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Biomedical subjects

E Suzuki

Publications and source records attributed to E Suzuki.

At least 19 recordsLinked to original sources

Localization and health effects of lanthanum chloride instilled intratracheally into rats.

Lanthanum (La) is one of the rare earths used in diverse high technology fields for which sufficient data for assessing its health effects have been lacking. The biological effects and metabolic behaviors of La were studied by instilling lanthanum chloride intratracheally into male Wistar rats. The distribution of La among tissues revealed that the metal remains mostly in the lung with a biological half-time of 244 days. The subcellular localization by transmission electron microscopy with an X-ray microanalyzer indicated that La localizes in macrophages as high electron-dense granular inclusions in lysosomes and on the cell surface and basement membranes of type I pneumocytes among lung cells. The pulmonary health effects were examined by biological indices of the bronchoalveolar lavage fluid (BALF) and lung tissue. The acute toxicity estimated by lactate dehydrogenase activity in BALF was comparable to those of yttrium and copper that had been determined under the same protocol. Microscopic examination of the lung indicated a characteristic increase in the number of eosinophils.

Animals

A gene homologous to chloroplast carbonic anhydrase (icfA) is essential to photosynthetic carbon dioxide fixation by Synechococcus PCC7942.

To understand the CO2-concentrating mechanism in cyanobacteria, a genomic DNA fragment that complements a temperature-sensitive high-CO2 (5%)-requiring mutant of Synechococcus PCC7942 has been isolated. An open reading frame (ORF272) encoding a polypeptide of 272 amino acids (Mr, 30,184) was found within the genomic region located 20 kilobases downstream from the genes for ribulose-1,5-bisphosphate carboxylase/oxygenase (rbcLS). Insertion of a kanamycin-resistance gene cartridge within the ORF272 in wild-type cells led to a high-CO2-requiring phenotype. Strains carrying a gene disabled by insertional mutagenesis accumulated inorganic carbon in the cells, but they could not fix it efficiently, even though ribulose-1,5-bisphosphate carboxylase activity was comparable to that of the wild-type strain. Therefore, the ORF272 was designated as a gene icfA, which is essential to inorganic carbon fixation. Furthermore, the predicted icfA gene product shared significant sequence similarities with plant chloroplast carbonic anhydrases (CAs) from pea (22%) and spinach (22%) and also with the Escherichia coli cynT gene product (31%), which was recently identified to be E. coli CA. These results indicate that the putative CA encoded by icfA is essential to photosynthetic carbon dioxide fixation in cyanobacteria and that plant chloroplast CAs may have evolved from a common ancestor of the prokaryotic CAs, which are distinct from mammalian CAs and Chlamydomonas periplasmic CAs.

Amino Acid Sequence

Dynamical structure of the antibody combining site as studied by 1H-15N shift correlation NMR spectroscopy.

The Fv fragment, which is a smallest antigen-binding unit of immunoglobulin, has been used for a 1H-15N shift correlation NMR study of the dynamical structure of the antibody combining site. Fv has been prepared by clostripain digestion of a mouse anti-dansyl IgG2a monoclonal antibody that lacks the entire CH1 domain. We have previously reported that of the six hypervariable regions, three each from the heavy chain (H1, H2, and H3) and the light chain (L1, L2, and L3), H3 is primarily responsible for the antigen binding in the anti-dansyl Fv fragment. The backbone amide nitrogens of all non-proline amino acid residues in H3 have been multiply labeled with 15N. [15N]T2 relaxation times and hydrogen-deuterium exchange rates of the amide groups of the main chain were measured in the absence and presence of epsilon-dansyl-L-lysine (DNS-Lys). It has been shown that (1) in the absence of DNS-Lys H3 displays a significant degree of internal motion and (2) antigen binding induces a significant change in the dynamical structure of H3.

Amino Acid Sequence

Molecular cloning and nucleotide sequence determination of the regulator region of mecA gene in methicillin-resistant Staphylococcus aureus (MRSA).

Molecular cloning and nucleotide sequence analysis were performed for the identification of the regulator genes of methicillin resistance in the genome of a MRSA strain N315. Two open reading frames (orfs) were identified in the 5'-flanking region of the mecA gene. Predicted amino acid sequences of these orfs showed extensive homology to the co-inducer and the repressor protein of the penicillinase (PCase) production in Staphylococcus aureus as well as in Bacillus licheniformis. These orfs are considered to encode putative co-inducer and repressor proteins specific for the regulation of methicillin resistance in MRSA.

Amino Acid Sequence

Plasma homovanillic acid, plasma anti-D1 and -D2 dopamine-receptor activity, and negative symptoms in chronically mediated schizophrenia.

We have investigated the relationship between the concentration of homovanillic acid in human plasma (pHVA) and plasma anti-D1 and anti-D2 dopamine receptor activity in chronic schizophrenic patients whose neuroleptic dosage was changed. The change in pHVA level correlated with that in anti-D1, not anti-D2 activity, thus suggesting that the neuroleptic-induced changes in pHVA concentration may be associated with the blocking of D1- as well as D2- receptors. The change of scores on the Scale for the Assessment of Negative Symptoms did not significantly correlate with changes in anti-D1 or anti-D2 activity, but did so correlated with the change in pHVA level.

Antipsychotic Agents

Characterization of atrial natriuretic peptide in urine from rats treated with a neutral endopeptidase inhibitor.

To explore the mechanisms for the natriuretic effects of a neutral endopeptidase inhibitor, candoxatril, the concentration of atrial natriuretic peptide (ANP) and its molecular forms in the urine of Dahl salt-sensitive (S) rats were examined. Candoxatril-induced natriuresis (+120%, p less than 0.05) was associated with a marked increase in the urinary ANP excretion (+1200%, p less than 0.05). Analysis by Sephadex G-50 gel filtration revealed that molecular weight of the major fraction of immunoreactive (ir-) ANP in the plasma of candoxatril-treated Dahl S rats was 3K, whereas that in the urine was 2.5 K. Further analysis by reverse phase high performance liquid chromatography showed that ir-ANP in the plasma of Dahl S rats was alpha-rANP (1-28), while that in the urine from rats treated with candoxatril was alpha-rANP (1-25). These results indicate that candoxatril inhibits the complete degradation of ANP in the kidney, thereby increasing the amount of biologically active ANP reaching the distal nephron and contributing to natriuresis.

Analysis of Variance

Interleukin-6 is antiproliferative to a mouse hybridoma cell line and promotive for its antibody productivity.

Monoclonal antibody production by hybridoma cells at moderately slowed growth states would be favorable for commercial scale production since cells can devote their resources to performing the differentiated function, immunoglobulin production. We found that a purified recombinant human interleukin-6, which had been reported to support or stimulate proliferation of B cell hybridoma/plasmacytoma cells, suppressed growth of a hybridoma cell line in serum-free medium. In the presence of the interleukin, the growth-suppressed cells were viable for remarkably long periods in batch culture, and after removal of the interleukin from the culture medium, they started to proliferate at their normal growth rate. As the concentration of the interleukin increased in the culture, the growth rate decreased and the specific antibody productivity (antibody production rate per cell) increased to 5-fold of control at 10 U ml-1 (2 ng ml-1) of the interleukin.

Animals

Efficient selection of mu m-mutants from mu m-expressing myeloma cells by treatment with ricin A-conjugated anti-mu antibody.

We have developed an efficient system for obtaining myeloma mutants defective in trans-acting factors required for immunoglobulin (Ig) gene expression. The system consists of a myeloma cell line designed for this purpose and an efficient method for selecting mutants from it. The cell line is X63.653 transfected with the mu gene, whose tailpiece sequence was replaced with the transmembrane sequence of human EGF receptor to hold mu on the cell surface and whose CH1 sequence was removed to prevent mu from being retained in the endoplasmic reticulum. It efficiently and stably expressed mu chains of IgM on the cell surface (mu m+) without light chains. To obtain mutants lacking mu m (mu m-) from the mu m+ cell line by selectively killing mu m+ cells, a method with ricin A-conjugated anti-mu antibody was more reliable than complement lysis mediated by anti-mu antibody. Applying the system, we obtained a variety of mu m- mutants.

Animals

Immunocytochemical studies on light-induced changes in phosphatidylinositol 4,5-bisphosphate immunoreactivity in the visual system of normal and norpA mutant of Drosophila.

The distribution of phosphatidylinositol 4,5-bisphosphate (PIP2) in the visual system of Drosophila was studied by indirect immunofluorescence staining using a monoclonal antibody against PIP2. The retina of the compound eye and the cortical regions of the optic lobe were heavily stained by the antibody. In the retina, photoreceptor cells were stained with the antibody, but non-neuronal cells such as pigment cells and cone cells were not stained. The staining intensity in the light-adapted photoreceptor cells was lower than that in dark-adapted cells in normal flies, whereas no such difference in immunoreactivity was observed in the norpAEE5 mutant, whose photoreceptor cells are deficient in phospholipase C. These results suggest that the PIP2 in the photoreceptor cells is hydrolyzed by phospholipase C coded by the norpA gene upon light stimulation.

Animals

Increased oxidized form of human serum albumin in patients with diabetes mellitus.

High-performance liquid chromatographic (HPLC) analysis of human serum albumin (HSA) on Asahipak GS-520H columns at neutral pH (6.87) showed a clear resolution of human mercaptalbumin (HMA) and nonmercaptalbumin (HNA), which are reduced and oxidized form of HSA, respectively. We studied the conversion of HMA to HNA (mercapt-nonmercapt conversion) as an index of oxidative change of the tissues and organs in 28 normal subjects and in a total of 47 patients with non-insulin dependent diabetes mellitus (NIDDM). Mean (+/- SD) values of the HMA fraction of HSA, f(HMA), [HMA/(HMA + HNA)], was significantly lower in NIDDM patients than in normal subjects (0.63 +/- 0.067 vs 0.75 +/- 0.028, P < 0.001). It was lower in poorly controlled NIDDM patients (0.63 +/- 0.058, n = 20) than in well controlled NIDDM patients (0.67 +/- 0.032, n = 9) (P < 0.05). Plasma glucose values sampled on occasions including overnight fasting and postprandial ones (r = -0.441, n = 47, P < 0.01), but not plasma glucose values sampled on overnight fasting (r = -0.345, n = 29) or postprandial (r = -0.467, n = 18) conditions and HbA1c (r = -0.211, n = 34), negatively correlated with the f(HMA) values, indicating that mercapt-nonmercapt conversion may not be due to cumulative hyperglycemia over a month, but due to short-term alteration in blood glucose level. The presence or absence of diabetic complications including nephropathy, retinopathy and neuropathy did not affect the f(HMA) values. In conclusion, decreased f(HMA) values in the diabetic patients suggested the presence of a rapidly altered oxidative change of albumin due to hyperglycemia.

Adult

Nortriptyline response in elderly depressed patients.

1. Depressed geriatric patients were treated with nortriptyline (NT) for 6 weeks. The authors measured serum levels of NT and 10-hydroxynortriptyline (10-OH-NT) using a column-switching HPLC method, and examined aging effects on NT steady-state levels to NT doses (doses/kg) ratios and NT levels to 10-OH-NT levels ratios as well as clinical response and propensity for side effects. 2. There was no significant relationship between the ages and the NT serum levels to NT doses (doses/kg) ratios, the ages and the 10-OH-NT levels to NT levels ratios, or the ages and the clinical response or the %improvement of Hamilton Scores. 3. Then the authors divided the subjects into two groups: a younger group and an elderly group with the cut off age of 60. The elderly group received significantly smaller doses of NT and had significantly lower serum levels of NT. The elderly group had tendency to have lower serum levels of 10-OH-NT. However, no significant difference was found in the improvement scores or the %improvement of depression. The elderly patients did not have higher propensity for unnegligible side effects.

Adult

Alteration in surface structure of Clara cells and pulmonary cytochrome P-450b level in rats exposed to ozone.

Changes in the surface structure of Clara cells in the terminal bronchioles following exposure of rats to 0.4 ppm ozone (O3) for 14 days were evaluated and compared to the content of pulmonary cytochrome P-450, an enzyme active in xenobiotic metabolism. Exposure to O3 caused a striking alteration of Clara cells in the terminal bronchiole. After 6 h exposure apical protrusions of Clara cells enlarged and these Clara cells formed clusters. However after 24 h exposure, the Clara cells decreased in number and flattened. They increased in number and enlarged again during the subsequent period of exposure. By the 14th day of O3 exposure the number of Clara cells had increased significantly. The content of cytochrome P-450b (IIB1), a main isozyme of pulmonary cytochromes P-450 of rats, was determined by an immuno-blotting method using anti-cytochrome P-450b antibody. The cytochrome P-450b in the rats exposed to O3 increased significantly to 1.37- and 1.81-times that of the control on the 7th and 14th days, respectively. Immuno-electron microscopy demonstrated that cytochrome P-450b was localized abundantly in endoplasmic reticulum of Clara cells. Morphological alterations in Clara cells appear to be closely related with changes in the cytochrome P-450b content of the lung.

Animals

Survey of methicillin-resistant clinical strains of coagulase-negative staphylococci for mecA gene distribution.

A total number of 125 methicillin-resistant (MIC, greater than or equal to 16) coagulase-negative Staphylococcus strains isolated in Japan were surveyed for the distribution of the mecA gene, the structural gene for penicillin-binding protein 2', which is the causative genetic element for the intrinsic resistance of methicillin-resistant Staphylococcus aureus. Screening with colony hybridization by using a cloned mecA gene probe revealed that 121 strains (96.8%) belonging to the nine coagulase-negative Staphylococcus species (S. epidermidis, S. haemolyticus, S. saprophyticus, S. sciuri, S. simulans, S. hominis, S. capitis, S. warneri, and S. caprae) carried mecA in their genome, indicating wide distribution of the gene among coagulase-negative Staphylococcus species. Most (93.4%) of the mecA-carrying strains were producers of penicillinase. Four strains, including two S. haemolyticus and two S. saprophyticus strains, did not carry mecA in spite of their resistance to methicillin. One of them was of low-level resistance (MIC, 16), but three of them had moderate- to high-level resistance to methicillin (MIC, 64). Analysis of gel electrophoretic banding patterns of penicillin-binding proteins of these strains showed absence of penicillin-binding protein 2' but some alterations in signal intensities of the other penicillin-binding proteins. The result indicated that about 3% of methicillin-resistant coagulase-negative staphylococci in these hospitals had a resistance mechanism different from that associated with the production of penicillin-binding protein 2', as has been reported in the case of a borderline methicillin-resistant strain of S. aureus.

Bacterial Proteins

Role of endogenous ANP in sodium excretion in rats with experimental pulmonary hypertension.

To explore the roles of endogenous atrial natriuretic peptide (ANP) in the volume regulation, we examined effects of antiserum for ANP or a neutral endopeptidase inhibitor, thiorphan, in rats with monocrotaline (MCT)-induced pulmonary hypertension. ANP concentrations in the plasma and right ventricle and ANP mRNA in the right ventricle of MCT-treated rats were higher than in vehicle-treated rats. The administration of the ANP antiserum into the MCT-treated rats did not affect the right atrial pressure or blood pressure but significantly decreased urinary excretion of Na by 60%. No decrease occurred in the control rats. Thiorphan dose dependently increased the urinary excretion of Na by 140% without influencing the right atrial pressure or blood pressure. This natriuresis was associated with 50 and 450% increases in ANP concentrations in the plasma and urine, respectively. The degrees of increases in urinary Na excretion, ANP, and guanosine 3',5'-cyclic monophosphate were significantly greater in the MCT-treated rats than in the control rats. Thus an increased secretion of ANP in pulmonary hypertension actually contributes to Na excretion. The augmentation of endogenous ANP activity may further potentiate the compensatory role of this peptide in the regulation of body fluid volume.

Animals

Electron-microscopic evidence for cytochrome P-450 in Clara cells and type I pneumocytes of the rat lung.

Using immunoelectron microscopy, we examined the distribution of cytochrome P-450 in rat lungs and determined the cell types in which P-450 was localized. To demonstrate P-450, a rabbit polyclonal antibody against P-450b (P-450 IIB1), the major constitutive isozyme of rat pulmonary P-450, was used. Strong reactivity for anti-P-450b antibody was observed in nonciliated bronchial and bronchiolar epithelial cells (Clara cells) and type I pneumocytes. Type II pneumocytes and Kultschitzky cells showed no reactivity. Most ciliated cells were negative, although a small number in peripheral bronchioles showed a weakly positive reaction. Endothelial cells and other mesenchymal cells were P-450b negative. Many studies have suggested that the Clara cell was metabolically very active and had P-450 containing mixed function oxidase systems. Our study shows that both Clara cells and type I pneumocytes have P-450 and may play an important role in the metabolism of xenobiotics. The metabolic significance of type I pneumocytes was not recognized, but it is reasonable to postulate that type I pneumocytes may have a xenobiotic metabolic function because they comprise most of the surface area of the lungs and are thus extensively exposed to the environment.

Animals

Cellular mechanisms for synthesis and secretion of atrial natriuretic peptide and brain natriuretic peptide in cultured rat atrial cells.

To investigate the cellular mechanism for the synthesis and secretion of atrial natriuretic peptide (ANP) and brain natriuretic peptide (BNP), we examined the effects of vasoactive agents on the secretion rates and gene expression of ANP and BNP in cultured rat atrial cells. Endothelin (10(-7) M, +61%), 12-O-tetradecanoylphorbol 13-acetate (TPA, 10(-6) M, +62%), the calcium ionophore A23187 (10(-6) M, +95%), and Bay K 8644 (10(-6) M, +34%) (p < 0.05 each) all increased the secretion of ANP into the culture media in a dose-dependent fashion. On the other hand, endothelin (10(-7) M, +57%) and TPA (10(-6) M, +55%) (p < 0.01 each) increased the secretion of BNP in a dose-dependent manner, whereas A23187 (10(-6) M, -45%, p < 0.001) suppressed the secretion of BNP in a dose-dependent manner, and Bay K 8644 caused no significant effects on BNP secretion. The molecular forms of intracellular ANP were exclusively gamma-ANP, whereas those of BNP were gamma-BNP and its carboxy terminal 45-amino-acid peptide, BNP-45. The ratio of media to cell contents was much higher in BNP than in ANP. Northern blot analysis revealed that both ANP mRNA and BNP mRNA levels were significantly increased by 10(-7) M endothelin (ANP mRNA, +52%; BNP mRNA, +36%; p < 0.05 each) and 5 x 10(-5) M 1-oleoyl-2-acetylglycerol (ANP mRNA, +296%; BNP mRNA, +133%; p < 0.01 each) but not by 10(-6) M A23187. Thus, the secretion of ANP is stimulated by both the elevation of [Ca2+]i and the activation of protein kinase C, whereas its synthesis is increased mainly by the activation of protein kinase C. The synthesis and secretion of BNP are augmented by the activation of protein kinase C rather than the elevation of [Ca2+]i. Furthermore, the processing and secretion of ANP and BNP may be regulated in different manners.

Animals

Vitamin C activity of 2-O-alpha-D-glucopyranosyl-L-ascorbic acid in guinea pigs.

The vitamin C activity of 2-O-alpha-D-glucopyranosyl-L-ascorbic acid (AA-2G), which is one of chemically stable derivatives of L-ascorbic acid (AsA), in guinea pigs was investigated. Male guinea pigs were divided into 9 groups and fed AsA-deficient diet for 24 days with the following supplement: AA-2G- or AsA-supplemented groups were orally supplemented with 0.96, 1.92, 9.6 and 192 AA-2G mg/animal/day or equimolar amounts of AsA (0.5, 1, 5 and 100 mg/animal/day, respectively); AsA-deficient group received neither of them. The body weight gain, serum alkaline phosphatase activity, and the concentration of AsA and AA-2G in the liver, adrenals and urine of the guinea pigs were measured at the end of the experimental period. The AA-2G-supplemented guinea pigs showed similar body weight gain to the animals supplemented with equimolar amount of AsA. Serum alkaline phosphatase activity in both AA-2G- and AsA-supplemented groups was significantly higher than that of AsA-deficient group. But there was no significant difference between the groups supplemented with AA-2G and the equimolar amount of AsA. AA-2G-supplemented guinea pigs showed no apparent symptoms of scurvy. In AA-2G-supplemented groups, AA-2G was not detected in the liver, adrenals and urine, but AsA was found and the AsA concentration increased with increasing AA-2G dosage. The AsA concentration in the tissues of each AA-2G-supplemented groups was higher than that of AsA-deficient group, which was similar to that of the groups supplemented with equimolar amount of AsA. These results showed that AA-2G has the same vitamin C activity as AsA on a molar basis for the orally supplemented guinea pigs.

Adrenal Glands