Kagemasa Kuribayashi

Parietal cell autoantigens involved in neonatal thymectomy-induced murine autoimmune gastritis. Studies using monoclonal autoantibodies

Autoimmune gastritis accompanied by autoantibodies to parietal cells was induced in BALB/c nu/+ mice by neonatal thymectomy 2-4 days after birth. Three monoclonal autoantibodies, designated as 2B6 (IgG1), 2G10 (IgG2b), and 1H9 (IgG1), were obtained from one of these mice. All three reacted specifically with parietal cells, 2G10 recognizing species-specific antigenic determinants and 2B6 and 1H9 recognizing interspecies-specific antigenic determinants. All three recognized antigens on the membrane of intracellular secretory canaliculi and the cytoplasmic tubulovesicular system of parietal cells. At least two different molecular groups were recognized by these antibodies; 2B6 recognizing a 65,000-79,000-mol wt group and 1H9 recognizing a 92,000-120,000-mol wt group. Sera of most mice with autoimmune gastritis reacted with either or both groups. Both groups were consistently coprecipitated by any of the three antibodies when solubilized in NP-40. Sera, from patients with pernicious anemia, containing anti-parietal cell antibodies could also precipitate these two groups of antigens. Competition assay and physicochemical studies showed that the epitopes recognized by the three monoclonal antibodies are different.

Retrovirus-specificity of regulatory T cells is neither present nor required in preventing retrovirus-induced bone marrow immune pathology.

Summary Chronic viral infections of the hematopoietic system are associated with bone marrow dysfunction, to which both virus-mediated and immune-mediated effects may contribute. Using unresolving noncytopathic Friend virus (FV) infection in mice, we showed that unregulated CD4+ T cell response to FV caused IFN-γ-mediated bone marrow pathology and anemia. Importantly, bone marrow pathology was triggered by relative insufficiency in regulatory T (Treg) cells and was prevented by added Treg cells, which suppressed the local IFN-γ production by FV-specific CD4+ T cells. We further showed that the T cell receptor (TCR) repertoire of transgenic Treg cells expressing the β chain of an FV-specific TCR was virtually devoid of FV-specific clones. Moreover, anemia induction by virus-specific CD4+ T cells was efficiently suppressed by virus-nonspecific Treg cells. Thus, sufficient numbers of polyclonal Treg cells may provide substantial protection against bone marrow pathology in chronic viral infections.

Reconstitution of anti-tumor effects of lentinan in nude mice : roles of delayed-type hypersensitivity reaction triggered by CD4-positive T cell clone in the infiltration of effector cells into tumor

Lentinan, an antitumor polysaccharide used clinically in Japan, requires the intact T cell compartment to manifest its antitumor effects. The aim of the current study was to clarify the mechanisms playing crucial roles in the T cell requirement in the expression of antitumor effects of lentinan. Lentinan treatment of BDF1 mice transplanted intradermally with FBL‐3 induced complete tumor regression and a marked increase in survival time. The antitumor action of lentinan was abolished in mice treated simultaneously with antibodies to CD4 and CD8 antigens, whereas antibody to CD4, CD8 or NK1.1 alone was ineffective. The natural killer, cytotoxic T lymphocyte, and helper T cell activities were already augmented in this FBL‐3/BDF1 system and thus further augmentation of these activities by lentinan was not observed. These activities did not correlate with the antitumor activity of lentinan, as was confirmed in lymphocyte subset depletion experiments. On the contrary, the delayed‐type hypersensitivity (DTH) response against tumor‐associated antigens was triggered by lentinan and was abrogated only in mice treated simultaneously with antibodies to CD4 and CD8 antigens. Furthermore, a non‐cytolytic tumor‐associated antigen‐specific CD4+ T cell clone able to induce the DTH response in concert with lentinan reconstituted the antitumor effects in B6 nude mice when administered with lentinan. These results suggest that, in addition to the augmentation of immune effector cell activity against tumors, infiltration of these cells into the tumor burden initiated by the DTH responses at tumor sites may be involved in eradication of tumors by lentinan.

Immunological properties of Fc receptor on lymphocytes: 5. Suppressive regulation of humoral immune response by Fc receptor bearing B lymphocytes

Abstract High anti-DNP PFC responses to DNP-DE or DNP-KLH were obtained by transferring normal or primed FcR − B cell fractions into irradiated syngeneic recipients. On the other hand, the FcR + B cell fraction showed a low precursor activity. Trypsinization of the FcR + B cells, to eliminate remaining antigen-antibody complexes on the surface, failed to augment the response in comparison with that of trypsin-untreated FcR + B cells. Therefore, the weak precursor activity of FcR + B cells seemed to be inherent. No synergistic interaction between the FcR + B and precursor FcR − B cells, to give rise to the maximum PFC response, was observed. On the contrary, the FcR + B cells significantly suppressed the PFC responses of FcR − B cells. This kind of suppression could be mediated by a factor released from the FcR + B cell, but not from the FcR − B or original-unrosetted spleen cell fraction. The factor was not attributable to macrophages, because the FcR + B cells isolated from normal spleen cells, of which macrophages were depleted by Sephadex G-10 columns, could produce the factor with the same activity. Stimulation by specific antigen is not necessary for the induction of the factor(s) as well as of the suppressing FcR + B cells. It seems to be necessary to stimulate FcR by antigen-antibody complexes to produce or release this factor.

Immunological properties of Fc receptor on lymphocytes. 1. Functional differences between Fc receptor-positive and negative lymphocytes in humoral immune responses.

Abstract Using an EA rosetting system, it was observed that Fc receptors (FcR) were present on the surface of T cells as well as B cells, and that functional differences existed between FcR-positive (FcR + ) and FcR-negative (FcR − ) cells in both T and B cells in in vivo humoral immune responses. Approximately 15% of splenic T cells obtained by nylon wool passage are FcR + . The number of surface immunoglobulinbearing cells as detected by immunofluorescent staining accounted for less than 10% of these FcR + cells. FcR + and FcR − T+B-cell populations obtained from spleens contain 60 and 20% of surface immunoglobulin-positive cells, respectively. In the adoptive primary response in which horse RBC and dinitrophenyl-conjugated dextran (DNP-DE) were used as T-dependent and T-independent antigens, respectively, the majority of precursor B cells were FcR − . In the secondary response using hapten-primed B cells and carrier-primed T cells, the majority of memory B cells for a haptenic determinant were also FcR − . Furthermore, the majority of functional cells exerting helper activity in the same hapten-carrier system are FcR − cells, and FcR + T cells collaborate much less effectively with either memory B cells or helper FcR − T cells.

Resistance to Friend Murine Leukemia Virus Infection Conferred by the Fv-4 Gene Is Recessive but Appears Dominant from the Effect of the Immune System

ABSTRACT Fv-4 is a mouse gene that dominantly confers resistance to infection with Friend murine leukemia virus (F-MuLV) (S. Suzuki, Jpn. J. Exp. Med. 45:473–478, 1975). However, the resistance caused byFv-4 is recessive in nude mice, which suggests that immunological effects play important roles in this resistance in vivo (K. Higo, Y. Kubo, Y. Iwatani, T. Ono, M. Maeda, H. Hiai, T. Masuda, K. Kuribayashi, F. Zhang, T. Lamin, A. Adachi, and A. Ishimoto, J. Virol. 71:750–754, 1997). To determine the immunological effect on the resistance in vivo, we infected immunologically immature newborn mice homozygous (Fv-4r/r) and heterozygous (Fv-4 r/−) for Fv-4. Although theFv-4 r/r mice showed complete resistance to F-MuLV whether infected neonatally or as adolescents, theFv-4 r/− mice showed high sensitivity to viral proliferation and disease induction when infected as newborns but complete resistance when infected as adolescent mice. To confirm the immunological effect on the resistance in adolescent mice with theFv-4 r/r and Fv-4 r/−genotypes, we examined the effect of an immunosuppressant drug, FK506, on the resistance. The mice with the Fv-4 r/rgenotype treated with FK506 still showed resistance, but the mice with the Fv-4 r/− genotype became highly sensitive to F-MuLV infection. Flow cytometric analysis to detect theFv-4 gene product showed that the Fv-4 gene product was expressed on the cells from newborn and adolescent mice. The Fv-4 gene product was also detected on the cells from the FK506-treated mice as well as on those from untreated mice. However, a quantitative difference in the gene product between the cells with the Fv-4 r/r andFv-4 r/− genotypes was detected by indirect staining for flow cytometry. These results show that the resistance to F-MuLV infection conferred by the Fv-4 gene is originally recessive, but it looks dominant in adolescent mice mainly because of the effect of the immune system.

Interferon‐γ‐producing Tumor Induces Host Tumor‐specific T Cell Responses

We investigated the mechanism of host immune responses against two interferon‐γ (IFN‐γ) gene‐transduced tumors, plasmacytoma MOPC104E(Muγ) and mammary cancer SC115(Kγ), which originally had weak immunogenicity. Both IFN‐γ‐producing tumor cells had reduced tumorigenicity and were rejected by syngeneic mice. The rejection was completely blocked by in vivo treatment with anti‐CD8 or anti‐IFN‐γ monoclonal antibodies. While anti‐CD4 monoclonal antibody also blocked the rejection of SC115(Kγ), it enhanced the initial tumor growth of MOPC104E(Muγ). Specific protection against subsequent challenge with the respective parental tumor cells was demonstrated in mice which rejected the IFN‐γ‐producing tumor cells. Cultured lymphocytes derived from immunized mouse spleens had cytotoxic T cell activity against parental tumor cells, as well as against cells that produced IFN‐γ, These findings indicate that the antitumor effects are mediated by cytotoxic T cells and, partly, by helper T cells, and that locally secreted IFN‐γ plays an important role in generating these effector cells.

Immunological properties of fc receptor on lymphocytes. 3. Stimulating activities of fc receptor-bearing and -nonbearing cells in mixed-lymphocyte reaction.

Abstract By separating FcR + and FcR − cells from stimulator spleens using an EA rosetting procedure, it was found that EA-rosetting (FcR + ) cells stimulate mixed-lymphocyte culture reaction (MLR) far more effectively than do non-EA-rosetting (FcR − ) cells. The difference in stimulatory activity is observed in MLR of both H-2 and M-locus different combinations and cannot be explained by the proportion of B cells and macrophages contained in each population. The finding that FcR + cells can stimulate allogeneic responding T cells more effectively than FcR − cells suggests a close association of FcR with Ia and Mls antigens on the cell surface.

Characterization of a CD4(L3T4)-positive cytotoxic T cell clone that is restricted by class I major histocompatibility complex antigen on FBL-3 tumor cell.

An L3T4(CD4)+ CTL clone specific for Friend virus-induced tumor FBL-3 was isolated, characterized and compared with a conventional Lyt-2(CD8)+ CTL clone. This clone L3.1 was obtained from the limiting dilution culture of splenic MLTC cells from a CB6F1 mouse whose CD8+ T cells had been suppressed by an in vivo injection of anti-Lyt-2.2 mAb. The phenotype of clone L3.1 was sIg-, Thy-1.2+, L3T4(CD4)+, Lyt-2 (CD8)-, and Ia- as determined by flow-cytometry. Northern blot analysis also confirmed that mRNA for L3T4(CD4), but not for Lyt-2 (CD8) was present in the total RNA of L3.1. The FBL-3-specific killing activity of L3.1 was inhibited by anti-H-2D6 mAb, and the tumor cells did not express class II MHC antigen, indicating that the recognition of tumor antigen by this CD4+ CTL clone was restricted by the class I MHC molecule on the tumor cells. Furthermore, the finding that anti-L3T4(CD4) mAb GK1.5 inhibited the specific and lectin-dependent non-specific cytotoxicity of L3.1 suggested that CD4 molecules on this CTL clone are not ligand (MHC class II)-binding proteins, but are involved in signal transduction.

Immunological properties of fc receptor on lymphocytes. 7. Suppression of mixed lymphocyte reaction by fc receptor- -bearing splenic lymphocytes.

Abstract In this report the possible functions of FcR + and FcR − lymphocytes in the MLR system when purified by an EA rosetting procedure were examined. It is shown that a large proportion of responding cells is confined to FcR − and not to FcR + T cells. When the FcR + T cells are recombined with syngeneic responding FcR − T cells at the start of culture, the response of the latter cells against MMC-treated allogeneic cells is significantly suppressed. This suppression is dose dependent. At the same time, a similar type of suppression was observed when FcR + B cells, which were depleted of macrophages, were used instead of FcR + T cells. Hence, it is conceivable that a common mechanism, such as binding of immune complexes (EA) on FcR of T as well as B cells, might be involved, resulting in the suppression in MLR. Furthermore, the FcR + lymphocytes, containing both T and B cells, depress not only the proliferating response of syngeneic FcR − T cells to allogeneic stimulation, but also the induction of cytotoxic lymphocytes in MLR. These results suggest that FcR-bearing cells after binding with immune complexes have a suppressive influence on cell-mediated immune responses.