L antibody or with anti-CD15s antibody. These results indicate that CD62L on MZ lymphocytes and CD15s on FDCs may play a role of the cluster formation, unlike the clusters in the other parts of LFs.     

CXCR3 chemokine receptor distribution in normal and inflamed tissues: expression on activated lymphocytes, endothelial cells, and dendritic cells

Using new human CXCR3 chemokine receptor-specific monoclonal antibodies, we studied human CXCR3 tissue distribution in lymphoid and nonlymphoid organs, as well as in inflammatory conditions, including rheumatoid arthritis, Hashimoto's thyroiditis, and dermal vasculitis. CXCR3 was expressed by certain dendritic cell subsets, specifically myeloid-derived CD11c positive cells, not only in those present in normal lymphoid organs, but also in germinal centers generated in inflammatory conditions. CXCR3 expression was also detected in some lymphocyte subsets such as intraepithelial lymphocytes of secondary lymphoid organs and infiltrating lymphocytes in inflammatory conditions. In addition, CXCR3 was constitutively expressed by endothelial cells (EC) of vessels of medium and large caliber but not in small vessels from different organs. Finally, enhanced CXCR3 expression was found in EC and in infiltrating lymphocytes with an activated phenotype in inflammatory diseases. The CXCR3 chemokine receptor may play a role in the regulation of leukocyte migration to inflammatory sites.     

Regulation of secondary lymphoid organ development by the nuclear factor-κB signal transduction pathway

Summary: In primary lymphoid organs, such as thymus and bone marrow, B and T lymphocytes differentiate from lymphoid stem cells into mature albeit naïve effector cells. In contrast, secondary lymphoid organs, such as the spleen, lymph nodes, and Peyer's patches (PPs), provide an environment that enable lymphocytes to interact with each other, with accessory cells, and with antigens, resulting in the initiation of antigen‐specific primary immune responses. Recently, the analysis of gene‐knockout mice has shed light on the signaling pathways, cellular requirements, and molecular mechanisms involved in secondary lymphoid organ development. In particular, signals that converge on the nuclear factor‐κB (NF‐κB) pathway have been demonstrated to play an important role in both early developmental steps as well as maintenance of secondary lymphoid organ structures. Analysis of the histopathological changes in secondary lymphoid tissues of mice lacking individual Rel/NF‐κB family members, upstream kinases, and receptors strongly indicates that activation of the recently described alternative NF‐κB pathway by membrane‐bound lymphotoxin, via p52–RelB heterodimers, plays a major role during initiation steps of secondary lymphoid organ development. Induction of the classical p50–RelA NF‐κB activity, as exemplified by tumor necrosis factor receptor signaling, clearly also contributes, but seems to be involved primarily in later developmental step, such as the proper cellular and structural organization of B‐cell follicles.     

Identification of CXCL13 as a new marker for follicular dendritic cell sarcoma

The homeostatic chemokine CXCL13 is preferentially produced in B-follicles and is crucial in the lymphoid organ development by attracting B-lymphocytes that express its selective receptor CXCR5. Follicular dendritic cells (FDCs) have been identified as the main cellular source of this chemokine in lymphoid organs. Recently, genome-wide approaches have suggested follicular CD4 T-helper cells (T(H)F) as additional CXCL13 producers in the germinal centre and the neoplastic counterpart of T(H)F (CD4+ tumour T-cells in angioimmunoblastic T-cell lymphoma) retains the capability of producing this chemokine. In contrast, no data are available on CXCL13 expression on FDC sarcoma (FDC-S) cells. By using multiple approaches, we investigated the expression of CXCL13 at mRNA and protein level in reactive and neoplastic FDCs. In reactive lymph nodes and tonsils, CXCL13 protein is mainly expressed by a subset of FDCs in B-cell follicles. CXCL13 is maintained during FDC transformation, since both dysplastic FDCs from 13 cases of Castleman's disease and neoplastic FDCs from ten cases of FDC-S strongly and diffusely express this chemokine. This observation was confirmed at mRNA level by using RT-PCR and in situ hybridization. Of note, no CXCL13 reactivity was observed in a cohort of epithelial and mesenchymal neoplasms potentially mimicking FDC-S. FDC-S are commonly associated with a dense intratumoural inflammatory infiltrate and immunohistochemistry showed that these lymphocytes express the CXCL13 receptor CXCR5 and are mainly of mantle zone B-cell derivation (IgD+ and TCL1+). In conclusion, this study demonstrates that CXCL13 is produced by dysplastic and neoplastic FDCs and can be instrumental in recruiting intratumoural CXCR5+ lymphocytes. In addition to the potential biological relevance of this expression, the use of reagents directed against CXCL13 can be useful to properly identify the origin of spindle cell and epithelioid neoplasms.     

CD4+ T cells of both the naive and the memory phenotype enter rat lymph nodes and Peyer's patches via high endothelial venules: Within the tissue their migratory behavior differs

It is thought that naive T cells predominantly enter lymphoid organs such as lymph nodes (LN) and Peyer's patches (PP) via high endothelial venules (HEV), whereas memory T cells migrate mainly into non-lymphoid organs. However, direct evidence for the existence of these distinct migration pathways in vivo is incomplete, and nothing is known about their migration through the different compartments of lymphoid organs. Such knowledge would be of considerable interest for understanding T cell memory in vivo. In the present study we separated naive and memory CD4⁺ T cells from the rat thoracic duct according to the expression of the high and low molecular weight isoforms of CD45R, respectively. At various time points after injection into congenic animals, these cells were identified by quantitative immunohistology in HEV, and T and B cell areas of different LN and PP. Three major findings emerged. First, both naive and memory CD4⁺ T cells enter lymphoid organs via the HEV in comparable numbers. Second, naive and memory CD4⁺ T cells migrate into the B cell area, although in small numbers and continuously enter established germinal centers (GC) with a bias for memory CD4⁺ T cells. Third, memory CD4⁺ T cells migrate faster through the T cell area of lymphoid organs than naive CD4⁺ T cells. Thus, our study shows that memory CD4⁺ T cells are not excluded from the HEV route. In addition, “memory” might depend in part on the ability of T cells to specifically enter the B cell area and GC and to screen large quantities of lymphoid tissues in a short time.     

The immune system in chickens

The most sophisticated feature of the immune system expressed in vertebrates is recognition of foreign molecules by distinct types of immunocompetent cells. Birds are the first vertebrates in which a clear dichotomy of the lymphoid system has been established: 1. Thymus-derived (T) lymphocytes, the effector cells in cell-mediated (immunity and 2. Bursa-derived (B) lymphocytes, the precursor cells of the antibody-synthesizing plasma cell. Lymphocyte differentiation begins with the migration of haemopoietic stem cells from yolk sac and liver into bursa and thymus early in embryonic life followed by clonal expansion within the central lymphoid tissues. The second stage of differentiation is considered to begin with the migration of lymphocytes to the peripheral lymphoid tissue (spleen and lymph nodes). Based on genetic information, B cells are capable of recognizing foreign antigens by specific immunoglobulin molecules whereas T cell receptors are presumed to be products of the immune response genes. Surface differences provide the basis for analyzing the population dynamics of T and B cells and the pathway of immunologic diseases as well. There is compelling evidence that antigen entering the body stimulates a conventional type of systemic immune response. Antigen which remains in the mucosa, however is apt to induce a local type of response. The responses include B cells, which remain in the peripheral lymphoid tissues for the most part, and secrete immunoglobulins of different classes (IgM, IgG, IgA). T cells, however, are represented by the circulating pool of lymphocytes, and do not synthesize antibodies but instead release various mediators upon interaction with the antigen, which play a role in cell-mediated immunity. The antibody response to most antigens requires cooperation between T and B cells and macrophages as well, but in some instances T cells can also suppress B cell function. There is some evidence that the immune response of chickens is genetically controlled, which is particularly pertinent to susceptibility and resistance to diseases. Since humoral and cell-mediated immunity can separately be affected by removal of central lymphoid organs, chickens may serve as a useful model to elucidate the function of the immune system in health and disease.     

Human decidual stromal cells secrete C-X-C motif chemokine 13, express B cell-activating factor and rescue B lymphocytes from apoptosis: distinctive characteristics of follicular dendritic cells

BACKGROUND Decidual stromal cells (DSCs) have classically been considered fibroblastic cells, although their function, cell lineage and origin are not fully understood. We previously demonstrated that human DSCs showed similarities with follicular dendritic cells (FDCs): DSCs expressed FDC-associated antigens, both types of cells are contractile and both are related to mesenchymal stem cells (MSCs). To further characterize DSCs, we investigated whether DSCs and FDCs share any distinctive phenotypical and functional characteristics. METHODS Human FDC lines were obtained from tonsillectomy samples, human DSC lines from elective termination of pregnancy samples and human MSC lines from bone marrow aspirates. We isolated DSC, FDC and MSC lines and compared their characteristics with flow cytometry and enzyme-linked immunosorbent assay. Cell lines were cultured with tumour necrosis factor (TNF) and lymphotoxin (LT)α(1)β(2), cytokines involved in FDC differentiation. Cell lines were also differentiated in culture after exposure to progesterone and cAMP, factors involved in the differentiation (decidualization) of DSC. RESULTS Like MSCs, DSCs and FDCs expressed MSC-associated antigens (CD10, CD29, CD54, CD73, CD106, α-smooth muscle actin and STRO-1) and lacked CD45 expression, and all three types of cell line showed increased expression of CD54 (ICAM-1) and CD106 (VCAM-1) when cultured TNF and LTα(1)β(2). DSCs and FDCs, however, exhibited characteristics not observed in MSCs: DSCs expressed FDC-associated antigens CD14, CD21 and CD23, B cell-activating factor and secreted C-X-C motif chemokine 13. Moreover, DSC lines but not MSC lines inhibited the spontaneous apoptosis of B lymphocytes, a typical functional attribute of FDC. During culture with progesterone and cAMP, FDCs, like DSCs but in contrast to MSCs, changed their morphology from a fibroblastic to a rounder shape, and cells secreted prolactin. CONCLUSIONS Our results suggest that DSCs and FDCs share a common precursor in MSCs but this precursor acquires new capacities when it homes to peripheral tissues. We discuss these shared properties in the context of immune-endocrine regulation during pregnancy.     

Tissue localization and retention of antigen in relation to the immune response

Antigen persists for months or even years in lymphoid tissues of immune animals and this antigen is believed to participate in the induction and maintenance of B-cell memory as well as in the maintenance of serum antibody levels. In the present report we describe the phenomenon of antigen localization and long-term retention on mouse follicular dendritic cells (FDCs). The antigens used were injected in the hind footpads of immune mice and the popliteal lymph nodes were the lymphoid organs generally studied. In addition to presenting the morphological features of mouse FDCs, we report the results of a study of the mechanism of antigen migration from the site of initial localization in the lymph node subcapsular sinus to the regions of follicular retention in the cortex. The migration was followed by light and electron microscopy. The results support the concepts that immune complexes are trapped in the subcapsular sinus and are transported by a group of nonphagocytic cells to follicular regions. The mechanism of transport may involve either migration of pre-FDCs with a concomitant maturation into FDCs, or cell-to-cell transport utilizing dendritic cell processes and membrane fluidity; or a combination of the two mechanisms may be in operation.     

Sequence analysis or rat integrin alphaE1 and alphaE2 subunits: tissue expression reveals phenotypic similarities between intraepithelial lymphocytes and dendritic cells in lymph

The integrin alphaOX-62 subunit is defined by the OX-62 monoclonal antibody that was raised against rat dendritic cells in lymph (veiled cells) and shows properties similar to those of human alphaE that is predominantly expressed on intraepithelial lymphocytes. To clone alphaOX-62, rat probes generated using primers specific for the human alphaE sequence were used to screen rat T cell cDNA libraries. cDNA clones encoding two similar but not identical alpha subunits that are closely related but distinct from human alphaE were isolated. alphaE1 is predicted to be the rat homolog of mouse alphaM290 and alphaE2 corresponds to rat alphaOX-62. Immunofluorescence analysis revealed that mouse alphaE1 and rat alphaE2 are expressed in dendritic epidermal T cells in the skin, intraepithelial lymphocytes in the small intestine and in cells with a dendritic morphology present at sites where gammadelta T cells occur in lymphoid organs. Unexpectedly, in veiled cells alphaE2 is co-expressed with intracellular CD3-delta and a 33-kDa CD3 chain but not the T cell receptor. These findings suggest that veiled cells may be derived from a lymphoid precursor. Furthermore, veiled cells show phenotypic similarities to intraepithelial lymphocytes.     

Selective attraction of naive and memory B cells by dendritic cells

In this study, we investigate whether dendritic cells (DC), known to interact directly with T and B cells, might also contribute to the recruitment of B cells through the production of chemotactic factors. We found that B cells responded to several chemokines (CXCL12, CCL19, CCL20, and CCL21), which can be produced by DC upon activation. In addition, supernatant from DC (SNDC) potently and selectively attracted naive and memory B cells but not germinal center (GC) B cells or other lymphocytes (CD4(+), CD8(+) T cells or NK cells). Production of this activity was restricted to DC and was not increased following DC activation by LPS or CD40 ligand. Surprisingly, the B-cell chemotactic response to SNDC was insensitive to pertussis toxin treatment. In addition, the chemotactic factor(s) appeared resistant to protease digestion and highly sensitive to heat. This suggested that the DC chemotactic factor(s) is different from classical chemoattractants and does not involve G(alpha(i)) proteins on the responding B lymphocytes. It is interesting that SNDC was able to synergize with several chemokines to induce massive migration of B lymphocytes. These observations show that DC spontaneously produce factors that, alone or in cooperation with chemokines, specifically regulate B-cell migration, suggesting a key role of DC in the recruitment or localization of B lymphocytes within secondary lymphoid organs.