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N Shinomiya

Publications and source records attributed to N Shinomiya.

At least 55 records · Page 3Linked to original sources
PubMed1993

Therapeutic effect of granulocyte colony-stimulating factor and cephem antibiotics against experimental infections in neutropenic mice induced by cyclophosphamide.

Effects of recombinant human granulocyte colony-stimulating factor (rhG-CSF) against severe infections in cyclophosphamide (CY)-induced neutropenic mice were investigated by its single use or by its combination with cephem antibiotics. Treatment with rhG-CSF increased the number of peripheral blood leucocytes and strikingly shortened the duration of the neutropenic state. Most of the regained population in the peripheral blood leucocytes were neutrophils. Functions of neutrophils, such as phagocytic activity, superoxide release, and expression of Mac-1 molecules, were remarkably enhanced by administration of rhG-CSF. When rhG-CSF was administered to CY-induced neutropenic mice before infection, protective effects against various kinds of bacteria were remarkable. On the other hand, such remarkable effects were not observed when rhG-CSF was administered after infections. However, even in the case of post-infectious treatment, a combination therapy of rhG-CSF with cephem antibiotics, particularly with Cefodizime (CDZM), showed a significant improvement in the survival rate and a decrease in the number of viable bacteria in the liver. These results suggest that a combination therapy of rhG-CSF with cephem antibiotics, especially with CDZM, is an efficient regime against severe infections in patients with neutropenia induced by a heavy anti-tumour chemotherapy.

Animals↗
PubMed1992

Increased mitochondrial uptake of rhodamine 123 by CDDP treatment.

Rhodamine 123 (R 123) is a positively charged dye at physiological pH that accumulates specifically in the mitochondria of living cells without cytotoxic effect. In the present study, the uptake of R 123 by EL-4 lymphoma cells in culture with anticancer agents was measured by flow cytometry. Changes in R 123 uptake during the cultivation period were compared with cell distribution at different phases of the cell cycle. According to the increase in the proportion of S phase cells, mitochondrial synthesis increased, giving rise to a maximal fluorescence intensity of about 1.3-fold. Synchronous cultures showed the same relationship between increased mitochondrial uptake of R 123 and the S phase fraction as was observed in normal cultures. After treatment with 10(-3) M 5-fluorouracil (5-FU) for 1 h, EL-4 cells showed an increased binding of R 123 per cell followed by an accumulation of early S phase cells transiently. However, uptake of R 123 decreased 24 h later. On the contrary, after treatment with 10 micrograms/ml of cis-diamminedichloroplatinum (CDDP), a G2 + M block was observed from 12 h of reseeding and accumulation of the G2 + M cells continued. In this case, high uptake of R 123 continued during the observation period. From these results, mitochondrial synthesis seemed to increase according to the increment in proportion of S phase when the acceleration of the cell cycle turnover was augmented or the cycle was blocked in S phase by 5-FU. CDDP inhibited the cell division at G2 + M phase and caused increased R 123 fluorescence per cell. The stainability of R 123 may indicate the activity of cell division and may be a good way of evaluating the efficacy of antitumor drugs on the cells.

Cell Cycle↗
PubMed1992

Intracellular hydrogen peroxide production by peripheral phagocytes from diabetic patients. Dissociation between polymorphonuclear leucocytes and monocytes.

Although the standard assays for reactive oxygen species have been based on the measurement of those released into the extracellular environment, the microbicidal capacity to the engulfed microorganisms is mainly dependent on those released into the intracellular environment, such as phagosomes. We studied intracellular oxidative activities of individual phagocytes by dichlorofluorescein (DCFH) oxidation assay to investigate the relationship between the reactive oxygen species released intracellularly and the impaired microbicidal capacity in diabetic patients. Time courses of intracellular production of hydrogen peroxide by polymorphonuclear leucocytes (PMNL) and monocytes were observed at the resting condition and after the stimulation with phorbol myristate acetate (PMA; 160 nM) by flow cytometry. Thirty-four patients with non-insulin-dependent diabetes mellitus (NIDDM) and 23 age-matched healthy volunteers were subjected to the studies. PMNL from patients with NIDDM showed a significantly decreased capacity to produce hydrogen peroxide after the stimulation (P less than 0.05 at 15 min, P less than 0.01 at 30 and 45 min). By contrast, intracellular hydrogen peroxide production by monocytes at the resting condition and an early stimulatory phase (8 min after the stimulation) was significantly (P less than 0.01) enhanced in patients with NIDDM compared with that in controls. Both the changes of intracellular hydrogen peroxide production observed in PMNL and monocytes from patients with NIDDM were in association with an increased haemoglobin Alc level in erythrocytes, but did not relate to total cholesterol and triglyceride levels in the serum. The possible mechanisms of these dissociated changes in hydrogen peroxide producing capacity of phagocytes from patients with NIDDM are discussed.

Adult↗
PubMed1992

[Production of reactive oxygen species by rat alveolar macrophages. Dissociation between the intracellular and extracellular release of hydrogen peroxide].

In order to clarify the features of reactive oxygen species produced by rat alveolar macrophages (AMs), the concentrations of intracellular and extracellular hydrogen peroxide were measured under various experimental conditions. Intracellular hydrogen peroxide was measured by DCFH method using a flow cytometer, while the extracellularly released fraction was measured by scopoletin method using a spectrophotometer. The concentration of intracellular hydrogen peroxide after stimulation with opsonized zymosan (10 micrograms/ml) was significantly higher than that after stimulation with phorbol myristate acetate (PMA; 100 ng/ml). On the other hand, hydrogen peroxide released extracellularly after stimulation with PMA was significantly greater that that after stimulation with opsonized zymosan. These results indicate that the soluble membrane stimulant and the phagocytic particles have different mechanisms in activating the production of hydrogen peroxide in AMs. That is, hydrogen peroxide induced by PMA was mainly released extracellularly, while that induced by zymosan was mainly released into the intracellular environment. At rest, the concentration of intracellular hydrogen peroxide in rat AMs was high. Potassium cyanate, a known mitochondrial inhibitor, suppressed the intracellular hydrogen peroxide in AMs not only at rest but also after stimulations, indicating that most of the reactive oxygen species released into the intracellular environment in AM are produced by mitochondria. From these results, in order to gain a closer insight into the function of AMs, it is very important to distinguish the oxidative metabolites produced intracellularly which are related to bactericidal function from those of the extracellularly released fraction which give rise to lung damage.

Animals↗
PubMed1991

HLA-D gene studies in relation to immune responsiveness to a grass allergen Lol p III.

The grass pollen allergen Lol p III (Mr 11,000) is a well-characterized antigen that has been found useful in immunogenetic studies of human immune responsiveness. Since immune responsiveness to this allergen is associated with HLA-DR3, we investigated whether there was any sequence in the HLA-D region that would render a person "susceptible" [antibody (Ab)-positive] to the allergen. By sequence-specific oligonucleotide (SSO) slot-blot and sequence analyses of polymerase chain reaction (PCR)-amplified genomic DNA from Lol p III responder and nonresponder subjects, Ab responsiveness was found to be strongly associated with the sequence Glu-Tyr-Ser-Thr-Ser (EYSTS), present in the first polymorphic regions of DR beta I polypeptide chains of DR3, DR11 (split of DR5), and DRw6. Of the 41 grass-allergic subjects investigated, 19 had the EYSTS sequence, of whom 18 (95%) were Lol p III immunoglobulin G (IgG) Ab responders; among the 22 EYSTS- subjects, ten were Lol p III responders (P = 0.001, relative risk = 21.6). No such association was found with any polymorphic sequences in other DR beta chains, or in DQ alpha I and DQ beta I chains. These findings suggest that the EYSTS sequence is important in the presentation of an epitope of Lol p III; other sequence(s) may be involved in the presentation of other epitope(s). To our knowledge, this is the first demonstration of a strong association between a specific HLA sequence and immune responsiveness to a well-defined antigen. The paper shows that presence of the EYSTS sequence classifies subjects as Lol p III responders in 18/19 cases.

Allergens↗
PubMed1991

Effects of macrophage colony-stimulating factor on reduction of viable bacteria and survival of mice during Listeria monocytogenes infection: characteristics of monocyte subpopulations.

Mice could well tolerate infection with a lethal dose of Listeria monocytogenes after intraperitoneal preinjections with 250 micrograms of macrophage colony-stimulating factor (M-CSF) per kg of body weight for 5 days. The characteristic changes in the surface markers (Mac-1, LFA-1, and F4/80) of peripheral monocytes were also investigated in order to analyze the mechanism of protection by M-CSF. This investigation shows the excellent effect of intraperitoneal preinjections of M-CSF on the reduction of viable Listeria organisms and the improvement of survival after an intravenous Listeria infection.

Animals↗
PubMed1991

Enhanced resistance against Listeria monocytogenes achieved by pretreatment with granulocyte colony-stimulating factor.

Phagocytosis, H2O2 production, Mac-1 expression, and in vivo elimination of Listeria monocytogenes were enhanced in granulocyte colony-stimulating factor (G-CSF)-treated mice. Transfer of polymorphonuclear leukocytes prolonged survival of mice infected with a lethal dose of L. monocytogenes. G-CSF augments the functions of polymorphonuclear leukocytes and thus plays a role in protection.

Animals↗
PubMed1991

Flowcytometric measurement of the cell cycle of experimental tumors: some devices for accurate measurement of proliferative activity.

In this paper we present the cell cycle analysis using the latest flowcytometer, FACScan and dedicated softwares. Although the 5-bromodeoxyuridine (BrdU) and anti-BrdU method developed by Gratzner has become popular for analysis of cell kinetics, contaminating interstitial cells and cell doublets are still factors responsible for some mistakes in measuring the cell cycle accurately. These hampering cells were almost gated out on the catogram consisting of the forward scatter and DNA content (propidium iodide, PI). After that, the bivariate distribution of BrdU and PI according to the tumor growth showed reliable patterns. In LLC tumor the population, of which DNA content was compatible with S phase cells, was detected in the cytogram. Thus, the two-dimensional analysis of the cell cycle can demonstrate each population clearly. The results obtained from the BrdU method are more beneficial than autoradiography or microspectrophotometry, and reproducible data of the cell cycle parameters can be acquired with some devices described here.

Adenocarcinoma↗
PubMed1991

Effects of combined therapies with protein-bound polysaccharide (PSK, Krestin) and fluorinated pyrimidine derivatives on experimental liver metastases and on the immunologic capacities of the hosts.

An experimental model is introduced for the study of liver metastases using intrasplenically injected EL-4 and Lewis lung tumor cells. Fluorinated pyrimidine derivatives, 1-(2-tetrahydrofuryl)-5-fluorouracil and 5-fluorouracil, showed inhibitory effects on the frequencies of liver metastases. Immunosuppressive effects of these drugs were compared at the doses capable of showing 50% inhibition of the development of metastatic nodules. These derivatives strongly suppressed the phagocytic activity and the number of Kupffer cells of the liver and then the humoral response against sheep red blood cells, the delayed hypersensitivity against picryl chloride. On the contrary, combined administration of protein-bound polysaccharide (PSK) and 1-(2-tetrahydrofuryl)-5-fluorouracil showed no inhibitory effect on these activities.

Adjuvants, Immunologic↗
PubMed1991

[Augmentation of activities of peripheral granulocytes and of resistance against bacterial infection after administration of G-CSF into mice].

We evaluated the metabolic capability of murine peripheral granulocytes after administration of recombinant human granulocyte colony-stimulating factor (rhG-CSF) by quantitative flow cytometric assay for H2O2-dependent oxidative product formation. Intraperitoneal administration of a daily dose of 10 micrograms of rhG-CSF for 5 days induced doubling of the leukocyte population. Differential counting of peripheral leukocytes and scattergram by flow cytometry showed an increased mature granulocyte population. After stimulation with phorbol myristate acetate, the granulocytes of the rhG-CSF-administered mice demonstrated some hyperresponsive population and an increased H2O2 production. The hyperresponsive population showed H2O2 production 4-6 times higher than did normal cells. Granulocytes from the G-CSF-treated mice revealed an augmented phagocytic activity and an increased expression of Mac-1 molecules. Moreover, mice treated with G-CSF showed an enhanced resistance against intravenous infection with a lethal dose of E. coli. Granulocytes showing such markedly increased oxidative metabolism may be a significant component of the host defence to various infective organisms.

Animals↗
PubMed1991

Thymic depletion in pregnancy: kinetics of thymocytes and immunologic capacities of the hosts.

In the present study we observed a significant depression of thymocytes during pregnancy and investigated the influences of this thymic change on the immunologic capacities of peripheral lymphocytes. Thymocytes in pregnant mice began to decrease in number from Day 10 and reached about 0.1-fold of the nonpregnant level at Day 19, just before parturition. At late stage of pregnancy, thymocyte subpopulation expressing CD4+CD8+ and Thy1.2+PNA+ was selectively depressed. On the contrary, peripheral lymphocytes including splenocytes, peripheral blood lymphocytes and peripheral lymph node cells showed no depression. As to the immunologic capacities of the pregnant hosts, delayed footpad reaction and phagocytic activity of fixed liver macrophages in vivo were remarkably suppressed, but MLR reactivity and antibody response to SRBC or haptens were well preserved. Transfer of pregnant sera or administration with steroid hormones especially E3 into nonpregnant mice induced similar changes in the thymus and peripheral lymphocytes in number and subsets but this could not mimic the immunologic reactivities of the pregnant mice. These results suggest that sex steroid hormones such as E3 play an important role in the changes in cell populations of each lymphoid organ and the immune reactivities of the hosts during pregnancy. However, other factors also contribute to the immunologic capacities of the maternal hosts.

Animals↗
PubMed1991

Adsorption and preparation of human viruses using hydroxyapatite column.

The adsorption and chromatographic properties of hydroxyapatite sorbents for application to different viruses have been investigated. The strong adsorption of viruses was observed on macroporous hydroxyapatite with hydrophilic properties of the sorbent surface. The viruses were purified on this sorbent without loss of biological activity. The column can be used for virus vaccine production.

Adsorption↗
PubMed1991

[Establishment and characterization of a human undifferentiated carcinoma cell line (HMG)].

The undifferentiated carcinoma cell line (HMG) was established from a nude mouse tumor which had been produced by transplantation of a intraperitoneal tumor of 27-year-old woman. The HMG cell line has the following biological properties. 1. The HMG cells are round to oval in shape and grow as floating cell aggregates like a rouleau or a cluster of grapes. 2. 100 passages have been carried out over a year, and the population doubling time is about 17 hrs. 3. In the original tumor, keratin and vimentin were expressed simultaneously, in HMG cells, however, only localization of vimentin was confirmed. 4. By chromosomal analysis, over 90% of the cells revealed 46, XX, with no karyological abnormalities, at passage 82. 5. When heterotransplanted into the subcutis of a nude mouse, HMG cells produced a undifferentiated carcinoma resembling the original tumor.

Adult↗
PubMed1991

[Analysis of tumor cell growth and effects of antitumor drugs on cell cycle regulation by flow cytometry].

Relationship between tumor cell growth and cell cycle and influence of antitumor drugs on the cell cycle regulation were investigated by the use of murine EL-4 tumor. Cell growth curve of EL-4 cells was well related to the cell cycle pattern analyzed by two color flow cytometry both in vivo and in vitro. The ratio of S phase rapidly increased in early log phase and decreased from late log to plateau phases. The ratio of G0/G1 phase showed a reciprocal change. Minimum effective doses of 5-fluorouracil (5-FU) and CDDP on EL-4 growth in vitro were 1 x 10(-6) M and 0.5 micrograms/ml, respectively. At such doses, 5-FU showed an accumulation in early S phase and CDDP showed a partial synchronization in S phase and subsequent accumulation in late S and G2 + M phase. In case of in vivo administration with 5-FU (20 mg/kg/day) ratio of S phase was higher than in untreated control mice at day 2-3 but decreased rapidly thereafter. Mice administered with CDDP (5 mg/kg/day) showed a decrease in S phase from day 2 and completely rejected the tumors by day 5. From these results, each phase of cell cycle was influenced by the cell growth characteristics and the cell cycle pattern was dynamically changed according to the mode of action of antitumor drugs. Moreover, in vivo effects of these drugs can be evaluated adequately by the analysis of the cell cycle.

Animals↗
PubMed1991

Simplified method for isolation of white cells from whole blood suitable for direct polymerase chain reaction.

A simple method for isolating mononuclear cells from whole blood is described. The procedure utilizes phytohemagglutinin to agglutinate the erythrocytes, separating white cells from whole blood in a very brief handling time. The isolated cells are readily subjected to DNA isolation simply by boiling, and the released DNA can be directly employed for the polymerase chain reaction analysis. The efficiency of this method is similar to other conventional methods, but less costly and less time-consuming. This method is particularly useful in analyzing DNA samples from the peripheral blood cells when the simplicity and low cost of the assay are preferable.

Base Sequence↗
PubMed1991

Depression of early protection against influenza virus infection by cyclophosphamide and its restoration by Y-19995 [2,4'-bis(1-methyl-2-dimethyl-aminoethoxyl)-3-benzoylpyridine dimaleate].

The relationship between depression of early protection against influenza virus infection and the decrease in the number of peripheral polymorphonuclear leukocytes in cyclophosphamide-treated mice was investigated by means of a novel synthetic compound, Y-19995 [2,4'-bis(1-methyl-2-dimethyl-aminoethoxyl)-3-benzoylpyridine dimaleate], which had been shown to exert a potent restorative effect on leukocytopenia in immunocompromised hosts. Following intranasal inoculation with influenza virus (1.5 x 10(3) plaque-forming units) into untreated mice, the pulmonary virus titer progressively increased during 3 days and decreased gradually from day 7 after infection. The treatment with cyclophosphamide 2 days before infection markedly enhanced the pulmonary virus multiplication from the early phase of infection, and the higher virus titer was maintained thereafter. When mice were given Y-19995 after cyclophosphamide treatment, virus titers from the early to late phases of infection were lower than those in untreated mice. The number of peripheral polymorphonuclear leukocytes in cyclophosphamide-treated mice rapidly decreased and returned to normal levels only 9 days after the treatment, while such leukocytopenia was prevented to some extent and the leukocyte count was restored completely up to 7 days by postcyclophosphamide treatment with Y-19995. Furthermore, the treatment with Y-19995 augmented the inactivation of virus by the polymorphonuclear leukocytes. However, the virus inactivation by alveolar macrophages was modified only slightly by Y-19995 treatment. In addition, Y-19995 treatment could potentiate antibody-dependent cell-mediated cytotoxicity of polymorphonuclear leukocytes against the virus-infected target cells, and the production of serum neutralizing antibody to influenza virus in untreated and cyclophosphamide-treated mice. Y-19995 revealed neither antiviral nor interferon-inducing activities.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals↗
PubMed1990

[Flow cytometric analysis of oxidative burst of phagocytes with small amount of peripheral blood].

We have developed a simple method for assessing the oxidative metabolic burst of peripheral blood leukocytes with a minute amount of whole peripheral blood by flow cytometry according to the method of Bass et al. with some modification. By this method, we can measure the H2O2 production by both granulocytes and monocytes in the same blood sample. The oxidative product formation by peripheral blood neutrophils can be monitored sequentially in the same mouse infected with E. coli. The mice infected intravenously with 0.1 LD50 of the bacteria showed increased basal activities from an early stage of infection; those infected intraperitoneally with the same dose of the bacteria showed a delayed enhancement. In case of infection with 0.01 LD50, the enhanced basal activities lasted for only a short period of time. The H2O2 production was correlated well with the clearance of the infected bacteria. These results demonstrated that the oxidative-product formation by peripheral blood neutrophils is affected by both the route and the dose of infection.

Animals↗