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R M Binns

Publications and source records attributed to R M Binns.

At least 19 recordsLinked to original sources
PubMed1992

Active lymphocyte traffic induced in the periphery by cytokines and phytohemagglutinin: three different mechanisms?

Phytohemagglutinin (PHA) injection induces transient protease-sensitive traffic of lymphocytes in skin and other tissues in several species. Examination of the possible roles of cytokines in such reactions showed that recombinant bovine and human tumor necrosis factor (TNF)-alpha potently induce dose-dependent lymphocyte traffic in pig skin (and in other tissues including the draining lymph nodes) with early kinetics and a morphology of the inflammatory reaction similar to that of PHA (peaking 9-12 h). Recombinant human interleukin (IL)-1 alpha also induces dose-dependent lymphocyte traffic, but it peaks at 4 h. Entry of labeled lymphocytes into inflammatory sites induced by PHA, TNF-alpha and IL-1 alpha, but not into normal skin, is inhibited by approximately 80% by their pretreatment with trypsin, indicative of the induction of endothelial determinants recognized by protease-sensitive surface molecules on the lymphocytes. Even the minimal lymphocyte traffic induced by interferon-gamma and lipopolysaccharide was similarly protease sensitive. At the earliest stage (approximately 2 h) of significant induction of lymphocyte entry by TNF-alpha and IL-1 alpha the inductive signal for each appears easily saturated. Thus lymphocyte entry is little increased by increasing low cytokine doses over 100-fold: However, these reactions are additive, and this was used to confirm that they are distinct from each other and from PHA. A further distinction was revealed by the homing of lymphocytes pretreated with pertussis toxin: such lymphocytes were greater than 90% inhibited in their homing to tissues through constitutive high endothelial venules (HEV) and greater than 60% inhibited in homing to TNF-alpha and IL-1 alpha skin sites, but unaffected in homing to PHA skin sites (like most non-HEV-mediated traffic). Moreover, potent chicken anti-TNF-alpha, which prevented TNF-induced lymphocyte entry, did not affect PHA-induced traffic. Thus, these three agents which induce peripheral lymphocyte traffic appear to involve different mechanisms as shown by differences in (i) their kinetics; (ii) the effect of anti-TNF-alpha and (iii) the effect of pertussis toxin treatment of the lymphocytes and by the fact that their inductive mechanisms are additive in effect.

Animals↗
PubMed1992

Homing of blood, splenic, and lung emigrant lymphoblasts: comparison with the behaviour of lymphocytes from these sources.

The distinctive homing patterns of [125I]deoxyuridine-labelled lymphoblasts and 51Cr-labelled lymphocytes from the three sites--splenic and pulmonary venous emigrants, and peripheral blood--were demonstrated by study of their entry into a large number of tissues. These included the major organs such as the lung, liver, bone marrow, spleen, skin, and muscle, and several lymph nodes (LNs), Peyer's patches (PPs), tonsils and thymus, portions throughout the gastrointestinal tract and immunologically-stimulated sites. Major differences in homing of blood blasts (up to 4-fold) were found in particular tissue types, such as the different LNs, three PP types, and the three tonsils, and their distribution was quite different from lymphocytes. While splenic blasts behaved like blood blasts, lung blasts showed even more distinctive homing, e.g. to tissues with particular blast uptake, including spleen, stomach, bladder, portal and stimulated LNs, and liver and lung. While blast recovery data showed that all types homed well to bone marrow (about 25%) and the gastrointestinal wall (about 15%), the roughly 50% found in blood, muscle, skin, lung, and liver showed some homing preferences: blood blasts tended to stay more in blood, splenic blasts to home to muscle, and lung blasts to liver and lung. These studies have demonstrated that physiologically-derived lymphoblasts are not predominantly mucosal-homing but distribute through several non-lymphoid organs and that, even considering only those in the blood compartments, they show differences in homing preference.

Animals↗
PubMed1992

Subsets of null and gamma delta T-cell receptor+ T lymphocytes in the blood of young pigs identified by specific monoclonal antibodies.

Rat monoclonal antibodies (mAb) against isolated pig Null T cells were derived using a novel two-colour cytofluorometric assay. One (MAC320) identified all blood CD2-sIg- 'Null' cells (present at up to approximately 6 x 10(6)/ml). Another type (MAC319 and MAC318) identified a subset comprising approximately 60% or approximately 30% of the Null cell population. This percentage appears genetically determined. This subset partially overlapped with a gamma delta T-cell receptor+ (TcR+) population which consisted of approximately 40% of Null T cells. The antibodies did not react with other leucocyte or lymphocyte populations. In non-reducing conditions, MAC320 precipitated two molecules at approximately 270,000-280,000 MW in SDS-PAGE; the larger of which was also precipitated by MAC319 (and MAC318, which binds to the same epitope). Under reducing conditions, MAC320 immunoprecipitated two or three polypeptide chains at approximately 130,000-160,000 MW; MAC319 precipitated only the largest of these polypeptides. The large MAC319+ MAC320+ molecule on one subset is removed by bromelain treatment; the smaller MAC319- MAC320+ molecule on the remaining Null cells is not bromelain sensitive. Several properties of this new antigen complex specific to pig Null T cells show that it is distinct from the ruminant T19 complex.

Animals↗
PubMed1990

Kinetics of the in vitro antibody response to transmissible gastroenteritis (TGE) virus from pig mesenteric lymph node cells, using the ELISASPOT and ELISA tests.

A method is described for in vitro studies of viral humoral immune responses in the pig. After oral immunization with transmissible gastroenteritis (TGE) coronavirus, antibody production from primed mesenteric lymph node cells was revealed by an in vitro boost with viral antigen. For the latter the leukocytes were co-cultured with UV-inactivated virus using a variety of different methods of antigenic stimulation. Enumeration of specific antibody-secreting cells (ASC) and titration of secreted anti-virus antibodies were performed with ELISASPOT (using 3-amino 9-ethyl carbazole as the peroxidase chromogen) and ELISA tests respectively, according to the Ig isotype. The results showed a close relationship between ASC numbers and secreted antibody titres. The best in vitro antibody synthesis was observed when the sensitized cells were maintained in contact with virus during the whole culture period. Antibody responses were defined by a kinetic profile characterized by a narrow peak, with a maximum occurring after 4 and 6 days of culture and with the IgA response appearing earlier than the IgG. This methodology, which analyses specific antibody responses at the cellular level, may permit studies on the mechanisms of Ig isotype regulation. Extended to leukocytes from other organs of the immune system, it may also constitute an in vitro model to study antibody responses expressed in different lymphoid tissues of the pig.

Animals↗
PubMed1990

Exit of recirculating lymphocytes from lymph nodes is directed by specific exit signals.

During recirculation, lymphocytes leave the peculiar structurally inverted lymph nodes (LN) of pigs via blood vessels instead of via efferent lymphatics, as in sheep and other mammals. This functional difference provided an opportunity to show the existence of signals directing lymphocyte exit from LN. The recirculation of pig peripheral blood lymphocytes was traced through fetal sheep LN and of sheep PBL into and out of unsuckled newborn piglet LN, using the lack of natural antibody or natural killer cell function in these immunologically mature young to compare foreign and homologous lymphocyte behavior. In spite of some 50 million years of evolutionary divergence, the detailed kinetics and route of recirculation of the xenogeneic PBL were essentially the same as those of the host species. Thus determinants guiding the anomalous blood exit from pig LN must involve conserved "exit" signals in a new site and not changes in pig lymphocyte homing receptors.

Animals↗
PubMed1990

Phytohemagglutinin induces major short-term protease-sensitive lymphocyte traffic involving high endothelium venule-like blood vessels in acute delayed-type hypersensitivity-like reactions in skin and other tissues.

Injection of phytohemagglutinin induces dose-dependent acute delayed-type hypersensitivity-like reactions in young pig skin which attract large numbers of labeled peripheral blood lymphocytes and lymphoblasts but not red cells. Similar reactions are induced in the gut wall, joints, and draining lymph nodes, and also in the skin of sheep and cattle, and by concanavalin A. Labeled peripheral blood lymphocyte entry starts within 2 h, is maximal at 6-18 h and is effectively over by 48 h and, both early and late, is markedly inhibited in a selective way by trypsinization, like high endothelium venule-mediated entry to lymphoid tissues. Electron microscopy showed development of high endothelium venule-like blood vessels with intramural lymphoid cells in mononuclear infiltrates. Inductive processes in this model short-term peripheral defense mechanism are being investigated.

Animals↗
PubMed1990

Lymphocyte subsets and their proliferation in a model for a delayed-type hypersensitivity reaction in the skin.

A delayed-type hypersensitivity (DTH) reaction was induced in the skin of young pigs, by local injection of phytohaemagglutinin, and evaluation was carried out on the resulting accumulation of lymphocyte subsets and lymphocyte production by incorporation of bromodeoxyuridine in the skin and the draining lymph node. There was a rapid increase in mononuclear cells, which were found in clusters around venules. These included very few B lymphocytes, and CD8+ lymphocytes far outnumbered CD4+ cells. Underlining the importance of determining absolute numbers, the relative and absolute numbers of lymphocyte subsets showed quite different patterns during the development of the skin reaction. Lymphocytes in the normal skin incorporated the DNA precursor bromodeoxyuridine at higher rates than have been found for peripheral lymphoid organs. After intradermal phytohaemagglutinin injections, all subsets showed high proliferation rates in the skin, with kinetics which differed from the reaction in the draining lymph node. The labelling indexes of cells labelled with bromodeoxyuridine in vitro and in vivo were comparable. The phytohaemagglutinin injections also caused a marked and rapid increase in the proliferation of the cells in the basal layer of the epidermis. This model DTH-like reaction in skin with major CD8+ T-cell accumulation and proliferation locally and in the lymph nodes provides a reliable model for study of such reactions and for investigation of the regulatory role of cytokines.

Animals↗
PubMed1989

The null T cell in pig blood is not an NK cell.

Up to 50% of the blood lymphocytes in young pigs are thymus-derived, lack all subset-specific markers and appear immunologically unresponsive, with no known functional role. In an examination of their possible role in natural killing, NK activity was found in unpurified mononuclear cells and in preparations of unselected and nylon non-adherent lymphocytes (T cells and Null cells). However, NK activity was abolished by removing the E rosette forming T cells using a rat IgM anti-pig CD2 monoclonal antibody and rabbit complement, but not by control treatments with a non-binding rat IgM monoclonal reagent and complement or with any other reagent alone. Thus the resting Null T cell appears not to play a significant role in natural killing.

Animals↗
PubMed1989

What is the function of peripheral lymphocytes migrating to the thymus and of B lymphocytes proliferating in the thymus?

In studies on the role of other lymphoid organs in lymphocyte production and lymphocyte migration in young pigs and lambs, lymphocytes from these tissues were always found in the thymus. There were no major differences in the entry of labelled cells when the spleen, lymph nodes, bone marrow or Peyer's patches were selectively labelled. The immigrants were both mature small lymphocytes and lymphocytes just produced in the peripheral lymphoid organs. They enter via specialized venules at the outer part of the thymic medulla and do not migrate into the cortex. The lymphocyte homing is markedly reduced by prior incubation with trypsin. The relative numbers of immigrants within the thymus were small but, due to the huge cellular content of the thymus, the absolute number entering per day totalled several million from each peripheral organ. Another unexpected finding was the high mitotic rate in the medulla of the thymus. Moreover, in the adult rat thymus although there were only 0.14% B cells, these proliferated at a higher rate than in peripheral organs. The functional meaning of these data is obscure so far and the studies should stimulate further work on these topics of lymphocyte entry and B lymphocyte production in the thymus.

Animals↗
PubMed1988

Subpopulations of T lymphocytes emigrating in venous blood draining pig thymus labelled in vivo with fluorochrome.

The emigration of labelled thymus cells in the pig was studied directly in blood draining the large right distal cervical lobe of the thymus after controlled labelling with FITC delivered through cannulated branches of a main thymic artery and vein by temporary ex vivo perfusion at body temperature. Roughly 1% of thymic cells emigrated per day. Unlike most thymocytes, which are small, the size spectrum of thymic emigrants is slightly larger than that of typical blood lymphocytes. Surface-marker studies show that the surface phenotypes of the emigrants differ from both typical thymus and peripheral blood lymphocytes. Although the emigrants resemble thymocytes in the high proportion of strong rosettes formed with sheep red blood cells (RBC), they rosette poorly with pig red cells, particularly in the unenhanced saline test, in this respect behaving like blood lymphocytes. The peripheral T-cell subset bearing a Fc receptor is almost absent in thymus, but is well represented among the emigrants which thus resemble corticosteroid-resistant thymocytes in the pig. The large population of thymus-dependent Null lymphocytes in young pig blood apparently arise in thymus since they constitute 1/3 of emigrants, although only forming less than 10% of thymus cells. This emigration of thymic cells is discussed in relation to its implications for the turnover of known functional peripheral T-cell populations.

Animals↗
PubMed1987

Staining of pig lymphocytes subpopulations with acid alpha-naphthyl acetate esterase.

The use of alpha-naphthyl acetate esterase (ANAE) as a T cell marker in some other species and the broad correlation of incidence of ANAE-positivity and E rosette-formation in the pig suggest that ANAE-staining may be a T-cell marker in the pig. However, by studying the staining of lymphocytes within a variety of rosettes in fixed preparations a similar incidence of pig blood lymphocytes were found to be ANAE+ among T cells (E rosettes formed in dextran), B cells (antiglobulin rosettes) and Fc-gamma receptor-bearing B and T cells (EA rosettes in saline and dextran): complement (C') receptor-bearing cells showed a higher incidence of staining than other lymphocytes. Analysis of staining morphology suggested that certain morphologies within the B and T lineages may be confined to subpopulations. Thus ANAE positivity is certainly not a marker identifying blood T lymphocytes but could be of some value indicating subpopulations of B and T lymphocytes.

Animals↗
PubMed1987

Evidence of a selective major vascular marginal pool of lymphocytes in the lung.

The size and mobilization of the pulmonary vascular pool of lymphocytes was studied in young pigs. Blood lymphocytes, labeled in vitro with a fluorescent dye, were injected either intravenously or into the left heart. Comparable high numbers of lymphocytes were found in the lung after 30 min, demonstrating a specific homing of lymphocytes to the lung vascular bed. When the left lung was perfused with a cell-free medium for as long as 4 h in vitro, a continuous venous release of lymphocytes was found that resulted in about 1.5 x 10(9) lymphocytes and 5.7 x 10(6)/min leukocytes. Peripheral blood lymphocytes were labeled with 51Cr in vitro and injected intravenously. Fifteen minutes or 1 day later the lungs were perfused, and the release of cell-bound radioactivity was measured. The data indicate the existence of a large intravascular pulmonary lymphocyte pool. The sequestration of lymphocytes in the lung in many lymphocyte migration studies is not a removal of effete cells but a physiologic phenomenon.

Animals↗
PubMed1985

Characterization of pig T cell growth factor and its species-restricted activity on human, mouse and sheep cells.

Crude T cell growth factor was prepared from pig blood cells in mixed lymphocyte culture together with Concanavalin A. The TCGF was recovered from the crude supernatant by ammonium sulphate precipitation and fractionated by gel exclusion chromatography to yield active fractions corresponding to an apparent molecular weight of 23,000d. The TCGF was further purified by isoelectric focussing and was found to migrate as a single peak of pI 5 - 5.5. The crude preparation was found to support the growth of mouse and sheep activated cells but had no effect on human activated cells. Human TCGF supported the continued growth of activated pig cells whereas mouse and sheep TCGF had no such effect.

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