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Patients affected by AIDS-related complex (ARC) show several immunological abnormalities which may lead to a disregulation of immunosurveillance against viral latent infections. In this paper evidence for Epstein-Barr virus (EBV) reactivation in seven out of eight affected by persistent generalized lymphadenopathy is given. These patients showed either elevated levels of circulating EBNA-positive transformed cells or depressed EBV-specific T cell cytotoxicity, as assessed by the regression assay, or both. A direct involvement of EBV in the pathogenesis of ARC is thus suggested. Natural killer cell activity was found decreased, correlating to the evidence of circulating EBV-infected cells and of impaired EBV-specific immune control. These data provide evidence that, when specific immune mechanisms lose control on ubiquitous latent viruses, the risk for reactivation becomes higher. In the case of EBV, direct evidence of this event is provided by the emergence in the peripheral blood of clones of infected cells with unlimited growth potential. [ABSTRACT FROM AUTHOR]

The intrinsic immunosuppressive properties of regulatory T (Treg) cells can be harnessed for therapeutic approaches aiming at down-modulating harmful immune reactions. In this context, expanded type 1 Treg cells (Tr1 cells) specific for ovalbumin (ova-Tr1 cells) have been tested for clinical efficacy in the treatment of autoimmune disorders such as refractory Crohn's disease (CD). The clinical use of these therapeutic products warrants exploration of their mechanism of action. Here, we identified a relationship between the CD activity index and the expression of lytic molecules by the ova-Tr1 cells administered in the previously reported First-in-Man study [Crohn's And Treg cells Study 1 (CATS1) study]. Accordingly, ova-Tr1 cells were found to carry granules containing high levels of lytic molecules, including multiple granzymes and granulysin. These cells displayed a T-cell receptor (TCR)-independent cytotoxic activity, which was preferentially directed toward myeloid cell lines and monocyte-derived dendritic cells. Upon contact with myeloid cells, ova-Tr1 cells induced their apoptosis via a perforin-independent and a granulysin/granzyme-dependent mechanism. As compared to CD8+ cytotoxic T cells, ova-Tr1 cells required more time to lyse target cells and displayed a more gradual lytic activity over time. Notably, this activity was sustained over days resulting in the control of myeloid cell populations at a relatively low ratio. Our study reveals that ova-Tr1 cells are endowed with a sustained cytotoxic activity that relies on a unique combination of granulysin and granzymes and that preferentially eliminates myeloid target cells in a TCR-independent manner. [ABSTRACT FROM AUTHOR]

Summary: Pneumococcal infections cause a large global health burden, and the search for serotype‐independent vaccines continues. Existing conjugate vaccines reduce nasopharyngeal colonization by target serotypes. Such mucosal effects of novel antigens may similarly be important. CD4+ Th17 cell‐dependent, antibody‐independent reductions in colonization and enhanced clearance have been described in mice. Here we describe the evaluation of T helper type 17 (Th17) cytokine responses to candidate pneumococcal protein vaccine antigens in human cell culture, using adenoidal and peripheral blood mononuclear cells. Optimal detection of interleukin (IL)‐17A was at day 7, and of IL‐22 at day 11, in these primary cell cultures. Removal of CD45RO+ memory T cells abolished these responses. Age‐associated increases in magnitude of responses were evident for IL‐17A, but not IL‐22, in adenoidal cells. There was a strong correlation between individual IL‐17A and IL‐22 responses after pneumococcal antigen stimulation (P < 0·015). Intracellular cytokine staining following phorbol myristate acetate (PMA)/ionomycin stimulation demonstrated that  > 30% CD4+ T cells positive for IL‐22 express the innate markers γδT cell receptor and/or CD56, with much lower proportions for IL‐17A+ cells (P < 0·001). Responses to several vaccine candidate antigens were observed but were consistently absent, particularly in blood, to PhtD (P < 0·0001), an antigen recently shown not to impact colonization in a clinical trial of a PhtD‐containing conjugate vaccine in infants. The data presented and approach discussed have the potential to assist in the identification of novel vaccine antigens aimed at reducing pneumococcal carriage and transmission, thus improving the design of empirical clinical trials. [ABSTRACT FROM AUTHOR]

Gimap5, a member of the GTPase of the immunity-associated protein family (Gimap), regulates T cell survival. A strong indication of this is found in the diabetes-prone BioBreeding rat (BBDP), where a frameshift mutation in Gimap5 results in T-cell lymphopenia. We have investigated the function of human Gimap5 in T cells. We found that reduction of Gimap5 by RNA interference in Jurkat cells did not affect the number of apoptotic cells whereas transient over-expression of Gimap5 resulted in a major increase in the number of apoptotic cells. The same effect of over-expression was found in naive human T cells purified from blood but not in activated human T cells. This suggests that the apoptosis-inducing effect of Gimap5 over-expression is dependent on the activation status of the cells. Since the apoptosis-inducing effect of Gimap5 was contrary to the expected function of Gimap5 based on the phenotype of BBDP rats, we over-expressed rat wt Gimap5 and Gimap5 with the mutation found in BBDP (Gimap5-lyp). Both versions of rat Gimap5 induced apoptosis when expressed in the rat T-cell line C58(NT)D.1.G.OVAR.1, however, Gimap5-lyp greatly exacerbated cell death. Finally, we detected the subcellular localization of Gimap5 to be at the endoplasmic reticulum and by quantitative PCR, we found that endogenous Gimap5 mRNA is up-regulated in activated T cells. [ABSTRACT FROM AUTHOR]

In unimmunized specific pathogen-free mice, there are unique memory-type CD8+ T cell populations expressing asialoGM1 (ASGM1). These cells were classified into central memory-type T cells (TCMT) judging from their expression profile of CD44, IL-2Rβ, CD62L and CCR7 cell-surface molecules. Among CD44highCD8+ so-called memory CD8+ T cell population, ASGM1+CD44highCD8+ TCMT, but not ASGM1−CD44highCD8+ memory T cells, produced IFN-γ by stimulation with anti-CD3 mAb. The physiological significance of ASGM1+CD8+ TCMT as early source of IFN-γ was also demonstrated in vivo. Namely, intravenous injection of anti-CD3 mAb (2 μg) resulted in early activation of IFN-γ-producing ASGM1+CD8+ TCMT cells as well as NKT and NK cells. Unexpectedly, however, few IFN-γ-producing CD4+ T cells were detected until 4 h after anti-CD3 mAb administration. Thus, ASGM1+CD8+ TCMT were demonstrated to be early IFN-γ producer, which may be crucial for Th1-dependent cellular immunity. Indeed, co-culture of naive CD4+ T cells with ASGM1+CD8+ TCMT but not ASGM1−CD8+ T cells caused a great acceleration of IFN-γ-producing Th1 cells in vitro. Finally, we found that Th1-prone C57BL/6 mice possessed higher percentage (10%) of ASGM1+CD8+ TCMT in CD8+ T cells compared with that (3%) of Th2-prone BALB/c mice. Moreover, ASGM1+CD8+ TCMT derived from C57BL/6 mice produced higher levels of IFN-γ compared with those from BALB/c mice. Thus, ASGM1+CD8+ TCMT, whose differentiation in vivo is genetically controlled, appear to play a critical role in the control of type 1 immunity, which is essential for therapy of tumors and infectious diseases. [ABSTRACT FROM AUTHOR]

Pepstatin A is well known to be an inhibitor of aspartic proteinases such as pepsin, cathepsins D and E. Except for its role as a proteinase inhibitor, however, the pharmacological action of pepstatin A upon cells remain unclear. In this study, we found that pepstatin A suppressed receptor activator of NF-κB ligand (RANKL)–induced osteoclast differentiation. Pepstatin A suppressed the formation of multinuclear osteoclasts dose-dependently. This inhibition of the formation only affected osteoclast cells, i.e., not osteoblast-like cells. Furthermore, pepstatin A also suppressed differentiation from pre-osteoclast cells to mononuclear osteoclast cells dose-dependently. This inhibition seems to be independent of the activities of proteinases such as cathepsin D, because the formation of osteoclasts was not suppressed with the concentration that inhibited the activity of cathepsin D. Cell signaling analysis indicated that the phosphorylation of ERK was inhibited in pepstatin A-treated cells, while the phosphorylation of IκB and Akt showed almost no change. Furthermore, pepstatin A decreased the expression of nuclear factor of activated T cells c1 (NFATc1). These results suggest that pepstatin A suppresses the differentiation of osteoclasts through the blockade of ERK signaling and the inhibition of NFATc1 expression. [ABSTRACT FROM PUBLISHER]

We investigated the co-distribution of lymphocyte subpopulations and non-lymphoid ‘accessory’ cells in 35 cases of cutaneous lymphoproliferative diseases (T-cell lymphoma, 10 cases; B-cell lymphoma, 17 cases; pseudolymphoma, 8 cases) using immunohistochemical methods. T-zone histiocytes and particularly Langerhans cells were abundant in all cutaneous T-cell lymphomas, but were also found in B-cell lymphomas. T-zone histiocytes were associated with T-lymphocytes, especially T-helper cells., but not with T-suppressor cells. Dendritic rcticulum cells were essentially confined to well differentiated germinal centres. Macrophages occurred in both lymphomas and pseudolymphomas without definite relationship with either B- or T-cells. In malignant lymphomas of high grade malignancy, macrophages represented the only non-lymphoid cell type. Our results indicate that malignant lymphoid cells, like normal lymphocytes, require definite micro-environments which are, at least in part, maintained by certain non-lymphoid cells. [ABSTRACT FROM AUTHOR]

The identification of mononuclear cells extracted from cutaneous tumours (basal cell carcinoma, squamous cell carcinoma and superficial spreading melanoma) has been investigated. The relative numbers of T cells and B cells have been determined using the E-rosette test and the EAC-rosette test. The results have been compared to those of delayed hypersensitivity type reactions. Different cell distribution patterns (E/EAC ratio) have been found in the infiltrates according to the type of tumour. An immunocytochemical technique has been developed for the identification in situ of immunoglobulin- producing cells in the inflammatory infiltrates. In each case the class of immunoglobulin (IgM, IgG or IgA) has been identified and the relative frequency of Ig-producing cells has been determined. The results indicate humoral and cellular immune responses with variations attributable to the type of tumour. In weakly malignant tumours, the infiltrate is characterized by an elevated number of T lymphocytes and numerous plasma cells which secrete all classes of Ig; in highly malignant tumours it is characterized by a reduced number of both T lymphocytes (E rosette) and plasma cells which do not secrete all classes of Ig. [ABSTRACT FROM AUTHOR]

Anti-endothelial and other cell membrane-reactive antibodies in scleroderma were characterized by immunoblotting sera with membrane and cytosol preparations of human umbilical vein endothelial cells (HUVEC). dermal fibroblasts and a T cell lymphoma HUT78. Antibodies reactive with HUVEC membranes were found in 17 of 20 patients with scleroderma (33 bands) in contrast to only two of 20 controls (two bands; P < 0.01) and three of 11 patients with myocardial infarction (four bands). Eleven of the 20 patients possessed antibodies that were specific for HUVEC membrane and did not cross-react with other cell lines. Analysis of patient subgroups showed that HUVEC membrane antibodies were present in nine of 11 patients with systemic sclerosis and in all nine with the CREST syndrome, and were HUVEC-specific in five and six of these cases, respectively. Although considerable heterogeneity was seen, antibodies to an 18-19-kD membrane epitope were found in 11 of the 20 patients but in none of the controls (P < 0.01). This antibody which reacted particularly with HUVEC (n = 9) and HUT78 membranes (n = 9) was associated with CREST syndrome rather than systemic sclerosis (9/9 versus 1 /11 ; P<0.01), and after elution was shown to possess anticentromere activity. In addition, antibodies reactive with both fibroblast (n = 11; 18 bands) and HUT78 membranes (n = 18; 42 bands) were detected and were specific for either fibroblast or HUT78 membranes in nine and 14 patients, respectively. There was no significant difference in the incidence of these fibroblasts and HUT78 membrane antibodies in the two patient subgroups. These findings support the concept that membranereactive antibodies, including anticentromeric antibodies, may play a central role in the pathogenesis of scleroderma, through their ability to react with endothelial cells. [ABSTRACT FROM AUTHOR]

T cells from patients with CVID have defects that may relate to the failure in vivo of B cell production of antibodies. Antigen-driven responses of T cells from CVID patients and normal subjects have been assessed by measuring DNA synthesis in vitro. Low density cells enriched for antigen-presenting dendritic cells were pulsed with purified protein derivative (PPD) and cultured with autologous T cells. Overall, T cells from CVID patients showed a significantly low mean response to PPD, although non-specific DNA synthesis induced in CVID T cells by IL-2 was within the normal range. However, mean PPD-specific T cell responses in CVID were not restored by IL-2 irrespective of the presence of monocytes. Depletion of CD8+ cells also failed to restore the mean PPD response of CVID CD4+ T cells. Limiting dilution analysis showed that in CVID there was a reduced frequency of antigen-specific cells within the T cell preparations. The mean frequency of the PPD-specific T cells in cultures from patients vaccinated with bacille Calmette Guérin (BCG) was reduced to 1 in 109000 T cells compared with 1 in 18600 T cells in BCG-vaccinated normal donors. These data show that the reduced PPD-specific response in CVID is due to a partial peripheral loss of antigen-specific cells. [ABSTRACT FROM AUTHOR]

The TCR repertoire of human CD8+ peripheral blood lymphocytes has been determined using MoAbs to the Vβ32, 3,5.1,5.2/5.3,6.7,8, 12 and 19(17) Vβ gene families. The CD8 T cell repertoire for Vβ2 and Vβ3 is shown to be skewed, with an excess of individuals having higher values than are consistent with a normal distribution. A significant majority of these individuals are over the age of 40. High values of Vβ CD8+ cells were found for each Vβ family studied except for 6.7a. Individual high values are stable for at least 12 months. In addition, the total percentage of CD4 and CD8 cells reacting with this panel of reagents was determined. There is a significant excess of Vβ CD4+ cells (33%) over CD8+ Vβ+ cells (21.9%). Thus the human CD8 Vβ repertoire differs from the human CD4 repertoire in a number of important ways. [ABSTRACT FROM AUTHOR]

Cytokines are important mediators involved in the development of effector cells and in the regulation of immune responses. The gene expression of these mediators in T cell subset has yet to be fully elucidated. Using sensitive reverse transcription-polymerase chain reaction (RT-PCR), the kinetics of cytokine gene expression in human CD4+ and CD8+ T cells were examined. CD4+ T cells were more readily activated by phorbol myristate acetate (PMA) and phytohaemagglutinin (PHA) than CD8+ T cells in terms of the IL-2 receptor (IL-2R) mRNA expression. Quantitative differences in cytokine gene expression between CD4+ and CD8+ T cells were confirmed and higher levels of cytokine mRNAs were induced in CD4+ than in CD8+ T cells. Early induction of IL-2 mRNA was observed in both T cell subsets. The demonstration of different kinetics of cytokine gene expression illustrates one of the examples of the complexity of immunoregulation. The differential response of cytokine gene expression in different T cell subsets should be taken into consideration when clinical studies in cytokine production by peripheral blood mononuclear cells are interpreted. [ABSTRACT FROM AUTHOR]

Normobaric O2 exposure decreased spleen cell (SC) response to T cell mitogen Con A. ³H-TdR incorporation of SC from O2 exposed mice (O2SC) compared to those of control mice (Air SC) decreased significantly after 72 and 87 h O2 exposure. The dose response kinetics to Con A were identical in O2SC or Air SC. Increasing SC number did not restore the response to Con A and the depressed hyperoxic effect was not related to suppressor cells in the spleen of O2 exposed mice. Response of O2SC to Con A was restored by the thiol compound 2-niercaptoethanol (2-ME). and the degree of restoration by 2-ME. was inversely proportional to the depressed response. Addition of intact peritoneal cells (PC) induced restoration within the same range as 2-ME. Restoration of the mitogenic response by 2-ME involved antioxidant properties and suggested that macrophages were functionally injured by O2 exposure. In cases where mitogen response was highly depressed, restoration was only partial; in these conditions in vivo O2 injury probably involved both macrophages and splenic T cells. The mechanisms of O2 toxicity have been discussed in terms of free radical generation under hyperoxic conditions. [ABSTRACT FROM AUTHOR]

Suppressor cell activity has been demonstrated in the peripheral blood of patients with leprosy. Cells bearing the suppressor/cytotoxic phenotype have been enumerated in both peripheral blood and tissues, and microanatomical differences in tissue distribution have been observed. This first generation of studies has been characterized by considerable disagreement, a not unusual circumstance in the study of leprosy. In the case of blood suppressor cell activity, there appears to be no doubt as to its existence, but much uncertainty regarding its distribution. Concerning peripheral blood phenotypic suppressor cells, the observed differences in lepromatous and ENL patients may well reflect differences in methods used. Concerning phenotypic suppressor cells in tissue, there is no agreement as to their numbers or microanatomical distribution across the spectrum of leprosy or in its reaction states. Although these observational differences make firm conclusions impossible, this first generation of studies has provided new ways of considering old problems. For example, lepromin unresponsiveness might be a consequence of active cellular suppression. Differences in the numbers (or percentages) of the suppressor phenotype in blood or tissues of lepromatous patients with or without ENL reopens the door to the possibility of cell-mediated immune mechanisms in the pathogenesis of ENL. The identification of defective suppressor cells as important in the pathogenesis of hypergammaglobulinaemia is of interest in and of itself, but also gives rise to the possibility that other kinds of phenomena may be a consequence of defective or effete suppressor mechanisms. The observation of microanatomical differences in the distribution of the suppressor phenotype in tuberculoid and lepromatous leprosy indicates that effective or ineffective immunity might be a sequela of particular interactions between the suppressor/cytotoxic and helper/inducer phenotypes, and that these interactions merit further study. These new perspectives may be subject to experimental testing by the next generation of studies, which will surely include the techniques of clonal expansion and limiting dilution, as well as the study of interleukins 1 and 2. [ABSTRACT FROM AUTHOR]

Protein energy malnutrition decreased the number of rosette forming T lymphocytes, of T4 positive cells and their ability to provide help to B cells in antibody synthesis. There was a reduction in serum thymic hormone activity and an elevation of leucocyte terminal transferase and plasma cortisol levels. The numerical and functional deficiency of T4+ helper cells may be important in the pathogenesis of some of the clinical and immunological manifestations of protein energy malnutrition. [ABSTRACT FROM AUTHOR]

Evidence is presented that T lymphocyte cytotoxicity is mediated by hydrogen peroxide (H2O2). Ata concentration of 5 ×10-4 MH2O2 induced 51Cr release from pre-labelled P815 mastocytoma cells. H2O2 was generated when T lymphocytes from mouse spleen were exposed to P815 cells. The concentration of H2O2 produced was apparently one thousand times lower than the concentration required to lyse the P815 cells. This suggests that the H2O2 is produced and acts at a highly localized site on the target cell. Sulphydryl groups on the target cell were particularly sensitive both to H2O2 and to spleen cell attack. The activity of the spleen cells was inhibited by cyanide and azide and by reducing agents which protected the target cells. Cytotoxicity was enhanced by agents which prevented H2O2 breakdown. [ABSTRACT FROM AUTHOR]

Purified human mononuclear cell subpopulations have been evaluated in the in vitro lymphocyte proliferation assay. Monocyte-depleted mononuclear cells had reduced or absent responses to mitogens and antigens which could be restored to the original mononuclear cell response by addition of purified plastic adherent cells. Purified T lymphocytes obtained by density gradient centrifugation of E-rosette-forming cells demonstrated low but significant proliferative responses to mitogens, but no significant response to antigens. The addition of monocytes potentiated the response of purified T cells to mitogens and antigens, but did not fully reconstitute the original mononuclear cell response unless non-T lymphocytes were also present. It is concluded that mononuclear cell proliferation represents a complex mechanism of cellular interaction involving multiple subpopulations of cells. [ABSTRACT FROM AUTHOR]

Blood lymphocytes from eight patients with chronic lymphatic leukaemia (CLL) were stimulated by phytohaemagglutinin (PHA). The effect was defined as a pronounced increase of ³H-thymidine incorporation, and a concomitant blast transformation of a large portion of surface Ig-positive, E-rosette-forming (E-RFC) negative cells. E-RFC depletion reduced these effects to nearly background levels, and the pooled data strongly suggest that malignant R-lymphocytes in CLL are capable of responding to a PHA-induced T cell factor. A possible use for this factor in characterizing different subpopulations of CLL cells is discussed. [ABSTRACT FROM AUTHOR]

Some properties of the receptor for IgM on human lymphocytes have been investigated. It was shown that the interaction of native IgM with the receptor present on T and B lymphocytes is not critical for its detection in the EAM-rosette assay. In fact, high values of EAM-RFC could be found on cell suspensions cultured overnight in either IgM-free or IgM-containing media. In addition, the inhibition of EAM-rosettes by human monoclonal IgM at 37°C was not as effective as at 4°C. Rabbit IgM showed a significantly greater ability to inhibit the binding of antigen-IgM antibody complexes than human IgM. The receptor for IgM was easily removed by handling procedures, the incubation of lymphocytes at 4°C and treatment of the cells with low concentrations of trypsin or pronase. After the enzymatic treatment, a rapid resynthesis occurred, which restored the number of EAM-rosettes formed by T cells and significantly increased the number formed by B cells. The interaction between the receptor and antigen-IgM antibody complexes stopped the spontaneous shedding of the receptor at 4°C. When the incubation of the cells with immune complexes was performed at 37°C, a .significantly different behaviour between T cells equipped with receptor for IgM and those possessing receptor for IgG was found. After the binding of EAG to the receptor for IgG, a process of rapid dissociation of rosettes occurred, whereas the incubation with EAM did not induce an irreversible loss of the receptor for IgM. [ABSTRACT FROM AUTHOR]

The mean electrophoretic mobility (EPM) of blood lymphocytes from a patient with Brutton type of agammaglobulinaemia was high with a unimodal distribution pattern. That of blood lymphocytes from a patient with severe thymic dysplasia was low, with a small faster group. The difference between the major groups was about 30%. Thymocytes had a mean EPM 25%, lower than that of normal T cells. Blood and spleen cells in 8 chronic lymphatic leukaemia of B-cell origin had a mean EPM 35% higher than that of the majority of normal B cells. [ABSTRACT FROM AUTHOR]

The article focuses on the pathogenesis of mycosis fungoides (MF). MF is a cutaneous lymphoma characterized by an initially polymorphous infiltrate composed largely of T-lymphocytes. The dermal infiltrate invades the epidermis, forming small accumulations of T-lymphocytes termed Pautrier's microabscesses. Atypical lymphocytes with hyperconvoluted nuclei, termed as Lutzner or mycosis cells, are found in the cutaneous infiltrates, lymph nodes, or peripheral blood of MF patients. A leukaemic variant of MF, the Sézary syndrome, is characterized by erythroderma and a large percentage of a typical lymphocytes in the peripheral blood and tissue infiltrates.

Isoglobotriaosylceramide (iGb3) is a stimulatory antigen for a unique type of T cell, Natural Killer T cells. Produced in the lysosomal compartment by mammalian antigen-presenting cells, iGb3 is one of the few clearly identified carbohydrate ligands for biological receptors. A major source of glycoconjugate structural diversity arises from the possibility of forming different linkages between the same monosaccharide units. Globotriaosylceramide (Gb3) exists as a natural isomer for iGb3, and both isomers are frequently found together in mixtures of glycosphingolipids extracted from mammalian cell membranes. Discriminating these isomers has been feasible using monoclonal antibodies raised against specific carbohydrate epitopes, or by unambiguous structural characterization, which requires relatively large amounts of pure compounds isolated from grams, or tens of grams, of biological samples. However, the precise detection of iGb3 from small amounts of biological samples, where it may be mixed with Gb3 present in much higher abundance, is a prerequisite for answering further important biological questions such as stimulation of NKT cells. Here we describe a specific and sensitive method based on ion trap mass spectrometry to discriminate iGb3 from Gb3. We also demonstrate its application to quantifying the amount of iGb3 in a prototype antigen-presenting cell, rat RBL-CD1d cells, using a chemically synthesized short N-acyl chain iGb3 as internal standard. This methodology may have wide implications for functional glycosphingolipidomics of immune cells and glycosphingolipid biomarker analysis. [ABSTRACT FROM AUTHOR]