Localization in Mouse Lymphoid Tissue of Receptors for Immunoglobulin

Cryostat sections of mouse spleen and lymph nodes adsorbed sheep erythrocytes coated with rabbit or mouse IgG antibody. The reaction was mediated by the Fe part of the immunoglobulin molecule. Haemadsorption was inhibited by mouse IgG and antigen-antibody complexes. Complement was not required.
Sensitized erythrocytes failed to bind to thymus and to the thymus dependent areas of the peripheral lymphoid tissue. Results of Inhibition experiments with rabbit anti-mouse lymphocyte serum and anti-Θ serum further suggested that binding of sensitized erythrocytes to lymphoid tissue sections can be used as a marker for B lymphocytes.     

Fate of Autogenous Canine Lymphocytes after Injection into an Afferent Lymphatic of the Popliteal Lymph Node

Dog thoracic duct lymphocytes were labeled in vitro with ³H-uridine and infused into an afferent lymphatic of the popliteal lymph node of the same dog. Ten minutes after infusion nearly all the injected radioactivity was recovered from the lymph node. An effect of infusion flow rate on the percentage of cells retained by the lymph node was observed ½ to 3½ hours after infusion, and was probably mediated by the tendency of the node to become edematous after infusions at a rate exceeding 0.045 ml/min. Edematous nodes retained 83.7% of the cells, as compared to 47.5% for nonedematous nodes. As early as 30 minutes after infusion a small amount of ³H-radioactivity was found in the spleen and thoracic duct lymph. The deep iliac and paraaortic nodes on the side of the infusion contained significant amounts of ³H-radioactivity, while negligible amounts were detected in the contralateral popliteal node at any time. The intranodal localization of the ³H-labeled cells was studied by radioautography. All labeled cells remained intrasinusoidal during the first 4 hours after infusion. At 9 and 21 hours some labeled cells were located in the extrasinusoidal parenchymal lymphoid tissue of the cortex and the medulla, but the majority still remained intrasinusoidal.     

Avian lymph node: Light and electron microscopic study

We have observed by light and transmission electron microscopy lymphoid accumulations (LA) in the chicken located along the posterior tibial-popliteal and lower femoral veins. Three types of LA were characterized: (1) LA on the wall of the lymphatic, (2) LA with germinal center, and (3) well-developed LA possessing germinal centers and an intricate lymphatic sinus system. The latter will be termed a lymph node and is perhaps the structure responding to foot-pad injection of antigen and/or phytohemagglutinin (PHA). After the injection of PHA into the foot-pad, the lymph node enlarged and revealed the intermingling of two distinct groups of cells consisting of either small lymphocytes or medium lymphocytes and lymphoblasts. Because our earlier immunological paper proved the presence of T and B cells in the node, the two histologically distinct groups of cells appearing after PHA injection could reflect compartmentalization of T and B cells in the avian lymph node. Lymphoid and adipose tissues are in the same compartment. After PHA or antigen injection into the foot pad, the lymphoid tissue proliferates and the amount of the adipose tissue rapidly decreases. This suggests that lymphoid and adipose tissue form a special complex which is separated from the surrounding tissue by delicate connective tissue capsule. The relationship of the lymphoid and adipose tissue is comparable with that of myeloid and adipose tissue in the bone marrow. The majority of the sinuses shows smooth endothelial lining while others contain “hairy” macrophages attached to the endothelium. The germinal centers are located at the periphery of the node, but a few occur inside. The cellular content of the germinal centers is not unusual except for the presence of plasma cells.     

Immunology of the lower respiratory tract. Serial morphologic changes in the lungs and tracheobronchial lymph nodes of dogs after intrapulmonary immunization with sheep erythrocytes.

Sequential histologic changes in immunized and contralateral lung lobes, and ipsilateral and contralateral tracheobronchial lymph nodes were evaluated from dogs after intra-pulmonary immunization with sheep erythrocytes. In addition, changes in the relative numbers of unstimulated lymphocytes, activated lymphocytes, and plasma cells were determined for immunized and contralateral lung lobes by the use of bronchopulmonary lavage. Histologic evidence of an immune response occurred 2 days after immunization in ipsilateral tracheobronchial lymph nodes and on Day 5 in immunized lung lobes. Pulmonary lymphoid infiltrates appeared initially around pulmonary venules and veins. There was expansion of these infiltrates into alveolar spaces, where mixed mononuclear aggregates were formed in association with alveolar macrophages 7 days after immunization. A similar but attenuated and delayed response occurred within contralateral control lung lobes, although mononuclear aggregates were not found. Activated lymphocytes in lavage samples increased prior to histologic evidence of pulmonary lymphoid infiltrates. These results suggest that after intrapulmonary immunization the lung recruits circulating immunocytes produced in ipsilateral lung-associated lymph nodes as the source of specific antibody-forming cells in bronchoalveolar air spaces.     

Cellular Reaction in the Footpad and Draining Lymph Nodes of Mice Induced by Mycobacterial Fractions and BCG Bacilli

Ten micrograms of trehalose-6, 6'-dimycolate (cord factor), injected into the footpad of mice, induced histological changes similar to those following injection of living BCG bacilli. Both materials induced in the draining lymph nodes the formation of granulomas composed of epitheloid cells, macrophages, and small numbers of lymphocytes. Apart from the granulomatous inflammatory process, marked hyperplasia of the lymphoid tissue in the paracortical zone of the nodes and accumulations of macrophages were evident. In some cases, the macrophages were very numerous and replaced part of the lymphoid tissue. Compared to cord factor, wax D showed weak granulomagenic activity. Only slight and transient inflammation was found in the footpads as well as transient slight lymphoid hyperplasia. Wax D also induced small accumulations of macrophages. Complete Freund's adjuvant induced, under the same experimental conditions, large accumulations of macrophages in the draining lymphnode and lymphoid hyperplasia in the paracortical zone. No cellular reaction was seen in the liver, spleen, and lungs after injection of cord factor and BCG into the footpads of the animals. The results and implications are discussed.     

Spontaneous infarction of superficial lymph nodes

Five cases of extensive infarction of lymph nodes were traced in just over 16 years' surgical material. All presented with painful swelling in a superficial lymph node chain. None was diagnosed clinically; two were interpreted as fibroadenoma of the axillary tail of the breast, and two as a femoral hernia. Microscopically the lymph nodes in the first three weeks after infarction were characterized by extensive necrosis of medullary and cortical lymphoid cells, but the central reticulin architecture and a narrow, incomplete rim of viable subcapsular lymphoid tissue were preserved. Reactive perinodal inflammation and the formation of granulation tissue resembled the reaction to myocardial infarction. The late stage of the lesion was characterized by incomplete regeneration of lymphoid tissue in the lymph nodes. The lesions appeared attributable to thrombosis of veins within the substance and the hila of the nodes.     

Development of lymphoid tissues of the stripe-faced dunnart (Sminthopsis macroura)

The development of the lymphoid tissues of a model marsupial, the stripe-faced dunnart, has been described from birth to weaning, a period of 2.5 months. At birth the lymphoid tissues, including the thymus, lymph nodes and mucosa-associated lymphoid tissues, were undeveloped. A thoracic thymus consisting primarily of stromal tissue was observed by day 4 after birth but by day 12, lymphocytes were observed in the thymus and some cortico-medullary differentiation was apparent. Lymph nodes were histologically mature by day 31, the earliest day investigated for this tissue. In gut tissue, lymphoid follicles were first observed by day 57 post-partum. No bronchus-associated lymphoid tissue was observed in any lung samples. The thymus, lymph nodes and gut-associated lymphoid tissues were all distinguishable before weaning (day 70) but not all were histologically mature. The sequence of development of the lymphoid tissues in the stripe-faced dunnart was similar to those observed in other marsupial species.     

Immunology of the lower respiratory tract. III. Concentrations of antigen and of antibody-forming cells in pulmonary and systemic lymphoid tissues of dogs after intrapulmonary or intravenous administration of sheep erythrocytes

The distribution of antibody-forming cells to sheep erythrocytes among canine pulmonary and systemic lymphoid tissues differs distinctively depending on whether antigen is administered by the intrapulmonary or the intravenous (i.v.) route. After local (intrapulmonary) immunization, antibody-forming cells are restricted to the lung and regional lymph nodes; after i.v., they are widespread. To test the hypothesis that the tissue distribution of antigen is an important determinant of the resultant distribution of antibody-forming cells, dogs were immunized with radioiodinated sheep erythrocytes, either intrapulmonary (i.p.) or i.v. After 4 days, the concentrations of tissue-bound radioactivity and of antibody-forming cells in various lymphoid tissues were compared. The distribution of tissue-bound radioactivity among lymphoid tissues was clearly determined and different depending on the route of immunization. After i.p. administration, radioactivity was bound to lung and hilar lymph nodes; after i.v., it was found in the lung, liver, spleen and occasionally hilar nodes. Antibody-forming cells appeared in those lymphoid preparations which contained tissue-bound radioactivity. The exception was that locally applied antigen elicited antibody-forming cells in the lung poorly, despite the fact that abundant antigen remained localized to the lung. Notably, i.v. immunization resulted in the appearance of both antibody forming cells and cell-bound radioactivity in the lung. It is concluded that the distribution of antigen among tissues is a major determinant of the distinctive patterns of appearance of antibody-forming cells after different routes of immunization.     

IgA antibodies in the bile of rats. III. The role of intrathoracic lymph nodes and the migration pattern of their blast cells.

Eight weeks after rats had had their mesenteric lymph nodes (MLN) removed surgically, they were found to be still able to generate substantial titres of biliary IgA-antibodies after antigens were injected into their Peyer's patches. This suggested that systemically significant IgA production could be induced in extra-abdominal lymphoid tissue. It was found that the intrathoracic lymph nodes (ITLN) were an important source of IgA production. These nodes could be stimulated to produce biliary antibody by introducing antigen either into the peritoneal cavity or directly into the thorax. Cells forming IgA were identified in the ITLNs by haemolytic plaque assays and immunoperoxidase techniques. In spite of this, immunoblasts obtained from the ITLNs, and labelled with 125IUdR did not localize in the gut after i.v. injection to anywhere near the extent that immunoblasts from the MLN did. Instead they seemed to have a predilection for localizing in the lungs.     

Haemadsorption by Lymphoid Tissue from Normal Mice

Tissue sections of normal mouse lymphoid organs were shown to adsorb erythrocytes of sheep and chicken origin. Only the marginal zone of spleen follicles and the subcapsular and medullary sinuses of lymph nodes were re-active. Sheep and chicken erythrocytes adhered to the same areas and formed mixed clusters when tested together. Haemadsorption by tissue sections was inhibited by anti-lymphocyte serum and anti-mouse IgM. It was also abolished by acid or heat elution. Haemadsorption was probably due to cytophilic antibodies of the IgM class adsorbed to tissue macrophages.     

Cell-mediated immunity and lymphocyte populations in experimental Argentine hemorrhagic fever (Junín Virus).

Guinea pigs infected with the XJ prototype strain of Junín virus reproduce the main features of Argentine hemorrhagic fever, showing hemorrhages, leukothrombocytopenia, and focal lymphoid tissue necrosis. Viral lymphotropism is shown by the presence of viral antigens, severe cytopathic effect, and high virus titers in lymphoid organs. A pronounced depression of humoral immune response to sheep erythrocytes as well as to the virus is described. This study was carried out to determine whether cellular immune response was also modified and which cell populations were affected. Delayed hypersensitivity skin reaction to purified protein derivative was found to be markedly depressed after infection. A noticeable decrease in both percentages and absolute T lymphocyte numbers, detected by E rosettes, in spleen and lymph nodes, together with a low absolute T cell number in peripheral blood, were observed. Total cell counts in spleen, lymph nodes, and peripheral blood were also reduced. On the contrary, no modification in percentages of B lymphocytes, as measured by EAC rosettes, was found. These results indicate that cell-mediated immunity is markedly impaired in guinea pigs infected with the XJ strain of Junín virus. Its relationship with the pathogenesis of the disease is discussed.     

Distribution of plaque-forming cells in the mouse for a protein antigen. Evidence for highly active parathymic lymph nodes following intraperitoneal injection of hen lysozyme.

The distribution of plaque-forming cells (PFC) throughout the lymphoid system of CBA mice was followed with time after a primary intraperitoneal injection of hen egg white lysozyme emulsified in Freund's complete adjuvant (HEL-CFA) and after a secondary soluble injection. Throughout the primary response (predominantly IgG) and during the first week of the secondary response (exclusively IgG), the highest density of PFC was found in the draining parathymic lymph nodes, followed by the local spleen and mesenteric lymph nodes. The antibody-forming activity of the bone marrow increased as the immune response progressed, so that by the 3rd week of the secondary response this compartment provided the majority of the PFC. PFC first appeared in the accessory axillary, brachial or inguinal lymph nodes and in the thymus a few days after the secondary injection but accounted for only 1-5% of the total activity during the entire course of the secondary response. The specificity of the antibody produced in the spleen, parathymic and mesenteric lymph nodes was identical as judged by plaque inhibition by seven chemically related lysozymes which implies that these PFC were well mixed. It is postulated, therefore, that the change in distribution of PFC from an early local response to a general systemic response, and finally to a predominantly bone marrow response, was due to the migration of memory cells from the draining parathymic lymph nodes and spleen throughout the lymphoid system with an ultimate settling of the cells in the bone marrow.     

Penetration of colloidal carbon through post-capillary venules in lymph nodes and peyer's patches of the guinea-pig: a potential immunogeneic route.

Colloidal carbon injected intravascularly provided a selective marker for post-capillary venules in lymph nodes and Peyer''s patches. In the first few minutes a denticulate outline to the lumen was formed by carbon capping the high columnar epithelium and carbon particles were deeply embedded between endothelial cells. Ten minutes after injection carbon had reached the basement membrane and was lying outside this membrane 30 min later, at first free but later engulfed inside macrophages. Carbon was retained in post-capillary venules for the duration of the experiment (8 h). Discontinuities were present in the basement membane in about one-fifth of venules, and lymphocyte and macrophage penetrating the basement membrane are demonstrated. It is postulated that carbon penetrates the post-capillary venular wall by increased intraluminal hydrostatic pressure arising from contraction of muscles that surrounded the lymphoid tissue in the case of the gut, or skeletal muscle compressing lymph nodes against a bony surface in the axilla, groin or neck. Secondly, if carbon is a model for particulate antigens, then post-capillary venules provide a potential immunogenic route whereby antigens can reach lymphoid tissues from the circulation, as proposed by Burwell (1962) for transplantation antigens.     

Transmigration of titanium dioxide (TiO2) particles in rats after inhalation exposure

Titanium dioxide (TiO2) has been used extensively in the manufacturing of white pigment and has generally been regarded as a nuisance dust in animals and man. After inhalation exposure, little is known about transmigration routes and potential toxic effects of translocated particles in other organs. In order to answer these questions, rats were exposed to TiO2 by inhalation exposure at concentrations of 0, 10, 50, and 250 mg/m3 for 2 years. A few free particles were retained in the nasal and tracheobronchial epithelium without any cellular damage, but aggregates of dust-laden macrophages (dust cells) were found in the lymphoid tissue of the submucosa. Inhaled particles were mostly engulfed by alveolar macrophages and confined sharply to the alveolar duct region at 10 and 50 mg/m3, while dust cells were scattered throughout alveoli at 250 mg/m3. A fraction of the inhaled particles was retained in the membranous pneumocytes and interstitial macrophages. A dense accumulation of dust cells was found in the perivascular and peribronchial lymphoid tissue. Some dust cells entered peribronchial lymphatics or pulmonary blood vessels and the general circulation. Dust cells in the hyperplastic peribronchial lymphoid tissue were exposed directly in the luminal surface of the airways and were subsequently eliminated via airways. Massive dust deposition was observed in the tracheobronchial lymph nodes. Dust transmigration was markedly reduced in the cervical lymph nodes, and only a trace amount of dust particles was found in the mesenteric lymph nodes. Some dust cells entered either blood or lymphatic vessels in the lymph nodes and then migrated into the general circulation. The incidence of extrapulmonary dust deposition in the liver or spleen was increased in a dose-related fashion similar to the lung dust burden. Since there was no tissue response to translocated particles in the lymph nodes, spleen, or liver, potential adverse health effects appear to be negligible.     

Histology and immunohistochemistry of the gut-associated lymphoid tissue of the eastern grey kangaroo, Macropus giganteus

Mesenteric lymph nodes and gut-associated lymphoid tissue (GALT) from juvenile eastern grey kangaroos were investigated. The mesenteric nodes had a similar structure to that described for eutherian mammals. They contained distinct regions of medulla and cortex, with prominent follicles and germinal centres. Gut associated lymphoid tissue consisted of areas of submucosal follicles. These varied from areas of densely packed lymphocytes with darkly staining, prominent coronas to areas with no defined follicles. The distribution of T cells in these tissues was documented by use of species-crossreactive antibodies to the surface markers CD3 and CD5; B cells were identified by antibodies to CD79b. Within the lymph nodes T cells were located mainly in the paracortex and cortex, with limited numbers observed in the follicles; B cells were located on the marginal zone of the follicles. In GALT, T cells were located in the peripheral regions of the germinal centres of secondary follicles, while B cells were abundant in primary follicles. These observations are consistent with those made in a range of other marsupials (metatherian) and eutherian mammals and are indicative of the capacity to respond to antigens entering via the mouth.     

A monocytogenic humoral factor released after lymph node stimulation

The injection of complete Freund's adjuvant or certain other preparations directly into the cervical lymph nodes of rats is followed by a striking, prolonged monocytosis. The monocytosis may be transmitted passively with serum and is thought to be due to a hormone elaborated by appropriately stimulated lymphoid tissue and acting on the marrow or elsewhere.

It is suggested that proliferation of mononuclear phagocytes in inflammation, delayed hypersensitivity, cellular immunity and antibody production may be associated with similar stimulation of lymphoid tissue followed by hormonal release.

Animals inoculated in neonatal life with extracts of lymph nodes (a procedure that suppresses certain delayed hypersensitivity reactions) failed to developed a monocytosis when stimulated in adulthood or when given active serum from stimulated normal rats. Neonatally treated animals, however, after lymph node stimulation in adulthood do produce a serum factor capable of transmitting monocytosis to normal rats.

The results imply that the action of the monocytogenic hormone may be largely dependent on a state of specific natural hypersensitivity whose development depends on absence of the hormone from the immature lymphoid tissue present in foetal and neonatal life. It is postulated that injection of the hormone in early life suppresses the proliferation of those clones that produce the antibody on which hypersensitivity in adulthood depends. The suggestion is made that a similar mechanism may explain the ability of various other hormones to act at very low concentrations.

     

Induction of secondary and tertiary lymphoid structures in the skin

During embryogenesis a developmental program leading to the formation of lymph nodes and Peyer's patches is initiated. We now show that lymph node-like structures as well as tertiary lymphoid structures can ectopically be induced by intradermal injection of newborn lymph node-derived cells. ICAM-1/VCAM-1-expressing stromal organizers, follicular dendritic cells, lymphatic endothelium, and HEVs in these structures are of donor origin, while all hematopoietic cells are host derived. Formation depends on lymphotoxin-expressing donor cells, whereas further organization requires lymphotoxin-expressing recipient cells. While induced secondary lymphoid structures develop a normal cellular architecture, the degree of organization in tertiary structures is correlated to the immune activation status of the host. These results indicate that the cellular and molecular requirements for the establishment of lymph nodes and tertiary structures are remarkably similar and that hyperactivated lymphocytes can fulfill the role of lymphoid tissue inducer cells during inflammatory responses.     

Reciprocal lymphoid cell homing studies using wild and LPR (lymphoproliferation) mutant C57BL mice

The injection of lymphoid cells from adult lpr mice into normal and athymic congenic mice does not transfer the lpr (lymphoproliferation) syndrome. We studied whether this phenomenon could be due to abnormal homing. The lymphoid cells from lpr donors do not show a marked deficiency of migration to lymphoid organs in comparison with cells from Wild donors and a T-cell lymphoma BL/VL3. The lymphoid organs of lpr recipients do not show an intrinsic abnormality as homing sites for lymphoid cells. The data reveal some minor migration preferences: the lpr cells migrate better than Wild cells into lpr lymph nodes (including athymic lpr hosts), whereas Wild cells migrate slightly better than lpr cells into Wild lymph nodes. In spite of such minor preferences, Wild cells can efficiently migrate into lpr lymphoid organs, as well as lpr cells into Wild lymphoid organs. Thus, the lack of transfer of lymphadenopathy in Wild recipients cannot be attributed to an alteration of lpr cell homing.     

Nucleases in immunity. I. The effect of immunization on RNase and DNase activity in lymphoid tissues

The level of RNase and DNase in the spleen, lymph nodes and thymus glands of mice immunized with sheep erythrocytes was determined. Within 12 hr after immunization there was a moderate decrease in the level of specific RNase activity in the spleen. The depression persisted for several days and then returned to normal. The level of DNase activity also decreased in the spleen of immunized animals, returning to near normal levels on day 4 to 5 and increasing moderately by day 6 and 8.

RNase activity in the lymph nodes and thymus increased rapidly after immunization, reaching a peak level several fold higher than in control animals on days 2 and 6. The level of DNase activity in lymph nodes and thymus was also elevated during the first few days after immunization, but to a lesser extent. The changes in total enzyme activity generally preceded the appearance of haemolytic plaque forming cells. Most of the antibody forming cells were present in the spleen, with peak numbers at day 4. Much fewer antibody forming cells were present in the lymph nodes, and even fewer in the thymus. The relationship between immunogenesis and nucleic acid metabolism in lymphoid tissue was discussed.

     

In-vivo modulation of thymus-derived lymphocytes with monoclonal antibodies in mice. I. Effect of anti-Thy-1 antibody on the tissue distribution of lymphocytes.

A procedure is described for the in vivo removal of all detectable T lymphocytes from spleen and lymph nodes in mice. A single intraperitoneal injection of monoclonal anti-Thy-1 antibody into mice leads to rapid depletion of functional T cells from peripheral lymphoid organs, but not thymus. The extent of T-cell depletion is dependent on the cytotoxic titre of the anti-Thy-1 antibody used. Antibody with a median cytotoxic titre greater than 10(6) causes the complete removal of cells bearing Thy-1, Lyt-1 and Lyt-2 surface antigens from peripheral lymphoid populations in 3 days. Eight days after treatment Thy-1+, Lyt-1+ and Lyt-2+ cells begin to reappear in these organs. Splenic B cells, assayed by the expression of surface immunoglobulin (sIg) and by mitogenic responsiveness to bacterial lipopolysaccharide (LPS), are not affected by this treatment. The monoclonal anti-Thy-1 antibody does not appear to penetrate thymus tissue and bind to thymocytes. Anti-Thy-1 antibody, but not F(ab')2 is required for in-vivo T-cell depletion. These findings indicate that anti-Thy-1 antibody causes the removal of Thy-1+ cells from peripheral lymphoid tissue, and as the circulating levels of anti-Thy-1 antibody decrease, cells from the thymus repopulate the thymus-dependent areas of the depleted lymphoid organs.