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New short-lived strains of mice (SAM-P), which have been developed by Takeda et al. (1981), show a defective antibody response to T dependent (TD) antigen in vitro, as demonstrated in the accompanying paper (see page 419). In the present study, we investigated the cellular site of the defect, using a cell culture system. In this paper, it is demonstrated that T-helper (Th) cell activity for the antibody response to TD antigen is impaired, while other cellular immune responses, e.g. mixed leucocyte reaction, cytotoxic T-lymphocyte response, and delayed-type hypersensitivity reaction, are normal. These results suggest that the defect in T-helper subset is limited in helper function for the antibody response, and that the helper function for the cell-mediated immune responses is intact. These two functions of the T-helper subset are apparently regulated in a different manner. The SAM-P strains of mice may thus serve as an appropriate model for studying functional heterogeneity in T-helper/inducer cell subsets. [ABSTRACT FROM AUTHOR]

The experiments described in this paper have examined the migration of three fluorochrome-labelled T-lymphocyte subsets (CD4+, CD8+ and γδ+T19+) on a single passage from blood to lymph, through prescapular lymph nodes. Lymphocytes obtained from prescapular efferent lymph were labelled in vitro with fluorochrome and returned to the blood of the same animal. Over the next 2 days, lymph was continuously monitored and the cells in all collections, including the one used for intravenous infusion, were phenotyped and analysed by flow cytometry. Significant differences in the subset ratios between the infused, starting population and the recirculated population iodicated that CD4+ and γδ+T19+ lymphocytes are extracted by a resting lymph node at the same rate and that both are extracted at a faster rate than CD8+ lymphocytes. The results presented here also suggest that a unique subset of γδ+T19+ lymphocytes may be present in blood that does not recirculate through peripheral lymph nodes. [ABSTRACT FROM AUTHOR]

The interest of the scientific community in regulatory CD4+ T cells has reached an enormously high level. Common agreement is that they inhibit not only the proliferation of CD4 and CD8 lymphocytes, but also the activities of natural killer cells and macrophages. However, very important issues concerning actual mechanism(s) and specificity of the action of regulatory T cells (Tregs) upon responder cells are still unsolved or vague. The best known marker for Tregs is the expression of transcription factor FoxP3, widely used for their enumeration. It is known that FoxP3 inhibits cytokine production so the most probable action of Tregs is direct. However, FoxP3 expression cannot be used for functional studies in humans. Therefore we identified human peripheral blood Tregs as a distinct, very well-defined population of peripheral blood T cells with reduced CD4 and high CD25 expression (CD4low CD25high), which fulfils the current phenotypic criteria identifying the Tregs by simultaneously expressing high amounts of FoxP3. We conclude that the definition of a CD4low CD25high phenotype is enough to unambiguously detect and study the regulatory function of these cells. On the functional level, the CD4low Tregs are able to non-specifically suppress the proliferation of autologous, previously polyclonally activated CD4+ and CD4− lymphocytes and to kill them by direct contact, probably utilizing intracellular granzyme B and perforin. [ABSTRACT FROM AUTHOR]

Background and Purpose:Protease-activated receptor-2 (PAR-2) is expressed on lymphocytes and endothelial cells, and plays a significant role in inflammatory reactions. Since leukocyte-endothelial cell interaction and reactive oxygen species (ROS) generation are hallmarks of the development of inflammation, the effects of PAR-2 activation by trypsin on lymphocyte adhesion and ROS generation was examined utilising PAR-2 wild type and knockout (PAR-2−/−) mice.Experimental Approach:Lymphocyte adhesion to the luminal surface of mouse isolated aortae was measured using 51Cr-labelled leukocytes and ROS generation from isolated lymphocytes was quantified using chemiluminescence.Key results:Trypsin induced adhesion of lymphocytes when added exogenously to the endothelial surface of the aorta for 30 min. Similarly, increased lymphocyte adhesion was also observed when mice were injected with trypsin intravenously 24 h prior to the adhesion assay, an effect which was partly ICAM-1 mediated. Trypsin also increased ROS generation from isolated mouse lymphocytes in a dose-dependent manner. The increase in lymphocyte adhesion and ROS production in response to trypsin were abolished in PAR-2−/− mice indicating a PAR-2 dependent mechanism. Superoxide dismutase had a greater inhibitory effect in PAR-2−/− mice compared to wild type mice when lymphocytes were stimulated with PMA but not trypsin.Conclusions and Implications:The present study indicates that activation of PAR-2 may be an important factor in modulating lymphocyte adhesion and ROS generation. The results have implications for developing anti-inflammatory strategies.British Journal of Pharmacology (2006) 149, 591–599. doi:10.1038/sj.bjp.0706905; published online 18 September 2006 [ABSTRACT FROM AUTHOR]

The distribution of T lymphocytes expressing the different polypeptides of the leucocyte common antigen (LCA) family detected by CD45R and UCHL1 antibodies has been studied in normal lymphoid tissues. In the thymus most cortical thymocytes express UCHL1 and co-express CD4 and CD8. The more mature membrane CD3+ (mainly medullary) T cells are heterogeneous and may express both UCHL1 and CD45R weakly or be restricted to display CD45R or UCHLI alone. In the medulla both the CD45R+ and UCHL1+ subpopulations contain single positive CD4 and CDS cells. In tonsils, germinal centre T cells are almost exclusively UCHL1+, CD4+ and a proportion also express HNK-l (Leu 7) antigen. In the paracortical areas approximately equal numbers of CD45R+ and UCHL1+ cells are found but these separately occupy nests of cells containing one or the other type. Again, both CD45R+ and UCHL1+ cells include single CD4+ and CD8+ lymphocytes. A small proportion (<5%) of strongly CD45R+, UCHL1+ double-stained cells are also seen, and these probably represent recently activated lymphocytes. In the gut, small clusters of such strongly double- labelled cells are in the submuscular mucosae while cells of the lamina propria are almost exclusively UCHL1+. Many intra-epithelial lymphocytes are only weakly positive or negative for UCHL l and appear to be CD45R+ . These results are consistent with the view that expression of different CD45 polypeptides identifies successive stages of thymocyte-T-cell maturation and that following their thymic education, unprimed T lymphocytes are CD45R+, while primed memory T cells arc UCHL1+. These populations occupy different microenvironments. [ABSTRACT FROM AUTHOR]

Primary and secondary bovine allogeneic mixed leucocyte cultures were examined for the generation of antigen-specific cytotoxic leucocytes. While optimal generation of murine and human cytotoxic T lymphocytes typically requires 4-8 days, alloantigen-specific cytotoxic bovine leucocytes were demonstrated consistently only after prolonged incubation periods, optimally found to be about 15 days. Restimulation of long-term bovine mixed leucocyte cultures with the original stimulator population revealed responder cells demonstrating augmented alloantigen-specific lytic activity. When placed into human recombinant interleukin-2, responder cells expanded and required passaging every 3 -4 days. The same was not true of cells placed into interleukin-2-free medium. Cells cultured in interleukin-2-containing medium retained alloantigen specificity after 10 weeks of culture. Moreover, they continued to display total dependence on human, simian or bovine interleukin-2 for growth. [ABSTRACT FROM AUTHOR]

Application of a contact-sensitizing agent to the skin of mice previously exposed to UV radiation at a different site results in the induction of hapten-specific suppressor T lymphocytes. When splenic lymphocytes from such mice were cultured with normal lymphocytes and hapten-conjugated splenic adherent cells, the primary proliferative response was suppressed. The cell responsible for the suppression in vitro was a T lymphocyte, and two signals were required for its induction, ultraviolet radiation and hapten sensitization. The T cell suppressing lymphoproliferation was specific for the hapten applied after UV radiation. The UV-induced T suppressor cell inhibited only primary lymphoproliferation; the response of lymphocytes from immunized mice was unaffected. The activity of the UV-induced suppressor cell was not affected by mitomycin C treatment. Thus, suppression of the primary proliferative response of lymphocytes to hapten-modified syngeneic cells in vitro correlates with in vivo suppression of contact hypersensitivity by these UV-induced suppressor cells. This suggests that the suppressor cells act by preventing the proliferation of hapten-specific responder clones. Use of this in vitro assay system should facilitate investigation of the characteristics of these cells and the mechanism by which these regulatory T lymphocytes inhibit contact sensitization. [ABSTRACT FROM AUTHOR]

The relative radiosensitivities of precursor (Tsp), activated (Ts) and memory (Tsm) suppressor T cells for delayed-type hypersensitivity (DTH) to sheep red blood cells (SRBC) were investigated in mice. Spleen cells from CBA mice, primed i.v. with 109 SRBC 3–4 days previously, contain specific Ts cells which substantially impair the induction of DTH to SRBC in normal syngeneic recipients. Exposure of mice to 400 rad irradiation 1 day before the priming completely eliminated the subsequent development of Ts cells. In contrast, 3 days after the priming injection, Ts cell activity in mice is resistant to doses higher than 600 rads. Mice primed 40 days previously with 109 SRBC contain Ts-cell memory which can be readily recalled by i.p. injection of 108 SRBC, The secondary Ts cells which specifically inhibit DTH induction can be demonstrated adoptively in normal recipients. Mice were exposed to various doses of irradiation 40 days after the priming and 1 day before the i.p. injection, Ts memory was significantly reduced by 300 rads and was completely abrogated by 400 rads. The relative radiosensitivities of the three subsets of suppressor T cells are in the order of Tsm = Tsp > Ts. [ABSTRACT FROM AUTHOR]

The accessory cell requirements of cytotoxic T-lymphocyte (CTL) responses directed at multiple minor alloantigens are examined using a short- term, combined in-vivo and in-vitro protocol. The development of cytotoxic activity in vitro from T cells sensitized in vivo requires tow-density accessory cells derived from the immunizing strain which have to be H-2 compatible with the responder population. The accessory activity of these cells can be by-passed by interleukin-2 (IL-2)-containing supernatant. Since IL-2 is a product of helper T (Th) cells secreted upon activation by antigen recognized in context of self H-2 and is known to stimulate antigen-activated cytotoxic T-lympifocyte precursors (CTL-P) non-specifically to effector CTL function, the results indicate that access- ory cells interact in an H-2 restricted fashion with helper rather than cytotoxic T-lymphocyte precursors. The low-density accessory cells active in this system do not express Fc receptors (FcR) and are present in both the adherent and non-adherent fractions of spleen or lymph nodes. [ABSTRACT FROM AUTHOR]

The syngeneic mixed lymphocyte reaction (SMLR) is the proliferative response of T lymphocytes cultured with syngeneic non-T lymphocytes. In previous studies, we found that the SMLR reaches adult level of activity at 4 weeks of age in BALB/c mice. We now report that the SMLR declines with age in this strain. The decline was first documented at 12 months of age, when non-T spleen cells were less able to stimulate young adult T cells than were non-T cells from 2 to 3 month-old mice. Splenic T cells from 12 month old mice were as responsive as splenic T cells from 2 to 3 month old mice. By 24 months of age, mice had no significant SMLR activity. Splenic T cells from 24 month old mice did not respond and splenic non-T cells did not stimulate SMLR when cultured with cells from young adult mice. Finally, suppressor cells were demonstrated in spleen cell preparations from 24 month old mice and may explain or contribute to the impaired SMLR in these animals. [ABSTRACT FROM AUTHOR]

Evidence is presented which indicates that recirculating lymphocytes originating from the intestine in pigs are returned to the blood circulation at the level of the mesenteric lymph nodes (MLN) and not via efferent intestinal lymph. This was demonstrated by three observations: (1) removal of all MLN resulted in a thirty-fold increase in lymphocyte numbers in efferent lymph of pigs, but not in rats: (2) there are about twenty-five times more lymphocytes in afferent intestinal lymph than efferent intestinal lymph in normal pigs; (3) 51Cr-labelled lymphocytes injected into afferent lymphatics are mostly recovered in the node tissue or efferent lymph of sheep, and very few in the venous drainage, whereas in pigs relatively few labelled cells are recovered in the node or in efferent lymph. [ABSTRACT FROM AUTHOR]

The article presents a study which assays x-ray resistant porcine suppressor T cells for their ability to suppress the proliferation of lymphocytes. The cellular and genetic requirements for the induction of such suppressor cells in pigs are investigated through the use of non-lymphoid cell cultures and cell mediated cytotoxicity assay.

Antigen-antibody complexes efficiently inhibit the induction of antibody formation. Using Mishell-Dutton cultures, it can be demonstrated that neither T cells nor their products are required for this inhibition of IgM PFC formation. The blockade is at the level of B cells and cannot be overcome by LPS or TRF. The data demonstrate that cross-linking of antigen- and Fc-receptors by antigen-antibody complexes is a blocking signal for B cells. [ABSTRACT FROM AUTHOR]

A 180,000 MW molecule has been identified on porcine leucocytes that is the homologue of the 215,000/300,000 MW WC1 (T19) leucocyte antigen previously considered to be restricted to ruminants. In ruminants the WC1 molecule is expressed by a T-cell subpopulation that is CD2-CD4-CD8-CD5+ and that is γδ T-cell receptor positive (TcR+). In pigs, the 180,000 MW molecule, identified by a new monoclonal antibody CC101, is expressed by a γδ TcR+ T-cell subpopulation that is also CD2-CD4 CD8. The p180+ cells are a major T-cell subpopulation comprising approximately 40% of the peripheral blood mononuclear cells from 6-9-month-old pigs. Expression of p180 identifies the majority of the CD2-CD4-CD8- T cells in porcine blood. The p180+ T cells have a distribution in lymphoid tissues that is distinct from that oft cells that express the CD2, CD4 or CD8 molecules. They are evident particularly in the thymic medulla, the epithelium, lamina propria and interfollicular areas of the small intestine, and the superficial dermis of the skin, but largely absent from conventional T-dependent areas of secondary lymphoid tissue. [ABSTRACT FROM AUTHOR]

Rat monoclonal antibodies (mAb) against isolated pig Null T cells were derived using a novel twocolour cytofluorometric assay. One (MAC320) identified all blood CD2- sIg- 'Null' cells (present at up to ∼6 × 106/ml). Another type (MAC319 and MAC318) identified a subset comprising ∼60% or ∼30% of the Null cell population. This percentage appears genetically determined. This subset partially overlapped with a γδ T-cell receptor+ (TcR+) population which consisted of ∼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 ∼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 ∼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. [ABSTRACT FROM AUTHOR]

African sleeping sickness is accompanied by a severe immunosuppression. As part of our efforts to examine the mechanisms by which this suppressive state is induced, we studied alterations in human T-lymphocyte function caused by Trypanosoma brucei rhodesiense. To this end, we used an in vitro system in which phytohaemagglutinin (PHA)-stimulated human peripheral blood mononuclear cells (PBMC) were cultured in a medium containing soluble, non-dialysable parasite products. We were able to demonstrate significant suppression of both lympho-proliferation and interleukin-2 receptor (IL-2R) expression. These effects were found to be dose-dependent and reversible after 48 hr of culture. The suppressive effects of living trypanosomes and the soluble parasite products on lympho- proliferation and interleukin-2 receptor expression were similar in that both precluded the entry of PHA-activated PBMC into the cell cycle. Eighty to ninety-eight per cent of the activated cells remained arrested in the G0/Gla (early G1) phase even 48 hr after stimulation, i.e. when last tested. Parasite-induced expression could not be overcome by the addition of recombinant human 11-2. These results suggest that immunosuppression associated with African trypanosomiasis may result from parasite-induced alteration of very early events during lymphocyte activation, leading to a virtually complete block in cell cycle progression and inhibition of IL-2R expression. [ABSTRACT FROM AUTHOR]

Immunohistological studies of human first trimester pregnancy decidua demonstrated the presence of the p75 interleukin-2 receptor (IL-2R) and the pI45 interleukin-4 receptor protein (IL-4R) on cells in the decidual stroma; there was no expression of CD25, the p55 IL-2R. The IL-4R was also expressed on the basal face of the glandular epithelial cells. Flow cytometric analysis of antibody-labelled decidual cell dispersions confirmed these results. Double antibody labelling demonstrated that p75 was expressed exclusively on the CD56-positive decidual large granular lymphocytes (LGL), whereas the IL4-R was expressed on some decidual LGL, and most decidual macrophages and T cells. In vitro incubation of decidual cells with IL-2 failed to induce expression of p55 or to increase the expression of either p75 or the pI45 IL-4R. Purified decidual LGL proliferated in vitro in response to IL-2, and IL-4 inhibited this IL-2-induced proliferation. [ABSTRACT FROM AUTHOR]