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Biomedical subjects

M Sawada

Publications and source records attributed to M Sawada.

At least 325 records · Page 18Linked to original sources
PubMed1992

Stable expression of monkey cytochrome P-450IA1 cDNA in Chinese hamster CHL cells and its application for detection of mutagenicity of aflatoxin B1.

A monkey cytochrome P-450IA1 cDNA (MKah1) was transfected into Chinese hamster CHL cells using a vector containing the SR alpha promoter, and sublines stably expressing P-450IA1 were established. The cells showed 25-fold higher sensitivity to the cytotoxic effect of aflatoxin B1 than the parental CHL cells. This hypersensitivity was almost completely suppressed by alpha-naphthoflavone, which is a known specific inhibitor of P-450IA. The cells expressing P-450IA1, but not CHL cells, showed a positive response to aflatoxin B1 in an assay for mutagenicity at the HGPRT locus.

Aflatoxin B1↗
PubMed1992

A PCR method for the quantitative assessment of mRNA for laminin A, B1, and B2 chains.

Laminin, a basement membrane glycoprotein, is involved in the development of normal kidney and its dysregulation contributes to glomerulosclerosis in renal disease. Studies designed to assess the regulation of this molecule at the level of transcription have been hindered by the relatively low abundance of the mRNA, making standard techniques such as Northern hybridization and RNase protection difficult and inaccurate. In this report, we have utilized the polymerase chain reaction (PCR) to quantitate differences in laminin mRNA expression during normal development of the mouse kidney. We have constructed a synthetic template to be used as an internal standard for mRNA quantitative of laminin chains A, B1 and B2, and beta-actin. This DNA template can be used to generate complementary RNA which can be reverse transcribed and amplified simultaneously with 0.5 microgram of total cellular mRNA allowing for accurate and absolute quantitation of laminin mRNA by PCR.

Animals↗
PubMed1992

Stimulation of phospholipase A2 by xanthine oxidase in the rat corpus luteum.

The ability of the superoxide radical (SOR) generated by xanthine oxidase to activate phospholipase A2 (PLA2) was examined in microsomes prepared from luteinized rat ovaries. Treatment of microsomes with xanthine oxidase resulted in a rapid burst in SOR formation followed by an increase in PLA2 activity. Stimulation of PLA2 activity was dose related and similar in microsomes prepared from control or prostaglandin F2 alpha (PGF2 alpha)-treated rats. Activation was inhibited by the antioxidants, vitamin E and nordihydroguaiaretic acid, and by superoxide dismutase and catalase, which metabolize SOR and H2O2 to remove reactive oxygen species from the cell. The stimulation of PLA2 activity by xanthine oxidase was dependent upon the addition of calcium ions, and it was highest in samples in which cytosol was added to membranes. These results indicate that the SOR and/or H2O2 may mediate PLA2 activation, which may be involved in the luteolytic process.

Acetophenones↗
PubMed1992

Establishment of mouse oligodendrocyte/type-2 astrocyte lineage cell line by transfection with origin-defective simian virus 40 DNA.

A permanent glial cell line has been established from the neonatal mouse primary mixed glial cell cultures by transfection with replication origin-defective simian virus 40 DNA. This cell line, designated OS3, has morphological similarity to type-2 astrocyte and expresses an astrocyte-specific marker, glial fibrillary acidic protein (GFAP), when cultured in the presence of 10% calf serum (CS). OS3 cells do not express the O4 antigen, galactocerebroside (GalC) and A2B5 under this culture condition. When cultured in a medium containing 2% CS or a chemically defined medium, these cells undergo morphological transformation. Some of these cells express O4 antigen and/or GalC, and the percentage of GFAP positive cells decreases under these conditions. Thus depending on the culture conditions, the OS3 cells display either type-2 astrocyte properties or immature oligodendrocyte characteristics. Furthermore, the OS3 cells show similar responses to the various growth factors as do oligodendrocyte/type-2 astrocyte (O-2A) progenitors. Therefore, the OS3 cell line is an unique mouse bipotential permanent O-2A lineage cell line which may be useful to analyze the developmental properties of these glial cells.

Animals↗
PubMed1992

An inhibitor of sweet taste response modulates glucose-induced phosphoinositide breakdown in rat pancreatic islets.

We studied the effect of a specific-competitive inhibitor of the sucrose taste response, p-nitrophenyl-D-glucopyranoside (PNP-Glu) on insulin release and phosphoinositide metabolism in rat pancreatic islets. The alpha-anomer, but not the beta-anomer, of PNP-Glu at a concentration of 5 mM inhibited insulin release induced by 10 mM glucose. Islets were labeled by exposure for 2 h to 10 uCi of myo-[2-3H] inositol solution supplemented with 2.8 mM glucose. Forty islets were then incubated in the presence of 10 mM LiCl, 1 mM inositol and 10 mM glucose with or without the anomers of PNP-Glu. [3H] radioactivity in the incubation medium remained significantly greater in the presence of the alpha-anomer of PNP-Glu than in the presence of glucose alone after 5- and 20-min incubation. The inositol monophosphate levels in the islets incubated with glucose alone were increased more than in the islets with alpha-anomer. The beta-anomer of PNP-Glu did not change either glucose-induced insulin release or phosphoinositide breakdown. A patch-clamp study revealed that neither anomer affected the glucose-dependent ATP-sensitive K(+)-channels. These results indicate that the anomeric preference for glucose in insulin release in the pancreatic islets is closely associated with phosphoinositide breakdown.

Animals↗
PubMed1992

[Immunohistochemical study of proliferating cell nuclear antigen (PCNA) in gynecological tumors and their related lesions].

We have investigated whether monoclonal antibody (PC10) of proliferating cell nuclear antigen (PCNA) could be useful as a marker of proliferating cells within formalin-fixed, paraffin-embedded tissue sections of 140 gynecological tumors and their related lesions. There was a positive correlation (r = 0.76) between the labelling index for PCNA and that for Ki67. Immunohistochemical staining for PC10 was confined to the nucleus and showed a diffuse or granular pattern or a mixture of both. The distribution of PC10 staining in non-neoplastic tissues was localized to proliferating cell compartments. In malignant tissues, the localization of the distribution of PCNA-positive cells came to be lost and the proportion of positive cells varied from case to case as well as from field to field within the same tissue section. The cases in which more than 31% of cells were positive for PCNA were as follows: Cervical squamous dysplasia 2/3, squamous carcinoma in situ 2/5, microinvasive squamous carcinoma 2/2, invasive squamous carcinoma 9/13, adenocarcinoma in situ 4/4, microinvasive adenocarcinoma 3/3, invasive adenocarcinoma 6/7, endometrial adenocarcinoma 6/25, ovarian epithelial malignant tumors 11/17, sex cord stromal tumors 2/14, and germ cell tumors 3/22. It is concluded that immunohistochemical staining for PC10 may be useful as a marker for proliferating activity of the cells both in normal and tumor tissues rather than for malignancy.

Antigens, Neoplasm↗
PubMed1992

[A case of secondary myelodysplastic syndrome following chemotherapy for lung cancer].

A case of secondary myelodysplastic syndrome (MDS) following chemotherapy for lung cancer is reported. A 78-year-old man, with a smoking history of 20 cigarettes/day for 55 years, was incidentally, diagnosed as having stage IV squamous cell carcinoma of the lung in 1987 during admission for transurethral resection of bladder cancer. He received combination chemotherapy of mitomycin C, vincristin, and cisplatin for his lung cancer between July and September 1988. His clinical course remained almost stable until October 1989, when his blood count showed severe anemia and thrombocytopenia. He was diagnosed as having secondary MDS induced by cytotoxic agents used for the treatment of lung cancer, based on the dysplastic findings of precursor cells in the bone marrow and the chromosome abnormality of 51XY, +8, +9, +21, 3p-, 5q-, +2mar. He died of infection with the progression of MDS in March 1990.

Aged↗
PubMed1992

[A case report of a massive pulmonary tumor embolism occurring during surgery for renal cell carcinoma].

A 59-year-old man, who suffered a massive pulmonary tumor embolism during surgery for renal cell carcinoma with vena caval invasion, was treated by emergency pulmonary embolectomy using cardiopulmonary bypass. Renal cell carcinoma occasionally extends into the inferior vena cava (IVC) as a tumor thrombus. In such patients, removal of the tumor thrombus from the IVC has to be performed in addition to radical nephrectomy. However, the massive pulmonary tumor embolism is a major potential hazard during radical surgical resection. To prevent intraoperative pulmonary embolisms from occurring, scheduled use of cardiopulmonary bypass with the cooperation of cardiovascular surgeons is recommended.

Carcinoma, Renal Cell↗
PubMed1992

[11-Dehydro-TXB2 and 2,3-dinor-TXB2 as new parameters of TXA2 generation].

Artifactual formation of thromboxane (TX) B2 during blood collection falsifies real value of TXB2 in plasma. A part (29.3%) of TXB2 is metabolized to 11-dehydro (DH)-TXB2 in several organs. 11-DH-TXB2 was not generated during blood collection or during serum formation. The peak amount of 11-DH-TXB2 after intravenous injection of TXB2 to rabbits was lower than that of TXB2, but the level of 11-DH-TXB2 was kept 2-3 times higher than that of TXB2 even after more than 5 min. A half life of 11-DH-TXB2 is 45-60 min in the human. Large species differences were found. In human urine, 11-DH-TXB2 was excreted 1.5-5.8 times more than 2,3-dinor-TXB2. Patients with ARDS and DIC, who received platelet transfusion, excreted increased amounts of 2,3-dinor-TXB2 and 11-DH-TXB2 in urine. 11-DH-TXB2 may be a useful parameter of TXA2 formation in pathological states.

Animals↗
PubMed1991

Tumor necrosis factor reduces the ACh-induced outward current in identified Aplysia neurons.

Effects of extracellularly applied recombinant human tumor necrosis factor (rhTNF) on the acetylcholine (ACh)-induced K+ current recorded from identified neurons (R9 and R10) of Aplysia kurodai were investigated with conventional voltage-clamp and pressure ejection techniques. Bath-applied rhTNF (200-500 U/ml) reduced the ACh-induced current in the neurons without affecting the holding current and resting membrane conductance. The suppressing effect of rhTNF on the current was completely reversible. Inhibition by rhTNF was non-competitive. Heat-inactivated rhTNF was without effect. Our results suggest that the immunomodulator TNF can act on the ACh receptor in the nervous system.

Acetylcholine↗
PubMed1991

Adhesion of immature thymocytes to thymic stromal cells through fibronectin molecules and its significance for the induction of thymocyte differentiation.

Only 10-15% of unseparated thymocytes adhered to culture plates precoated with fibronectin (FN), but 60-70% of the CD4-8- (double-negative) thymocyte population did. This population bound to FN but not to collagen, laminin, or vitronectin. Its binding to FN was inhibited by anti-FN antibody or a mixture of synthetic peptides corresponding to two different sites of FN, termed the V10 sequence and the RGDS (Arg-Gly-Asp-Ser) sequence, which interact, respectively, with the VLA-4 and VLA-5 FN receptors expressed on T-lineage cells. CD4-8- thymocytes also adhered to a monolayer of a thymic stromal cell clone, MRL104.8a, that induces growth-maintenance and differentiation of such thymocytes. The involvement of FN-FN receptor interaction in this adhesion was demonstrated by the following lines of evidence: (i) the MRL104.8a cells expressed FN molecules on their surface and (ii) the adhesion of CD4-8- thymocytes to MRL104.8a monolayers was almost completely inhibited by simultaneous addition of anti-FN antibody and a mixture of peptides (V10 plus RGDS) capable of binding to anti-FN receptors (VLA-4 and -5). Most important, blocking the adhesion of CD4-8- thymocytes to the thymic stromal cell monolayer resulted in potent inhibition of the differentiation of these thymocytes, which was otherwise induced toward the expression of CD4 and/or CD8 molecules. These results indicate that immature CD4-8- thymocytes adhere to thymic stromal cells preferentially through FN-FN receptor interaction and that such adhesion has a critical role in inducing and/or supporting the differentiation of these thymocytes.

Animals↗
PubMed1991

[Interferon-alpha and gamma product in peripheral blood of patients with thyroid diseases].

To investigate the immunological process in various thyroid disease, we measured interferon-alpha, -gamma (IFN-alpha,-gamma, natural killer (NK) activity, and lymphocyte subsets in the peripheral blood of 27 patients with Basedow's disease (BD) (M:F = 9:18), 8 with Hashimoto's thyroiditis (HT) (2:6), 5 with idiopathic hypothyroidism (1HT) (1:4), and normal controls (C). IFN-alpha, -gamma levels were measured by bioassay with Dye-uptake method, and NK activity was measured by the LDH method. The mean +/- SD levels of IFN-gamma in BD, HT, IHT, and C (N = 217) were 173.9 +/- 88.0, 288.0 +/- 134.9, 120.4 +/- 38.0 and 173.9 +/- 88.01U/ml, respectively. The IFN-gamma level was higher in HT (p less than 0.05) than in controls, and lower in BD (p less than 0.02). Moreover, this IFN-gamma level did not correspond with the titers of thyroid hormones; TSH, anti-microsome antibody, and anti-TSH-receptor antibody in peripheral blood. However, IFN-alpha levels and NK activity in the patients of every group were similar to those in controls. The ratios of lymphocyte subsets of peripheral blood were measured by cytofluorometry with monoclonal antibodies. The mean +/- SD levels of T cells in BD, HT, IHT and C were 75.9 +/- 7.0, 83.4 +/- 7.2, 85.2 +/- 5.7, 82.3 +/- 5.8%, and those of B cells were 17.5 +/- 6.1, 10.9 +/- 4.2, 8.0 +/- 5.8, 10.9 +/- 5.0%, respectively. The ratio of T cells was higher in IHT (p less than 0.05) and lower in BD (p less than 0.01) than in controls.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1991

Establishment and characterization of a new cell line TC-YIK originating from argyrophil small cell carcinoma of the uterine cervix integrating HPV16 DNA.

A new cell line, designated TC-YIK, was established from YIK-1 tumor cells, derived from argyrophil small cell carcinoma (ASCC) of the uterine cervix, and serially heterotransplanted into nude mice, integrating human papillomavirus type 16 (HPV16) DNA. The population doubling time of TC-YIK was approximately 21.6 hours at the 119th subculture. Subcutaneous injection of 1 x 10(8) TC-YIK cells into nude mice yielded a solid tumor. The cytologic appearance of TC-YIK was similar to that of YIK-1. The TC-YIK cells contained argyrophil granules and neurosecretory granules in the cytoplasm and showed positive immunohistochemical staining for neuron-specific enolase, serotonin, and chromogranin. Thus, TC-YIK retained the histochemical characteristics of ASCC. The TC-YIK cells contained HPV16 DNA in a multiple-copy integrated form and actively transcribed the integrated HPV16 genome. Amplification of the c-myc oncogene was observed in the TC-YIK cells. These data suggest that TC-YIK is a useful in vitro experimental model of ASCC and that HPV16 and c-myc may play some role in the genesis of this malignant tumor and/or maintenance of the transformed TC-YIK phenotype.

Animals↗
PubMed1991

Ionic mechanism of the outward current induced by extracellular ejection of interleukin-1 onto identified neurons of Aplysia.

The ionic mechanism of the effect of extracellularly ejected recombinant human interleukin-1-beta (rhIL-1) on the membrane of identified neurons R9 and R10 of Aplysia was investigated with voltage-clamp, micropressure-ejection, and ion substitution techniques. Micropressure-ejected rhIL-1 caused a marked hyperpolarization in the unclamped neuron. Clamping the same neuron at its resting potential level (-60 mV) and reejecting rhIL-1 with the same dose produced a slow outward current (I0(IL-1), 20-30 s in duration, 3-5 nA in amplitude) associated with a decrease in input membrane conductance. I0(IL-1) was decreased by depolarization and increased by hyperpolarization. The extrapolated reversal potential of I0(IL-1) was approximately +15 mV. I0(IL-1) was sensitive to changes in the external Na+ concentration but not to changes in K+, Ca2+ and Cl- concentrations, and was resistant to tetraethylammonium (5 mM) and 4-aminopyridine (5 mM). Neither perfusion of the neuron with 50 microM tetrodotoxin nor perfusion with 10 mM Co2+ seawater caused any changes in I0(IL-1). I0(IL-1) was partially reduced by 50 microM ouabain. These results suggest that extracellular IL-1 can induce a slow outward current associated with a decrease in Na+ conductance and the immunomodulator IL-1 can act directly on the nervous system as well as on the immune system.

Animals↗
PubMed1991

Analysis of a decreased Na+ conductance by tumor necrosis factor in identified neurons of Aplysia kurodai.

The ionic mechanisms of the effect of extracellularly ejected recombinant human tumor necrosis factor-alpha (rhTNF-alpha) on the membrane of identified neurons R9 and R10 of Aplysia kurodai was investigated with conventional voltage-clamp, micropressure ejection, and ion substitution techniques. Micropressure-ejected rhTNF caused a marked hyperpolarization in the unclamped neuron. Clamping the same neuron at it resting potential level (-60 mV) and reejecting rhTNF-alpha with the same dose produced a slow outward current [Io (TNF)] associated with a decrease in input membrane conductance. Io (TNF) was decreased by depolarization and increased by hyperpolarization. The extrapolated reversal potential of Io (TNF) was approximately +10 mV. Ion substitution and pharmacological experiments suggest that Io (TNF) in identified neurons R9 and R10 of A. kurodai is due to a decreased Na+ conductance but not due to an activation of the Na(+)-K+ pump. Our results demonstrate that the immunomodulator TNF can act directly on the nervous system as well as on the immune system.

Animals↗
PubMed1991

The activation of Ca(2+)-dependent K+ conductance by adrenaline in mouse peritoneal macrophages.

Responses to adrenaline in mouse peritoneal macrophages were investigated with perforated and cell-attached patch-clamp recording, and with a combination of the perforated-patch recording and fura-2 fluorescence measurements. Extracellularly applied adrenaline induced a transient outward current (4-10 s in duration, 100-500 pA in amplitude) at -40 mV associated with a marked increase in conductance. The adrenaline-induced current [Io (Adr)] reversed polarity near -80 mV. The reversal potential depended distinctly on the external K+ concentration but not on external Cl- concentration. Removal of external Ca2+ did not affect Io(Adr) within 2-4 min but subsequent responses to adrenaline were progressively depressed. In contrast, treatment with an intracellular Ca2+ chelator, the acetoxymethyl ester of 1,2-bis-(2-aminophenoxy)ethane-N,N,N',N'-tetraacetic acid completely abolished Io(Adr). Furthermore, Io(Adr) was blocked by bath-applied quinidine and charybdotoxin, but not by tetraethylammonium or apamin. Extracellular application of an alpha 1-adrenoceptor agonist phenylephrine and of noradrenaline mimicked Io(Adr). On the other hand, Io(Adr) was antagonized by a non-selective alpha-adrenoceptor antagonist phentolamine (0.2 microM) and an alpha 1-adrenoceptor antagonist prazosin (0.2 microM), but was not affected by an alpha 2-adrenoceptor antagonist yohimbine (1 microM) or a beta-adrenoceptor antagonist propranolol (1 microM). Cell-attached single-channel recordings with the pipette solution containing 145 mM KCl revealed the activation of single-channel currents with a conductance of 40 pS during application of adrenaline outside the patch. Parallel measurements of membrane current and fura-2 fluorescence in the same cell demonstrated a correlation between the rise in [Ca2+]i and an increase in K+ conductance. Therefore, it is concluded that adrenaline activates a Ca(2+)-dependent K+ conductance by release of Ca2+ from internal stores through an activation of an alpha 1-adrenoceptor.

Animals↗
PubMed1991

Isolation of a menadione-resistant subclone from Chinese hamster lung (CHL) cells in culture.

Menadione-resistant subclones were selected from cultured Chinese hamster lung (CHL) cells which had been mutagenized with MNNG or ENU. The frequency of surviving colonies and the level of resistance were higher in mutagenized cells than in non-mutagenized cells. A subclone (designated MM1) was isolated from MNNG-treated cells and showed the highest level of resistance, 3 times higher than the parental CHL cells. The level of resistance was stable in non-selective medium over 3 months. The MM1 cells were also 2-3 times more resistant to other naphthoquinones. The activity of NADPH-cytochrome P-450 reductase, which is thought to play an important role in activation of menadione, was reduced in the MM1 cells to half that in the parental CHL cells. On the other hand, no differences between MM1 and CHL cells were found in the activity of superoxide dismutase and catalase which are assumed to defend against the cytotoxicity of menadione. Karyotype analyses indicated that one small chromosome was lost in the MM1 cells. The MM1 cells showed a 3-fold resistance to menadione in the chromosomal aberration test. The frequencies of chromosomal aberrations induced by adriamycin and mitomycin C which could be activated by NADPH-cytochrome P-450 reductase were almost the same in the MM1 and CHL cells, suggesting that the reductive activation of these compounds by this enzyme in microsomes may not be involved in the induction of chromosomal aberrations.

Animals↗