K Hozumi

Tohoku University, Sendai-shi, Miyagi-ken, Japan

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Publications (13)40.12 Total impact

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    ABSTRACT: It has been reported that the three-dimensional structure of thymic epithelial cells (TECs) is responsible for thymic positive selection but that this ability disappears when TECs are cultured in monolayer. These results have supported the hypothesis that certain TEC-specific molecules are extinguished during monolayer culture. In this study, using MHC class II-restricted T-cell receptor transgenic mice, we demonstrated that preselected CD4(+)8(+) (DP) thymocytes were inhibited from developing into CD4(+)8(-) (CD4SP) cells in reaggregate thymus organ culture with monolayer-cultured TECs, but this inhibition was removed when TECs were cultured in monolayer with protein synthesis inhibitor or when the cultured TECs were treated with fixative. These results seem to be inconsistent with the previous hypothesis and indicate that monolayer culture allows TECs to retain the surface molecules necessary for positive selection but interferes with their function, which must be sustained for three dimensional structure.
    Cellular Immunology 08/2001; 211(1):71-9. · 1.74 Impact Factor
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    ABSTRACT: To overcome low efficiency of retroviral infection into immature T cells, we modified reaggregation fetal thymus organ culture by closely packed co-culture with virus-producing cells (VPC). The viral vector was constructed in chimeric vector, pMX, with IRES and tailless-rat CD2 as a surface marker of infected cells. A rearranged TCR beta gene (Vbeta8.2) was further inserted into the construct for investigating effect of the introduced gene in T cell development. Using this system, we succeeded to transfer the viral vector into immature thymocytes at a remarkably higher efficiency compared to conventional methods using medium containing retrovirus. Moreover, the introduced TCR beta gene was expressed on thymocytes of RAG2-deficient mice to induce in the transition of CD4-CD8- double-negative (DN) into CD4+CD8+ double-positive (DP) cells by transducing beta-selection signaling. Thus, our modified reaggregation culture system is useful for studying the molecular mechanism of T cell development due to a highly efficient gene transfer into immature T cells.
    Immunology Letters 02/2000; 71(1):61-6. · 2.34 Impact Factor
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    ABSTRACT: The differentiation process from CD4-CD8- double-negative (DN) thymocytes to CD4+CD8+ double-positive (DP) stage is accompanied by vigorous proliferation. The resulting DP cells contain a sizable proportion of large cycling cells, but most DP cells are small resting cells. To explore the molecular mechanisms which regulate cell proliferation of DP thymocytes prior to further development, we used TCR-transgenic (Tg) mice with non-selecting MHC (Tg-Neut), which contain almost exclusively DP thymocytes that are not subject to either positive or negative selection. In Tg-Neut, the thymus contained DP cells of relatively large size, which showed higher extracellular signal-regulated kinase activity and enhanced responsiveness to mitogen compared to small DP cells. This indicates that all the large DP cells in the thymus are not positively selected and that they possess proliferative potential. When Tg-Neut mice were backcrossed with CD45 knockout mice (CD454-/- Tg-Neut), the thymus showed an increase of large DP cells and cycling cells, but a decrease of apoptotic cells. Furthermore, Bcl-2 expression and Jun N-terminal kinase activity, which are associated with resistance to apoptosis, were enhanced. These observations suggest that thymocyte proliferation in the DP stage is suppressed by a CD45-related process with regulation of mitogen-activated protein kinase and Bcl-2 unless DP cells receive TCR-mediated signals.
    International Immunology 02/1999; 11(1):89-97. · 3.14 Impact Factor
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    ABSTRACT: Antigen stimulation via TCR in mature T cells provides rapid induction of tyrosine phosphorylation of intracellular substrates including ZAP-70. To study the potential involvement of tyrosine phosphorylation in CD4+CD8+ [double-positive (DP)] thymocytes in the positive selection process in vivo, we isolated and analyzed them in the presence of phosphatase inhibitor. DP thymocytes were obtained from TCR transgenic mice (TCR-Tg) expressing MHC class I- or class II-restricted TCR in selecting and non-selecting MHC backgrounds respectively. The phosphorylation of ZAP-70 in DP thymocytes of class I-restricted TCR-Tg was significantly higher in the positively selecting background than in the non-selecting one. However, such a phosphorylation difference between selecting and non-selecting TCR-Tg was found to be considerably less in class II-restricted TCR-Tg. A similar bias for ZAP-70 phosphorylation was also observed on selecting DP thymocytes when I-A(beta) deficient- and beta2-microglobulin-deficient mice were compared. These ex vivo studies suggest that TCR-mediated signaling on DP thymocytes induces ZAP-70 phosphorylation under a different manner of engagement of TCR to class I and class II molecules in the positive selection process.
    International Immunology 09/1998; 10(8):1203-10. · 3.14 Impact Factor
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    ABSTRACT: T cell receptor (TCR) genes are rearranged and expressed in an ordered manner during T cell development. The basic mechanism regulating this stepwise DNA alteration is poorly understood. To address this issue, we explored the presence of a stage-specific element for germ-line transcription of the TCR alpha gene which is closely associated with gene rearrangement. First, germ-line transcription of the TCR alpha gene including the first segment of the J alpha locus, J alpha49, was delayed compared to that of the TCR beta gene in both normal and TCR-transgenic (Tg) mice. Furthermore, expression of this transcript could be induced by CD3epsilon-mediated signals in recombination-activating gene (RAG)-2-deficient mice. In TCR-Tg mice, the endogenous J alpha49 germ-line transcript could not yet be observed at the CD25+ double-negative (DN) stage when the TCR alpha transgene was expressed. Of immature T cell hybridomas derived from either scid thymocytes (CD25+ DN) or immature CD8-single positive (ISP) thymocytes, only the latter hybridoma expressed the J alpha49 germ-line transcript. These data indicate that the J alpha49 germ-line transcription occurs only at a specific developmental stage. Second, to determine which elements may be regulating stage specificity, we performed transient transfection analysis with a reporter gene and demonstrated that the upstream region of the J alpha49 locus possesses promoter activity in correlation with germ-line transcription in ISP-derived but not in SCID-derived hybridomas. These results indicate that the expression of TCR alpha germ-line transcripts is regulated in a stage-specific manner by a cis-element located within the J alpha locus.
    European Journal of Immunology 05/1998; 28(4):1368-78. · 4.97 Impact Factor
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    ABSTRACT: It was found that freshly isolated BALB/c CD4+ T cells produced high levels of IL-4 and IL-10 in response to immobilized anti-CD3 mAb, while C57BL/6 CD4+ T cells produced low amounts of IL-4 and IL-10. The high IL-4-producing ability of BALB/c mice was demonstrated to be genetically dominant and it was controlled by non-MHC gene (or genes). The cells responsible for IL-4 production in BALB/c mice were defined as TCRVbeta8.2+ CD4+ CD62L- CD45RB- memory-type T cells, which were distinct from NK1.1+ CD4+ NKT cells. Although these memory-type T cells were also detected in C57BL/6 mouse spleen at the same frequency, they showed a functionally different property from BALB/c CD4+ CD62L- CD45RB- T cells in terms of IL-4 production. The fact that germfree BALB/c mouse spleen cells also produced high levels of IL-4 suggested that the IL-4 producer in BALB/c mice might be developed under the influence of unknown factors other than environmental Ags. The CD4+ CD62L- CD45RB- T cells obtained from BALB/c mice accelerated the development of IL-4-producing memory-type CD4+ T cells from CD4+ CD62L+ CD45RB+ naive T cells prepared from OVA-specific TCR-transgenic mice. Therefore, IL-4-producing CD4+ CD62L- CD45RB- T cells might play an important role in the preferential induction of Th2-dominant immunity in BALB/c mouse strain.
    The Journal of Immunology 07/1997; 158(12):5698-706. · 5.52 Impact Factor
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    ABSTRACT: Immature CD4+CD8+ double-positive (DP) thymocytes are positively selected for further development if they express TCR reacting with thymic ligands of low affinity. However, the majority of DP thymocytes express low TCR levels. This low level of TCR may be insufficient to recognize thymic ligands. To understand the basis for the low expression of TCR on DP thymocytes, we determined the density of TCR expression at various stages of their development using TCR transgenic (TCR-Tg) mice. We found that TCR expression was high in the thymocytes that had recently transited into the DP stage but then gradually decreased on DP cells if they were not selected by TCR interaction with MHC molecules. However, such TCR suppression was not observed in positively selected DP cells and in the non-selected DP cells obtained from CD45 deficient mice or from mice receiving anti-CD4 mAb. These findings suggest that the once highly expressed TCR at the DP stage is suppressed by CD45 and/or CD4 on non-selected thymocytes. Furthermore, TCR suppression is prevented by TCR-mediated signals. The maintenance of high TCR levels on positively selected DP thymocytes may facilitate their selection.
    International Immunology 11/1996; 8(10):1529-35. · 3.14 Impact Factor
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    ABSTRACT: Haloperidol (HPD) is a dopamine receptor blocker and a major causative agent of neuroleptic malignant syndrome. To investigate the influence of HPD on immune responsiveness, the natural killer (NK) cell activity of mice was examined after intraperitoneal administration of HPD for 5 days. NK cell activity was markedly decreased without a depletion of NK cells. Bromocriptine (BROMO), which is used for the treatment of neuroleptic malignant syndrome, also decreased NK cell activity. The suppressive effect on NK cell activity was inhibited by injecting both HPD and BROMO simultaneously. Serum levels of prolactin (PRL) decreased after BROMO injection, although serum PRL level was not decreased after the combined administration of HPD and BROMO agents. The reduction of NK cell activity caused by BROMO and by HPD was prevented by the co-injection of PRL and by a beta-adrenergic blocker, respectively. These results indicate that HPD decreases NK cell activity in a PRL-independent manner and that BROMO decreases it via PRL reduction. It seems that the PRL-independent suppressive effect of HPD on NK cells, which is neutralized by BROMO, is mediated by splenic sympathetic function via the beta-adrenergic receptor system. Therefore, BROMO helps to alleviate the depressed NK cell activity caused by HPD therapy.
    Brain Behavior and Immunity 04/1996; 10(1):17-26. · 5.61 Impact Factor
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    ABSTRACT: The effects of IL-7 on the growth and differentiation of thymocytes were analyzed using murine fetal thymus organ cultures (FTOC) in the presence of mAbs specific for the conventional IL-7 receptor (IL-7R) and for the common gamma (gamma c) chain. In FTOC, the development of CD4-CD8- double-negative thymocytes to CD4+CD8+ double-positive (DP) and CD4+ or CD8+ single-positive (SP) cells was not completely blocked by adding these mAbs, although cell growth was reduced by the treatment. To define a developing stage sensitive to the mAbs, most immature thymocytes, Pgp-1+ c-kit+ cells, were cultured in 2-deoxyguanosine treated fetal thymus. In the presence of both mAbs in the culture, neither DP nor SP thymocytes developed whereas either of the mAbs partially blocked their development. These results indicate that the gamma c chain is involved in early T cell development as an indispensable subunit of the functional IL-7 receptor complex.
    International Immunology 10/1994; 6(9):1451-4. · 3.14 Impact Factor
  • K Hozumi, T Masuko, Y Hashimoto
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    ABSTRACT: Nonparenchymal cells isolated from Fischer rat liver were separated into subpopulations by passage through a nylon wool column and/or culturing in a plastic plate. Besides typical Kupffer cells, we detected a unique population of cells (called PKu cells) which were plastic adherent but did not spread on a short term culture, were nonadherent on nylon wool, and were nonphagocytic, as opposed to Kupffer cells. Both Kupffer cells and PKu cells expressed CD4 (recognized with W3/25 mAb), CD8 (OX-8), a NK cell antigen (3.2.3), and a monocyte antigen (OX-41), as assessed by flow cytofluorometry. However, the proportion of cells bearing high densities of CD8 and 3.2.3 antigens was much larger for PKu cells than for Kupffer cells. Reverse transcriptase-PCR analysis of CD8 mRNA revealed that PKu cells expressed the CD8 alpha chain but not the beta chain. When PKu cells were treated with phorbol 12-myristate 13-acetate (PMA), they exhibited spreading and phagocytic activities, and showed a similar morphology to Kupffer cells in the spread form. Moreover, PMA treatment decreased the high density of CD8 antigen on PKu cells. These findings indicated that PKu cells present in normal rat liver are precursors of Kupffer cells.
    Journal of Biochemistry 06/1994; 115(5):904-8. · 2.72 Impact Factor
  • K Hozumi, T Masuko, Y Hashimoto
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    ABSTRACT: Expression of Thy-1 and CD3 antigens on thymocytes and the nylon wool-non-adherent (nylon-nonad) fractions of peripheral lymphocytes in F344 rats was studied by flow cytometry using anti-Thy-1 and anti-CD3 antibodies. In terms of the expression and density of Thy-1 and CD3, the thymus and peripheral lymphoid tissues in rats contained different populations. Nearly all thymocytes showed a high density of Thy-1 (Thy-1hi); a small population with a low density of Thy-1 (Thy-1lo) was found in CD3 bright-positive (CD3hi) thymocytes. Three-color analysis with CD4, CD8 and Thy-1 revealed Thy-1lo thymocytes to have mainly CD4 or CD8; Thy-1hi thymocytes also contained CD4 or CD8 single-positive (SP) cells as a minor population. Thus, SP thymocytes bearing CD3 were divided into two populations by evaluation of the density of Thy-1. Of peripheral CD3+ T cells, 15% showed a low density of Thy-1+ and the others were Thy-1-. Both T cells responded to anti-CD3 monoclonal antibody (mAb). However, no Thy-1hi T cells were found in peripheral lymphoid tissues. These results suggested that SP thymocytes in rats develop with the reduction of Thy-1, but not with its loss, and are newly supplied to peripheral lymphoid tissues as the phenotype, Thy-1lo, and are altered to Thy-1- T cells. Analysis of each cell size and disappearance of peripheral Thy-1lo T cells with age supported the above conclusion.(ABSTRACT TRUNCATED AT 250 WORDS)
    Immunology Letters 03/1994; 39(2):179-85. · 2.34 Impact Factor
  • K HOZUMI, T MASUKO, Y HASHIMOTO
    Immunology Letters - IMMUNOL LETT. 01/1994; 39(2):179-185.
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    ABSTRACT: In this study, we determined the characteristics of CD3-positive (CD3+) T cells existing in rat bone marrow (BM). In contrast to splenic T cells, BM CD3+ T cells are composed of a higher proportion of CD8+ T cells, and the number of both cell types increased with age. Such CD3+ T cells in aged rats showed a similar usage of TCR V beta as splenic T cells, suggesting that BM CD3+ T cells are thymus-dependent and composed of an ordinary population in view of the expression of the TCR beta-chain. Purified T cells obtained from aged rat BM showed a markedly proliferative response by stimulation with immobilized anti-CD3 mAb, as did splenic T cells. However, the addition of BM non-T cells completely inhibited the response of both BM and splenic T cells in vitro. These results suggest that T cells in rat BM are negatively regulated by BM non-T cells in their response to the TCR-mediated signal not to disrupt the microenvironment of the BM.
    Immunology Letters 06/1993; 36(2):137-43. · 2.34 Impact Factor