Differentiation and Maturation of Functional T Lymphocyte Subsets in the Thymus I. Effect of Irradiation of the Thymus

この論文は国立情報学研究所の学術雑誌公開支援事業により電子化されました。Effect of the irradiation of the thymus on the development of T cells was investigated. C57BL/6 (H-2^b, Thy-1.2) mice were irradiated without shielding or with shielding over the thymus or over the right leg and tail, and they were transferred with bone marrow cells from B10. Thy-1.1 (H-2^b, Thy-1.1) mice. After the reconstitution, the proportion of donor-type Thy-1.1 bearing cells were analysed by flow cytofluorometry. In the whole-body irradiated mice thymus cells were nearly completely replaced by donor derived cells within 21 days. In thymus-shielded mice and right-leg-and-tail-shielded mice, about half of the thymus cells were donor-type cells at 28 days after the reconstitution. The development of donor-derived cytotoxic T cell precursors (CTLp) and helper T cells (Th) in the thymus of these recipient mice was investigated. In order to increase the sensitivity of detection, concanavalin A-mediated polyclonal assay system was used for both CTLp and Th, and the PNA^- cell fraction of thymocytes was used as a source of functional cells. In whole-body irradiated mice and in right-leg-and-tail-shielded and irradiated mice, CTLp developed before Th did, whereas the reverse was true in thymus-shielded and irradiated mice. These results strongly suggested that the capacity of the thymus to allow the proliferation of donor type Thy-1 bearing cells and the development of CTLp in the thymus was independent of whether or not this organ was irradiated. On the other hand, the development of functional Th was obviously retarded by the irradiation on the thymus, suggesting that the radiosensitive thymic element plays a critical role for the generation of Th

Differentiation and maturation of functional T lymphocyte subsets in the thymus. II. Generation of T cell specificities and functions from a single stem cell.

この論文は国立情報学研究所の学術雑誌公開支援事業により電子化されました。Functional differentiation of T cells in the thymus from a single stem cell was investigated by transferring bone marrow cells dirrectly into the thymus (i.t.) of irradiated and reconstituted mice, and assaying the immunologic function of donor drived T cells. To assure the clonality of donor derived T cells, bone marrow cells from two mutually indentifiable strains were 1 : 1 mixed and the limited number of the mixture was transferred i.t. into the recipient mice. As the donor of bone marrow cells, C57BL/6 (H-2^b, Thy-1.2, Lyt-2.2) and B6. Lyt-2.1 (H-2^b, Thy-1.2, Lyt-2.1) mice were used, and B10. Thy-1.1 (H-2^b, Thy-1.1, Lyt-2.2) mice were used as the recipients. Recipient mice were either thymus-shielded and irradiated or whole-body irradiated and reconstituted with 10^7 syngeneic bone marrow cells. The recipient's thymus which contained Thy-1.2^+ cells on day 28 were regarded as positive for donor derived cells, and among these positive thymuses those which contained either Lyt-2.1^+ cells or Lyt-2.2^+ cells were regarded as seeded by a single stem cell (single positive thymus cells). Such single positive thymus cells were treated with anti-Thy-1.1 plus complement to remove recipient-type T cells and the recovered cells were assayed for proliferative response and cytotoxic response in mixed lymphocyte culture against allogeneic lymphocytes. Cells were also assayed for concanavalin A-mediated polyclonal helper T cell (Th) and cytotoxic T cell (CTL) functions. Results indicated that proliferative response to alloantigens was observed only in thymus-shielded recipients, whereas CTL to alloantigens was generated in both thymus-shielded recipients and whole-body irradiated recipients. No general rule of preference for any allogeneic cells was found. Both Th and CTL activities, detectable by polyclonal assays, were generated from a single stem cell in thymus-shielded recipients. However, only CTL but not Th was generated in whole-body irradiated recipients. Moreover, a clone of T cells which responded with CTL to a given allogeneic cells did not necessarily respond with a marked proliferation to the same stimulator cells

T Cell Regulation of Pokeweed Mitogen-induced Polyclonal Immunoglobulin Production in Mice : Role of Virus-replicating T Cells in Suppression

この論文は国立情報学研究所の学術雑誌公開支援事業により電子化されました。Role of virus-replicating T cells in the regulation of pokeweed mitogen (PWM)-induced polyclonal immunoglobulin (Ig) production was studied in mice. A large number of cells which can replicate vesicular stomatitis virus (VSV) were generated in PWM-stimulated cultures of spleen cells. The optimal dose of PWM for the generation of VSV-replicating cells was within the range 25 to 50μg/ml, which were also shown to be optimal for the induction of suppressor T cells. In spleen cell cultures, helper activity was attributed not only to Lyt-1^+2^- but also to Lyt-1^+2^+ T cells and both the precursor and effector suppressor T cells were found to be Lyt-1^-2^+. The VSV-replicating cells in this system were shown to be Thy-1^+ and Lyt-1^+2^-. These cells, however, were unlikely to be helper T cells, but may be a subset of T cells involved in suppression. Thus, inoculation of VSV into the PWM-stimulated cultures of spleen cells resulted in the marked augmentation of Ig production, and the suppressive activity of T cells which had been precultured with a high dose of PWM was abolished by the infection with this virus while the helper activity remained unaffected. Further, it was shown that although the Lyt-1^-2^+ T cell subset could be directly stimulated by PWM to generate suppressor cells, the presence of Lyt-1^+2^- cells in the culture markedly enhanced the generation of Lyt-1^-2^+ suppressor cells

Lymphocyte-stromal cell interaction induces IL-7 expression by interferon regulatory factors.

The interaction between lymphocytes and stromal cells plays important roles in coordinated development of early lymphocytes. IL-7 is an essential cytokine for early lymphocyte development produced by stromal cells in the thymus and bone marrow. Although IL-7 is induced by interaction of early lymphocytes and stromal cells, its molecular basis is still unknown. To address this question, we employed co-culture system with an IL-7-dependent pre-B cell line, DW34, and a thymic stromal cell line, TSt-4. Co-culture with DW34 cells enhanced the levels of IL-7 transcripts in TSt-4 cells. Interestingly, the co-culture also induced transcripts of IFN-α and IFN-β but not of IFN-γ. In addition, exogenous IFN-β stimulation increased the levels of IL-7 transcripts in TSt-4 cells. Next, to elucidate the molecular mechanism of IL-7 induction, we analyzed the IL-7 promoter activity by reporter assay. The IL-7 promoter showed specific transcriptional activity in TSt-4 cells. An interferon-stimulated response element (ISRE) in the IL-7 promoter was essential for the induction of IL-7 transcription by both co-culture and IFN-β stimulation. Finally, overexpression of wild-type and dominant-negative forms of interferon regulatory factors (IRFs) activated and repressed, respectively, the IL-7 promoter in TSt-4 cells. Collectively, these results suggested that IRFs activated by lymphocyte adhesion induce IL-7 transcription through ISRE in stromal cells and that type I IFNs may be involved in the activation of IRFs. Thus, this study implied a physiological function of the IFN/IRF signal during lymphocyte development

Selective increase of autoimmune epitope expression on aged erythrocytes in mice: implications in anti-erythrocyte autoimmune responses

We investigated the impact of changes occurring during red blood cell (RBC) ageing on the RBC-binding activity of pathogenic anti-erythrocyte monoclonal antibodies derived from autoimmune-prone New Zealand black (NZB) mice. As assessed by flow cytometric analysis on in vivo biotinylated RBCs, all five NZB-derived anti-RBC mAb exhibited more efficient binding to aged RBCs than to young RBCs, and resulted in a selective elimination of more aged RBCs from the circulating blood. In addition, treatment of RBCs with proteases markedly enhanced the binding of all five anti-RBC mAb, raising the possibility that increased exposure of autoimmune epitopes on aged RBCs may be in part, a result of contacts with proteolytic enzymes during the lifetime of circulating RBCs. In marked contrast, the binding activity of mAb raised in non-autoimmune animals against antigens expressed on RBCs, such as CD44, CD47, CD147 and TER-119, was either decreased or unchanged with RBC ageing, and these epitopes, except for that recognized by anti-CD47 mAb, were highly sensitive to mild treatment with proteases. Our data unravel the unique molecular feature of RBC epitopes involved in autoimmune haemolytic anaemia, suggesting that membrane alterations in aged RBCs might play a significant role in the development of the autoantibody response to RBCs