The paracortical area in dermatopathic lymphadenitis and other reactive conditions of the lymph node

We compared the paracortical area in 4 cases of dermatopathic lymphadenitis (DL) with the same area in 11 cases of various other reactive conditions of the lymph node by immuno- and enzymehistochemical techniques. In addition, electron microscopy was performed on three cases of DL. The paracortical nodules in DL proved to be composed of a variable number of dendritic, OKT6+ OKIa + ATPase+ cells, admixed with helper T-lymphocytes. All other lymph nodes studied lacked dendritic OKT6+ cells, whereas OKIa positivity was found in the cortical (follicular centers and mantle zones) and paracortical area (lymphocytes and scattered dendritic cells). Short incubation for ATPase revealed a paracortical, pericellular staining pattern in cases of DL, whereas in all other cases this staining pattern was observed only after long incubation times. On electron microscopy, three types of dendritic cells were found in DL, namely interdigitating reticulum cells ( IDRC ). Langerhans cells (LC) and macrophages. Intermediate forms between IDRC and LC, containing a few Birbeck granules and a well developed rough endoplasmic reticulum, were found. It is suggested that immunoreactivity for the monoclonal antibody OKT6 is restricted to cases of DL, and is due to the appearance of dendritic cells that have LC-characteristics. These cells either arrive from the skin along afferent lymph vessels, or are the result of a local transformation process of IDRC that acquire LC-characteristics, i.e. OKT6 immunoreactivity and Birbeck granules.     

Langerhans cells in the lymph node: mirror section and immunoelectron microscopic studies

Cells immunostained with antibodies against both OKT-6 and S-100 protein were observed only in superficial and hilar lymph nodes draining tissues with predominantly squamous epithelia. In contrast, in mesenteric lymph nodes and the spleen, only S-100 protein-positive, but OKT-6-negative cells were found. We suspect that the S-100 and OKT-6-positive cells might be Langerhans cells (LC) and the S-100-positive, OKT-6-negative cells, interdigitating reticulum cells (IDC). We further postulate that the LC in superficial and hilar lymph nodes might migrate from squamous epithelia, with which contact is required for the formation of Birbeck granules.     

Langerhans cells in the lymph node: mirror section and immunoelectron microscopic studies.

Cells immunostained with antibodies against both OKT-6 and S-100 protein were observed only in superficial and hilar lymph nodes draining tissues with predominantly squamous epithelia. In contrast, in mesenteric lymph nodes and the spleen, only S-100 protein-positive, but OKT-6-negative cells were found. We suspect that the S-100 and OKT-6-positive cells might be Langerhans cells (LC) and the S-100-positive, OKT-6-negative cells, interdigitating reticulum cells (IDC). We further postulate that the LC in superficial and hilar lymph nodes might migrate from squamous epithelia, with which contact is required for the formation of Birbeck granules.     

Lymph node accessory cells in the immune response

Summary The immune response in the rat parathymic lymph node was studied after administration of antigen into the peritoneal cavity. Special attention was paid to the accessory cells,which might induce the response. During the induction phase of the response a heterogeneous population of non-lymphoid mononuclear cells was present in the subcapsular sinus and the cortex of the node. These cells resemble veiled cells described in skin draining lymph and interdigitating cells in the paracortex of skin draining lymph nodes, but they do not contain Birbeck granules. It is concluded that the appearance of these granules depends on the site of the exudate provocation and that the presence of the organelles in these accessory cells is not obligatory for lymphocyte stimulation.     

Structure of the non-lymphoid cells during the postnatal development of the rat lymph nodes

This study describes the postnatal development of the nonlymphoid cells with special reference to the fibroblastic reticulum cells (FRCs) and interdigitating cells (IDCs). The first lymphocytes of the neonatal lymph nodes are located in the developing deep cortex units (DCUs) identified by the Gomori's technique for reticulin fibres. Ultrastructural studies demonstrate that FRCs form the stroma of the DCUs. By light and electron microscopy, it is demonstrated that FRCs occupy the outer cortex in the following stages of development of the lymph nodes. Thus, FRCs form the stroma of the primary follicles and, later, are transformed in follicular dendritic cells (FDCs) of the germinal centres. Immature or pro-IDCs appear as migrating elements in the deep cortex of lymph nodes of the neonatal rats. The ultrastructure of the pro-IDCs resembles that of the mature IDCs but not that of the phagocytic cells. Pro-IDCs are transformed into mature IDCs whose cytoplasmic expansions contact lymphocytes via tight junctions. Some of these lymphocytes are likely apposed to FRCs of the DCUs. No cells containing Birbeck granules were found in the parenchyma of the lymph nodes during the postnatal development. The role of these nonlymphoid cells is discussed with respect to the immunologic function of mammalian lymph nodes.     

The paracortical area in reactive lymph nodes demonstrating sinushistiocytosis

The enzyme and immunohistochemical features of lymphnodes showing sinus histiocytosis have been studied. Sinus histiocytes with phenotype OKM1+ OKT4+ Leu3a+ To5+ OKIal- showed strong acid phosphatase and non-specific esterase, weak endogenous peroxidase and no ATPase activities. In nine out of ten lymph nodes, paracortical collections of dendritic OKT6+ OKIal+ cells were observed. In two of the four cases studied these dendritic cells showed strong ATPase activity. We suggest that the dendritic OKT6+ OKIal+ ATPase+ interfollicular cells represent newly arrived veiled cells (VC) which have entered the lymph node by the afferent lymph, settled in the interfollicular area and are probably involved in the induction of a cellular immune response. OKT6+ OKIal+ ATPase+ VC may subsequently transform into mature, OKT6- OKIal+ ATPase+ interdigitating reticulum cells which are involved in the negative feedback of the cellular immune response. The association with sinus histiocytosis is probably related to the fact that an increase in mononuclear phagocytes in the afferent lymph is accompanied by a relative increase in VC. Our results demonstrate that in lymph nodes showing sinus histiocytosis, two cell types increase in number, i.e. an Ia- sinusoidal cell, engaged in phagocytosis of foreign material, and an Ia+ dendritic cell in the interfollicular area, probably involved in the induction of a cellular immune response.     

Langerhans cell-like dendritic cells in the cornea,tongue and oesophagus of the chicken (Gallus gallus)

Langerhans cells are dendritic leucocytes which reside mainly within stratified squamous epithelia of skin and mucosa. Their visualization requires the use of ATPase histochemistry, electron microscopy for identifying the unique trilaminar cytoplasmic organelles (the Langerhans cell granules or Birbeck granules), and the expression of major histocompatibility complex class II molecules. Following uptake of antigen, Langerhans cells migrate via the afferent lymphatics to the lymph nodes and undergo differentiation from an antigen-processing cell to an antigen-presenting cell. Using the same approach as that employed in previous studies for the identification of chicken epidermal Langerhans cells, we show here the presence of ATPase-positive and major histocompatibility complex class II-positive Langerhans cell-like dendritic cells at the mucosal surface of the eye, tongue and oesophagus of the chicken. Ultrastructurally, these cells qualified as Langerhans cells except that they lack Langerhans cell granules. Thus, as in mammalian skin and mucosa, chicken mucosa contains mucosal dendritic cells with morphological and phenotypical features for the engagement of incoming antigens within epithelium and lamina propria.     

Differentiation of Langerhans Cells from Interdigitating Cells Using CD1a and S-100 Protein Antibodies

The present study shows that Langerhans cells can be differentiated from Interdigitating cells at the light microscopic level. Superficial lymph nodes and skin taken from necropsies and the lymph nodes of dermatopathic lymphadenopathy (DPL) were used for this experiment. Sections of lymph node and skin were embedded using the acetone, methyl benzoate and xylene (AMeX) method and dendritic cells were immunostained with anti S-100 protein antibody (S-100, and OKT-6 (CD1a) using the restaining method. Langerhans cells in the skin were positive for both CD1a and S-100. Dendritic cells positive for both CD1a and S-100, and dendritic cells positive for S-100, but not for CD1a were observed in superficial lymph nodes. In normal superficial lymph nodes, there were more interdigitating cells than Langerhans cells. The majority of the dendritic cells in the DPL were Langerhans cells. We conclude that the S-100 and CD1a positive cells are Langerhans cells, and the S-100 positive-CD1a negative cells are interdigitating cells.     

Differentiation of Langerhans Cells from Interdigitating Cells Using CD1a and S-100 Protein Antibodies

The present study shows that Langerhans cells can be differentiated from Interdigitating cells at the light microscopic level. Superficial lymph nodes and skin taken from necropsies and the lymph nodes of dermatopathic lymphadenopathy (DPL) were used for this experiment. Sections of lymph node and skin were embedded using the acetone, methyl benzoate and xylene (AMeX) method and dendritic cells were immunostained with anti S-100 protein antibody (S-100, and OKT-6 (CD1a) using the restaining method. Langerhans cells in the skin were positive for both CD1a and S-100. Dendritic cells positive for both CD1a and S-100, and dendritic cells positive for S-100, but not for CD1a were observed in superficial lymph nodes. In normal superficial lymph nodes, there were more interdigitating cells than Langerhans cells. The majority of the dendritic cells in the DPL were Langerhans cells. We conclude that the S-100 and CD1a positive cells are Langerhans cells, and the S-100 positive-CD1a negative cells are interdigitating cells.     

The ultrastructure of the abdominal lymph nodes cortex in rats during primary and secondary immune response (a special reference to dendritic reticulum cells and interdigitating cells)

The stimulation of coeliac rat lymph nodes was performed by intraperitoneal injections of typhoid vaccine and was unique for the primary immune response and repeated after 6 weeks for the secondary response. The light and electron microscopic observations showed for the primary response, an early germinal center reaction, which might be accounted for by a background of continuous stimulation of the coeliac nodes, stemming from the digestive tract. The dendritic reticulum cells (DRC), considered typical for the B area, were located at the borderline between the germinal center and the mantle zone. Their cytoplasmic extensions penetrated the lymphocyte-lymphoblastic center, surrounding most of the germinal center cells. The marginal zone and the paracortex reacted as a whole, the interdigitating cells (IDC) being the dominant feature. An explanation would be that the marginal zone can be penetrated by T cells and connected IDCs, thus, the B and T areas seem to be largely interspersed. The results suggest that IDCs are cells of direct monocytic origin.     

Role of tumor cell apoptosis in tumor antigen migration to the draining lymph nodes

Establishment of an immune response against cancer may depend on the capacity of dendritic cells to transfer tumor Ags into T cell-rich areas. To check this possibility, we used a colon cancer cell variant that yields tumors undergoing complete T cell-dependent rejection when injected into syngeneic rats. We previously demonstrated that immunogenicity of these tumors depended on the early apoptosis of a part of these tumor cells. In this paper we show that fluorescent tumor cell proteins are released from FITC-labeled tumor cells and undergo engulfment by tumor-infiltrating monocytes without a phenotype of mature dendritic cells or macrophages. Fluorescence-labeled mononuclear cells with a phenotype of MHC class II+ dendritic cells are also found in the T cell areas of the draining lymph nodes. Interestingly, no fluorescent cell can be found in lymph nodes after a s.c. injection of Bcl2-transfected apoptosis-resistant tumor cells that yielded progressive tumors. Proliferation of tumor-immune T lymphocytes was induced by dendritic cells isolated from the draining lymph nodes recovered after a s.c. injection of apoptosis-sensitive, but not apoptosis-resistant, tumor cells. These results show that tumor cell apoptosis releases proteins that are engulfed by inflammatory cells in the tumor, then transported to lymph node T cell areas where they can induce a specific immune response leading to tumor rejection.     

Evidence that cutaneous antigen-presenting cells migrate to regional lymph nodes during contact sensitization

These studies address the hypothesis that Ag-bearing epidermal Langerhans cells migrate to the regional lymph node during contact sensitization and function as APC. Skin from C3H mice was grafted onto BALB/c nude mice, and 7 or 14 days later, the recipients were sensitized with FITC through the grafts. APC from lymph nodes draining the site of sensitization were capable of sensitizing C3H recipients to FITC. Because sensitization is MHC restricted, only cells reaching the lymph node from the grafted skin could have induced contact hypersensitivity in C3H mice. Examination of the FITC+ draining lymph node cells by immunofluorescence and immunoelectron microscopy demonstrated that all were Ia+, most were F4/80+, and some contained Birbeck granules. These studies demonstrate that Ia+, FITC+ cells from the skin, at least some of which are Langerhans cells, leave the skin after epicutaneous sensitization with FITC and participate in the initiation of the contact hypersensitivity response within the regional lymph node.     

Dynamic nature and function of epidermal Langerhans cells in vivo and in vitro: a review, with emphasis on human Langerhans cells.

Summary Epidermal Langerhans cells (LC) are Birbeck granule-containing bone-marrow-derived cells, which are located mainly in the suprabasal layer of the epidermis. They can be readily identified by their strong expression of CDIa and MHC class II molecules. In addition to these classical properties, an extensive phenotypic profile of normal human LC, summarized in this review, is now available. The powerful capacity of LC to activate T lymphocytes is clearly documented and, to date, LC are recognized as the prominent antigen-presenting cells of the skin immune system. They are generally believed to pick up antigens encountered in the epidermis and to migrate subsequently from the epidermis to the skin-draining lymph nodes. Upon arrival in the paracortex of lymph nodes, the antigen-laden LC transform into interdigitating cells and they present antigen to naive T lymphocytes in a MHC class II-restricted fashion; this results in the generation of antigen-specific immune responses. It has also been demonstrated that transformation of LC into interdigitating cells occurs when LC are culturedin vitro. Bothin vivo andin vitro studies have indicated that properties of LC, such as phenotype, morphology and the stimulatory potential to activate T lymphocytes, are dependent on the local microenvironment in which the LC reside. The essential role of LC in the induction of contact allergic skin reactions and skin transplant rejection is well established.     

Phagocytosis of erythrocytes in the subarachnoid space at spinal nerve exits

Summary Interdigitating cells (IDC) in the thymus of the spotless starling, Sturnus unicolor, were examined by electron microscopy. They occur principally in the thymic medulla and corticomedullary border. They possess an irregular nucleus and a perinuclear area of cytoplasm, containing most of the membranous organelles, surrounded by a peripheral electron-lucent zone. Clusters of smooth Golgi vesicles and complicated labyrinthine membrane-membrane contacts are the most characteristic cytological features. Birbeck granules are absent. Lymphocytes, plasma cells and even myoid cells can be found embedded in the cytoplasm. Immature elements, intermediate between epithelial-reticular cells and interdigitating cells, are tentatively identified as prointerdigitating cells. The functional significance of IDCs, and their phylogenetic significance in the vertebrate immune system, is discussed.     

Dermal dendritic cells, and not Langerhans cells, play an essential role in inducing an immune response

Langerhans cells (LCs) serve as epidermal sentinels of the adaptive immune system. Conventional wisdom suggests that LCs encounter Ag in the skin and then migrate to the draining lymph nodes, where the Ag is presented to T cells, thus initiating an immune response. Platelet-activating factor (PAF) is a phospholipid mediator with potent biological effects. During inflammation, PAF mediates recruitment of leukocytes to inflammatory sites. We herein tested a hypothesis that PAF induces LC migration. Applying 2,4-dinitro-1-fluorobenzene (DNFB) to wild-type mice activated LC migration. In contrast, applying DNFB to PAF receptor-deficient mice or mice injected with PAF receptor antagonists failed to induce LC migration. Moreover, after FITC application the appearance of hapten-laden LCs (FITC+, CD11c+, Langerin+) in the lymph nodes of PAF receptor-deficient mice was significantly depressed compared with that found in wild-type mice. LC chimerism indicates that the PAF receptor on keratinocytes but not LCs is responsible for LC migration. Contrary to the diminution of LC migration in PAF receptor-deficient mice, we did not observe any difference in the migration of hapten-laden dermal dendritic cells (FITC+, CD11c+, Langerin-) into the lymph nodes of PAF receptor-deficient mice. Additionally, the contact hypersensitivity response generated in wild-type or PAF receptor-deficient mice was identical. Finally, dermal dendritic cells, but not LCs isolated from the draining lymph nodes after hapten application, activated T cell proliferation. These findings suggest that LC migration may not be responsible for the generation of contact hypersensitivity and that dermal dendritic cells may play a more important role.     

The early postnatal development of the primary immune response in TNP-KLH-stimulated popliteal lymph node in the rat

Summary The popliteal lymph nodes were removed from young rats of various ages five days after a single immunization with TNP-KLH in the hind footpads. Cryostat sections of the lymph nodes were investigated by means of enzyme and immunohistochemical techniques at the light-microscopical level.The presence and localization of anti-TNP antibody-containing cells were examined using a new technique to visualize specific antibodies. Moreover, the development of the lymph nodes following exogenous antigenic stimulation was compared with that of unstimulated lymph nodes.Specific antibody-containing cells could not be found before day 15 after birth, in rats immunized at day 10. From that time these lymphoid cells were located primarily at the border between cortex and medulla. Younger popliteal lymph nodes showed only aspecific immunoglobulin-containing lymphoid cells. With age, the number of specific antibody-containing cells tended to increase. These cells were more mature, according to morphological criteria and were located nearer the medulla.The first primary follicles were seen at day 19, as was the case in unstimulated animals. The first secondary follicles, containing germinal centers, were detected at day 23, whereas in unstimulated popliteal lymph nodes they were never found.Trapping of immune complexes could not be demonstrated before day 33 after birth. The later appearance of this phenomenon might be a consequence of the techniques applied to demonstrate specific antibody-containing cells.Abbreviations PLN popliteal lymph node - FDC follicular dendritic cell - IDC interdigitating cell - HEV high endothelial venule - TNP trinitrophenyl - KLH keyhole limpet hemocyanin - PBS phosphate-buffered saline - GCPC germinal center precursor cell - sIg surface immunoglobulin - cIg cytoplasmic immunoglobulin     

Lymph node macrophages and reticulum cells in the immune response

Summary The morphology and kinetics of macrophages and reticulum cells of rat lymph nodes have been studied in relation to the immune response to a second exposure to antigen. During the first 24 h after stimulation monocyte-like exudate macrophages, including some scattered interdigitating cells (IDC), contain granules similar to those present in epidermal Langerhans cells and lymph-borne veiled cells. In this induction phase these macrophages migrate from the marginal sinus into the paracortex and during the migration they gradually transform into IDC. In the proliferation phase the paracortex is mainly populated by transitional macrophages and there are almost no typical IDC present between the lymphoblasts. In the memory phase the relative number of IDC again rapidly increases. During this period in the paracortex there are often typical IDC which contain partially digested necrotic lymphocytes, thus resembling tingible body macrophages (TBM) of the germinal centre in this respect.It is suggested that the newly arrived macrophages induce the lymphoblast reaction, while mature IDC may have an inhibitory function in the memory phase of the immune response. In this phase the phagocytic potential of IDC is clearly shown.     

Initial lentivirus-host interactions within lymph nodes: a study of maedi-visna virus infection in sheep.

Reactive changes occurring within lymph nodes draining the subcutaneous site of acute infection with maedi-visna virus (MVV) were studied, and the appearance of infected cells correlated with the immune response. Cells infected with virus were detected in the node by cocultivation from day 4 postinfection (p.i.), with maximum numbers being seen between days 7 and 14, but even then infected cells were rare, with a maximum frequency of 23 50% tissue culture infective doses (TCID50) in 10(6) lymph node cells. At later times, infected cells were still detected, but their numbers fell to 1 to 2 TCID50 per 10(6) cells. Virus-specific CD8+ cytotoxic T-cell precursors (CTLp) were isolated from infected nodes from day 10 p.i. onwards, and T-cell proliferative responses to MVV were first detected on day 7 and consistently detected after day 18. Histological analysis showed a vigorous immune response in the node. There was a marked blast reaction in the T-cell-rich zones, which was greatest at the time when the number of virally infected cells was at its height. At this stage, large numbers of plasma cells were seen in the medullary cords, indicating that extensive T-cell-dependent B-cell activation was occurring in the T-cell-rich zones. Germinal centers were prominent shortly after the onset of the T-zone response and were still present at 40 days p.i. Phenotype studies of isolated lymph node cells failed to detect major changes in the proportion or phenotype of macrophages, CD1+ interdigitating cells, and CD4+ or CD8+ T cells despite the fact that CD8+ lymphoblasts form a major population leaving the node in efferent lymph. This suggests that there is a balanced increase in the number of all cell types in response to the virus within the node and selective migration of CD8+ lymphoblasts containing virus-specific CTLp from the node. Virus-specific immune responses are therefore present within the node when infectious virus isolation is maximal, but cellular immunity may act to control the level of infection from day 18 onwards.     

The role of dendritic cells in the initiation of immune responses to contact sensitizers. I. In vivo exposure to antigen

Twenty-four hours after skin painting mice with picryl chloride (PIC) there was a four- to fivefold increase in the numbers of dendritic cells (DC) isolated from the lymph nodes. These DC initiated primary proliferative and cytotoxic responses when added to cultures of normal syngeneic lymph node cells. The proliferative response was enhanced when the donors of the responding lymph node cells were sensitized with the same antigen. Contact sensitivity developed in syngeneic mice injected into the footpads with 30,000-50,000 DC from lymph nodes of mice painted with picryl chloride 1 day previously. Thus, 1 day after skin painting mice, there were dendritic cells in the draining lymph nodes which were able both to initiate primary stimulation of lymphocytes in vitro and to sensitize recipient mice to give specific delayed hypersensitivity reactions.     

Avian epidermis contains ATPase- and Ia-positive Langerhans-like cells

Summary The occurrence of cells resembling mammalian Langerhans cells in the avian epidermis was studied by ATPase histochemistry, Ia immunoreactivity and electron microscopy. The existence of MHC class II antigen-(Ia) expressing, ATPase-positive dendritic cells, which are ultrastructurally similar to mammalian Langerhans cells except for the absence of Birbeck granules, was demonstrated. These cells may be a basic component of the immune system of birds.