Search PubMed⌕ Search

Biomedical subjects

I Negishi

Publications and source records attributed to I Negishi.

35 records · Page 2Linked to original sources
PubMed1995

Naive CD28-deficient T cells can initiate but not sustain an in vitro antigen-specific immune response.

Naive T cells require an Ag-specific signal, as well as a costimulatory signal to mount a primary Ag-specific response. Because of their low precursor frequency, it has been difficult to study costimulatory requirements of these Ag-specific T cells. We have generated a CD28-deficient mouse that has been bred to a TCR transgenic (Tg) mouse to better study the function of CD28 during CD4+ T cell responses to Ag. In the absence of CD28, naive TCR Tg T cells responded vigorously to peptide, but responded poorly to mitogen activation. Comparison of activation-induced cell-surface molecules, including CD25, CD44, CD69, and CD71, showed no significant differences between CD28+ and CD28- TCR Tg T cells during the first 24 to 48 h after Ag stimulation. Despite relatively normal surface phenotype and normal proliferative response to Ag, CD28- T cells produced little IL-2, had a decreased sensitivity to lower Ag concentrations, and were unable to maintain their proliferative response. These results suggest that naive T cells are able to utilize other costimulatory signals to initiate a primary Ag-specific response, but require CD28 for optimal, sustained proliferation.

Abatacept↗
PubMed1995

Massive cell death of immature hematopoietic cells and neurons in Bcl-x-deficient mice.

bcl-x is a member of the bcl-2 gene family, which may regulate programmed cell death. Mice were generated that lacked Bcl-x. The Bcl-x-deficient mice died around embryonic day 13. Extensive apoptotic cell death was evident in postmitotic immature neurons of the developing brain, spinal cord, and dorsal root ganglia. Hematopoietic cells in the liver were also apoptotic. Analyses of bcl-x double-knockout chimeric mice showed that the maturation of Bcl-x-deficient lymphocytes was diminished. The life-span of immature lymphocytes, but not mature lymphocytes, was shortened. Thus, Bcl-x functions to support the viability of immature cells during the development of the nervous and hematopoietic systems.

Animals↗
PubMed1994

Targeted disruption of Bcl-2 alpha beta in mice: occurrence of gray hair, polycystic kidney disease, and lymphocytopenia.

Mice carrying ablated coding regions of the bcl-2 alpha and bcl-2 beta transcripts have been made. bcl-2-/- mutants are smaller but viable, although about half of them die by 6 weeks of age. As shown earlier with somatic bcl-2 gene-targeted mice, the number of lymphocytes markedly decreased within few weeks after birth while other hematopoietic lineages remained unaffected. Among lymphocytes, CD8+ T cells disappeared most quickly followed by CD4+ T cells, whereas B cells were least affected. bcl-2-/- lymphocytes, however, could respond normally to various stimuli including anti-CD3, Con A, phorbol 12-myristate 13-acetate plus ionomycin, interleukin 2, lipopolysaccharide, and anti-IgM antibody. Abnormalities among nonlymphoid organs include smaller auricles, hair color turning gray at 4-5 weeks of age, and polycystic kidney disease-like change of renal tubules. These results suggest that Bcl-2 may be involved during morphogenesis where inductive interactions between epithelium and mesenchyme are important such as in the kidneys, hair follicles, and perichondrium of auricles. Surprisingly, the nervous system, intestines, and skin appear normal despite the fact that these organs show high levels of endogenous Bcl-2 expression in normal mice.

Animals↗
PubMed1994

Requirement for CD8 beta chain in positive selection of CD8-lineage T cells.

CD8 is either an alpha alpha homodimer or an alpha beta heterodimer, although most peripheral CD8-lineage T cells express only the CD8 alpha beta heterodimer. The physiological function of CD8 beta was elucidated with mice that were chimeric for the homozygous disruption of the CD8 beta gene. The CD8 beta-1- T cells developed normally to CD4+CD8+ stage, but did not efficiently differentiate further, which resulted in few peripheral CD8+ T cells. The number of peripheral CD8+ T cells was restored by transfer of an exogenous CD8 beta gene into CD8 beta-deficient T cells. Thus, CD8 beta is necessary for the maturation of CD8+ T cells.

Animals↗
PubMed1993

Disappearance of the lymphoid system in Bcl-2 homozygous mutant chimeric mice.

The bcl-2 proto-oncogene can prevent the death of many cell types. Mice were generated that were chimeric for the homozygous inactivation of bcl-2. Lymphocytes without Bcl-2 differentiated into phenotypically mature cells. However, in vitro, the mature T cells that lacked Bcl-2 had shorter life-spans and increased sensitivity to glucocorticoids and gamma-irradiation. In contrast, stimulation of CD3 inhibited the death of these cells. T and B cells with no Bcl-2 disappeared from the bone marrow, thymus, and periphery by 4 weeks of age. Thus, Bcl-2 was dispensable for lymphocyte maturation, but was required for a stable immune system after birth.

Animals↗
PubMed1992

[Positive selection of a T cell repertoire during intrathymic differentiation].

In SWR mice most of thymocytes bearing a high density V beta chain of T cell antigen receptor (TCR), V beta 17a brightly positive cells, were biased to CD4+8- subpopulation, whereas such thymocytes from SJL mice resided in both CD4+8- and CD4-8+ subpopulations. To elucidate the mechanism underlying the apparent difference in the thymocyte population, CD4/CD8 phenotype of thymocytes bearing high V beta 17a was analyzed in bone marrow (BM) chimeras in which SWR or SJL mice were used as BM donors and various strains of mice as recipients. When SWR precursor cells developed in SJL or B10 thymus, the proportion of V beta 17a+CD4-8+ thymocytes was expanded to the level of that seen in normal SJL mice. By contrast, the proportion of SJL derived V beta 17a+ CD4-8+ thymocytes which had developed in SWR or DBA/1 thymus was reduced to the level of that in normal SWR mice. An intermediate proportion was observed when SWR precursor cells had developed in the thymus of B10.A(4R). It was revealed that H-2K molecules expressed on thymic epithelial cells is one of the determinants which influence the proportion of V beta 17a+CD4-8+ thymocytes. In addition, a hierarchy of selective pressure among the different H-2K haplotypes was present, such as Ks,b greater than Ks/q,k greater than Kq. Fine analysis of the H-2K molecules in which several kinds of H-2Kb mutant (bm) mice were used as recipients to prepare bone marrow chimeras demonstrated that substitutions of amino acids at a region on the beta-pleated floor of antigen recognition site reduced significantly the proportion of V beta 17a+CD4-8+ cells. According to the three-dimensional class I structure, TCR are unable to access directly to this region. Thus, the present finding suggests that the substitutions of amino acids at this site alter the shape and charge of peptide binding site of H-2K molecules and eventually influence the positive selection of the V beta 17a+ T cell repertoire during differentiation.

Animals↗
PubMed1991

Deficiency in early development of the thymus-dependent cells in irradiation chimeras attributable to recipient's environment.

Allogeneic bone marrow chimeras were prepared using reciprocal combinations of AKR and C3H mice. When C3H mice were recipients, the number of thymocytes recoverable from such chimeras (C3H recipient chimeras) was small as compared with that from chimeras for which AKR mice were used as recipients (AKR recipient chimeras) regardless of donor strain. The thymocytes from C3H recipient chimeras showed a profound deficiency in generating proliferative responses to stimulation by anti-CD3 mAb (2C11) or anti-TCR (alpha, beta) mAb (H57-597), even though the expression of CD3 and TCR molecules fell within the same range as that in AKR recipient chimeras. Furthermore, after stimulation with immobilized 2C11, the proportion of IL-2R+ cells in the thymocytes from C3H recipient chimeras was much less than that in AKR recipient chimeras. However, no significant difference in proliferative responses to 2C11 plus PMA, in influx of Ca2+ after stimulation with 2C11 or IL-2 production in response to 2C11 plus PMA or PMA plus A23187 was demonstrated between C3H and AKR recipient chimeras. These findings suggest that the thymocytes from C3H recipient chimeras have a deficiency in the signal transduction system as compared with chimeras for which AKR mice are the recipients. The thymic stromal component involved in this difference in the C3H recipient chimeras is discussed.

Animals↗
PubMed1991

H-2K molecules positively select V beta 17a+ CD4(-)8+ T cells in bone marrow and thymic chimeras.

Population size of V beta 17a brightly positive cells among CD4(-)8+ thymocytes was analyzed in thymic chimeras as well as bone marrow (BM) chimeras in which SWR/J mice were used as BM donors and various strains of mice including H-2Kb mutant (bm) mice as recipients. It was shown that the proportion of V beta 17a+ CD4(-)8+ thymocytes was determined by H-2K molecules expressed on thymic epithelial cells. The highest proportion was observed in Ks and Kb thymuses, the intermediate proportion in Ks/q and Kk, and the lowest in Kq thymuses. Fine analysis of the H-2Kbm molecules involved in the positive selection revealed that the region important to the selection was located on the beta-pleated floor of antigen recognition site. According to the three-dimensional class I structure, this site appears not to be directly accessible to the T cell antigen receptor. Thus, the present finding suggests that the substitutions of amino acids at this site alter the shape and charge of the peptide binding site and eventually influence the positive selection of the V beta 17a+ T cell repertoire during differentiation.

Animals↗
PubMed1991

Distribution of MEL-14+ cells in various lymphoid tissues.

The distribution of MEL-14+ lymphocytes was investigated by both fluorocytometric analysis and complement-dependent-cellular-cytotoxicity (CDCC) tests in which rabbit anti-rat Ig was added with complement at a secondary step. When CDCC was employed to detect MEL-14+ cells, almost half of the thymocytes were found to be MEL-14+ in various strains of mice. This high proportion of MEL-14+ cells stands in striking contrast to prior reports. Furthermore, when determined by fluorocytometric analysis, MEL-14+ cells were found to comprise more than 80% of the cells in the thymus. The MEL-14+ thymocytes comprised both immature subsets (CD4-8-, CD4+8+) and mature subsets (CD+8-, CD4-8+). MEL-14 brightly positive (MEL-14high) cells, however, were located mainly in mature T cell subpopulations within the thymus. The MEL-14high thymocytes appeared to be susceptible to the CDCC method. Most of MEL-14+ cells present in spleens and lymph nodes were shown to be included in the MEL-14high population. The MEL-14+ cells susceptible to treatment with MEL-14, rabbit anti-rat Ig plus complement in the spleen and lymph node were restricted to cells of the T-lineage. These data suggest that T cells may change from cells with low expression of the MEL-14 antigens at their surface to cells with high MEL-14 antigens in the process of differentiation. Furthermore, these findings indicate that MEL-14 molecules may be used as a surface marker to characterize an important T cell subpopulation.

Animals↗
PubMed1990

Sequential analysis of the thymocyte differentiation in fully allogeneic bone marrow chimera in mice. II. Further characterization of the CD4+ or CD8+ single positive thymocytes.

Differentiation of CD4+8- and CD4-8+ single-positive (SP) thymocytes in fully allogeneic bone marrow chimeras were investigated using multicolor cytometric analysis. The proportion of CD3+ cells in CD4+ SP population derived from donor mice considerably increased between day 12 and 14 after bone marrow transplantation (BMT), and gradually increased thereafter. The proportion of V beta 8+ cells in the CD3+CD4+ population remained constant (around 20%) at each period, suggesting that alpha and beta chains were used as TCR. The proportion of J11d+ cells in the CD4+ SP thymocytes transiently increased from day 12 to 14 and decreased thereafter, even though almost half of CD4+ SP cells were still dull J11d+ at day 35 after BMT. When CD8+ SP populations were analyzed, the proportion of CD3+ cells was very small until day 18. Thereafter, the proportion considerably increased and reached a maximum (83.2%) at day 21. The proportion of V beta 8+ cells in the CD3+ CD8+ SP population fell within range between 20 and 30%. However, before day 18, most of the V beta 8+ cells were dull positive, while after day 21 the majority were bright V beta 8+. Further, CD8+ SP cells at day 12, 14 and 18 were largely bright J11d+. After day 21, however, the proportion of bright J11d+ cells rapidly decreased. Similar results were obtained when the sequence of appearance of CD4+ and CD8+ SP cells was compared among bright CD3+, bright V beta 8+ or J11d- mature populations. The CD4+ SP cells regularly appeared earlier than CD8+ SP cells in the mature populations. These findings indicate that a considerable heterogeneity exists within both CD4+ and CD8+ SP populations and that the differentiation process for CD4+ SP cells precedes that for CD8+ SP cells.

Animals↗
PubMed1989

Donor and recipient specific tolerance in cells from semi-allogeneic, H-2 subregion compatible or fully allogeneic bone marrow chimeras attributable to clonal deletion.

Specificities of tolerance induced in allogeneic bone marrow (BM) chimeras which had been established by injecting allogeneic BM cells pretreated with anti-Thy-1 mAb alone (without complement (C)) were analyzed using Simonsen's splenomegaly assay. Lymphocytes from fully allogeneic, semi-allogeneic and H-2 subregion compatible BM chimeras were specifically unresponsive to donor and recipient antigens (Ag). However, cells from H-2 subregion compatible chimeras initiated as vigorously a GVHR in F1 recipient mice, which were disparate at H-2K and I-A regions, as did spleen cells of donor mice, which were incompatible at the entire H-2 and minor histocompatibility regions of the recipients. The donor cells from such chimeras that initiated these considerable GVHR were either CD4+ or CD8+ T cells. Furthermore, synergistic effects by the CD4+ and CD8+ T lymphocytes were also observed. We found no evidence for a suppressive mechanism(s) in maintenance of the specific tolerance in allogeneic chimeras. Further, when lymphoid cells from these chimeras were adoptively transferred to irradiated mice of the donor strain and maintained for 5 days in the absence of recipient Ag (tolerogen), the adoptively transferred cells were shown to retain their unresponsiveness to the recipient Ag. These results reveal that T lymphocytes from allogeneic BM chimeras prepared by our method had been specifically induced to a tolerant state to both donor and recipient Ag and that the major mechanism of induction and maintenance of long-lasting tolerance is attributable to clonal deletion of both CD4+ and CD8+ T cell subsets rather than to the development of a population of suppressor cells of any sort.

Animals↗
PubMed1989

Positive selection of a T-cell subpopulation in the thymus in which it develops.

In SWR mice the expression with high-density V beta 17a (high V beta 17a) of the T-cell antigen receptors correlates with the CD4+8- subpopulation of thymocytes. By contrast, in thymocytes of SJL mice the expression of high V beta 17a is observed on the CD4+8- or CD4-8+ subpopulation. However, when the thymocytes from SWR mice have been developed in the SJL or B10 thymus but not in the H-2 compatible DBA/1 thymus, a greater proportion of thymocytes that express high V beta 17a was found to be CD4-8+. By contrast, only a small proportion of KJ23a+ thymocytes from SJL mice that had differentiated in the thymus of SWR or DBA/1 mice was CD4-8+, whereas a high proportion of CD4+8- cells expressed V beta 17a. Further, an intermediate proportion of KJ23a+ thymocytes that had derived from SJL donor mice was present on CD4-8+ thymocytes that had developed in B10.A(4R) thymus. These findings demonstrate that the appearance of a particular subpopulation of thymocytes (CD4-8+ with a beta chain of T-cell antigen receptor identified as V beta 17a) is determined by the histocompatibility complex products that are expressed in the thymic microenvironment in which the T cells develop.

Animals↗
PubMed1975

Sex difference in the development of fatty liver by orotic acid.

Effects of orotic acid on liver lipid accumulation and incorporation of methionine [methyl-14C] into liver phosphatidylcholine and protein, and into serum beta-lipoprotein were studied. Male and female rats of Wistar strain were fed a semisynthetic diet supplemented with 1 per cent orotic acid for 7 days. Feeding of orotic acid induced a marked fatty liver in female rats, but not in males. In female rats, radioactivity in liver phosphatidylcholine was significantly decreased by orotic acid, and that in liver protein was slightly decreased. In male rats, incorporation of methionine [methyl-14C] into liver phosphatidylcholine and protein was unchanged between the control and the rats fed orotic acid. Radioactivity in serum beta-lipoprotein was decreased to a greater extent in female rats than in males. These results suggest that sex difference in the development of fatty liver may be due to the difference in the effect of orotic acid on liver phosphatidylcholine biosynthesis.

Acetates↗