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Biomedical subjects

K Shimada

Publications and source records attributed to K Shimada.

At least 775 records · Page 43Linked to original sources
PubMed1994

Nitric oxide release from rat aortic smooth muscle cells is not attenuated by angiotensin converting enzyme inhibitors.

We investigated the effects of angiotensin converting enzyme inhibitors on nitric oxide (NO) synthesis in cultured rat vascular smooth muscle cells. We measured the production of nitrite, a stable metabolite of NO, and the expression of inducible NO synthase mRNA by vascular smooth muscle cells. Incubation of the culture with interleukin-1 beta (10 ng/ml) for 24 h caused a significant increase in nitrite levels. The basal and interleukin-1 beta-induced nitrite production by vascular smooth muscle cells were not affected by the presence of angiotensin converting enzyme inhibitors (0.1 approximately 10 microM), enalaprilat, cilazaprilat or captopril. Unstimulated vascular smooth muscle cells expressed no inducible NO synthase transcripts, whereas incubation with interleukin-1 beta for 24 h induced marked inducible NO synthase mRNA expression. The angiotensin converting enzyme inhibitors, however, had no effects on the interleukin-1 beta-induced inducible NO synthase mRNA expression. These results indicate that angiotensin converting enzyme inhibitors do not attenuate NO synthesis by vascular smooth muscle cells under basal and interleukin-1 beta-stimulated conditions.

Amino Acid Oxidoreductases↗
PubMed1994

Application of the desulfurization of phenothiazines for a sensitive detection method by high-performance liquid chromatography.

A simple and sensitive high-performance liquid chromatographic (HPLC) method for the determination of phenothiazine (PHE) is described. PHE is converted to diphenylamine (DIP) by desulfurization with Raney nickel catalyst. DIP is highly sensitive to electrochemical detection. The calibration graph for PHE quantification after desulfurization was linear between 0.1 and 2.0 ng per injection. The detection limit (signal-to-noise ratio = 3) of PHE after desulfurization was 10 pg, which is twenty times higher than that of the parent compound PHE. The proposed desulfurization technique was applied to other PHE-related compounds. The structural confirmation of the desulfurized product of PHE was carried out by LC-MS using atmospheric pressure chemical ionization.

Calibration↗
PubMed1994

Energy migration in allophycocyanin-B trimer with a linker polypeptide: analysis by the principal multi-component spectral estimation (PMSE) method.

Energy migration processes in allophycocyanin-B trimer with a linker polypeptide were analyzed using the principal multi-component spectral estimation (PMSE) method, which does not require assumption of component number, decay function, or the spectral band shape. We determined the number of spectral components showing independent kinetic behavior by the eigen-value of an auto-correlation matrix, and further the spectra of the components and their rise and decay curves. Two decay components were resolved at 20 degrees C: one corresponded to the decay of one type of beta-84 chromophore, and the other to the decay from the thermally equilibrated state between another type of beta-84 chromophore and the alpha-allophycocyanin B chromophore. An additional slow decay process was resolved at -196 degrees C. We also compared the component spectra obtained using the PMSE method with the decay-associated spectra obtained using the global analysis.

Cyanobacteria↗
PubMed1994

cDNA cloning and expression of bovine endothelin converting enzyme.

We have cloned cDNA encoding bovine endothelin converting enzyme (ECE). The predicted amino acid sequence of bovine ECE consisted of 758 amino acid residues. Northern blot analysis revealed that ECE mRNA was abundantly expressed in lung. Co-expression of the cloned cDNA of bovine ECE with human preproET-1 cDNA in CHO-K1 cells resulted in the production of mature ET-1 and this production was inhibited by phosphoramidon.

Amino Acid Sequence↗
PubMed1994

Involvement of LDL receptor in the proliferation of vascular smooth muscle cells.

To study the involvement of the low-density lipoprotein (LDL) receptor in the growth of vascular smooth muscle cells (VSMC), we compared the proliferation of cultured VSMC from Watanabe heritable hyperlipidemic (WHHL) rabbits, which lack the LDL receptor, and VSMC from normal Japanese white rabbits in response to platelet derived growth factor (PDGF). The increase in the number of VSMC from WHHL rabbits in response to PDGF (10(-8) M) was significantly lower than that of VSMC from normal rabbits. PDGF stimulated the synthesis of DNA in VSMC from both normal rabbits and WHHL rabbits, but the response was significantly lower in the latter. To determine the involvement of the LDL receptor in the decreased mitogenic response of WHHL rabbit VSMC, we used an anti-LDL receptor monoclonal antibody (MAb) to normal rabbit VSMC; DNA synthesis of VSMC was stimulated by PDGF, but the effect was significantly blocked by the anti-LDL receptor MAb. Mitogen-activated protein (MAP) kinase activity in normal rabbit VSMC was increased by exposure to PDGF, but the effect was significantly suppressed in the presence of the MAb. The anti-LDL receptor MAb markedly inhibited LDL binding to the surface of normal rabbit VSMC. These results suggest that the LDL receptor influences the proliferation of VSMC and thus might be involved in the pathogenesis of atherosclerosis.

Animals↗
PubMed1994

Derivatization of thiol-containing compounds.

The determination of thiol-containing compounds in biological fluids is important in biochemistry and clinical chemistry. In this paper, derivatization reagents for thiols are reviewed with respect to their reactivity, selectivity, spectroscopic characteristics and their applicability especially to high-performance liquid chromatography. Derivatization used in ultraviolet and electrochemical detection. The derivatization reagents contain a functional group, e.g. an N-substituted maleimide, active halogen or aziridine, which react with the thiol group. Derivatization for use in flow injection analysis, thin-layer chromatography or gas chromatography-mass spectrometry is also described.

Chromatography, High Pressure Liquid↗
PubMed1994

Tissue-specific expression of calbindin-D28K gene during ontogeny of the chicken.

The vitamin D3-dependent calcium binding protein, calbindin-D28K (CaBP-D28K), plays an important role in transepithelial calcium translocation. To evaluate its role in chick embryonic calcium metabolism, steady-state levels of CaBP-D28K mRNA in various tissues of the chick embryo were determined by Northern blot and slot blot analyses, and CaBP-D28K concentrations in the examined tissues and plasma were estimated by RIA. High levels of CaBP-D28K mRNA were found in the embryonic kidney (mesonephros) on embryonic day (E) 10 and E12 and thereafter gradually decreased until hatching. CaBP-D28K mRNA levels were low in the yolk sac until E16 but increased on E18 and reached a maximum on E20. A steady increase in CaBP-D28K mRNA levels was observed in the cerebellum during the development from E10 to post-hatching. CaBP-D28K mRNA levels in the intestine were very low during the incubation period but significantly increased on days 1 and 7 after hatching. By Northern blot analysis, CaBP-D28K mRNA was barely detectable in liver, heart, and chorioallantoic membrane of the embryonic chick. Changes in immunoreactive CaBP-D28K of each tissue paralleled observed changes in mRNA levels. In plasma, measurable levels of CaBP-D28K were found as early as E8 and were stable until E18, when 6.5-fold increase was observed compared to E16. The highest level of CaBP-D28K in plasma was found on E20 and decreased after hatching. These temporal profiles of CaBP-D28K suggest that it may play an important role in the regulation of chick embryonic calcium homeostasis.

Aging↗
PubMed1994

Cloning and functional expression of endothelin-converting enzyme from rat endothelial cells.

Endothelin (ET) is a 21-residue potent vasoconstrictive peptide produced by vascular endothelial cells and formed from its precursor, big endothelin (big ET), by endothelin-converting enzyme (ECE). Here we report the cloning and functional expression of a complementary DNA encoding a rat ECE from endothelial cells. Rat ECE is a highly glycosylated protein consisting of 10 possible N-linked glycosylation sites, a zinc-binding domain, and a single membrane-spanning region. It has structural and sequence homology to neutral endopeptidase and Kell blood group protein, a putative neutral endopeptidase. Monoclonal antibody risen against purified rat lung ECE recognized broad glycosylated bands in membrane fractions prepared from both rat lung and COS cells transfected with a rat ECE expression vector. Expressed ECE was inhibited by phosphoramidon, but not by thiorphan. It also cleaved big ET-1 efficiently but not big ET-2 or big ET-3. Northern blot analysis revealed that ECE messenger RNA is widely expressed in many tissues.

Amino Acid Sequence↗
PubMed1994

One of the retinoic acid-inducible cDNA clones in mouse embryonal carcinoma F9 cells encodes a novel isoenzyme of fructose 1,6-bisphosphatase.

Rae-30, one of the retinoic acid (RA)-inducible cDNA clones in mouse embryonal carcinoma F9 cells, was sequenced and the deduced RAE-30 protein showed about a 70% homology to mammalian fructose 1,6-bisphosphatase (EC 3.1.3.11) (FBPase), in comparison to over 85% homology observed among the previously documented rat liver, pig kidney and human leukemic HL-60 cell FBPases. The Rae-30 mRNAs were not detected in various tissues of adult mice, including the liver and kidney, but were detected in a placenta and predominantly in the intestine of adult mice. These findings indicate that the Rae-30 cDNA encodes a novel isoenzyme of FBPase, which is likely to be involved in early differentiation in mammalian cells.

Amino Acid Sequence↗
PubMed1994

Conjugation of recombinant reverse transcriptase of HIV-1 to beta-D-galactosidase from Escherichia coli for ultrasensitive enzyme immunoassay (immune complex transfer enzyme immunoassay) of anti-HIV-1 IgG.

Recombinant reverse transcriptase (RT) of HIV-1 was conjugated to beta-D-galactosidase from Escherichia coli in three different ways. Maleimide groups were introduced into beta-D-galactosidase molecules using N,N'-o-phenylenedimaleimide in the absence (method I) or presence (method II) of N-ethylmaleimide or into beta-D-galactosidase molecules, which had been treated with excess of 4,4'-dithiodipyridine to block thiol groups, using N-succinimidyl-6-maleimidohexanoate (method III). Subsequently, the maleimide groups were reacted with thiol groups introduced into recombinant RT molecules using N-succinimidyl-S-acetylmercaptoacetate. The conjugates were tested by a sensitive enzyme immunoassay (immune complex transfer enzyme immunoassay). The immune complex consisting of 2,4-dinitrophenyl-bovine serum albumin-recombinant RT conjugate, anti-HIV-1 IgG and recombinant RT-beta-D-galactosidase conjugate was captured by polystyrene beads coated with (anti-2,4-dinitrophenyl group) IgG, eluted with N epsilon-2,4-dinitrophenyl-L-lysine and transferred to polystyrene beads with (anti-human IgG gamma chain) IgG. The conjugate prepared by method III, which showed the least polymerization, the least loss of the specific enzyme activity and the lowest nonspecific binding, improved the sensitivity of the enzyme immunoassay for anti-HIV-1 IgG approximately 30-fold compared with RT-horseradish peroxidase conjugate.

Animals↗
PubMed1994

Close relation between lipoprotein (a) levels and atherothrombotic disease in Japanese subjects > 75 years of age.

Levels of lipoprotein (a) (Lp[a]) and various hemostatic factors were studied in 132 Japanese aged > 75 years (mean 83). The group consisted of 50 healthy persons, 36 hypertensive subjects, 31 patients with chronic cerebral infarction, and 15 with coronary artery disease. Lp(a) levels were slightly lower in the healthy "old old" subjects than in the 184 healthy younger adults (mean +/- SD: 10.7 +/- 7.9 vs 12.1 +/- 10.1 mg/dl). There were no gender-related differences in the Lp(a) levels of healthy adults and healthy old old subjects. Lp(a) levels were higher in the hypertensive old old subjects (14.6 +/- 15.4 mg/dl) and the old old patients with cerebral infarction (21.3 +/- 16.2 mg/dl) and coronary artery disease (26.5 +/- 20.4 mg/dl). The prevalence of subjects with high Lp(a) levels (> 30 mg/dl) was the greatest among old old patients with coronary artery disease (27%). Lp(a) levels in the 132 old old subjects showed positive correlations with sialic acid, fibrinogen, factor VII activity, and D-dimer levels. These results indicate a close association between Lp(a) levels and atherothrombotic disease as well as the characteristics of Lp(a) as an acute phase reactant in old old Japanese. Subjects with higher Lp(a) levels may develop cardiovascular disease later in life, whereas the remaining healthy old old subjects have lower Lp(a) levels.

Aged↗
PubMed1994

Amperometric determination of oxalate in plasma and urine by liquid chromatography with immobilized oxalate oxidase.

A detection system for plasma and urinary oxalate involving a postcolumn enzymatic reaction and electrochemical detection is described. Oxalate oxidase was immobilized on AF-Tresyl Toyopearl 650 gel. The immobilized oxalate oxidase was packed into a stainless-steel column (10 x 4 mm I.D.) and used on-line as an immobilized enzyme reactor (IMER). The hydrogen peroxide produced by enzymatic reaction was detected amperometrically at a platinum electrode maintained at +0.5 V vs. Ag/AgCl. The HPLC separation of oxalate was carried out using a Capcell Pak C8 column and an isocratic mobile phase containing 80 mM KH2PO4 and 5 mM tetra-n-butylmammonium phosphate as an ion-pair reagent. The IMER was active and stable in the mobile phase employed. The plot of peak height against concentration of oxalate was linear in the range 0.1-1.6 mumol/ml with a correlation coefficient of 0.9989. The detection limit was 10 nmol/ml at a signal-to-noise ratio of 3. The within-day and between-day coefficients of variation were 5.3 and 7.9%, respectively.

Chromatography, High Pressure Liquid↗
PubMed1994

Detection of point mutations in the K-ras oncogene at codon 12 in pure pancreatic juice for diagnosis of pancreatic carcinoma.

BACKGROUND: Mutations in the K-ras oncogene at codon 12 are detected at a remarkably high frequency in pancreatic carcinomas and are believed to be a critical event in oncogenesis. The authors attempted to detect K-ras mutations in DNA obtained from pure pancreatic juice collected endoscopically, as a novel diagnostic approach to pancreatic carcinoma. METHODS: K-ras mutations were examined using the two-step polymerase chain reaction (PCR) combined with restriction enzyme digestion, followed by nonradioisotopic single-strand conformation polymorphism (SSCP) analysis. RESULTS: Specific mutations of the K-ras gene at codon 12 were found in six of nine (67%) duct cell carcinomas, all of which were negative by cytodiagnosis of the same pure pancreatic juice. K-ras mutations were not detected in the pancreatic juice from 14 healthy control subjects, 10 patients with chronic pancreatitis, or 3 patients with islet cell tumors. CONCLUSIONS: Detection of K-ras mutation at codon 12 in pancreatic juice is highly specific for diagnosing pancreatic duct cell carcinoma and may be a valuable diagnostic modality for pancreatic carcinoma and for differentiating chronic pancreatitis from carcinoma.

Adenocarcinoma↗