Biomedical subjects
Publications and source records attributed to D Kabelitz.
Programmed cell death (apoptosis) is triggered in immature thymocytes and T-cell hybridomas by signalling via the CD3-T-cell receptor pathway. In this paper, Dieter Kabelitz and colleagues catalogue the recently accumulating evidence that apoptosis can also be initiated in mature peripheral T cells; this may constitute an important aspect of cellular immune regulation.
The frequencies of circulating donor-reactive cytotoxic lymphocyte precursors (CLP) and Il-2-producing helper lymphocyte precursors (HLP) were determined by limiting dilution analysis in 19 kidney allograft recipients before and at various intervals (up to 2 years) after transplantation. A significant, selective, and stable reduction of the frequencies of donor-reactive (but not of third party-reactive) CLP and/or HLP was observed in some patients beginning 3 to 6 months after transplantation. One patient developed reduced frequency of CLP only, 3 patients reduced frequencies of HLP only, and 2 patients reduced frequencies of both CLP and HLP. The selective reduction of donor-reactive CLP and/or HLP frequencies ranged from 5-25-fold when compared with the pretransplantation level and was associated with stable graft function. These data indicate that functional deletion of circulating donor-reactive T cells can occur at the level of cytotoxic T lymphocytes, Il-2-producing helper T lymphocytes, or both. Implications of these findings for the individualization of immunosuppressive regimens will be discussed.
We investigated the in vitro responsiveness of peripheral blood lymphocytes from two patients with T-cell chronic lymphocytic leukaemia (T-CLL) to Staphylococcus aureus enterotoxin (SE) superantigens. T-cell receptor (TcR) alpha beta (V beta 7.1)-expressing CD4+ leukaemic T cells from patient HE (white blood cell count 480,000/microliters) proliferated in response to SEA and, only at 1000-fold higher concentrations, to SEB, SED, and SEE. CD4+CD8+ TcR alpha beta (V beta 12.1)-expressing leukaemic T cells from patient KO (white blood cell count 120,000/microliters) were activated by SEB but not by the other tested SEs. In both instances, the activation of leukaemic T cells by SE was dependent on the presence of HLA-DR+ cells. Southern blot analysis of TcR beta gene rearrangement confirmed that the proliferating cells were derived from the leukaemic T-cell clone and not from contaminating normal T cells. These data indicate that leukaemic T cells from patients with T-CLL exert a clonally variable responsiveness to SE superantigens. We conclude that recognition of specific antigen and subsequent signal transduction can be initiated via the TcR of leukaemic T-CLL cells.
Peripheral blood gamma delta T cells expressing a V gamma 9/V delta 2 T-cell receptor are stimulated by killed bacteria including Mycobacterium tuberculosis (m.tb.) and group-A streptococci (strep A). In addition, recent data indicate that V gamma 9/V delta 2 T cells from unexposed individuals also respond to Plasmodium falciparum (P. falcip.) merozoites. Here we analyzed the reactivity to these ligands of 23 V gamma 9/V delta 2, 3 V gamma 9/V delta 1, and 4 V gamma 9-/V delta 1 clones derived from 8 healthy individuals after phytohemagglutinin stimulation of cell sorter-selected gamma delta T cells. Upon restimulation in the presence of irradiated antigen-presenting cells, the majority of V gamma 9/V delta 2 clones recognized m.tb. and strep A (but not strep D), and about one third of the clones also recognized P. falcip. Some clones, however, recognized only one or two of the tested ligands, and 4 V gamma 9/V delta 2 clones did not react at all. Interestingly, 2 of 3 V gamma 9/V delta 1 clones proliferated in response to m.tb., P. falcip., strep A and strep D, while V gamma 9-/V delta 1 clones were not activated by any of the tested ligands. Nucleotide sequence analysis indicated a broad diversity of V gamma 9 N regions in V gamma 9/V delta 2 clones. At the clonal level, our results demonstrate that individual V gamma 9/V delta 2 T cells can recognize m.tb., strep A, as well as P. falcip.-infected erythrocytes, with no influence of the expressed V gamma 9 N region.
The elements of allergic inflammation and the involvement of helper T lymphocytes are increasingly being recognized in the immunopathogenesis of asthma. Allergen exposure leading to the activation of allergen-specific T cells present in the lung can result in the release of cytokines which in turn can locally stimulate the cellular constituents of the lung. The airway epithelial cells may be the key participants in such an interaction. Therefore, we examined the ability of T-cell-derived IL-4 to modulate the production of C3 and C5 by the human type-II pneumocyte cell line A549, which is known to produce all the components and the regulatory proteins of the complement system. For estimation of C3 an ELISA detecting native C3 was used. Following stimulation of A549 with hrIL-4 a dose-dependent (1-50 U/ml) enhancement of C3 production was observed, which reached its maximum (5-fold of unstimulated cells) at 48 h and gradually declined thereafter. Concentrations of hrIL-4 higher than 50 U/ml did not further increase C3 production. In parallel experiments hrIFN-gamma at concentrations between 10 and 50 U/ml stimulated the C3 production to more than twice the quiescent state level within 24 h. In the pneumocyte cell line A549 we demonstrated the expression of a gene for the IL-4 receptor which appears to mediate the biological effect of this lymphokine. A diminution in the functionally active C5, estimated by ELISA at the same time, was observed in supernatants of A549 cultures following stimulation with hrIL-4 as well as with hrIFN-gamma.(ABSTRACT TRUNCATED AT 250 WORDS)
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Highly purified CD4-8- double-negative (DN) human thymocytes proliferated significantly more in response to interleukin (IL)-7 than to IL-2 in a dose-dependent fashion. Both IL-7 and IL-2 promoted the appearance of T-cell receptor (TCR) gamma delta-expressing cells in single cell suspension culture of DN thymocytes. While IL-7 exerted a vigorous growth-stimulating effect on TCR gamma delta-bearing DN thymocytes, IL-2 preferentially maintained the viability of gamma delta thymocytes without promoting their strong proliferation. In both instances, the major subset of gamma delta thymocytes expressed V delta 1 rather than V delta 2 as a variable delta-chain element on their surface. These results reveal differential effects of IL-7 and IL-2 on TCR gamma delta-bearing human thymocytes and underline the importance of both ILs in the development of DN thymocytes.
The in vitro effects of therapeutically administered mistletoe extracts (ABNOBAviscum) and pure mistletoe lectins (mainly mistletoe lectin I) on a variety of human and murine tumor cell lines have been investigated. Mistletoe extracts and purified mistletoe lectins inhibited in vitro the growth of all tumor cell lines tested including B cell hybridomas, P815, EL-4, Ke37, MOLT-4 and U937. The mechanism of growth arrest was shown to be due to the induction of programmed cell death (apoptosis). Thus, fragmentation of genomic DNA into oligonucleosomal bands of approximately 200 base pairs in length was observed within 20 h when tumor cells were incubated with mistletoe extracts or lectins. These data point to a rational basis for the direct cytotoxic effects of mistletoe extracts and lectins apart from the postulated immunostimulatory properties of these agents.
Two color immunofluorescence analysis of lymphocyte cell surface antigen expression using an unconjugated plus a biotinylated monoclonal antibody (mAb) requires four incubation steps: (1) unconjugated mAb; (2) fluorochrome-labelled goat anti-mouse Ig; (3) biotinylated mAb; (4) fluorochrome-labelled avidin or streptavidin. We describe a time-saving modification of this procedure which requires only two incubation steps: (1) simultaneous unconjugated and biotinylated mAbs; (2) fluorochrome-labelled avidin/streptavidin followed by fluorochrome-labelled goat anti-mouse Ig. The slightly delayed (5 min) addition of the goat anti-mouse Ig prevents it from binding to the mAb which has already interacted with avidin/streptavidin. Both procedures yield identical results with a variety of different mAbs.
Leukemic T cells from patient BU (WBC 22,000/microliter) resembled morphologically large granular lymphocytes, and expressed a V delta 1-encoded gamma delta T-cell receptor (TCR) on their surface. Upon in vitro activation with various mitogens, IL-2-dependent V delta 1+ cell lines were only obtained if V delta 1+ cells had been positively selected on a cell sorter before culture. However, even under these conditions, the V delta 1+ IL-2-dependent cell lines showed a TCR beta and gamma gene-rearrangement different from that of the freshly isolated leukemic cell population, indicating that they were not derived from the leukemic clone. Thus, known T-cell growth factors (IL-2, IL-4, IL-7) in concert with mitogenic signals failed to induce in vitro proliferation of the V delta 1+ leukemic clone. Interestingly, the patient responded to treatment with interferon-alpha 2c (daily dose of 10(6) I.U.). After 2 1/2 years of continuous interferon treatment, the patient is in partial remission with WBC around 8000/microliter.
We report that a human CD4+ T cell clone with specificity for staphylococcal enterotoxin (SE) superantigens A, D, and E can respond to SEs in two seemingly opposite ways. In the absence of antigen presenting cells (APC), SEA, D, and E (but not SEB or C1) strongly inhibited in a dose-dependent manner the responsiveness of clone D894/25 to exogenous IL-2. Growth inhibition was due to SE-induced programmed cell death (apoptosis) as shown by propidium iodide staining and the appearance of the characteristic ladder pattern of DNA fragmentation. Apoptotic cell death was accompanied by significant cell lysis after 4 and 8 h as measured in a 51Cr release assay. In contrast (but as expected), a proliferative response of clone D894/25 was triggered by SEA, D, and E in the absence of exogenous IL-2 but presence of HLA class II-positive lymphoblastoid cell line (LCL) as APC. Moreover, the addition of LCL feeder cells partially prevented the suppression of IL-2 responsiveness by SEs. Surprisingly, however, the latter two culture conditions (i.e. presence of LCL feeder cells with or without exogenous IL-2) were associated with similar levels of induced cell death as in the absence of LCL. At the clonal level, these data demonstrate that SE superantigens induce programmed cell death in a fraction (40-50%) of responsive mature T cells, irrespective of the presence or absence of MHC class II-positive APC. We conclude that the proliferative response of clone D894/25 which is triggered by SEs in the presence of APC and absence of IL-2 must originate from the fraction (50-60%) of clone T cells surviving SE-induced cell death.
Previous studies have established that inactivated mycobacteria are potent and selective activators of V gamma 9+/V delta 2+ human gamma/delta T cells. Here we have analysed the proliferative response of human gamma/delta T cells to five serologically distinct groups of streptococci. While heat-inactivated streptococci of all five serogroups tested (A, B, C, D and F) induced a strong proliferative response in peripheral blood mononuclear cells (PBMC), only groups A, B and C elicited a selective activation of V gamma 9+ gamma/delta T cells in 10 (serogroup B) or 11 (serogroups A and C) of 11 tested healthy individuals. In striking contrast, groups D and F streptococci failed to activate gamma/delta T cells in nine of 11 donors and induced only a weak gamma/delta T cell response in two additional individuals. Depletion of V gamma 9+ T cells before culture completely eliminated all gamma/delta T cell responses to streptococci. These data indicate that groups A, B and C (but not D or F) streptococci can be included in the growing list of selective ligands for V gamma 9+/V delta 2+ human gamma/delta T cells.
We have investigated the frequencies of Dermatophagoides pteronyssinus (Dpt)-reactive circulating T cells in individuals allergic to this house dust mite and in asymptomatic controls. In allergic patients the median for frequencies of Dpt-reactive T cells (80/10,000 T cells) was significantly higher (p = 0.0001) than that in the controls (19/10,000 T cells). Frequencies of spontaneously proliferating T cells were also determined. Here too, marked differences (p = 0.0001) were observed between the allergics (median = 32/10,000 T cells) and the non-allergics (median = 7.5/10,000 T cells). The allergen-reactive T cells tended to occur at a higher frequency with increasing serum level of Dpt-specific IgE. Thus the median value for Dpt-reactive T cells in patients showing RAST class greater than or equal to 4 (176.5/10,000 T cells) was higher than that shown by patients with RAST class 3 (62/10,000 T cells). Frequency estimations of allergen-reactive T cells before, during and after the completion of hyposensitization therapy might help to clarify their possible role in the allergic symptoms.
Like the endometrial stroma, the decidua contains lymphoreticular cells, and these are probably involved in immunological interactions between the conceptus and the mother. Lymphoreticular cells in decidual tissue obtained from 12 patients undergoing therapeutic abortion of an intact pregnancy at 6-10 weeks' gestation were investigated in this study. Immunophenotyping with a broad panel of monoclonal antibodies revealed various subpopulations of lymphoreticular cells. Macrophages (Ki-M6+, Ki-M7+, Ki-M8+, KP1+, MAC 387+ and Ki-M1P+) represented the largest fraction of intradecidual lymphoreticular cells. CD3+ and CD8+ lymphocytes were found in moderate numbers and CD4+ cells in small numbers. The majority of the intradecidual lymphoid cells exhibited an unusual phenotype [CD7+, CD2+, CD56+, triple negative (CD3-, CD4-, CD8)]. The distribution of these unusual lymphocytes mirrored that of the so-called endometrial stromal granulocytes. A few of these stromal granulocytes reacted with the macrophage-associated antibody KP1, but not with Ki-M1P, another macrophage marker. This was confirmed by immuno-electron microscopy. The finding that intradecidual CD3+ lymphocytes express neither the alpha/beta nor the gamma/delta heterodimer of the T cell antigen receptor was unexpected. However, these cells did express the alpha/beta heterodimer after in vitro culture with PHA-P and recombinant exogenous interleukin-2. No stimulated T lymphocytes expressing activation antigens could be detected. B lymphocytes, T and B immune accessory cells and CD15+ granulocytes were found only in small numbers or were absent. Amongst cells expressing NK cell markers, CD57+ and CD16+ cells were found in small to moderate numbers, while CD56+ cells were detected in large numbers.(ABSTRACT TRUNCATED AT 250 WORDS)
gamma/delta+ T-cells are a recently identified subpopulation of T-lymphocytes expressing an "alternative" T-cell receptor (TCR) molecule consisting of disulfate-linked or nonlinked gamma and delta chains. Despite a limited number of V gamma and V delta genes in the germ line, there is a large and diverse gamma delta TCR repertoire due to extensive N region variability. Recently developed monoclonal antibodies against V gamma and V delta gene products are useful reagents for the identification and isolation of gamma/delta+ T-cell subpopulations. The physiological significance of gamma/delta+ T-cells is still unknown. However, accumulating evidence indicates that human gamma/delta+ T-cells frequently recognize bacterial ligands as well as certain tumor cells. Interestingly, reactivity towards microbial antigens is usually restricted to a subpopulation of gamma/delta+ T-cells expressing a V gamma 9/V delta 2 TCR. However, different bacteria-reactive V gamma 9+/V delta 2+ gamma/delta+ T-cells display extensive N region variability, suggesting the involvement of a gamma/delta-specific superantigen in these responses. Little is known about the role of gamma/delta+ T-cells under pathological conditions. Rare cases of gamma/delta+ T-cell leukemias and lymphomas have been described. In addition, discrete changes in the distribution of gamma/delta+ T-cell subpopulations have been observed during HIV infection. Current thinking favors the interpretation that gamma/delta+ T-cells play a role in the immune reaction during infection and in the regulation of physiological or pathophysiological autoimmune responses.
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Staphylococcal enterotoxins (SE) are potent T-lymphocyte activators that stimulate T cells by directly cross-linking HLA-DR molecules on antigen-presenting cells with the V beta gene products of the T-cell receptor. The different SE activate all T cells expressing a given V beta, and, therefore, have been termed 'superantigens'. Here we show that SE are potent activators of leukaemic B cells from patients with chronic lymphocytic leukaemia (CLL). Purified B cells from seven of eight CLL patients with high WBC counts (greater than 80,000/microliters) responded to one or several of the tested SE (SEA, SEB, SEC1, SED, SEE) by proliferation ([3H]TdR incorporation) and/or Ig secretion. In several instances, the response of leukaemic B cells to SE was much stronger than was the response to other known B-cell activators including EBV, pokeweed mitogen (PWM), phorbolester (TPA), and Staphylococcus aureus Cowan I (SAC). The activation of leukaemic B cells by SE was strictly dependent on the addition of irradiated T cells isolated from healthy donors. FACS analysis of cultured cells ensured that the proliferating cells were indeed B cells. Taken together, these results demonstrate that SE are strong T-cell-dependent B-cell activators that, in some cases, can stimulate maturation of leukaemic B cells which are refractory to other activation signals.
The clinical success of organ transplantation depends to a large degree on the immunological acceptance of the grafted organ. This paper summarizes from an immunological point of view the recent progress that has been made to improve graft acceptance, and discusses some future aspects in the field. Over the last few years, major emphasis has been put on the development of new immunosuppressive drugs, including FK 506, rapamycin, and Deoxyspergualin. Together with monoclonal antibodies against defined T-cell surface antigens, there are now new and effective means available to prevent or treat rejection episodes. Progress has also been made in the field of HLA typing, where the introduction of molecular biology-based methods significantly increased the accuracy of HLA class II typing. The ultimate goal of transplantation immunology is the induction of (donor-) specific tolerance. While some protocols are effective in inducing peripheral tolerance in experimental animals, these regimens are at present not yet applicable in the clinical situation. To overcome the shortage of donor organs, alternative strategies are currently being considered. Among these, xenotransplantation may eventually prove successful, despite the massive immunological problems such as, e.g., the presence of preformed xenoreactive antibodies.