Search

Effective control of tuberculosis (TB) includes discrimination of subjects with active TB from individuals with latent TB infection (LTBI). As distinct interferon (IFN)-gamma and interleukin (IL)-2 profiles of antigen-specific T-cells have been associated with different clinical stages and antigen loads in several viral and bacterial diseases, we analysed these cytokines in TB using a modified QuantiFERON-TB Gold In Tube test. Detection of IL-2 in addition to IFN-gamma distinguishes not only Mycobacterium tuberculosis-infected subjects from healthy controls, but also individuals with LTBI from active TB patients. This may help to improve diagnostic tests for TB.

Flow cytometry and fluo-3/AM have been used to track cytosolic Ca++ modulation in human polymorphonuclear leukocytes (PMN) and T lymphocytes. The chemotactic peptide N-formylmethionyl-phenylalanine (FMLP) but not the phorbol ester PMA induced cytosolic Ca++ modulation in PMN along with forward and side scatter modification. PMA inhibited FMLP activity when preincubated with PMN. T lymphocytes were antigen specific T cell clones and were stimulated with various amounts of diverse superantigens or PHA. Data show that superantigens can induce either activation or anergy depending on culture conditions. The biological significance of these data are discussed.

Superantigens (SAg) interact with T lymphocytes bearing particular V beta sequences as part of their T cell receptor (TcR). The interaction, however, requires the presence of major histocompatibility complex (MHC) class II molecules on antigen-presenting cell (APC). In peculiar circumstances, MHC class II+ T cell clones (TCC) have been shown to present peptides and selected antigens interacting with antigen-specific TCC in the absence of APC. In this report we studied the capacity of SAg to mediate a T-T cell interaction, investigating the TCC ability to present a panel of staphylococcal enteroxins (SE) independently of the presence of added APC. Upon exposure to a broad range of SE concentrations, MHC class II+ TCC showed an intense proliferative response even in the absence of professional APC. Diverse SE optimally stimulated responder TCC at different concentrations. The proliferation was inhibited by anti-DR monoclonal antibodies, both in the presence and in the absence of APC. The SE activation of TCC in the absence of APC induced the same series of phenotypic variations as that observed following the TCC stimulation with APC. Irradiated TCC efficiently presented membrane-bound SE to responder TCC as well as professional APC. These results show that a single cell of a given clone effectively presents the SE to other cells of the same clone, and provide evidence that SAg can efficiently mediate T-T cell interaction. In addition, the possibility also exists that one cell of the clone can actually undergo an auto-stimulation via SAg-mediated interactions between its own TcR and MHC class II molecule. It has recently been suggested that the V beta-selective depletion of T cells observed in acquired immunodeficiency syndrome (AIDS) patients might be a consequence of the interaction between a human immunodeficiency virus (HIV)-encoded SAg and T cells expressing a SAg complementary V beta. We suggest that the hypothesized HIV-encoded SAg might mediate T-T cell interactions that could play a relevant role in the V beta-selective depletion of T lymphocytes observed in HIV-infected patients.

In this report we have analysed the kinetics of modulation of human peripheral blood T lymphocyte membrane molecules upon activation with optimal amounts of phytohaemagglutinin (PHA) and concanavalin A (ConA). The following activation-related and differentiation/adhesion molecules were selectively and concomitantly investigated on CD4+ and CD8+ subsets by dual colour flow cytometry: CD69, CD25 and CD71; CD2, CD45RA and L-selectin. Cultures were assayed after 24, 48, 72, 120 and 168 h of incubation with PHA and ConA. This approach allowed a comprehensive evaluation of membrane phenomena occurring during activation of normal resting human T lymphocytes. Data show that the kinetics of expression of these molecules follows a precise and consistent time-course with no major differences between CD4 and CD8 subsets. CD69 expression peaked at 24 h, whereas CD25 and CD71 expression peaked at 48/72 h with some differences between PHA and ConA activation. L-selectin expression started an evident decrease in step with culture time whose magnitude was dependent on the lectin used, being higher with PHA than with ConA. Conversely, the expression of CD45RA remained stable for 72 h and then briskly decreased with no major differences between PHA and ConA activation. CD2 molecules increased with time in number and density, although the percentage of positive cells remained essentially constant (greater than 85%). After 48/72 h of stimulation about 10% of cells co-expressed CD4 and CD8 molecules. To ascertain whether the phenomenon was restricted to cells in a particular activation state, the phenotype of cells in the diverse phases of the cell cycle was established. Results obtained show that only actively proliferating cells, that is cells in S and G2-M phases, co-expressed the two molecules, suggesting that such a phenomenon reflects a momentary dysregulation of the normal sequence of gene expression. The present data are also discussed in the light of the dynamic role of T lymphocyte activation and adhesion molecules in regulating cell-cell interactions, tissue localization and eventual immunological function.

The coordinate expression of CD4 and CD8 antigens defines four major subsets of adult mouse thymocytes (CD4+CD8-, CD4+CD8+, CD4-CD8+ and CD4-CD8-) that represent definite steps in the differentiative pathway of immature T-lymphocytes. Thymocytes from adult C57BL/6 mice were cultured in the presence of ST 789, an L-Arg synthetic derivative of hypoxanthine, (1 to 100 micrograms/ml) or PHA 60.1 to 1 microgram/ml), or both for 48 hours, and then stained with monoclonal antibodies to CD4 and CD8 antigens for dual color flow cytometry analysis. The expression of CD25, was also investigated. ST 789 was ineffective in modifying the distribution of the four thymocyte subsets and did not induce the appearance of CD25 on cortical thymocytes. PHA induced a dramatic dose-dependent decrease of the CD4+CD8+ subset that, however, was neither enhanced nor antagonized by ST 789. We also tested ST 789 for its ability to stimulate intracellular free Ca++ rise in thymocytes. The compound was ineffective in this assay. Conversely, thymocytes promptly responded to PHA stimulation. It is concluded that ST 789 has no effect on normal adult thymocyte differentiation/activation pathway.