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T Suda

Publications and source records attributed to T Suda.

At least 649 records · Page 36Linked to original sources
PubMed1989

Responses to corticotropin-releasing hormone and its bound and free forms in pregnant and nonpregnant women.

Plasma CRH levels are considerably higher in women during the third trimester of pregnancy than in non-pregnant women. Most of plasma CRH in pregnant women is bound to CRH-binding protein (CRH-BP). To gain further insight into CRH physiology during pregnancy, we measured the responses of plasma ACTH and cortisol and the changes in bound and free forms of CRH in plasma after human CRH administration (2 micrograms/kg) in five pregnant (39-40 weeks of pregnancy) and five nonpregnant women. The mean basal plasma ACTH and cortisol levels in the pregnant women were higher than those in the nonpregnant women. However, the maximum increments in plasma ACTH and cortisol levels and the integrated ACTH and cortisol responses, after subtraction of the basal levels after CRH administration, were similar in the two groups. The plasma CRH half-time in the pregnant group was similar to that in the nonpregnant group. The mean basal plasma CRH level in the nonpregnant women was 1.5 +/- 0.2 (+/- SE) pmol/L, and that in the pregnant women was 360 +/- 35 pmol/L. On gel filtration chromatography, almost all of the CRH in the plasma was protein bound (320 +/- 30 pmol/L) in the pregnant women; no CRH peaks were detected in nonpregnant women because of the low plasma CRH levels. After CRH administration, the level of the bound form of plasma CRH was highest at 5 min, and then declined to a plateau at 15 min and 30 min in the pregnant women. In the nonpregnant women, protein-bound CRH also was highest at 5 min, but it progressively declined thereafter. The disappearance rate of the bound CRH in plasma from the nonpregnant women was similar to that of the second compartment of the plasma decay curves of the free CRH from both groups. We conclude that the plasma ACTH and cortisol responses to exogenous CRH are similar in pregnant and nonpregnant women, the effect of CRH-BP on the disappearance of plasma CRH is minimal, and plasma CRH-BP in pregnant women has the capacity to bind additional CRH.

Adrenocorticotropic Hormone↗
PubMed1989

[Comparison of non-HPLC 1 alpha, 25(OH)2D assay system with the HPLC method].

Recently, a method not requiring high performance liquid chromatography (HPLC) has been developed (Amersham Inc., U.K) for the measurement of levels of 1 alpha, 25(OH)2D, which demonstrates the most potent physiological effect among the metabolites of vitamin D. Until recently, purification of 1 alpha, 25(OH)2D fraction with HPLC has been considered indispensable prior to radioreceptor assay of this metabolite. Therefore, we used the Amersham assay system to determine whether it is possible to correctly measure the level of 1 alpha, 25(OH)2D without the HPLC process. The procedures we used were to extract the serum with an organic solvent, followed by fractionation of vitamin D metabolites through an LH-20 column. The products with or without the use of HPLC were submitted to radioreceptor assay using chick embryo intestinal receptors. As a result, it was found that (1) Separation and yield were improved by changing the column size, (1) Serum values 1 alpha, 25 (OH)2D in healthy subjects obtained by HPLC and non-HPLC methods were in good agreement with the theoretical values. Although the values thus obtained in patients with chronic renal insufficiency (with or without dialysis) were somewhat lower than the theoretical values, the result was consistent because there was no significant difference between the tow sets of measured values, and the correlation coefficient was r = 0.990. (3) Variations within the assay were CV = 16.5% for HPLC group and CV = 21.4% for non-HPLC group.(ABSTRACT TRUNCATED AT 250 WORDS)

Calcitriol↗
PubMed1989

[Biochemical characterization of tumor rejection antigen (TRA) and genetic control in anti-TRA immune responses].

A tumor antigen capable of inducing tumor resistance (tumor rejection antigen; TRA) was obtained in a solubilized form by sodium dodecyl sulfate (SDS)-extraction of plasma membrane fraction from Rous sarcoma virus (RSV)-induced CSA1M fibrosarcoma cells (BALB/c origin). Analyses by Sephacryl S-300 gel filtration and SDS-polyacrylamide gel electrophoresis revealed that TRA activity was recovered in the fraction of an approximate molecular weight (m. w.) of 60kD. Such TRA was also found to exist in plasma membrane fraction from various RSV-induced tumor cells but not from tumor cells induced with agents other than RSV. Unfractionated crude SDS-solubilized preparation contained gp 70 as detected by rabbit anti-gp 70 antiserum, whereas this reactivity was lost in the fraction exhibiting an m.w. of about 60 kD. Since this fraction retained pp60src activity, the relation of TRA to pp60src was further investigated. pp60src was also obtained from the lysate of v-src-expressing yeast transformant. Immunization of BALB/c mice with such pp60v-src-containing lysate failed to induce any significant tumor protection. The above 60 kD fraction of CSA1M solubilized antigens was allowed to bind to Sepharose beads coupled with anti-pp60src monoclonal antibody and was depleted of pp60src. Immunization with the fraction depleted of pp60src activity (bead-unbound fraction) resulted in potent tumor protection. These results indicate that the CSA1M solubilized membranous component(s) which has an approximate m.w. of 60kD but is (are) distinct from functional pp60src functions as the TRA against RSV-induced CSA1M tumor cells. The results are discussed in the context of (1) the relationship between the src gene and the expression of TRA and (2) genetic control in immune responses to the TRA common to RSV-induced tumor cells.

Animals↗
PubMed1989

Metabolism of vitamin D in human promyelocytic leukemia cells (HL-60) II. Does the differentiation of the cells alter the metabolism?

The metabolism of 25-hydroxyvitamin D3 (25-OH-D3) in human promyelocytic leukemia cells (HL-60) which had been exposed to 1 alpha, 25-dihydroxyvitamin D3 (1,25-(OH)2D3) was compared with that in untreated cells. 8 alpha, 25-Dihydroxy-9,10-seco-4,6,10(19)-cholestatrien-3-one (8,25-(OH)2-3-oxoneo-D3) was the only major metabolite in untreated HL-60 cells but exposure of the cells to 1,25-(OH)2D3 for 1 day significantly changed the metabolism: 24R,25-dihydroxyvitamin D3 (24R,25-(OH)2D3) was produced as the major metabolite together with 8,25-(OH)2-3-oxoneo-D3 whose production was also increased. The 1,25-(OH)2D3 treatment did not stimulate the production of 25-hydroxyvitamin D3 26,23-lactone by the HL-60 cells. The metabolism of 25-OH-D3 by HL-60 cells was also significantly affected by fetal bovine serum added to the incubation medium: production of 24R,25-(OH)2D3 was significantly reduced and formation of 8,25-(OH)2-3-oxoneo-D3 was completely abolished.

Calcifediol↗
PubMed1989

Growth and differentiation of a human megakaryoblastic cell line, CMK.

Recently, a human megakaryoblastic cell line, CMK, was established from the peripheral blood of a megakaryoblastic leukemia patient with Down syndrome. Using this cell line, we studied the proliferation and differentiation of megakaryocytic cells in the presence of highly purified human hematopoietic factors and phorbol 12-myristate-13-acetate (PMA). In a methylcellulose culture system, interleukin-3 (IL-3) and granulocyte-macrophage colony-stimulating factor (GM-CSF) facilitated colony formation by CMK cells in a dose-dependent manner. The maximum stimulating doses of these factors were 10 and 200 U/mL, respectively. These concentrations were comparable to those that stimulate activity in normal hematopoietic cells. In contrast, granulocyte-colony stimulating factor (G-CSF), macrophage-colony stimulating factor (M-CSF), and erythropoietin (EPO) had no effects on the colony formation of CMK cells. In a liquid culture system, 20% of the CMK cells expressed glycoprotein IIb/IIIa (GPIIb/IIIa) antigen without hematopoietic factors, whereas 40% of the cells expressed GPIIb/IIIa with the addition of IL-3 and GM-CSF. EPO also slightly enhanced expression of GPIIb/IIIa. On the other hand, PMA inhibited growth of CMK cells and induced most of them to express the GPIIb/IIIa antigen. Furthermore, PMA induced CMK cells to produce growth activity toward new inocula of CMK cells. This growth factor (GF) contained colony-stimulating activity (CSA) in normal bone marrow (BM) cells. The activity was believed to be attributable mainly to GM-CSF, since 64% of this activity was neutralized by anti-GM-CSF antibodies and a transcript of GM-CSF was detected in mRNA from PMA-treated CMK cells by Northern blot analysis. These observations suggest that GM-CSF, as well as IL-3, should play an important role in megakaryocytopoiesis.

Blotting, Northern↗
PubMed1989

Generation of osteoclasts from isolated hematopoietic progenitor cells.

A variety of studies have shown that osteoclasts originate from bone marrow, but their exact progenitors and differentiation pathway remain unclear. The treatment of mice with a high dose of 5-fluorouracil (5-FU) results in an enrichment for primitive hematopoietic progenitors; using this procedure, we prepared a new class of murine hematopoietic colonies that had very high secondary plating efficiencies in vitro. When spleen cells from mice pretreated in vivo with 5-FU were cultured in the presence of methylcellulose medium containing recombinant interleukin-3 (rIL-3), small colonies consisting of blast cells with little sign of differentiation developed on day 7 of culture. We lifted these blast colonies, pooled them, and replated them as secondary methylcellulose cultures in the presence of rIL-3 and erythropoietin. Approximately 60% of the cells formed colonies comprising various combinations of neutrophils, macrophages, eosinophils, mast cells, megakaryocytes, and erythroblasts. We replated such blast cells into microtiter wells and cultured them in the presence of rIL-3 (100 U/mL) or recombinant granulocyte-macrophage colony stimulating factor (GM-CSF) (100 U/mL) plus 1.25(OH)2D3 (10(-7) mol/L). Multinucleated cells appeared from day 14 of culture and approximately 100 giant cells per well were scored on day 21 of culture. Parathyroid hormone (1 U/mL) also induced the multinucleated cell formation. May-Grunwald-Giemsa staining revealed the large cells containing many nuclei in their cytoplasm, which is characteristic of bone-resorbing cells or osteoclasts. These cells showed a tartrate-resistant acid phosphatase (TRAP) activity. Calcitonin caused a striking shape change in these cells and suppressed the formation of multinucleated cells. Moreover, electron microscopy shows that these cells were able to resorb fetal calvariae. In the presence of r granulocyte-colony stimulating factor, r macrophage-colony stimulating factor, or r interleukin-6 plus 1.25(OH)2D3, formation of TRAP-positive multinucleated cells was lower compared with the support of rIL-3 or rGM-CSF. Mature macrophages collected from colonies did not form the multinucleated cells as described above, even in the presence of rIL-3 and 1.25(OH)2D3. Moreover, to exclude the possibility that osteoclasts generated from non-blast cells, we performed a cloning experiment from one isolated blast cell and demonstrated that single cells differentiate into osteoclasts or macrophages in the presence of rIL-3 with or without 1.25(OH)2D3. This system will provide a useful model for further analysis of osteoclast formation in vitro.

Animals↗
PubMed1989

Bone cell physiology.

Physiologic regulation of bone cell function is achieved by the combined actions of some circulating hormones and several locally produced cytokines. Interactions among these agents are important, and the regulated generation of proteolytic activities is likely to play a major role in determining net effects on bone cell function. It is likely that the most important local factors in regulating osteoclast formation and activation have yet to be chemically characterized.

Animals↗
PubMed1988

Recombinant human interleukin 6 (B-cell stimulatory factor 2) is a potent inducer of differentiation of mouse myeloid leukemia cells (M1).

Recombinant human interleukin 6 (IL-6), a lymphokine involved in the final differentiation of activated B-cells into antibody-forming cells, greatly suppressed proliferation and induced differentiation of murine myeloid leukemia cells (M1) into mature macrophage-like cells. When M1 cells were treated with IL-6, their growth was completely arrested as early as on day 2, and they were induced to differentiate morphologically into macrophage-like cells. Differentiation-associated properties such as phagocytic activity, adherence to the dish surface, Fc and C3 receptors, were also induced within 24 h by IL-6, and they reached their respective maximal levels on day 2 or 3. The potency of IL-6 in suppressing proliferation and inducing differentiation was much greater than that of 1 alpha,25-dihydroxyvitamin D3 one of the most potent inducers of M1 cells. The present report indicates that IL-6 is involved in the differentiation of not only B-cells but also myeloid leukemia cells.

Animals↗
PubMed1988

Differentiation in vitro of T3+ large granular lymphocytes with characteristic cytotoxic activity from an isolated hematopoietic progenitor colony.

Blast colonies were developed by rIL-3 from the spleen cells of mice pretreated with 5-fluorouracil (5-FU) in the methylcellulose cultures. When such IL-3-induced blast colonies were individually lifted up and recultured in the presence of rIL-3 and recombinant erythropoietin (rEpo), a variety of hematopoietic colonies developed from every single colony, including neutrophils, macrophages, eosinophils, megakaryocytes, mast cells, and erythroblasts. The results indicated that IL-3-induced blast colonies consisted of multipotential hematopoietic progenitor cells. By culturing individual IL-3-induced blast colonies in the presence of rIL-2 and irradiated peritoneal macrophages, on the other hand, the proliferation of homogeneous lymphoid cells was observed in 5 of 24 wells, each of which received a single blast colony. Morphologically, they were typical large granular lymphocytes (LGL), and thus it was indicated that LGL could be differentiated directly from the hematopoietic progenitor cells utterly in vitro by rIL-2 and accessory macrophages. From one of these culture wells, a continuous LGL line, IL3B1, was successfully obtained. The proliferation of IL3B1 was dependent on IL-2 in the presence of accessory macrophages, but they no longer responded to IL-3, nor to another T cell growth factor, IL-4. Flow cytometric analysis indicated that the phenotype of IL3B1 was Thy-1+,T3+,L3T4-,Lyt-2-,T200+ Asialo GM1+, whereas that of original IL-3-induced blast cells was Thy-1+,T3-,L3T4-,Lyt-2-,B220-. The results suggested that the expression of T3 molecules was induced in the process of LGL differentiation from the hematopoietic progenitor cells in vitro. Conforming to this, it was revealed that both gamma and beta chain genes of the TCR were rearranged in IL3B1. Northern blot analysis indicated that IL3B1 had abundant mRNA for gamma chain, while mRNA for beta chain was rather faint. Functionally, IL3B1 exhibited typical NK-patterned cytotoxic activity against a panel of tumor cell targets. In addition, they showed significant cytotoxic activity against normal bone marrow cells, as well as various factor-dependent myelogenous progenitor cell lines, all of which were resistant to endogenous NK activity of the fresh spleen cells. These results indicated that at least a set of T3+ LGL with rearranged TCR genes could be directly differentiated from isolated hematopoietic progenitor cells in vitro. Results also suggested that such a prethymically differentiated subset of T-lineage lymphocytes, namely T3+ double-negative LGL, had particular cytotoxic activity in addition to conventional NK activity, which might well contribute to feedback regulation of hematopoiesis.

Animals↗
PubMed1988

Studies on the induction of tolerance to alloantigens. III. Induction of antibodies directed against alloantigen-specific delayed-type hypersensitivity T cells by a single injection of allogeneic lymphocytes via portal venous route.

BALB/c mice were inoculated with normal C3H/He spleen cells via the portal venous (p.v.) route. Intravenous injection of serum from these BALB/c mice into naive syngeneic mice resulted in almost complete abrogation of their ability to generate anti-C3H/He delayed-type hypersensitivity (DTH) responses as induced by s.c. immunization with C3H/He cells. Since a portion of the same serum did not inhibit the development of anti-C57BL/6 DTH responses, the suppressive effect of the transferred serum was alloantigen-specific. Such serum factor(s) was produced in normal but not in nude mice and the suppressive activity was transferred in H-2- or immunoglobulin allotype-incompatible combinations. Immunochemical analyses of this serum suppressive factor have revealed that its m.w. was approximately 150,000, corresponding to the size of immunoglobulin (Ig)G, and that the activity was trapped by protein A or by an anti-immunoglobulin column. Although the absorption of the serum from anti-C3H/He-tolerant BALB/c mice with C3H/He target spleen cells did not abrogate the suppressive activity, the additional absorption with spleen cells from anti-C3H/He hyperimmune BALB/c mice almost completely eliminated the suppressive potential. Moreover, pretreatment of BALB/c anti-C3H/He DTH effector spleen cells with the above serum from tolerant mice induced the inhibition of anti-C3H/He DTH responses. Taken together, these results indicate that a single injection of allogeneic cells via the p.v. route results in the production of antibody capable of inhibiting the capacity of DTH effector cells specific for alloantigens used for the p.v. presensitization.

Animals↗