Antibody-independent complement activation by a subpopulation of mitogen-induced human T lymphoblasts.

A study was made of the complement activation (CA) capacity of human T lymphoblasts obtained in vitro by PHA stimulation of highly purified T cells. The CA was studied by immunofluorescent detection of the C3 fragments deposited onto the cell membrane following incubation in autologous normal human serum (NHS). Whereas resting T cells did not stain, a mean of 12% of the blast cells were positive for monospecific anti-human C3 fluorescent serum. The possible mechanism through which C3 was deposited on the cell membrane may be a consequence of CA, because complement membrane fluorescence (CMF) was abolished by chelation of divalent cations with EDTA, complement inactivation by heating or incubation at low temperature. This reaction appears to occur in the absence of antibodies, because similar results were found when human agammaglobulinaemic serum was used as the complement source. The addition of ethyleneglycolbis-(aminoethylether)-tetraacetate (EGTA), supplemented with MgCl2 to the NHS failed to abolish the fluorescence, indicating that the alternative pathway was involved in the phenomenon. It is likely that the capacity to activate complement may distinguish a functional subpopulation of human T lymphoblasts.     

T and B-cell activation by mitogenic factor generated from antigen-stimulated lymphocytes.

Guinea-pig mitogenic factor(s) were found capable of stimulating both normal T and B cells. Preliminary physicochemical characterization could not dissociate the mitogenic factors for respective cell populations. Furthermore, both immune T and B cells could produce mitogenic factors upon stimulation with antigen in contrast to some other lymphokines which are known to be produced only by immune T cells. While T-cell-derived mitogenic factor was active on both allogenic T and B cells, it was only mitogenic to syngeneic B cells and not to syngeneic T cells. B-cell-derived factor, on the other hand, could stimulate both T and B cells either of syngeneic or allogeneic origin.     

Age-related changes in the expression of T cell activation antigens following phytohaemagglutinin stimulation.

The role of accessory cells (AC) in the temporal expression of several key T-cell-activation-associated antigens has been studied in healthy aged subjects. Compared to responses seen in young adults, phytohaemagglutinin (PHA) induced weak proliferation in peripheral blood mononuclear cells from the aged and lower numbers of T cells expressing CD71, CD25, CD38 or HLA-DR. T cell responses to the monoclonal antibody OKT3, however, were normal. Whereas HLA-DR+ T cell numbers could be increased by raising the AC content (up to 50%) in cultures comprising purified T cells and graded numbers of autologous AC, CD25+ T cell numbers remained largely unaltered. Co-stimulation with PHA + phorbol myristate acetate in the absence of AC restored both proliferation and CD25 expression in the aged. These results indicate that T cells from the healthy aged show selective deficiencies in their capacity to respond to mitogenic stimuli and suggest that impaired PHA responsiveness is due, at least in part, to defective AC-derived signals.     

Age-related differences in brain activation during emotional face processing

Advancing age is associated with significant declines on neurobehavioral tasks that demand substantial mental effort. Functional imaging studies of mental abilities indicate that older adults faced with cognitive challenges tend to activate more regions, particularly frontal, than their younger counterparts, and that this recruitment of additional regions may reflect an attempt to compensate for inefficiency in cortical networks. The neural basis of emotion processing in aging has received little attention, and the goal of the present study was to use functional magnetic resonance imaging (fMRI) to examine the influence of age on facial emotion processing and activation in cortical and limbic regions. Participants (eight old and eight young adults) viewed facial displays of happiness, sadness, anger, fear, disgust, and neutrality in alternating blocks of emotion and age discrimination. We predicted that in response to an emotion discrimination task, older adults would demonstrate increased use of frontal regions relative to younger adults, perhaps combined with diminished use of regions recruited by younger adults, such as temporo-limbic regions. During the emotion discrimination task, young participants activated, visual, frontal and limbic regions, whereas older participants activated parietal, temporal and frontal regions. A direct comparison between emotion and age discrimination revealed that while younger adults activated the amygdala and surrounding temporo-limbic regions, older adults activated left frontal regions. The results of this study suggest that older adults may rely on different cortical networks to perceive emotional facial expressions than do their younger counterparts.     

Activation of human T lymphocytes II. Involvement of the T3 antigen in polyclonal T cell activation by mitogenic lectins and oxidation

The effect of monoclonal antibodies against the T cell surface antigens T3 and T4 on accessory cell-dependent mitogen-induced proliferation of human antigen-specific T4⁺ T lymphocyte clones and purified peripheral blood T cells was studied. To avoid the Fc receptor-dependent mitogenic effects of the OKT3 antibodies, monocytes were replaced by Epstein-Barr virus-transformed B lymphoblastoid cells as accessory cells. Monoclonal antibody OKT3 but not OKT4 inhibited the response of both T lymphocyte clones and purified T cells to mitogenic lectins and oxidation. The inhibition was not due to nonspecific effects of the monoclonal antibodies since it affected only the initial triggering but not the proliferation of activated T cells and it could be overcome by higher concentrations of lectin indicating a competition between OKT3 antibody and lectin. Furthermore, OKT3 antibody at the same concentration that was inhibitory in this system was mitogenic in the presence of Fc receptor-bearing monocytes. Surface modulation of T3 but not of T4 antigens led to unresponsiveness to a mitogen pulse given directly after modulation. These findings suggest that antigen-specific and mitogen-induced T cell triggering are due to interaction with the same receptors of the T lymphocyte.     

Age-related changes in brain activation during a delayed item recognition task

To test competing models of age-related changes in brain functioning (capacity limitation, neural efficiency, compensatory reorganization, and dedifferentiation), young (n=40; mean age=25.1 years) and elderly (n=18; mean age=74.4 years) subjects performed a delayed item recognition task for visually presented letters with three set sizes (1, 3, or 6 letters) while being scanned with BOLD fMRI. Spatial patterns of brain activity corresponding to either the slope or y-intercept of fMRI signal with respect to set size during memory set encoding, retention delay, or probe stimulus presentation trial phases were compared between elder and young populations. Age effects on fMRI slope during encoding and on fMRI y-intercept during retention delay were consistent with neural inefficiency; age effects on fMRI slope during retention delay were consistent with dedifferentiation. None of the other fMRI signal components showed any detectable age effects. These results suggest that, even within the same task, the nature of brain activation changes with aging can vary based on cognitive process engaged.     

Changes in the T cell receptor macromolecular signaling complex and membrane microdomains during T cell development and activation

Initiation and propagation of T cell receptor signaling pathways involves the mobilization and aggregation of a variety of signaling intermediates with the T cell receptor and associated molecules into specialized signaling complexes. Accumulating evidence suggests that differential regulation of the formation and composition of the T cell receptor macromolecular signaling complex may affect the different biological consequences of T cell activation. The regulatory mechanisms involved in the assembly of these complexes remains poorly understood, but in part is affected by the avidity of the T cell receptor ligand, co-stimulatory signals, and by the differentiation state of the T cell.     

Age-related defects in CD4+ T cell activation reversed by glycoprotein endopeptidase

CD4(+) T cells from old mice show defects in the activation process including deficiency in the formation of immunosynapses with antigen-presenting cells. We show that CD4(+) T cells from old mice express unusually high levels of glycosylated forms of the bulky T cell glycoprotein CD43, particularly on a subset of functionally anergic cells expressing P-glycoprotein. T cells from old donors also show a decline in the association of CD43 with cytoskeletal matrix and in the proportion of T cells that can exclude CD43 from the synapse. O-sialoglycoprotein endopeptidase, which removes the external domain of CD43 and other O-sialoglycoproteins from the aged naive CD4(+) T cells of TCR-transgenic mice, restores early agonist-independent stages and later agonist-dependent stages of synapse formation as well as expression of the activation markers CD69 and CD25 to the levels found in the young mice. These data support a model in which O-glycosylated forms of T cell surface molecules, including CD43, are largely responsible for age-related defects in TCR signaling and function.     

Age-related changes in cerebral blood flow activation during a Card Sorting Test

 To determine the age-related changes in the neural processing involved in the Modified Card Sorting Test (MCST), we measured cerebral blood flow (CBF) during performance of the MCST and of the number-matching task in young and elderly subjects using positron emission tomography. Compared with that during the number-matching task, CBF during the MCST was increased in the left dorsolateral prefrontal cortex (DLPFC), left inferior parietal lobule, and left striate and prestriate cortices in both age groups. However, CBF activation in these areas was significantly lower in the elderly subjects than the young subjects. Furthermore, CBF activation was reduced in the left DLPFC, right parahippocampal gyrus, and prestriate cortex in proportion to the increase in the number of perseverative errors with aging. These results suggest that the impaired MCST performance in elderly subjects may be due, in part, to dysfunction of the network involving certain cortical areas such as the prefrontal and parahippocampal cortices, although the essential neural circuits for MCST performance were still preserved even in the elderly subjects. Received: 10 September 1996 / Accepted: 13 January 1997     

Targeting early events in T cell activation to construct improved vaccines

Live, attenuated vaccines currently offer the best protection against virulent pathogens. Recent advances in Immunology and Molecular Biology provide an opportunity to design vaccines that will be more effective and safer than existing ones. Immunologists are rapidly developing the capacity to identify and construct the minimal immunogenic units from pathogens. The molecular signals required to fully activate antigen presenting cells (APCs) and responder T cells are becoming apparent. Improved vaccine delivery systems are being designed which will mimic the actions of pathogens in vivo. These vaccines will incorporate protective epitopes fused to immunoregulatory cytokines in chimeric proteins. They will be encapsulated in formulations which allow for the slow release of these chimeric proteins thereby inducing the memory T cells required for long-lived immunity. These vaccine formulations will target receptors present on the most active APCs. Here we discuss how these advances will allow us to rationally construct "virtual pathogens" which will provide improved protection against new and old microbial foes.     

Preferential activation of helper/inducer T lymphocytes in autoimmune chronic active hepatitis.

We found a significant increase of activated circulating T lymphocytes expressing interleukin 2 receptor (IL-2r) (mean +/- s.e.m. 11.0 +/- 1.1%) or DR antigen (5.0 +/- 0.49%) in patients with autoimmune chronic active hepatitis (CAH) starting in childhood when compared to healthy controls (0.14 +/- 0.09%, P less than 0.001 and 2.8 +/- 0.06%, P less than 0.01). Patients with liver disorders due to Wilson's disease (IL-2r 0.64 +/- 0.25%, DR 3.5 +/- 0.22%) or alpha-1-antitrypsin deficiency (IL-2r 0.1 +/- 0.06%, DR 2.8 +/- 0.35%) had levels similar to controls. Levels of both IL-2r and DR positive T lymphocytes were higher in patients with uncontrolled CAH (IL-2r 18.0 +/- 1.01%; DR 6.3 +/- 0.78%) than in patients with inactive disease (IL-2r 3.2 +/- 1.4%, P less than 0.001; DR 3.0 +/- 0.13%, P less than 0.01). In patients with active disease levels of IL-2r positive cells were higher than DR positive cells (P less than 0.001). Only 21% of activated T cells coexpressed the two markers of activation. Sixty-seven percent of IL-2r positive T lymphocytes were helper/inducer and 25% suppressor/cytotoxic, while 66% of the DR positive T cells were suppressor/cytotoxic and 31% helper/inducer. The finding that the highest levels of activated T lymphocytes are present in patients with uncontrolled CAH suggests that these cells are involved in its pathogenesis. The preferential increase of activated helper/inducer cells might explain the enhanced immune reactivity characteristic of autoimmune CAH.     

T cell subpopulation dynamics following insulin-induced hypoglycaemia in normal subjects.

Using monoclonal antibodies OKT3, OKT4 and OKT8, T lymphocyte subpopulations were determined in eight normal male volunteers. One month later, the T cell populations were again measured before and during an insulin stress test. Compared to the month before, there was a statistically significant reduction in the numbers of OKT4 cells (P less than 0.01) in the basal sample. Administration of insulin produced a statistically significant rise in the numbers of total lymphocytes and in each of the T cell subpopulations at 30 and/or 60 min (P less than 0.01) when compared with the basal values. It was also noted that in some of the subjects, the sum of OKT4 and OKT8 cells was greater than the number of OKT3 cells after insulin administration. This suggests that under certain circumstances T cells in circulation may express both the helper and suppressor cell antigen. Insulin stress test is associated with increased production of stress hormones in response to the hypoglycaemia, and the observed lymphocyte changes may be mediated via these hormonal alterations.     

Analysis of CD4-positive T cell subpopulation in sarcoidosis.

Double-labelling immunofluorescence analysis within the CD4+ cell subset was carried out in 27 bronchoalveolar lavage fluids and 11 peripheral blood samples of sarcoidosis patients with anti-TQ1, anti-2H4 and anti-4B4 monoclonal antibodies. Helper/inducer CD4+TQ1-/4B4+ cells were strongly increased in the lung and slightly, but significantly, decreased in the blood of sarcoidosis patients with respect to normal controls. No differences were found in the number of both lung and blood CD4+2H4+ cells between sarcoidosis patients and controls. The findings are further evidence for a compartmentalization of T cell subsets in sarcoidosis.     

Some early events in the primary mitogenic stimulation of lymphocytes differ from later interleukin stimulation and other quiescence to growth activation systems.

The requirement for activity of the enzyme ADP-ribosyl transferase (ADPRT) and changes in single-strand DNA breaks were assessed during the initial stimulation of quiescent murine splenic lymphocytes with mitogen alone, the stimulation of activated blasts with IL-2-containing medium and, for comparison, the serum stimulation of quiescent fibroblasts and the induction of haemoglobin synthesis in an erythromyeloid cell line K562. Inhibitors of ADPRT, at concentrations previously found to have no effect on the proliferation of lymphoblastoid cell lines, blocked the stimulation of spleen cells by Con A or LPS; non-inhibitory analogues had much less effect. No early increase in ADPRT activity after mitogenic stimulation was detectable. The rejoining of single-strand breaks was observed after stimulation of splenic lymphocytes with Con A, but not consistently with LPS. Conversely, ADPRT inhibitors had only little effect on the IL-2-induced stimulation of Con A blasts, and no effect on the stimulation of fibroblasts or K562. Neither were any changes in strand breaks associated with these systems. These findings implicate ADPRT activity and the rejoining of strand breaks in the early mitogenic response as being distinct from later IL-2 activation and changes from quiescence to growth in other cell types.     

Age-related changes of expression of IL-2 receptor subunits and kinetics of IL-2 internalization in T cells after mitogenic stimulation

The age-associated changes of the expression of IL-2 binding molecules p55/Tac(alpha chain) and p70/75(beta chain) were examined after phytohemagglutinin (PHA) stimulation. The expressions of both p55/Tac molecules and p70/75 molecules were significantly reduced in the aged compared with those in the young persons. The amounts of p55/Tac and p70/75 molecules on T cells from the aged were 55% and 59% of those on young ones, respectively. The ratio of the amount of p70/75 to that of p55/Tac in aged T cells was 0.28 and that in young ones was 0.26. The ratio was somewhat higher in the aged but not significantly. We also examined the kinetics of IL-2 internalization mediated by its receptor. The calculated t1/2 of receptor-mediated IL-2 internalization was 17 min in the aged and 16 min in the young, respectively. There was no kinetic difference between the 2 groups. The percentage of the internalized IL-2 to the sum total was 58.2% in the aged and 73.4% in the young (P less than 0.02). the amount of internalized IL-2 in T cells from the aged was 48.6% of that from the young (P less than 0.01).     

The pattern of activation antigen expression on T-lymphocyte subpopulation in infectious mononucleosis.

The peripheral blood mononuclear cells of patients with infectious mononucleosis (IM) have been characterized by the determination of activation antigens using a panel of 11 monoclonal antibodies (MoAbs) belonging to 9 clusters. The activation of CD8+ peripheral blood mononuclear cells of IM patients was found to be determined by HLA-DR and CD45RO MoAbs. The antibodies of the CD30, CD40 and CD70 termed antigens also showed an increased expression as compared to the controls. In contrast, the CD25, CD69, CD71, and CD45 antigens were expressed at a low rate on the surface of the CD8+ cells. We found, on the other hand, a very low percentage of B lymphocytes in the peripheral blood, which reflects on the virus caused antigen modulation and on the effectivity of activated CD8 cells with the characteristic expression of the above outlined markers in IM. In six asymptomatic patients, the percentage of CD8+HLA-DR+, as well as of CD8+CD45RO+ cells proved to be lower than that in the active phase of the disease. The number of B cells showed normal value in the clinically asymptomatic cases.     

Induction of high-affinity paf receptor expression during T cell activation.

Activated human T cells via the CD2 or the CD3 pathways exhibited a higher capacity than resting T lymphocytes to incorporate and metabolize [3H]pafacether (paf) at 37 degrees C. Resting T lymphocytes lacked specific binding capacity for paf, yet high-affinity paf receptors (paf-R) were induced on CD3- or CD2-dependent activation. This up-regulation in the number of paf-R became apparent by day 1 of culture, reached a maximum of about 25,000 sites cell by days 4 to 6 and subsequently declined. Interestingly, human recombinant interleukin-2 in a dose-dependent manner prevented the decrease of high-affinity paf-R expression on T cells. By contrast, the receptor affinity was constant throughout the culture period. Thus, paf-R at different stages of T cell activation were indistinguishable with respect to receptor-ligand interaction, and differed only in their number. Together, these data demonstrate that after activation human T cells develop membrane high-affinity paf-binding sites. They also suggest for the first time that expression of the paf-R are coupled to T cell activation and/or differentiation.