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Biomedical subjects

T Hamaoka

Publications and source records attributed to T Hamaoka.

At least 19 recordsLinked to original sources
PubMed1995

Modulation of TCR-mediated signaling pathway by thymic shared antigen-1 (TSA-1)/stem cell antigen-2 (Sca-2).

Thymic shared antigen-1 (TSA-1) is a glycosyl-phosphatidylinositol (GPI)-anchored differentiation Ag expressed on murine lymphocytes, and is identical to stem cell Ag-2 (Sca-2). Using newly established mAb against TSA-1/Sca-2, we have previously shown that surface TSA-1 expression is induced upon activation in T cells, and that anti-TSA-1 inhibits IL-2 production induced by anti-CD3 stimulation in T cell hybridomas. In the present study, we have analyzed the functional role of TSA-1 during T cell activation using normal T cells, T cell hybridomas, and transfected Jurkat cell lines that expressed either GPI-anchored or transmembrane form of TSA-1. Anti-TSA-1 inhibited IL-2 production from normal T cells stimulated with soluble anti-CD3 plus accessory cells. Anti-TSA-1 exhibited the inhibitory effect on T cells, but not on accessory cells, because anti-TSA-1 inhibited IL-2 production in Jurkat cells transfected with TSA-1 cDNA, but not in control transfectant. A transmembrane form of TSA-1 was expressed in Jurkat cells by fusing the extracellular portion of TSA-1 to the transmembrane and cytoplasmic regions of the class 1 Db. The analysis using this transfectant revealed that anti-TSA-1-mediated inhibition of IL-2 production did not require the GPI anchor of TSA-1. Finally, in addition to the inhibition of IL-2 production, tyrosine phosphorylation of CD3 zeta-chains observed following TCR stimulation, one of the important early activation events, was markedly reduced by anti-TSA-1. These results imply that TSA-1/Sca-2 plays an important regulatory role in the TCR signaling pathway of activated T cells in addition to its role in T cell differentiation.

Animals↗
PubMed1995

Administration of recombinant interleukin 12 prevents outgrowth of tumor cells metastasizing spontaneously to lung and lymph nodes.

The present study investigates the ability of recombinant interleukin 12 (rIL-12) to modulate the growth of a primary tumor as well as the outgrowth of metastatic tumor cells in an ovarian carcinoma (OV-HM) model. This aggressive tumor displayed rapid growth of the primary tumor mass, high incidence of metastases to lung and lymph nodes, and invasion from the primary s.c. site to the peritoneal cavity. Starting 12 days after s.c. tumor cell implantation, several i.p. injections of rIL-12 at 2-3 day intervals resulted in regression of growing tumors. These treated mice did not show signs of metastases or tumor recurrence at the original site. One month after tumor implantation, untreated mice did not have visible lung metastasis, but some did have palpable lymph nodes. At this stage, the primary tumors of animals without palpable lymph nodes were surgically resected. When examined 2 months later, most animals had developed lymph node and lung metastases. In contrast, rIL-12 injections after tumor resection inhibited the development of metastases in both lung and lymph nodes. This contrasted with the failure of IL-2 to prevent metastases. Even for mice already showing signs of lymph node metastases or invasion of the abdominal wall, rIL-12 administration after tumor resection prevented further invasion to the peritoneal cavity and growth of metastatic tumor cells in lung. It was somewhat surprising that the IL-12 treatment of animals after 1 month of tumor growth without resection also resulted in complete tumor regression, as well as eradication of micrometastasis that would have occurred before the treatment. Moreover, they exhibited resistance to a rechallenge with the same tumor but not with a second tumor. Thus, this tumor system provides a relevant model to clinical situations in terms of treatment of advanced tumors and metastases. These results also indicate that IL-12 can induce a curative immune response, even in the face of an aggressive micrometastasizing tumor.

Animals↗
PubMed1995

cDNA cloning and characterization of a novel receptor-type protein tyrosine phosphatase expressed predominantly in the brain.

Protein tyrosine phosphatase has the potential to control various cellular events by negatively regulating the extent of tyrosine phosphorylation. Here, we report the isolation of a murine receptor protein tyrosine phosphatase, PTPBR7, which is expressed almost exclusively in the brain. Though the cytoplasmic portion of PTPBR7 reveals high similarity to HePTP/LC-PTP and STEP, these are, unlike PTPBR7, non-receptor protein tyrosine phosphatases. Unlike most receptor protein tyrosine phosphatases, PTPBR7 has only one cytoplasmic phosphatase domain, and its extracellular domain reveals no obvious structural similarity to known molecules. Thus, PTPBR7 defines a new subfamily of receptor-type protein tyrosine phosphatases. The putative extracellular domain of PTPBR7 was expressed in COS-7 cells as a chimeric fusion protein with an immunoglobulin Fc portion (PTPBR7-Fc). PTPBR7-Fc was secreted in the culture supernatant, confirming the capability of the extracellular domain of PTPBR7 to translocate across the cytoplasmic membrane. The cytoplasmic portion of PTPBR7 was expressed as a fusion protein in bacteria and was demonstrated to have catalytic activity. The expression of PTPBR7 was detectable in brain and especially in cerebellum but undetectable in liver, lung, heart, kidney, thymus, bone marrow, and spleen. In situ hybridization analysis revealed the most prominent signal in Purkinje cells. The predominant expression of PTPBR7 in the brain suggests that PTPBR7 may have role(s) in neuronal cells.

Amino Acid Sequence↗
PubMed1995

Do structural changes of T cell receptor complex occur in tumor-bearing state?

T cells in tumor-bearing mice and cancer patients were recently shown to be devoid of CD3-zeta chain, a signal-transducing invariant chain in T cell receptor (TCR) complex, and p56lck tyrosine kinase. In the present study, we investigated the structure and function of TCR complex in T cells from BALB/c mice bearing CSA1M fibrosarcoma. The expressions of TCR chains and p56lck in a T cell-enriched population from spleen were analyzed. Almost complete loss of CD3-zeta and p56lck was observed in the preparation from tumor-bearing mice as assessed by immunoblotting analysis using whole cell lysates, whereas the amounts of other TCR chains were relatively unchanged. However, these changes were due to the increase of contaminating Mac-1+ cells in the spleen of tumor-bearing mice because: 1) the removal of Mac-1+ cells led to the restoration of CD3-zeta and p56lck; and 2) CD3-zeta was clearly present when the preparation was solubilized with ionic detergent. Fc receptor gamma chain detected in the preparation from tumor-bearing mice disappeared along with the removal of Mac-1+ cells. These observations were further supported by the finding that addition of Mac-1+ cells from tumor-bearing mice to normal T cells resulted in loss of CD3-zeta, leaving CD3-epsilon largely intact. When T cells from tumor-bearing mice were highly purified by depletion of Mac-1+ cells, these T cells contained normal amounts of CD3-zeta at mRNA, protein, and surface levels, and expressed the properly assembled TCR complex on their cell surface. Moreover, stimulation of the TCR in these T cells by anti-TCR antibodies resulted in a comparable Ca2+ mobilization to that observed in normal T cells. These results suggest that no structural changes occur in TCR complex in our tumor-bearing mice, and that complete depletion of Mac-1+ cells in important to assess the structure of TCR complex.

Animals↗
PubMed1995

Leukomyelopathy in ataxic calves.

Seventy-three fattening calves ranging from 6 to 15 months old on a farm were affected by ataxia over a period of four months. About 10 ataxic animals recovered after the administration of various kinds of vitamins and needle puncture therapy. The principal lesions in 14 ataxic animals examined were characterized by bilateral vacuolation with axonal degeneration confined to the spinal white matter. Ultrastructurally, the vacuoles were present in the periaxonal areas and bounded by dissociated myelin lamellae. No pathogen was isolated from the visceral organs of the animals. The serum copper remained within the normal range. Possible etiological factors remain unknown in the ataxic disease.

Animals↗
PubMed1995

Modulation of B-cell abnormalities in lupus-prone (NZB x NZW)F1 mice by normal bone marrow-derived B-lineage cells.

(NZB x NZW)F1(NZB/WF1) mice spontaneously develop an autoimmune disease characterized by abnormality of haemopoietic stem cells. The present study examined a possible regulatory cell interaction between NZB/WF1 and normal bone marrow cells using radiation-induced chimeras. We demonstrated that the ability of NZB/WF1 bone marrow cells to transfer the typical disease with hypergammaglobulinemia including autoantibodies into lethally irradiated normal recipients was prevented by cotransfer of bone marrow from normal CBA/J mice but not from xid CBA/N mice carrying a selective defect in B-cell function. Flow cytometric analysis revealed that the generation of NZB/WF1 cells was reduced in the mixed chimeras given CBA/J but not CBA/N bone marrow cells. Interestingly, radiation chimeras reconstituted with a mixture of NZB/WF1 bone marrow and CBA/J splenic B cells did not show elevation of serum immunoglobulin levels, although most of the spleen cells were dominated by NZB/WF1 cells. On the other hand, NZB/WF1 B cells maturated in vivo in the presence of CBA/J bone marrow or splenic B cells lost the hyper-responsiveness to lipopolysaccharide (LPS) in the autoantibody production in vitro. These results suggest that radiosensitive normal B-lineage cells have the regulatory activity to ameliorate the hypergammaglobulinemia of NZB/WF1 mice by reducing the generation of NZB/WF1 B cells and/or by correcting their hyper-responsiveness, and that NZB/WF1 mice may have a defect(s) in the regulatory cell function. In addition, CBA/J splenic B cells were shown to modulate the B-cell abnormality even when injected into non-irradiated NZB/WF1 mice manifesting autoimmunity.

Animals↗
PubMed1995

[Shortening of hospitalization period for patients with pulmonary tuberculosis].

The development of specific chemotherapeutic agents revolutionally improved the prognosis of tuberculosis and markedly shortened the duration of the treatment. Evidence of successful treatment with short-course regimens for pulmonary tuberculosis has been accumulated. But, the duration of hospitalization tends to be determined empirically and varys among hospitals and clinicians. Generally speaking, the duration of hospitalization is longer in Japan than in the United States and Europian countries. We investigated retrospectively the duration of hospitalization, treatment period, and recurrence rate of the disease for patients with pulmonary tuberculosis who admitted to the Tokyo Metropolitan Fuchu Hospital in the years 1984, 1988, and 1991. Patients who had been previously treated for more than 2 weeks and/or, had severe complications, such as malignancy and miliary tuberculosis were excluded from the study. Cases who died during the hospitalization and discharged without permission were also excluded. Numbers of subjects in each year were 114, 114, and 115 in 1984, 1988, and 1991, respectively. A total of 343 patients (M: 254, F: 89) were enrolled to the study and their average age was 49.4 yr old. A number of patients with both smear and culture positive in sputum bacteriology was 210 (61.2%), and 42 cases (12.2%) were smear negative and culture positive. The results are as follows: (1) Mean duration of hospitalization in the years 1984, 1988, and 1991 were 141, 102, and 72 days, respectively. That is, a mean hospitalization period was shortened to about half during these seven years. (2) According to the chest x-ray classification (Group A: I, II3, B: II1, II2, III3, C: III1, III2), mean durations of hospitalization were 186 days for group A, 143 days for group B, and 108 days for group C in 1984, and 96 days, 76 days, and 59 days in 1991, respectively. (3) According to sputum bacteriology, mean durations of hospitalization were 169 days for (S+/C+) cases, 105 days for (S-/C+) cases, and 90 days for (S-/C-) cases in 1984, and 83 days, 67 days, and 41 days in 1991, respectively.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1995

Characterization of defectiveness in endogenous antigen presentation of novel murine cells established from methylcholanthrene-induced fibrosarcomas.

Three cell lines (4A1, 4C2 and 6D1 cells) derived from fibrosarcoma induced by the inoculation of 3-methylcholanthrene into C3H/HeN (H-2k) mice were examined for their ability to present antigens to CD8+ cytotoxic T lymphocytes (CTL). 6D1 and 4C2 cells were deficient in presenting endogenously synthesized influenza virus antigens to CTL, but they were able to present antigens when they were sensitized with a synthetic epitope peptide. The expression of the H-2 Kk gene in 4C2 and 6D1 cells was much reduced and was detectable only with Northern blot hybridization. The expression of two transporter genes (TAP1 and TAP2), examined by Northern hybridization, was also reduced in both cells, and negligible particularly in 4C2 cells. Interferon-gamma (IFN-gamma) treatment of these cells induced expression of Kk, TAP1 and TAP2 genes and rescued the defect of class I-restricted antigen presentation in 4C2 and 6D1 cells. Even after this treatment, however, antigen-presentation capability of 4C2 cells was still much lower than that of normal 4A1 cells. This finding suggests that 4C2 cells might have an additional defective gene(s), whose products are involved in the processing of class I-restricted antigen, besides the Kk and TAP genes, and this may explain the difficulty of 4C2 cells to induce tumour-specific immunity, as described previously. To our knowledge, the 4C2 cell is the first tumour cell postulated to have more than three defective genes involved in class I-restricted antigen presentation.

ATP Binding Cassette Transporter, Subfamily B, Mem↗
PubMed1994

Developmentally regulated expression of a murine receptor-type protein tyrosine phosphatase in the thymus.

Reversible phosphorylation of tyrosine residues plays a crucial regulatory role in various cellular events, including differentiation and proliferation of lymphocytes. Here, we report the isolation of a murine receptor-type protein tyrosine phosphatase (PTP), mRPTP-sigma, which is expressed in both immature thymocytes and stroma cells. At least two alternatively spliced transcripts of mRPTP-sigma (T and B) were observed. mRPTP-sigma T was the dominant form in the thymus and had three Ig-like and eight fibronectin type III-like domains in the extracellular portion. mRPTP-sigma T was almost identical with RPTP-sigma/PTP NE-3/PTP-P1/CPTP1, a PTP recently cloned from rat brain, except that RPTP-sigma/PTP NE-3/PTP-P1 was about 400 amino acids shorter than mRPTP-sigma T. mRPTP-sigma B, the second form of mRPTP-sigma, was dominant in the brain and was most likely the murine counterpart of RPTP-sigma/PTP NE-3/PTP-P1. In the developing thymocytes, the expression of mRPTP-sigma was high in double negative (CD4-CD8-), low in double positive (CD4+CD8+), and marginal in single positive (SP; CD4+CD8- or CD4-CD8+) subpopulations. No upregulation of mRPTP-sigma was observed in the spleen cells stimulated with Con A. Developmental regulation of mRPTP-sigma expression suggests its involvement in the control of T lymphocyte differentiation.

Alternative Splicing↗
PubMed1994

Identification and characterization of IL-2 hyper-responsive NZB/WF1 CD5- B cells.

The responsiveness of CD5- and CD5+ B cells of BALB/c and NZB/WF1 mice to various cytokines was examined with respect to their growth and differentiation. BALB/c splenic CD5- B cells required longer incubation with IL-2 or pretreatment with IL-4 to respond to IL-2 by DNA synthesis, whereas NZB/WF1 CD5- B cells were highly competent to IL-2. Flow cytometric analysis demonstrated that NZB/WF1 and BALB/c CD5- B cells had higher and intermediate proportions of B cells positive for IL-2R beta, respectively. On the other hand, BALB/c and NZB/WF1 splenic CD5+ B cells consisted of lower proportion of B cells positive for IL-2R beta than did their corresponding CD5- B cells and grew meagerly in response to IL-2. Peritoneal exudative CD5+ B cells of NZB/WF1 mice lacked IL-2R beta mRNA expression and failed to respond to IL-2. Although both BALB/c and NZB/WF1 CD5- B cells pretreated with anti-IgM, IL-4, and IL-5 responded to IL-2 by DNA synthesis, only BALB/c CD5- B cells developed into IgM-producing cells. Furthermore, BALB/c, but not NZB/WF1 CD5-, B cells pretreated with anti-IgM and IL-5 responded to IL-2 by IgM production without DNA synthesis. Thus, cross-talk between IL-4 and IL-2 operated in the growth responses of both BALB/c and NZB/WF1 splenic CD5- B cells, whereas cross-talk between IL-5 and IL-2 operated only in the differentiation of BALB/c CD5- B cells, providing us with another intriguing functional abnormality of NZB/WF1 B cells.

Animals↗
PubMed1994

Recovery of antitumor CD4+ T cell responsiveness, suppressed in the tumor-bearing state, by release from tumor burden.

The present study investigates the recovery of antitumor CD4+ T cell responsiveness, suppressed in the tumor-bearing state, following release of tumor burden. Spleen cells from BALB/c mice bearing a syngeneic tumor (CSA1M) 1-3 weeks after the inoculation with CSA1M cells produced interleukin-2 (IL-2) and IL-4 upon in vitro cultures without addition of exogenous tumor antigens. This lymphokine production was achieved through collaboration between anti-CSA1M CD4+ T cells and antigen-presenting cells (APC) that has been pulsed with CSA1M tumor antigens in vivo in the tumor-bearing state. The lymphokine-producing capacity gradually decreased as the tumor-bearing period increased, and spleen cells from mice at late (8-10 week) tumor-bearing stages produced reduced levels of lymphokines. Because APC in these cells exhibited enhanced capacities to present tumor antigens, the reduced responsiveness was ascribed to the dysfunction of CD4+ T cells themselves. However, removal of a tumor after 8 weeks resulted in a remarkable recovery of the lymphokine-producing capacities of whole spleen cells. In contrast to the reduction in CSA1M-antigen-presenting activity of APC following tumor resection, CD4+ T cells exhibited a reciprocal increase in their responsiveness to CSA1M antigens. The recovery of antitumor responsiveness was also observed in the in vitro cultures free from tumor burden; when spleen cells from mice at late tumor-bearing stages were pre-incubated for 1-2 days and re-cultured in fresh medium, they produced potent amounts of IL-2 and IL-4. These results indicate that the immunodysfunction of antitumor CD4+ T cells induced in the tumor-bearing state is not irreversible, and release from tumor burden results in almost complete recovery of the potent antitumor responsiveness they previously expressed.

Animals↗
PubMed1994

Suppression of allograft responses by combining alloantigen-specific i.v. pre-sensitization with suboptimal doses of rapamycin.

C57BL/6 (B6) mice were injected i.v. with class I H-2-disparate B10.QBR spleen cells (10(7)/mouse). This regimen, termed donor alloantigen-specific i.v. pre-sensitization (DSP), induced almost complete reduction of the anti-B10.QBR mixed lymphocyte reaction (MLR) that has been regarded to represent a cytotoxic T lymphocyte (CTL)-independent graft rejection pathway. Because the DSP regimen failed to affect the generation of CTL responses, it did not prolong graft survival. Repeated (four or 11 times) administration in vivo (during 5 or 18 days) of rapamycin at suboptimal doses (0.5-2.0 mg/kg/day) failed to eliminate the capacities to exhibit MLR as well as to generate CTL responses. The suppression of CTL responses was achieved only through the combination of these two treatments. It was also shown that prolongation of skin graft survival was not induced by either of a single DSP or rapamycin treatment alone, but by the combination of these. Potent suppression of CTL responses was also induced when a single DSP was combined with repeated injection of a suboptimal dose (0.75 mg/kg/day) of another immunosuppressive drug, FK506, instead of rapamycin. However, there was a substantial difference in cellular mechanisms underlying the suppression of CTL responses by the above two different combinations. Under conditions in which lymphoid cells from mice receiving the treatments with DSP plus FK506 failed to generate CTL responses, the addition of recombinant IL-2 (rIL-2) to cultures restored the CTL generation, suggesting that CTL precursors themselves are not attenuated by the combined treatment.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1994

Activation-induced expression of thymic shared antigen-1 on T lymphocytes and its inhibitory role for TCR-mediated IL-2 production.

We have produced a hamster mAb, PRST1, which reacts with thymic shared Ag-1 (TSA-1), a product of the Ly6 gene family. By cross-blocking experiments, we found that TSA-1 is identical to stem cell Ag-2 (Sca-2). Using PRST1, the changes of TSA-1/Sca-2 expression on mature T cells during the activation process were analyzed. Although freshly isolated T cells did not express detectable TSA-1 on their cell surface, in vitro stimulation of T cells with concanavalin A induced a marked increase of surface TSA-1 expression. The increased expression of TSA-1 on T cells was detected from 12 h after stimulation and was associated with the increase of TSA-1 mRNA. In vivo injection of mice with staphylococcal enterotoxin B (SEB) resulted in the enhanced TSA-1 expression in splenic V beta 8+ T cells. This antigen-specific induction of TSA-1 expression in vivo preceded a detectable increase in numbers of V beta 8- T cells after SEB injection. Functionally, whereas anti-TSA-1 mAb was not mitogenic to T cells, it inhibited anti-CD3-induced IL-2 production by T cell hybridomas. These results indicate that TSA-1/Sca-2 is a unique marker for T cell activation and a signal through this molecule may have a negative feedback role to limit IL-2 production from activated T cells stimulated through the TCR.

Animals↗
PubMed1994

Diversity in molecular mass of the common EDTA-soluble antigens of Clostridium chauvoei and Clostridium septicum.

Common EDTA-soluble antigens of Clostridium chauvoei and C. septicum were examined by indirect-immunofluorescence (IFA) and immunoblot analysis. The monoclonal antibodies (mAbs) specific for the 35 kDa antigen of C. chauvoei strain ATCC 10092 were used. These mAbs reacted with all 11 strains, 6 of C. chauvoei and 5 of C. septicum, in IFA. In immunoblot analysis with the mAbs, the bands at molecular mass of 35 kDa were found in all C. chauvoei strains, while the bands at 36 kDa were found in 4 of 5 strains of C. septicum. These results indicate that the 35 kDa antigen of C. chauvoei and the 36 kDa antigen of C. septicum possess a similar epitope recognized by the mAb.

Animals↗
PubMed1993

Suppression of allograft responses by combining donor alloantigen-specific intravenous presensitization with suboptimal doses of FK506.

C57BL/6 (B6) mice were injected i.v. with class I H-2-disparate B10.QBR spleen cells (10(7)/mouse). This regimen, termed "donor alloantigen-specific i.v. presensitization" (DSP), induced almost complete elimination of anti-B10.QBR mixed lymphocyte reaction/IL-2 production but did not affect the generation of CTL responses. Repeated (5 or 11 times) administration in vivo (over 7 or 18 days) of FK506 at suboptimal doses (0.75-1.0 mg/kg/day) failed to eliminate the capacities to exhibit MLR/IL-2 production and to generate CTL responses. Prolongation of skin graft survival was not induced by either of a single DSP or FK treatment (0.75-1.0 mg/kg/day, 11 times during 18 days) alone, but by the combination of these. Such combined treatment also resulted in almost complete reduction of CTL responses before (5 rounds of FK injection) or after (11 rounds of FK injection) recipients were engrafted with B10.QBR skin grafts. Under conditions in which lymphoid cells from mice receiving both treatments failed to generate CTL responses, the addition of recombinant IL-2 to cultures restored the CTL generation, suggesting that CTL precursors themselves are not attenuated by the combined treatment. Both prolongation of graft survival and suppression of CTL responses were obtained when the administration of FK506 was started before but not after DSP. Prolongation of graft survival could also be obtained in class I and II MHC-disparate combinations when the combined treatment was performed in a particular protocol. These results indicate that (1) DSP alone fails to prolong graft survival in class I and class I and II MHC-disparate combinations; (2) such failure is ascribed to the induction of CTL responses by CTL precursors and CTL helpers, both of which are DSP-resistant; (3) the administration of suboptimal doses of FK506 is not sufficient for the suppression of CTL responses, but is effective selectively for suppressing DSP-resistant CTL helpers; and (4) the combination of DSP with FK506 treatment in an appropriate protocol can thus prolong graft survival through the suppression of CTL-involved as well as CTL-independent graft rejection pathways.

Animals↗