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Direct cellular contact between thymocytes and thymus stromal cells within the thymus appears to contribute to the maturation of thymocytes. Thymocyte-stromal cell complexes, formed in vivo, have been isolated by others and postulated to play a role in T-cell differentiation. These previous studies have been hampered, however, by a time-consuming isolation procedure from which only small numbers of these complexes are recovered. We have examined a model to study thymocyte-stromal cell complexes in vitro in which thymocytes are added to primary cultures of thymus stromal cells. In the present study, we found that thymocytes were histotypically selective in their attachment to thymus stromal cells. We also investigated the kinetics of thymocyte attachment to these thymus stromal cells. Cultures were examined at selected time intervals from 5 min through 3 days of incubation. Thymocyte attachment to stromal cells was a biphasic interaction, with maximum surface attachment at 15 min of cocultivation, followed by migration of thymocytes into the cultures. Morphological studies were confirmed by using 3H-leucine-labeled thymocytes and liquid scintigraphy. With increased time in culture, thymocytes became amoeboid and migrated between the layers of stromal cells where thymocyte mitotic figures were seen at 4 and 8 hr. In some cases it appeared that stromal cells, which often grew two to three cell layers deep, played an active role in enclosing thymocytes within the cultures. Large numbers of viable thymocytes were observed in the cultures at 24 hr. The number of thymocytes then decreased progressively on days 2 and 3, when relatively few were found within the layers of the culture.
Anat Rec 1989 May
PMID:Kinetic analysis of thymocyte attachment to thymus stromal cells in culture by using phase-contrast and scanning electron microscopy. 265 86

Mice were immunized subcutaneously with thymus-independent (TI)-type 1 antigen trinitrophenylated lipopolysaccharide (TNP-LPS), TI-type 2 antigen TNP-Ficoll or thymus-dependent (TD) antigen TNP-keyhole limpet haemocyanin (TNP-KLH) in order to study the primary in situ immune response in popliteal lymph nodes (PLN) and spleen. The spleen responded more rapidly in developing specific antibody-forming cells (AFC) than the lymph nodes did, in spite of the fact that antigens reach the spleen only after passing several lymph node stations. This difference between lymph nodes and spleen in developing AFC was particularly significant with respect to the responses to TI (both type 1 and type 2) antigens. No differences in the distribution of specific AFC in PLN and spleen were observed after immunization with TI and TD antigens. Results are discussed with respect to the relative contributions of lymph nodes and spleen to immune responses to antigens injected subcutaneously.
Anat Rec 1989 Feb
PMID:Primary in situ immune response in popliteal lymph nodes and spleen of mice after subcutaneous immunization with thymus-dependent or thymus-independent (type 1 and 2) antigens. 271 42

Previous studies have indicated that the human thymus is composed of several discrete compartments. Cortical thymocytes are reactive with the monoclonal antibody anti-T6, whereas most medullary cells, unreactive with anti-T6, stain brightly with anti-T3 antibody, which defines mature T cell populations. By using an indirect immune rosette method, we isolated the minor thymocyte population (1 to 2% of all thymocytes) lacking both T3 and T6 but expressing T11 antigens. These cells could be maintained in culture supplemented with recombinant IL 2 (Rec-IL 2) for several days. Under these conditions, T3-T6- cells were shown to undergo phenotypic changes. In the absence of thymic macrophage (Mo), T3+ and T8+ thymocytes appeared in culture, whereas the development of T4+ cells strictly required the presence of Mo. The expression of T4 antigen could be largely prevented by the addition of anti-HLA-DR antibody, further indicating that Ia+ accessory cells had the ability to promote in vitro development of T4+ thymocytes. In the presence of Mo, not only T4+ but also T8+ cells were obtained. Double fluorescence staining with anti-T8-FITC and anti-T4-biotin demonstrated that after 12 days of culture, T4 and T8 antigens were mutually exclusive. Furthermore, during the course of these studies, we observed that under the culture conditions utilized (e.g., presence or absence of Mo), T3-T6-thymocytes failed to express the T6 antigen. Thus, the in vitro development of T cells bearing a mature phenotype could be obtained in the absence of intermediate expression of cortical (T6+) thymocytes.
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PMID:T cell surface markers expression by immature human thymocytes in in vitro culture: role of Ia+ accessory cells. 310 65

A population of adult CBA/J mouse bone marrow (BM) cells enriched by in vitro migration to supernatant prepared from neonatal thymus was labeled with a DNA-binding fluorochrome, Hoechst dye No. 33342 (H33342). Labeled cells were injected into irradiated recipients in order to compare the in vivo localization of the migration-enriched BM (MEBM) cells to the localization of injected nonenriched BM (NEBM) cell controls. A characteristic difference in the distribution of localized cells was observed in the spleen but not in other lymphoid organs. At 2 hr after injection the MEBM cells were located in the marginal zones surrounding the periarterial lymphoid sheaths (PALS) of the splenic white pulp. At 6 hr after injection the MEBM cells were seen distributed between marginal zones and the PALS and by 16 hr they had localized almost exclusively in the white pulp. In contrast, the NEBM cells were located in the marginal zones or red pulp for the duration of the experiment. These observations show that the MEBM cells home selectively to T-cell areas of the spleen. Direct immunofluorescent monoclonal antibody staining of H33342-labeled cells obtained from the recipient spleens at 16 hr demonstrated that the MEBM cells were negative for Thy-1 antigen, indicating that acquisition of Thy-1 was not prerequisite to the observed homing. The results are compared to known localization patterns of mature lymphocytes.
Anat Rec 1988 Jul
PMID:In vivo homing of thymus-enriched bone marrow cells. 318 66

For many years data on the development of specific antibody-forming cells in lymph nodes were incomplete, fragmentary, and even contradictory. A number of recent studies have been performed, concerning 1) their overall architecture; 2) migration of B-lymphocytes; 3) localization of accessory cells and T-lymphocytes which are believed to be involved in humoral immune responses; and 4) localization patterns of specific antibody-forming cells developing during thymus dependent and thymus independent immune responses. Comparison of these new results with those of earlier studies suggests a single route of migration followed by all cells which will differentiate into antibody-forming cells. During their differentiation into antibody-forming plasma cells, antigen reactive B-cells migrate along the required accessory cells and/or T-lymphocytes.
Anat Rec 1987 Aug
PMID:The "in situ" immune response in lymph nodes: a review. 331 Jul 34

Here we describe a nodule of lymphoid tissue which was consistently located in the proximal colon of mice approximately 25% of the distance from the cecum to the rectum. Immunohistochemical characterization of this nodule demonstrated that the majority of lymphocytes were relatively immature 14.8+ (B220+), IgM+, Ia+ (specificity 20) B cells some of which were also Ly-1+. These nodules also possessed an occasional T cell (Thy-1+, Ly-1+, Lyt-2+) aggregate at the periphery. Rare, small areas did not stain for either T or B cell markers. These lymphoid nodules were associated with epithelial cells which stained positively with the ER-TR4 monoclonal antibody (which also recognizes thymic cortical epithelial cells) and also with ER-TR6, which has been reported to recognize thymic macrophages or dendritic cells. The overlying colonic epithelium stained intensely with the ER-TR4 monoclonal antibody. Proximal colonic lymphoid tissue was extremely sensitive to steroid treatment, losing approximately 80% of its mass within 24 hours in response to a single intraperitoneal injection of 2 mg hydrocortisone acetate. This response was similar to that of the thymus and to that reported for the bursa of Fabricius, but unlike that of other gastrointestinal lymphoid aggregates. These results indicated that proximal colonic lymphoid tissue contains a high frequency of relatively immature B cells and may be a primary site of their generation, possibly including some of the Ly-1+ phenotype. These observations correlate with new evidence suggesting that the allantois participates in the formation of the distal midgut, including its lymphoid components.
Anat Rec 1988 Mar
PMID:Characterization of proximal colonic lymphoid tissue in the mouse. 336 57

Rabbits were intravenously primed and boosted with trinitrophenyl-keyhole limpet hemocyanin (TNP-KLH) and human serum albumin (HSA); both antigens were injected simultaneously. The localization of anti-TNP-antibody-forming cells (AFCs) and anti-HSA-AFCs was determined in various lymphoid organs of the rabbit. In all lymphoid organs of primed rabbits anti-TNP-AFCs outnumbered anti-HSA-AFCs, with the exception of the thymus, in which neither of them was encountered. In the spleen the antibody-forming cells were mainly situated in the periphery of the periarteriolar lymphocyte sheaths (PALS) and in the coaxial sheaths of lymphoid tissue surrounding the terminal arterioles. In the lymphoepithelial organs AFCs were almost exclusively situated in the interfollicular areas, and in the lymph nodes largely in the medulla. An intravenous booster injection led to a secondary immune response (i.e., increase of AFCs) in the spleen. No visible change in the number of specific AFCs was observed in the lymphoepithelial organs. However, in the mesenteric and popliteal lymph node the number of anti-TNP-AFCs had increased tremendously.
Anat Rec 1987 Jan
PMID:Development of specific antibody-forming cells in various lymphoid organs of rabbit after intravenous antigen administration. 345 65

An Escherichia coli derived somatomedin-C/IGF-I preparation (rec-IGF-I) with an amino acid sequence identical to the natural IGF-I derived from human plasma, increases body length and weight, as well as the growth of several organs of Snell dwarf mice, when administered for 4 wk. After 2 wk of treatment rec-IGF-I (22.2 micrograms/day) induced a significant increase over buffer treated controls, to a comparable degree as obtained with bacterially synthesized human growth hormone (bhGH; 8.4 micrograms/day). The weight/length ratio of rec-IGF-I and bhGH-treated dwarf mice after 4 wk of treatment were not significantly different. A significant increase over controls was obtained with both preparations. Organs with increased weights after bhGH treatment (brain; submandibular salivary glands; heart, liver, kidneys, thymus, and spleen) were also heavier after rec-IGF-I. Significance was only reached for the kidneys and the spleen and the musculus quadriceps femoris. Organ weights expressed as a percentage of body weight of bhGH and rec-IGF-I treated dwarfs were similar except for the relative weight of the heart of the bhGH group, which was significantly increased compared to the controls and the rec-IGF-I group. These data resolve the issue as to whether or not pure SM-C/IGF-I will induce growth in length and demonstrate the usefulness of recombinant IGF-I in the studies of growth regulation.
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PMID:Biosynthetic somatomedin C (SM-C/IGF-I) increases the length and weight of Snell dwarf mice. 374 55

Electron microscopic examination of the extrahepatic distribution of pit cells, a cell type found in the liver, revealed their existence in several other organs of the rat. They were relatively frequent in lungs, spleen (red pulp), small intestine, epididymis, trachea, and peripheral blood; much fewer in bone marrow and thymus (medulla); and nonexistent in lymph nodes, spleen (white pulp), and thymus (cortex). The pit cells in these organs, as well as in the liver, contained characteristic dense granules and rod-cored vesicles in the cytoplasm. Our observations suggest that pit cells circulating in the peripheral blood adhere to the endothelium of capillaries in the various organs and migrate into the tissue, where they have some special immunological function.
Anat Rec 1985 Feb
PMID:Pit cells in extrahepatic organs of the rat. 397 86

Fluorescence microscopy of thymus and spleen from four strains of mice (C3H and ICR controls, AKR spontaneously leukemic and NZB autoimmune) revealed varicose noradrenergic (NE) fibers in perivascular and parenchymal regions of both organs. Thymic innervation was largely perivascular, but isolated islands and strings of free NE fibers were noted among thymic parenchymal cells. A morphological proximity between NE fibers in the thymus and mast cells was noted in all strains studied, but was exceptionally prominent in the NZB thymus. Perivascular plexuses within the splenic white pulp sent single NE fibers between the surrounding lymphocytes. Catecholamines and histamine have been shown to modulate lymphocyte development and activity in vitro. The present study provides morphological evidence that both NE and histamine are available to lymphocytes in thymus and spleen, and thus provides morphological evidence for neural modulation of immune activity in vivo.
Anat Rec 1981 Apr
PMID:Sympathetic innervation of murine thymus and spleen: a comparative histofluorescence study. 616 12


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