PubMed Health⌕ Search

Biomedical subjects

T Tsubata

Publications and source records attributed to T Tsubata.

52 records · Page 3Linked to original sources
PubMed1992

A transgenic model of autoimmune hemolytic anemia.

We made double transgenic mice bearing immunoglobulin heavy and light chain genes encoding an autoantibody against the mouse erythrocyte by the cross of C57BL/6 mice carrying the transgene for each chain of the immunoglobulin. Although no obvious disorders were found in the single-chain transgenic mice, severely anemic symptoms were found in some of the double transgenic mice, in which most B cells express, at least on their surface, the autoantibody reactive to self-antigens on the erythrocyte. Individual double-transgenic mice showed a wide variation of phenotypes between severe anemia and no symptoms. Both deletion and anergy of autoreactive B cells were seen in each individual mouse, but their relative contribution to self-tolerance was variable and not directly related to the severity of anemia or the amount of the autoantibody produced. This transgenic system provides a good autoimmune disease model for exploring its onset mechanism, and means of its treatment and prevention.

Anemia, Hemolytic↗
PubMed1992

Crosslinking of the cell surface immunoglobulin (mu-surrogate light chains complex) on pre-B cells induces activation of V gene rearrangements at the immunoglobulin kappa locus.

We constructed an expression vector encoding a truncated Ig mu chain that lacks both VH and CH1 domains (mu delta m chain) and introduced the mu delta m vector into the Ig negative Abelson pre-B cell line P17-27. The transfectants expressed a large amount of the mu delta m chain on their surface, which was not complexed with the lambda 5 and VpreB surrogate light chain molecules. While P17-27 transfected with a vector for the intact micron chain (P17-27 micron) shows V kappa rearrangements in culture, V kappa rearrangements were not detected in P17-27 mu delta m cells. When the mu delta m chains on the cell surface were crosslinked by anti-mu antibodies, V kappa gene rearrangements were induced in P17-27 mu delta m. These results strongly suggest that crosslinking of the micron-lambda 5-VpreB complex on the pre-B cell surface generates a signal that activates V kappa gene rearrangement, and that the lambda 5 and VpreB molecules are necessary for the spontaneous crosslinking of surface Ig on pre-B cells.

B-Lymphocytes↗
PubMed1991

Cell surface expression of the short immunoglobulin mu chain (D mu protein) in murine pre-B cells is differently regulated from that of the intact mu chain.

Pre-B cells carrying DJH rearrangements in an appropriate reading frame produce a short mu protein consisting of a DJH and the C mu & sequence (D mu protein). We analyzed a D mu-producing Abelson-murine leukemia virus-transformed murine pre-B line, 300-19, and demonstrated that D mu proteins are expressed on the cell surface in association with surrogate L chain proteins (lambda 5 and VpreB). However, when we introduced an expression vector coding for the D mu protein into the null pre-B line P17-27, which produce lambda 5 and VpreB but no Ig molecules, most of the cells did not express D mu proteins on the cell surface although D mu proteins were produced intracellularly. On the other hand, P17-27 brings intact mu chains on the cell surface, when a vector coding for the intact mu chain is introduced. Thus, cell surface expression of the D mu protein has different requirements from that of the intact mu chain. A possible role of the VH protein encoded by germ-line VH transcripts is discussed.

Animals↗
PubMed1991

Molecular and cellular aspects of early B-cell development.

The molecular mechanisms underlying pre-B cell growth and immunoglobulin gene rearrangement have been actively investigated. Some growth factors for B-cell precursors as well as some genes activating V(D)J recombination have been identified. Furthermore, the molecular structure and signaling capacity of the surface immunoglobulin of pre-B cells have been characterized.

Animals↗
PubMed1991

Characterization of bullous pemphigoid antibodies by use of recombinant bullous pemphigoid antigen proteins.

The seroreactivity of patients with bullous pemphigoid (BP) to recombinant proteins representing sequences in the carboxyl domain of the murine 230-kD BP antigen (BPA) was determined. Sera from 133 patients with BP, 20 patients with pemphigus, and 21 normal subjects were examined by Western blotting by using two recombinant proteins: RP120 (MW = 120 kD), representing the C-terminal half of the 230-kD BPA, and RP60 (MW = 60 kD), representing the C-terminal quarter. These RP120 and RP60 were recognized by 84% and 61%, respectively, of the BP sera that reacted with the 230-kD BPA in epidermal extract, and not by any of pemphigus or normal sera. Furthermore, these RP120 and RP60 were not recognized by any BP sera that reacted only with the 170-kD BPA, which is known to be another major BPA. These findings indicate that one or more of the major antigenic regions localizes in the carboxyl-half domain of the 230-kD BPA, and also suggest that the 230-kD BPA may be distinct from the 170-kD BPA.

Animals↗
PubMed1991

Qualitative difference of anti-DNA antibody-producing cell precursors in the pre-immune B cell repertoire between normal and lupus-prone mice.

The precursor frequency for anti-DNA antibody-producing cells in the pre-immune B cell repertoire was investigated in young female BALB/c and NZW mice, and in young and aged female NZB x NZWF1 (B/WF1) mice. Spleen cells from these mice were diluted serially and stimulated polyclonally in vitro with lipopolysaccharide (LPS) and IL-4 to induce both IgM and IgG1 production. The results demonstrated that there existed virtually no difference in precursor frequency for IgM anti-DNA antibody-producing cells between normal and lupus mice, confirming previous observations made by other investigators. In contrast, the number of precursors for IgG1 anti-DNA antibody-producing cells was much higher in young and old B/WF1 mice than in normal mice. These results suggest that the high frequency of precursors for IgG1 anti-DNA antibody-producing cells in the pre-immune B cell repertoire of B/WF1 mice is a crucial factor for the pathogenesis of systemic lupus erythematosus.

Animals↗
PubMed1990

The products of pre-B cell-specific genes (lambda 5 and VpreB) and the immunoglobulin mu chain form a complex that is transported onto the cell surface.

We constructed expression vectors coding for the two pre-B-specific genes, VpreB and lambda 5, and transfected them together with a mu vector (mu tm) into Ig- myeloma cells. In a transfectant expressing all three introduced genes, the mu tm chain is transported on the cell surface. A biochemical analysis demonstrated that, in these cells, the mu tm chain is associated noncovalently with an 18-kD protein and covalently with a 22-kD protein, which are most likely the products of VpreB and lambda 5, respectively. Our results, thus, strongly suggest that the products of lambda 5 and VpreB bind to mu chains and have the same capacity as conventional Ig L chains to allow surface expression of mu chains.

Animals↗
PubMed1990

Molecular components of the B-cell antigen receptor complex of the IgM class.

The antigen receptors on mature B lymphocytes are membrane-bound immunoglobulins of the IgM and IgD classes whose cross-linking by polyvalent antigens results in B-cell proliferation and differentiation. How these membrane-bound immunoglobulin chains, which lack a cytoplasmic tail, generate a cell activation signal is not at present known. We now show that the IgM molecule is non-covalently associated in the membrane of B cells with two proteins of relative molecular mass 34,000 (Mr 34 K; IgM-alpha) and 39 K (Ig-beta) which form a disulphide-linked heterodimer. Surface expression of IgM seems to require the formation of an appropriate complex between IgM and the heterodimer. A transfection experiment indicates that IgM-alpha is the product of mb-1, a B-cell specific gene encoding a transmembrane protein with sequence homology to proteins of the T-cell antigen receptor-CD3 complex.

Animals↗
PubMed1989

Limited proteolysis of bovine myelin basic protein by calcium-dependent proteinase from bovine spinal cord.

In order to shed some light on the role of proteinases in the process of demyelination in the central nervous system, a calcium-dependent proteinase was purified to homogeneity from bovine spinal cord, and its action on myelin basic protein purified from the same tissue was investigated. Among the three major myelin basic protein fractions, Fraction I was resistant to the action of the enzyme whereas Fractions II and III were degraded in the same manner, giving two major bands on SDS-polyacrylamide gel electrophoresis. The hydrolysis products were fractionated by high-performance liquid chromatography and characterized. The results showed that the myelin basic protein (Fractions II and III) was rather selectively cleaved at the Val93-Thr94 bond and the Arg96-Thr97 bond in mutually exclusive ways, with minor cleavages at the Ala16-Ser17 and Gly68-Ser69 bonds, suggesting the implication in vivo of calcium-dependent proteinase in the limited proteolysis of myelin basic protein.

Animals↗
PubMed1989

Possible different mechanisms of B cell activation in systemic lupus erythematosus and rheumatoid arthritis: opposite expression of low-affinity receptors for IgE (CD23) on their peripheral B cells.

To clarify the differential state of B cell activation in patients with systemic lupus erythematosus (SLE) and rheumatoid arthritis (RA), we investigated the expression of low-affinity receptor for IgE (Fc epsilon RII; CD23) on their peripheral B cells by a cytofluorometry using H107 (CD23) and Leu-16 (CD20) monoclonal antibodies. The percentage of CD23-negative B cells in total lymphocytes was significantly greater in both groups of patients than in normal subjects, suggesting the hyperactivity of late-phase B cells in both diseases. However, the increase of CD23-negative B cells in RA was brought about by the increased number of total B cells, although that in SLE was mainly based on the relative decrease of CD23-positive B cells. The number of IgD-positive B cells was decreased, and the number of colony-forming B cells was markedly increased in SLE patients. These observations indicate that a B cell abnormality is mainly qualitative in SLE but quantitative in RA.

Adult↗
PubMed1988

Differentiation of an interleukin 3-dependent precursor B-cell clone into immunoglobulin-producing cells in vitro.

Precursors to B-cell lines with immunoglobulin genes in the germ-line context have been shown to be capable of generating mature B cells in vivo. We report here that an interleukin 3-dependent precursor B-cell line, LyD9, differentiated in vitro into mature B cells, producing IgM and IgG by coculture with bone marrow accessory (or stroma) cells or with dendritic cells and T cells. Up to 50% of IgM-positive cells, but no Thy-1-positive cells, appeared after the 7- to 10-day coculture. Induced LyD9 cells underwent heterogenous immunoglobulin gene rearrangement and synthesized mRNAs encoding immunoglobulin mu, gamma, and kappa chains. However, these cells did not show any rearrangement of genes encoding the alpha and beta chains of the T-cell receptor. The induction of differentiation by coculture with bone marrow stroma cells was blocked by anti-lymphocyte function-associated antigen 1 or anti-interleukin 4 antibody. These systems are useful for molecular biological studies on regulation of differentiation of bone marrow-derived cells into the B-cell lineage.

Animals↗
PubMed1988

B cell repertoire for anti-DNA antibody in normal and lupus mice: differential expression of precursor cells for high and low affinity anti-DNA antibodies.

The precursor frequency for anti-DNA antibody producing cells and the affinity of antibodies secreted by these cells in both immature prereceptor B cell populations and mature B cell populations were compared between 8-week-old C57BL/6 female mice and 9-month-old B/WF1 female mice by producing a large collection of IgM secreting hybridomas from LPS-stimulated B cells. The data indicate that precursor cells for high affinity anti-DNA antibody are eliminated as they mature in C57BL/6 mice, while a sizable number of such clones are present in mature splenic B cells of aged B/WF1 mice. These results suggest that the emergence of precursors for high affinity anti-DNA producing cells in mature B cell population is an important factor in the pathogenesis of SLE.

Animals↗
PubMed1986

Limiting dilution analysis of the stem cells for T cell lineage.

Stem cell activities of bone marrow, spleen, thymus, and fetal liver cells for T cell lineage were studied comparatively by transferring the cells from these organs through i.v. or intrathymus (i.t.) route into right leg- and tail-shielded (L-T-shielded) and 900 R-irradiated recipient mice, which were able to survive without supplying hemopoietic stem cells. Cells from B10.Thy-1.1 (H-2b, Thy-1.1) mice were serially diluted and were transferred into L-T-shielded and irradiated C57BL/6 (H-2b, Thy-1.2) mice, and 21 days later the thymus cells of recipient mice were assayed for Thy-1.1+ cells by flow cytofluorometry. The percentage of recipient mice possessing donor-type T cells was plotted against the number of cells transferred, and the stem cell activity in each cell source was expressed as the 50% positive value, the number of donor cells required for generating donor-type T cells in the thymuses of 50% of recipient mice. In i.v. transfer experiments, the activity of bone marrow cells was similar to that of fetal liver cells, and about 100 times and nearly 1000 times higher than those of spleen cells and thymus cells, respectively. In i.t. transfer experiments, the number of cells required for generating donor-type T cells was much lower than that in i.v. transfer experiments, although the ratio in 50% positive values between i.v. and i.t. transfers differed among cell sources. In i.t. transfers, the 50% positive value of bone marrow cells was five times, 400 times, and 500 times higher than that of fetal liver cells, spleen cells, and thymus cells, respectively. Our previous finding that stem cells are enriched in the spleens of mice which were whole body-irradiated and marrow-reconstituted 7 days earlier was confirmed also by the present limiting dilution assay carried out in i.v. as well as i.t. transfers.

Animals↗
PubMed1985

Tac antigen-positive T cells activated in autologous mixed lymphocyte reaction regulate the generation of killer T cells against hapten-modified autologous cells.

T cells that proliferate in the autologous mixed lymphocyte reaction (auto-MLR) have been shown to acquire some suppressor or regulatory activities. In the present study, we examined the suppressive effects of T cells activated in the auto-MLR on the induction of hapten-specific cytotoxic T cells. NRFT (depletion of ARFT from UT) were used as the responder cells of TNP-MLR. After primary and secondary TNP-MLR, the cells were harvested and tested for their cytotoxic activities against TNP-modified autologous cells by 51Cr-release assay. When UT cells cultured for 1 wk in auto-MLR were added to primary TNP-MLR at the beginning of culture, the cytotoxic activity tested at the end of the culture was suppressed from 15.6% +/- 2.7 to 5.8% +/- 1.1 (percent cytotoxicity, mean +/- SE). However, these auto-MLR-activated UT cells had little suppressive activity against cytotoxic T cells when they were added to the final assay of TNP-CTR. Suppressive activities of these cells on the generation of cytotoxic T cells during secondary TNP-MLR were also tested. The addition of auto-MLR-activated UT cells to the secondary TNP-MLR at the beginning of the culture reduced the cytotoxic activities of NRFT from 23.8% +/- 2.3 to 9.7% +/- 1.7 after secondary TNP-MLR. Allo-activated T cells, PHA blasts, and fresh autologous T cells were used as the controls, but none of the cells had suppressive effects on the generation of CTL. Characteristics of these suppressor cells were examined. Auto-MLR-activated cells from ARFT fractions exhibit very powerful suppressor activity. Treatment of the auto-MLR-activated T cells with mitomycin C eliminated their suppressive effects on the generation of CTL; 21.2% +/- 6.3 of UT cells became anti-Tac positive after 1 wk of auto-MLR. Treatment of auto-MLR-activated UT cells with anti-Tac antibody plus complement eliminated their suppressive activities on the induction of CTL. Thus, T cells stimulated in auto-MLR were shown to have suppressive effects on the induction of cytotoxic T cells against TNP-modified autologous cells. These cells were mitomycin C sensitive. Because anti-Tac antibody is reactive to activated T cells, activation of T cells during auto-MLR was thought to be necessary for the acquisition of the suppressive activity.

Antibodies, Monoclonal↗