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Biomedical subjects

C Hayashi

Publications and source records attributed to C Hayashi.

At least 91 records · Page 5Linked to original sources

Determination of complement breakdown fragments C3d and its subfragments in health and disease.

The breakdown products of the third component of complement in approximately 400 samples were measured by rocket immunoelectrophoresis and two-dimensional electrophoresis using the method of Brandslund et al [3]. It was confirmed that the measurement of the C3d level provides useful information on increased C3 consumption irrespective of the synthetic rate. Furthermore, three subfragments with C3d but without C3c antigenicity were distinguished, which were designated as C3d1, C3d2, and C3d3. The subfragment C3d3 which migrated to the most anodal side was a predominant component in the plasma from patients with autoimmune diseases. Little C3d3 subfragment was detected in normal plasma and in normal sera incubated in vitro for 24 hr. Even in the normal sera converted completely in vitro which contained little intact C3, only a limited amount of C3d3 was detected. In the plasma from postsurgical patients in whom activation of the complement system was considered to be in an acute phase, C3d3 was detected, but the C3d2 level was higher than the C3d3 level. In the plasma from patients with systemic lupus erythematosus having the normal C3d level, C3d3 was a major fragment. It is predicted that the preponderant presence of C3d3 in plasma could be the result of chronic continuous complement activation by immune complexes.

Arthritis, Rheumatoid↗

Poor response of Wv/Wx mice to a grafted neutrophilia-inducing, colony-stimulating-factor-producing tumor.

Although mice possessing two mutant genes at the W locus have a defect in multipotential hematopoietic stem cells that form macroscopic colonies in the spleen of irradiated mice, the number of neutrophils in the blood of these mutant mice is normal or nearly normal. We investigated neutrophil production using the NFSA fibrosarcoma of C3H mouse origin, which induces neutrophilia accompanied by production of a neutrophil-macrophage colony-stimulating factor by the tumor. When the NFSA tumor was transplanted to (C57BL/6 X C3H/He)F1-Wv/Wx or to congenic +/+ mice, neutrophilia developed in mice of both genotypes. However, there was a significant difference between the degree of neutrophilia that developed in them; there was a 107-fold increase in the +/+ mice, but only a 28-fold increase in the Wv/Wx mice four weeks after tumor transplantation. This result is consistent with the concept that doubly heterozygous W mice have multipotential stem cells with diminished ability to respond to stimulation. The unperturbed condition may not provide a sufficient stimulus to demonstrate the defect in neutrophil production in doubly heterozygous W mutant mice.

Anemia↗

Simple enzymatic detection method for urinary sulfated 7 alpha-hydroxy bile acids in normal subjects and in patients with acute hepatitis.

Urinary sulfated primary bile acids, 7 alpha-hydroxy bile acids, are detected by an enzymatic method using 7 alpha-hydroxysteroid dehydrogenase (EC 1.1.1.-, 7 alpha-HSD) after chromatographic fractionation on Sephadex G-25. Urinary sulfated or glucuronated bile acids are hydrolyzed by beta-glucuronidase/sulfatase (EC 3.2.1.31/EC 3.1.6.1) from Helix pomatia and then released 7 alpha-hydroxy bile acids are detected with 7 alpha-HSD in the presence of beta-ND+, diaphorase (EC 1.6.99.2, from Clostridium kluyveri) and 2-p-iodophenyl-3-p-nitrophenyl-5-phenyltetrazolium chloride. The absorbance of formazan formed during the enzymic reaction is measured at 500 nm. Excretion values of 7 alpha-hydroxy bile acids in normal subjects and in patients with acute hepatitis were compared. This enzymatic detection method for the excretion pattern of urinary 7 alpha-hydroxy bile acids may be useful for clinical diagnosis.

Bile Acids and Salts↗

Presence of mast cell precursors in the yolk sac of mice.

Concentration of mast-cell precursors in hematopoietic tissues of mouse embryos was evaluated by a limiting dilution method. Cells from yolk sacs, livers, and bodies of (WB x C57BL/6)F1 (hereafter called WBB6F1)- +/+ embryos were injected directly into the skin of adult WBB6F1-W/Wv mice which were genetically depleted of tissue mast cells. Concentration of mast-cell precursors was calculated from the proportion of injection sites at which mast cells did not appear. Since the concentration of mast-cell precursors in the yolk sac was about 30 times as great as that of embryonic body at Day 9.5 of the pregnancy, the mast-cell precursors seemed to be generated within the yolk sac. The concentration in the yolk sac reached the maximum level at Day 11, and then dropped markedly at Day 13. In contrast, mast-cell precursors increased from Day 11 to Day 15 in the fetal liver. As a result, the concentration of 11-day yolk sacs was comparable to that of 15-day fetal liver. Although intravenous injection of 15-day fetal liver cells (2 x 10(6)) rescued the general mast-cell depletion of WBB6F1-W/Wv mice, the intravenous injection of the same number of 11-day yolk sac cells did not rescue it. In contrast with fetal livers, yolk sacs scarcely contained hematopoietic stem cells which were measured by spleen colony formation. Therefore, the mast-cell precursors of the yolk sac may not originate from such stem cells.

Animals↗

[Susceptibility of clinical isolates to cefotaxime].

Susceptibilities of 737 strains of 19 species of bacteria to cefotaxime (CTX) were determined based on the inhibition zone diameter obtained by the single-disc method. Four categories were assessed. 1. Susceptibility of clinical isolates to CTX and 6 other antibiotics Against most strains, CTX showed higher antibacterial activity than other drugs (CET, ABPC, SBPC, CMZ, GM, AMK), especially for S. pneumoniae, S. pyogenes and S. agalactiae. Furthermore, CTX was more active than the other antibiotics against E. coli, Indole (+) Proteus, P. mirabilis, Klebsiella sp., S. marcescens, H. influenzae and E. cloacae. 2. Susceptibility of strains isolated from different clinical materials CTX showed the highest antibacterial activity against most strains isolated from sputum, urine, pus, blood and cerebrospinal fluid. However, CTX was occasionally less than potent AMK and GM against strains isolated from bile. Against P. aeruginosa strains derived from clinical materials, the following results were obtained: AMK greater than CFS, FOM greater than CTX greater than GM greater than SBPC 3. Susceptibility of clinical isolates in 7 different fields CTX was the most active antibiotic tested in the fields of internal medicine, pediatrics, urology, obstetrics & gynecology, dermatology and otorhinolaryngology. But in surgery, CTX was less potent than GM and AMK. 4. Susceptibility of clinical isolates of inpatients and outpatients CTX showed excellent activity against many beta-lactamase resistant strains isolated from patients.

Bacteria↗

Semi-automated continuous-flow enzyme immunoassay for antiepileptic drugs in serum.

We have developed a semi-automated method for measuring five kinds of antiepileptic drugs in serum by successfully adapting commercial competitive-binding enzyme immunoassay kits (MARKIT; Dainippon) for use with a continuous-flow analyzer (Technicon AutoAnalyzer II equipped with a dialyzer). The free enzyme-labeled drug is automatically separated by a microfilter from the competitive immunoreaction mixture between labeled and unlabeled drug for anti-drug immunoglobulin coupled to bacterial cell walls. The concentrations of the antiepileptic drugs in serum samples can be determined by automated measurement of enzyme activity of the enzyme-labeled drugs. Results of the semi-automated method correlated well with those obtained by manual enzyme immunoassay, gas-liquid chromatography, and "high-pressure" liquid chromatography. The correlation coefficients were all greater than 0.95, showing the practicality of this method for therapeutic monitoring of antiepileptic drugs.

Anticonvulsants↗

Bone marrow origin of mast cell precursors in mesenteric lymph nodes of mice.

Concentration of mast-cell precursors in the mesenteric lymph node of (WB X C57BL/6)F1 hybrid mice (WBB6F1) were evaluated by a limiting dilution method. Cells from WBB6F1-+/+ mice were injected directly into the skin of WBB6F1-W/Wv mice which genetically lack tissue mast cells. Concentrations of mast-cell precursors were calculated from the proportion of injection sites at which mast cells appeared. Although immunization with horse serum significantly increased the concentration of mast-cell precursors in the mesenteric lymph node, the concentration in the lymph node remained about 10% that observed in the peripheral blood mononuclear cells. Since the bone marrow origin of mast-cell precursors in the mesenteric lymph node was demonstrated by using giant granules of beige (C57BL/6-bgj/bgj) mice as a marker, the immunization seemed to increase the migration of bone-marrow-derived mast-cell precursors from the peripheral blood to lymph node.

Animals↗

A new technique in echocardiology: B-mode to M-mode transformation.

In conventional two-dimensional Echocardiography with sector scanning, M-mode is simultaneously displayed on TV-monitor as the time based ultrasound signals. With conventional system, one end of the cursor-line in B-mode image which indicates the beam direction of M-mode, is always fixed at the point corresponding to the transducer tip. This system has the capability to transform B-mode to M-mode images in any desired direction. However, the time resolution in M-mode is limited to about 33 milli-seconds.

Biometry↗

Spectrophotometric method for selective assay of the five isoenzymes of human lactate dehydrogenase, based on their different stabilities at alkaline pH.

After preincubation of human lactate dehydrogenase (EC 1.1.1.27) isoenzymes 1 through 5 with lactate at pH 9.4, 9.8, 10.25, and 10.6 at 30 degrees C for 10 min, the reduction of NAD+ was measured at the same pH values and temperature during the interval 1-2 min after adding the coenzyme. Relative to the reference activities measured at pH 8.7 by the method of Buhl et al. (Clin Chem 23: 1289-1295, 1977), the respective activities of isoenzymes 1 through 5 thus measured were 111, 104, 96, 68, and 0% at pH 9.4; 123, 108, 88, 0, and 0% at pH 9.8; 140, 100, 0, 0, and 0% at pH 10.25; and 138, 0, 0, 0, and 0% at pH 10.6. These relations allow selective assay of the respective isoenzymes in samples containing mixtures of them. Optimal conditions for such selective assay at 37 degrees C are reported.

Electrophoresis↗

Plasma cholecystokinin responses after ingestion of liquid meal and intraduodenal infusion of fat, amino acids, or hydrochloric acid in man: analysis with region specific radioimmunoassay.

A highly sensitive and precise radioimmunoassay system for plasma cholecystokinin (CCK) was developed with the anti-CCK-8 specific antiserum which raised against N-terminal amino acids residue of sulfated CCK-8 and reacted with CCK-8, CCK-33, and CCK-39 but not with gastrin and its related peptides. Mean concentration of the fasting plasma CCK determined with this method using CCK-8 as standard was 12.9 +/- 5.9 pg/ml in normal subjects (n = 26), and in patients with hepatic cirrhosis it was significantly higher (36.7 +/- 16.9 pg/ml, n = 9, p less than 0.01) than in normal subjects. In six young healthy volunteers, intraduodenal infusion of fat caused a significant increase (p less than 0.05) of plasma CCK from a basal level of 8.0 pg/ml to a peak of 43.0 +/- 12.0 pg/ml at 20 min after starting of infusion. In the same subjects, a significant increase of plasma CCK was also observed by amino acids infusion, but no elevation of plasma CCK level was found during intraduodenal acidification.

Adult↗

Enzymatic detection of urinary conjugated steroids after gel chromatography.

An enzymatic detection method is described for urinary conjugated steroids after chromatographic fractionation with Sephadex G-25. The principle of the method is as follows. Part of a 24-h urine sample, (1-2 ml of urine) is applied directly, to a short column of Sephadex G-25 and eluted with acetate buffer solution. Steroid conjugates in each fraction are hydrolyzed with steroid sulfatase--beta-glucuronidase. After enzymatic hydrolysis, an enzymatic color development reagent for steroids, either 3 alpha-hydroxysteroid dehydrogenase or 3 beta-hydroxysteroid oxidase, are added and the dye formed is measured spectrophotometrically. Excretion patterns of steroid-3 beta-sulfates, and steroid-3 alpha-glucuronides and steroid-3 alpha-sulfates ae shown with some patients' samples. A precision of the assay values for steroid-3 alpha-glucuronide, steroid-3 alpha-sulfate and steroid-3 beta-sulfates in urine samples and assay values for normal subjects are also studied. This simple enzymatic method for detecting the excretion patterns of urinary conjugated steroids may have a diagnostic value for clinical tests.

Adrenal Gland Neoplasms↗

A new enzymatic method to determine creatine.

A new enzymatic method is described for the determination of creatine in serum and urine using creatine amidinohydrolase (EC 3.5.3.3), sarcosine oxidase (EC 1.5.99.1) and formaldehyde dehydrogenase (EC 1.2.1.1). The principle of the method is as follows. Creatine is degraded to sarcosine and urea, and the sarcosine formed is measured with sarcosine oxidase in the presence of formaldehyde dehydrogenase and NAD+. The NADH + H+ produced is measured at 340 nm. Creatine concentration can be calculated directly from the absorptivity of NADH + H+ generated in the reaction or from creatine standard solutions. The assay takes less than 20 min. The standard curve is linear up to 50 mg creatine/1 (serum) and 800 mg creatine/1 (urine). Fifty random samples were assayed by this method (y) and simultaneously by the Folin method (x). The correlation coefficients were 0.995 for serum samples, 0.994 for urine samples, and the regression equations were y = 0.979x - 0.01 (serum) and y = 0.978x - 0.01 (urine).

Aldehyde Oxidoreductases↗

Assays of serum lipase by the "BALB-DTNB method" mechanized for use with discrete and continuous-flow analyzers.

We successfully adapted the dimercaprol (BAL) tributyrate-5,5'-dithiobis(2-nitrobenzoic acid) method (J. Biochem. 81: 361, 1977) for assay of lipase in human serum to a discrete analyzer (the TBA 880) (I) or a continuous-flow analyzer (AutoAnalyzer, Type II) (II). In both, BAL-tributyrate is used as substrate, in combination with serum esterase inhibitors and a chromogenic reagent for the SH group of the liberated BAL. Serum lipase activities of patients with pancreatic diseases, measured at 90 or 40 samples per hour by I or II, respectively, correlated well with those measured by the corresponding manual method or by Kaplan's radioassay (Anal. Biochem. 33: 213, 1970). The correlation coefficients were all greater than 0.95, and the coefficients of variation were less than 8%, showing the practical usefulness of these procedures.

Autoanalysis↗