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In order to elucidate the role of HLA class II molecules in generation of self-nonself discrimination of human T cells, we have analyzed T cell functions in an HLA class II-negative severe combined immunodeficiency patient. Patient PBL expressed no HLA-DR, -DQ, and -DP antigens as judged by immunofluorescence using mAb, and failed to elicit MLR responses from unrelated controls. Patient PBL contained mature T cells (CD3+ TCR alpha beta+) of the CD4 and CD8 subset, showing an apparently normal TCR diversity, as judged by use of anti-V beta 5, -V beta 6, -V beta 8, -V beta 12, and -V alpha 2 mAb. Patient PBL proliferated in response to anti-TCR/CD3 mAb and PHA, but not against recall Ag, despite immunization, and mounted proliferative, but not cytotoxic, responses against allogeneic cells. To find out whether the MLR responses were a consequence of self-nonself discrimination, the patient HLA-DR and -DQ genotype was determined using sequence specific oligonucleotide probes, revealing DRB1*0401 DQB1*0301 alleles, and MLR were set up against a panel of HLA-DR4 DQw3 stimulators matched or mismatched for DRB1*0401 DQB1*0301. Results showed no MLR against DRB1*0401 DQB1*0301 stimulators, but significant responses against stimulators expressing DRB1*0408 and/or DQB1*0302 alleles. Moreover, the DRB1*0401 DQB1*0301 APC reconstituted proliferation of patient PBL against PPD; this response was completely blocked by an anti-IL-2R (p55) mAb and partially also by anti-HLA-DR and -DQ mAb, indicating recognition of these molecules as restriction element presenting Ag--i.e., as self--by patient T cells. In conclusion, the novel demonstration of self-nonself discrimination by T cells from an HLA class II-negative SCID patient suggests that it may not be absolutely dependent on regular HLA class II expression within the differentiation environment in humans.

Background: We report on 9 infants with severe combined immunodeficiency (SCID), who additionally showed signs of Omenn syndrome with an exfoliative dermatopathy, alopecia, enlarged lymph nodes, a hepatomegalia and a striking blood eosinophilia. The immunological evaluation revealed the characteristic abnormalities of SCID with cellular and humoral immunodeficiency. All patients however had the unusual finding of mature T cells in the peripheral blood. By HLA typing these cells were noted to be of maternal origin in 5 patients. In the other 4 patients the T cells were of host origin. We asked for additional differences between both patient groups., Method: Both patient groups were analyzed and compared with regard to case histories, clinical, laboratory and histopathological parameters., Results: No clinical or laboratory differences could be detected. The histomorphologic analysis of patients with or without maternal T cells was identical. The skin biopsies showed dense cell infiltrations of lymphocytes, histiocytes and eosinophils, in the enlarged lymph nodes the latter two cell types predominated. Therefore the only difference between the 2 patient groups was the presence or absence of maternal T cells., Conclusion: Since the Omenn syndrome is found in association with maternal as well as patient derived T cells, we postulate that the peculiar symptoms of this syndrome are the result of a T cell induced inflammatory reaction, similar but not identical to a graft versus host reaction, occurring on the basis of an inborn SCID.

We have analyzed activation of resting human T cells by anti-T cell receptor (TCR) monoclonal antibody (mAb) BMA031, a murine mAb of the G2b isotype. Human peripheral blood lymphocytes (PBL) respond to anti-TCR mAb by short-term proliferation in vitro and by acquisition of responsiveness to interleukin 2 (rIL-2) in the absence of detectable IL-2 production. Cell depletion and limiting dilution experiments indicate that anti-TCR mAb +/- rIL-2 stimulation covers a substantial portion of human T cells, including CD4+ and CD8+ cells. Enhancement by rIL-2 of anti-TCR mAb-induced proliferation is blocked by anti-IL-2 receptor (IL-2R, p55) mAb, while anti-TCR mAb-induced proliferation is not. In contrast, anti-TCR mAb-induced proliferation is blocked by anti-lymphocyte function antigen 1 (LFA-1, CD11a) mAb and is not demonstrable in PBL from two patients with severe congenital LFA-1 deficiency, not even in the presence of irradiated LFA-1+ PBL. We conclude that stimulation of resting human T cells by anti-TCR mAb BMA031 enables dissociation of distinct steps in T cell activation that specifically require participation of IL-2R (p55) and LFA-1 cell surface molecules in a mutually exclusive way.

We have analyzed allorecognition, HLA restriction and T cell receptor (TcR) diversity in an HLA-heterozygous (HLA-DRw6,7) severe combined immunodeficiency (SCID) patient whose T cell system had been repopulated by HLA-homozygous (HLA-DRw6) paternal T cells following T cell-depleted bone marrow transplantation (BMT). Donor origin of T cells and host origin of antigen-presenting cells (APC) in peripheral blood and BM is shown by HLA typing of separated cell populations and two-color immunofluorescence using an anti-HLA monoclonal antibody (mAb). Peripheral blood lymphocytes (PBL) from the chimeric patient proliferate normally against PHA, anti-TcR/CD3 mAb, pooled allogeneic PBL, and also against the recall antigen (Ag) tetanus toxoid and purified protein derivative of tuberculin (PPD) following immunization, suggesting recognition by donor (DRw6) T cells of Ag presented by host (DRw6,7) APC. PPD-specific cytotoxic T lymphocytes generated in vitro from patient PBL post-BMT display specific cytotoxicity against targets expressing DRw6 and DR7, but not against DR-mismatched targets, suggesting that HLA restriction of Ag recognition may occur through determinants expressed by the host and not by the donor. Donor T cells differentiated in the HLA-semiallogeneic host show specific proliferative and cytotoxic responses against HLA-mismatched stimulators, but not against stimulators taken from the host, expressing the host-specific HLA-haplotype, or expressing the host-specific HLA-DR7 antigens. Compared to T cells directly taken from the donor, differentiation of donor T cells in the host is associated with a significant decrease of T cells expressing TcR V beta 5 and V alpha 2 determinants, while no differences in the abundance of of TcR V beta 6, V beta 8 and V beta 12 subsets were noticed. We conclude that allorecognition, major histocompatibility complex (MHC) restriction and TcR diversity generation of human T cells can be modulated through differentiation in an MHC-different environment, as had been previously shown to be the case in murine model systems.

We report the outcome of a non-T-cell-depleted bone marrow transplant from an HLA partially incompatible, MLR-positive, parental donor in a patient with an unusual form of immunodeficiency characterized by a lack of CD8 T cells and a failure of the CD4 cells to display functional activity in vitro. Without conditioning, and following a mild and transient GVHD, donor T cells persist in trace amounts in the host, where they coexist with the nonfunctional host T cells and cooperate with host APC in antigen recognition, thereby leading to a reconstitution of T cell functions in vitro and in vivo and development of a stable, so far unprecedented, human T-T split chimera across MHC barriers.

Circulating maternal T lymphocytes were noted in the peripheral blood of six patients with severe combined immunodeficiency. Phenotypical analyses revealed the presence of both CD4 and CD8 subsets in some but not all cases. The maternal T cells could be stimulated by anti-TCR/CD3 mAb +/- rIL-2, but were virtually silent in the MLR and against the recall Ag purified protein derivative of tuberculin and tetanus toxoid, even in immunized patients engrafted with T cells from a responding mother. Using a panel of mAb against TCR V region gene encoded epitopes including V beta 5, V beta 6, V beta 8, V beta 12, and V alpha 2, we show that maternal T cells displayed a profoundly reduced TCR diversity, characterized by a lack of one or even several TCR V subsets in all six cases and a dramatic (5- to 25-fold) expansion of other TCR V subsets in three cases. In one patient analyzed, limited TCR diversity was also seen in T cells cultured from bone marrow and skin; restimulation experiments of these cells against cells expressing host MHC Ag were unsuccessful, as were attempts to exclusively allocate anti-host proliferative responses of maternal control T cells to the TCR V subsets that had undergone expansion in vivo. We conclude that a severely reduced TCR diversity is a common feature of maternal T cells engrafted in severe combined immunodeficiency patients. These novel findings provide a structural basis to understand the failure of these cells to protect the host from infections and may also help to understand their relative inefficiency to induce lethal, multi-organ, graft vs host disease. Moreover, as an experiment of nature, the reported phenomenon clearly illustrates the functional consequences in vivo of an insufficient TCR diversity.

We have analyzed the immunocompetence of blood donor-derived, allogeneic T cells that had engrafted in a patient with severe combined immunodeficiency following transfusion with unirradiated blood. The blood donor T cells multiply and persist in the patients' bone marrow and peripheral blood over several months without leading to a graft-versus-host disease. The allogeneic T cells can be expanded in vitro and restimulated by cells expressing host HLA-DR antigens. The presence of blood donor-derived T cells prevented engraftment of a bone marrow graft from an HLA-identical sibling; in contrast, engraftment and immunological reconstitution by bone marrow donor-derived cells was seen following elimination of the allogeneic T cells in vivo. We conclude that the allogeneic T cells sensitized against host histocompatibility antigens were responsible for bone marrow graft failure, even though this sensitization did not lead to a graft-versus-host disease.

We have studied a recently described method to purge human marrow of T cells using magnetic beads conjugated to various anti-T cell monoclonal antibodies. Using anti-CD2 and anti-T cell receptor/CD3 monoclonal antibodies, we show that up to 2.76 log T cell purge can be achieved while recovery of hemopoietic activity is 82%. As a control, our clinically established T cell depletion protocol, namely soybean agglutination combined with E-rosetting, provided 3.39 log T cell purge and 59% recovery of hemopoietic activity. The impact of these results on T cell depletion strategies in HLA-nonidentical bone marrow transplantation is discussed.

Peripheral blood (PBL) and synovial fluid lymphocytes (SFL) from 18 patients with definite rheumatoid arthritis (RA) and from one patient with Reiter's disease (RD) were examined for their capacity to absorb quantitatively interleukin-2 (IL-2) from a standardized, lectin-free IL-2 source. For comparison normal ConA blasts and PBL from various inflammatory and noninflammatory diseases as well as from healthy control persons were studied. IL-2 activity was quantitated by measuring 3H-thymidine-2-deoxyriboside uptake in the IL-2 dependent murine T cell line CTL6. ConA blasts exhibited a high IL-2 absorption capacity and served as a positive control for calibrating the absorption assay. In a population of normal PBL at least 5-10% of ConA blasts were required to detect IL-2 absorption. Significant absorption was assumed if more than 50% of IL-2 activity was removed from 200 microliters of a lectin-free IL-2 standard following incubation with 5 X 10(6) lymphoid cells for 2 h at 4 degrees C; this criterion was fulfilled with 8 out of 20 SFL and 4 out of 14 PBL preparations from RA patients. As a rule SFL absorbed more IL-2 than PBL. Control PBL did not absorb significant quantities of IL-2. PBL from the RD patient apparently produced an IL-2 inhibitor during incubation with the IL-2 standard. IL-2 absorption by ConA blasts and SFL was fully inhibited by preincubation of the absorbing cells with monoclonal anti-TAC antibody, a reagent known to react with the human IL-2 receptor. The results are discussed in view of current concepts of antigen/mitogen induced T cell activation.

Acute rejection of allogeneic and semiallogeneic marrow grafts has long been considered to be a function of the natural immune system because it shares many features with NK activity in mice. With the use of a recently developed in vivo adoptive transfer assay in which spleen cells are transferred from mice able to reject a particular marrow graft into mice that fail to do so, we show that the cells responsible for induction of marrow graft rejection indeed display the phenotype of NK cells: they lack the T cell Ag CD4 and CD8 but express the NK Ag NK1 and ASGM1. The rejection induced by adoptively transferred cells is exquisitely specific--a feature that points to a specific recognition process by the transferred cells. To elucidate what the recognition structure on these cells may be we found that they express CD3 and most likely the beta-chain of the TCR. Highly purified responder cells with the NK1+, CD3+, CD4-, CD8- phenotype, when transferred into nonresponder recipients, cause specific marrow graft rejection. We conclude that the acute rejection of bone marrow grafts is caused by a cell that expresses NK phenotype but is of T cell lineage. This may suggest the specificity of acute marrow graft rejection is caused by a specific recognition process that involves TCR.