Regional graft vs host reaction to H-Y antigen (immunological mechanisms)

Structural changes in the regional popliteal lymph nodes have been studied in C57Bl/6 male mice at the peak of the reaction "graft-versus-host" to H-Y antigen. Morphological and morphometrical investigations have been carried out in three groups of males (10 animals in each group). The first group includes intact animals (the first control group). To the males of the second group (the second control group) lymphoid cells are inoculated from intact C57Bl/6 females. To the males of the third group (experimental group) lymphoid cells are inoculated from H-Y antigen immunized C57Bl/6 females (anti-H-Y effector lymphocytes). The popliteal lymph nodes of the male mice from the third group twice increase in their size comparing to those in the control (the first and the second groups). Miotic activity increases in them 4.5 times, amount of cellular blast forms in medullary cords--4 times and 10 times--in the light zone of the cortical substance. Ratio of macrophages and eosinophils in structural components of the lymph nodes studied changes; this is, evidently, connected with massive destructive progresses, that take place in the lymph nodes of the animals from the third group. The results of the morphological investigations are in agreement with the hypothesis suggested, explaining the mechanism of development of the regional reaction "graft-versus-host" to H-Y antigen, basing on idiotype-antiidiotype interaction (the idiotypic network in the immune system).     

Characteristics of the cellular reactions of the T- and B-dependent areas of the regional lymph nodes in dogs to the administration of immunodepressants

Mesenteric, bifurcational, axillary and popliteal lymph nodes have been studied in 22 healthy mature male dogs. Amount of blast cells, small lymphocytes, plasma cells and macrophages has been taken into account in the paracortical zone, in the germinative centers and in the medullary cords. For two weeks to one group of the animals every day imuran in turn with aurantin (10 mg/kg and 25 mg/kg) are injected, or antilymphocytic serum (ALS) intraperitoneally every other day (0.1 ml/kg). The combined injection of imuran and aurantin produces a more pronounced toxic effect to the hemopoietic organs than ALS. ALS is more specific for T-dependent zones of the lymph nodes. In the dose and interval mentioned ALS is an immunostimulating preparation for the immunocompetent cells of the germinative centers of the lymph nodes. The reaction of the lymph nodes depends on their regional belonging.     

Comparing the MRI appearance of the lymph nodes and spleen in wild-type and immuno-deficient mouse strains

The goal of this study was to investigate the normal MRI appearance of lymphoid organs in immuno-competent and immuno-deficient mice commonly used in research. Four mice from each of four different mouse strains (nude, NOG, C57BL/6, CB-17 SCID (SCID)) were imaged weekly for one month. Images were acquired with a 3D balanced steady state free precession (bSSFP) sequence. The volume of the lymph nodes and spleens were measured from MR images. In images of nude and SCID mice, lymph nodes sometimes contained a hyperintense region visible on MRI images. Volumes of the nodes were highly variable in nude mice. Nodes in SCID mice were smaller than in nude or C57Bl/6 mice (p<0.0001). Lymph node volumes changed slightly over time in all strains. The spleens of C57Bl/6 and nude mice were similar in size and appearance. Spleens of SCID and NOG mice were significantly smaller (p<0.0001) and abnormal in appearance. The MRI appearance of the normal lymph nodes and spleen varies considerably in the various mouse strains examined in this study. This is important to recognize in order to avoid the misinterpretation of MRI findings as abnormal when these strains are used in MRI imaging studies.     

Interrelationships of B- and T-cell areas in peripheral immune organs of the albino rat after hind limb autotransplantation

Using 68 3-month-old male albino rats, it was established that the pattern of the changing interrelationships of B- and T-cell areas in the spleen and in the popliteal, inguinal and medial iliac lymph nodes regional for the experimental limb during the initial stage after hind limb autotransplantation, with or without sciatic nerve alloplasty, represents a universal type of initial response of the peripheral immune organs to external challenge which takes place in three steps: 1) an increase in the number and size of the lymph nodules, 2) enlargement of T-cell areas, 3) an increase in the number of structures containing antibody-forming cells (the medullary cords in the lymph nodes and the splenic cords). Sciatic nerve alloplasty gives rise to expansion of the medullary cords in the lymph nodes and the marginal zone and cords in the spleen, with parallel significant enlargement of T-cell areas.     

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     

Retention of intact HSA for prolonged periods in the popliteal lymph nodes of specifically immunized mice.

The rate of clearance of radiolabel was studied from the popliteal lymph nodes of either nonimmune or specifically immunized mice following footpad injection of ¹²⁵I-labeled HSA. The ¹²⁵I was more rapidly and more completely eliminated from the nodes of nonimmune mice than from the nodes of specifically immunized animals. Autoradiographs showed that, whereas the radiolabel was strictly confined for long periods to the follicles of the nodes of immune mice, no such localizaton was present in nonimmune animals. Solubilization of the whole nodes with 5 M guanidine-HCl released the labeled material which could then be specifically precipitated with anti-HSA but not with anti-EA. Such specifically precipitable material could be extracted from the draining popliteal lymph nodes of the immune mice even 12 weeks after challenge. Analysis of the solubilized radioiodinated lymph node extract on Sephacryl S-200 gels showed that the majority of the material was indistinguishable from the ¹²⁵I-labeled HSA used for challenge. The radiolabel therefore remains attached to intact HSA for long periods in vivo in immune animals which have a highly developed mechanism for clearance of simple protein antigens. These data suggest that intact antigenic determinants can persist for prolonged periods and may play a role in the long-term maintenance and regulation of immune responses.     

Cyclic kinetics and mathematical expression of the primary immune response to soluble antigen

The kinetics and the distribution of antigen and antibody were shown to be similar in four species of experimental animals and in two species of wild rodents immunized with the protein-polysaccharide capsular plague antigen. Serologically active antigen and antibody were detected in homologous conjugating serological tests. Soluble antigen persists at the injection site for as long as a week and adsorbed antigen for two weeks or more. Antigen persists in the blood of animals for 2–4 days. In regional popliteal lymph nodes, antigen was detected for the first days, followed by antibody in both lymph node and blood. Plasma cell response was more intensive in animals inoculated with adsorbed antigen. The gradual decrease of antigen at the injection site shows superimposed up-and-down changes, mostly parallel with the antibody in the popliteal lymph node and blood, as well as with plasma cell response in the regional lymph node. Serological cycles were related to the resistance of immunized white mice to plague infection. Cyclic kinetics of specific polysaccharide in the faeces of dysentery patients was found.     

Replacement of the helper function of T cells by an RNA-containing antigen-specific lysis factor]

The lymph node viable cells suspension of immunized mice was centrifugated. The supernatant was chromatographed in Sephadex G-200, and fractions were deproteinized. The deproteinized third fraction (Mol wt 30000) stimulated specifically the plaque-forming cells of intact mice immunized by SRBC. It restored the capacity to antibody production in the lethally irradiated intact mice protected by the syngeneic bone marrow. The activity of this fraction disappeared following treatment with RNA-ase, but not with DNA-ase or trypsin. The first and the second deproteinized fractions of the supernatant inhibited non-specifically the viable lymph node cells of the immunized animals in the intact mice immunized with SRBC.     

Comparative study of the changes in microanatomical organization of lymph nodes located and functioning in the area of venous congestion

By the morphometric method the microanatomical organization of the popliteal and kidney lymph nodes of the type Vi was studied in venous congestion. In the first experimental series (lymph nodes localize in venous congestion area) the venous congestion in the popliteal lymph nodes was created by the ligature of caudal vena cava. In the second experimental series (lymph nodes, which clean the venous congestion area--the kidney vena was tied up. The small and short time increase of the marginal sinus volume and hypertrophy of the medullary cords, which seemed about medullary substance B-zone stimulation were characteristically for the lymph nodes, localize in the venous congestion area. The sharp increase of the marginal sinus volume during the long experimental time and paracortical T-zone hyperplasia were characteristically for the lymph nodes, which clean the venous congestion area.     

Degree of chromatin activity and the RNA level in lymphocytes of popliteal lymph nodes of dogs after administration of an antigen]

In the experiment performed on 127 dogs by means of cytospectrofluorometric analysis, using fluorochrome acridine orange in dynamics up to 1 year, changes in the level of chromatin activation and RNA content have been studied in lymphocytes of the germinative centers and the crown of lymphoid nodules, in the paracortical zone and medullary cords of the regional and contralateral popliteal lymph nodes, after subcutaneous injection of antigen (BCG vaccine, 0.2 mg/kg) into the lateral area of the foot of the left pelvic extremity. The immune response is accompanied with a periodical increase in the level of chromatin activation and RNA content in populations of lymphocytes in the regional and contralateral popliteal lymph nodes with maximum in 6 h, 3-7 days, 1-3 months after the antigen injection. The intensity of these processes has an unequal level in lymphoid cells of various structural components; it is higher in lymphocytes of the contralateral lymph node.     

Staining patterns for the anti-horseradish peroxidase antibody reaction in proplasma cells developing in the medulla of rat popliteal lymph nodes during the secondary response

The development of plasma cells from lymphocytes was studied in the medulla of popliteal lymph nodes of rats during the secondary response to horseradish peroxidase (HRP). Changes in the microscopic appearance of proplasma cells were compared with changes in the intensity of the anti-HRP antibody reaction in these cells. Early proplasma cells, appearing 2 to 3 days after the injection of HRP into the footpads, were relatively small cells similar in size to lymphocytes. Their small nuclei were eccentrically located due to the one-sided enlargement of the pyroninophilic cytoplasm. The reaction for the anti HRP antibody in these cells was weak or negative. Other proplasma cells located in the same medullary cord regions showed a more intense antibody reaction. This change was correlated, in many cases, with an enlargement of the nucleus, giving the cells a blast-like appearance. Three to 6 days after the reinjection of the antigen, the medullary cords contained many mature plasma cells characterized by an intense antibody reaction. The mature plasma cells were always accompanied by proplasma cells, the latter varying in microscopic appearance (stage of development) asd staining intensities (antibody contents). The staining intensities and the microscopic appearance of proplasma cells, and the proportion of proplasma cells to plasma cells, varied in different medullary cord regions of the same lymph nodes. The staining patterns, together with the microscopic appearance of the cells, seemed to show whether antibody formation was inhibited or stimulated.     

Structure of the arterial bed in human lymph nodes

Blood vessels, that bring blood to various areas of the human superficial inguinal lymph nodes are predominantly arterioles and precapillaries. They are often arranged radially from the hilus to the capsule and from the capsule towards the portal thickening. The arteries and arterioles of the portal and capsular trabeculae reach the paracortical zone, occupying an intermediate position between the medullary cords and the cortex of the lymph node. The arterioles of the paracortical zone, passing between the cortex and the medullary cords, acquire an arcuate appearance. In both directions from them (into central and peripheral areas of the node) precapillaries branch off at a right angle. The cortex is supplied with blood by the arteriolar branches of the paracortical zone and the capsule of the node. The cortical precapillaries branch into capillaries either within the lymphoid nodules, or along their periphery. In the medullary cords those arterioles branch, that get from the portal thickening, portal trabeculae and paracortical zone.     

Cellular composition of various structural zones of the iliac and mesenteric lymph node of pregnant rats

The dynamic of cellular reactions demonstrates certain changes in functional activity of all structures of the node during pregnancy. A similar trend of processes in the iliac (regional for the uterus) and mesenteric lymph nodes has been defined. At early stages of pregnancy, lymph nodule are the most active, this is demonstrated as an increasing portion of lymphoblasts, macrophages and dividing cells. During this period, cell composition of the cortical plateau is relatively stable. For the paracortical zone of the mesenteric lymph nodes a rather significant decrease in the portion of middle lymphocytes and reticular cells is characteristic. There is not any significant change in the relative amount of the cells in the same functional zone of the iliac lymph nodes during the same period of pregnancy. The medullar cords demonstrate an increasing number of blast forms and young plasmocytes. However, as the pregnancy develops, the structure of the paracortical zone undergoes an essential change--progressively increases the portion of lymphoblasts and large lymphocytes. The blastic reaction in the mesenteric lymph nodes is proved to depend, to some extent, on that in the iliac lymph nodes of the same animal. Mature plasma cells become the dominating cellular element in the medullary cords. At the end of the pregnancy a relative amount of the reticular cells increases in all structural zones of the node.     

Ontogeny of the antigen-reactive lymph follicle-forming capacity of the popliteal lymph node in neonatal mice

The ontogenetic development of the reactive lymph follicle-forming capacity of the popliteal lymph node was investigated immunohistochemically in young mice which had received a single injection of hemocyanin (KLH) in a rear footpad at a predetermined age (between 1 and 21 days). The mice were sacrificed at various intervals after injection. In non-stimulated young mice, primary lymph follicles first appeared in the popliteal node at 11 days of age. When KLH was given to 7-day-old or older mice, each draining popliteal node showed a marked increase in B lymphocytes in the extrafollicular zone 3 days after injection and produced a number of "new" lymph follicles outside the pre-existing follicles over the next few days. In mice injected at 2-4 days of age, these nodes showed an increase in B lymphocytes in the outer cortex and had produced several lymph follicles by 8 days of age. The number of lymph follicles produced by each node tended to increase in line with age at injection. These results indicate that neonatal popliteal nodes become able to produce lymph follicles in response to exogenous antigens some time before ontogenetically developing follicles appear. The formation of new lymph follicles observed in draining popliteal nodes after KLH injection at an early postnatal age is discussed in relation to the ontogenetic development of stromal cells (precursors of follicular dendritic cells) that are capable of interacting with B lymphocytes and the extent of B lymphocyte influx into the node induced by KLH stimulation.     

Catecholamine and serotonin content in the lymph nodes following deafferentation

Using Falck fluorescent technique in combination with microspectrofluorometry the measurement of catecholamine and 5-hydroxytriptamine level was performed in popliteal lymph nodes of normal dogs and 2 weeks after deafferentation. Maximal catecholamine and 5-hydroxytriptamine content was observed in adrenergic nerve fibers, elements of trabecular-elastic complex and monoaminocytes of medullary substance of intact animals. Biogenic amine level decreased deafferented limb lymph nodes. Monoamine imbalance according to divergent type was registered in a colateral limb, i. e. 5-hydroxytriptamine level was increased and catecholamine level was decreased.     

Peritoneal macrophages introduced into mouse foot pads enter the germinal center of regional lymph nodes nonspecifically.

Male mice were injected into their foot pads with sheep erythrocytes (SRBC) to form lymph follicles in the germinal centers in the popliteal lymph nodes. 4 weeks later, peritoneal macrophages labeled with carbon from syngeneic donors sensitized with SRBC or typhoid-paratyphoid bacilli (TAB) were separately injected into the foot pads as well. The popliteal lymph nodes were histologically examined at 6 h to 5 days after injection. Labeled macrophages appeared in the marginal sinus, migrated straight across the cortex from the marginal sinus to the lymph follicles and then entered the germinal centers. There was no difference in the mode of appearance, migration and localization of labeled macrophages in the regional lymph nodes between the mice given labeled macrophages from SRBC-sensitized donors and those given macrophages from TAB-sensitized donors. The entrance of lymph macrophages into the germinal centers of the regional lymph nodes would be immunologically nonspecific. After the injection of Pelikan ink into the foot pads, the macrophages which have taken up carbon in the peripheral tissue reached the regional lymph nodes via the afferent lymphatics and then entered the germinal centers, mainly through the medullary pole of the lymph follicles, after migrating along their immediate exterior from their marginal sinus to their medullary pole.     

The reaction of somatic lymph nodes to experimental exposure to sulfide baths]

Influence of strong sulfide baths of the sanatorium "Talgi" to the popliteal and superficial inguinal lymph nodes (LN) has been studied in 35 rats (140-150 g of body mass) and 15 animals have been used as the control. The slices are stained with hematoxyli-eosin, after van Gieson, Romanovsky-Giemsa, Kurnik and silver nitrate impregnation after Foot. The section area of the LN decreases, but amount of lymphoid nodules increases (especially in deep layers of the cortex). In the germinative centers amount of middle lymphocytes and mitoses becomes larger. In the internodular zones of the popliteal nodes amount of macrophages and eosinophiles enlarges significantly, and in the inguinal-amount of mast cells and eosinophiles. In the medullary sinuses amount of macrophages declines. In the medullary cords amount of immature plasma cells and blasts increases, while amount of mature cells decreases.     

Changes in the H-alloantigen-recognizing function of mouse lymphoid cells following hydrocortisone administration]

The capacity of spleen, thymus, and bone marrow cells of intact (control) and of hydrocortisone-treated mice CBA to induce the lymph node type of graft-v-host reaction (GVHR) in hybrids F1 (CBA X c57bl) was studied. After hydrocortisone injection (2.5 mg per mouse) the donor spleen cells became more active in GVHR, considering the value of lymph node indices and immunoblast content in the regional lymph node as compared with a control group. Following transplantation of thymus cells taken from the hydrocortison-treated donors the immunoblast count was higher, although the lymph node weight remained the same as in the control group. On the contrary, following the transfer of the bone marrow cells from the hydrocortisone-treated mice the lymph nodes enlarged, while the immunoblast count remained as low as in control. Consequently, exogenously conditioned increase in the hydrocortisone level was accompanied by an enrichment of the spleen and thymus cell populations with T-lymphocytes, proliferating in response to H-alloantigens.     

Adult thymectomy promotes the manifestation of autoreactive lymphocytes.

Spleen cells from adult thymectomized mice (ATX) were assayed in a syngeneic graft vs host (GVH) model based upon enlargement of the draining popliteal lymph node following syngeneic cell inoculation into the hind footpad. Spleen cells from ATX mice have been found to induce a significantly higher increase in the weight of the regional lymph node than that induced by the injection of normal spleen cells. Irradiated spleen cells from ATX donors did not cause a similar increase, suggesting either that proliferation of the transferred cells was required at some stage of the reaction or that autoreactive cells are radiosensitive. Autoreactive cells were found in the spleen of mice 2 to 3 months after the thymectomy but were never found in the lymph nodes of such animals or in the thymus of intact mice. They are not phagocytic adherent cells and are not retained on nylon wool columns, which suggests that they belong to the T-cell lineage. Autoreactivity is lost when spleen cells from ATX donors are depleted of autologous rosette-forming cells (A-RFC) by centrifugation on a Ficoll-Hypaque gradient after rosette formation. Autoreactive spleen lymphocytes might belong to the population of A-RFC previously characterized as a population of immature T cells.     

Isolation of monoclonal antibodies to antigen Lyt-3.2

A hybridoma producing monoclonal antibodies (McAb) NATF9.9 (F9) was obtained from fusion of murine myeloma X63 and splenocytes of AKR mice immunized with a single intravenous injection of 5 X 10(7) thymocytes of CBA mice. F9 McAb were cytotoxic for 80% thymocytes, 10% splenocytes, 20% lymph node cells, 85% cortical and 32% medullary thymocytes of CBA, C57BL/6, BALB/c, DBA/2 and SJL but not for the cells of C58 and AKR mice. F9 McAb reacted only with T cells and did not react with B cells and EL4 thymoma cells (Thy-1.2+, Lyt-1+2-3-). The proportion of F9+ cells accounts for about 40% among T lymphocytes of the lymph nodes and spleen as tested by flow-type cytometry. Lymph node cells treated with F9 McAb plus complement completely lost their reactivity with rat anti-Lyt-2 McAb and only partly (by 30%) with anti-Lyt-1 McAb. The reactivity pattern of F9 McAb attests to their specificity for Lyt-3.2 antigen.