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Biomedical subjects

H Inoue

Publications and source records attributed to H Inoue.

At least 1,009 records · Page 56Linked to original sources
PubMed1995

Determination of free hydroxyproline and proline in human serum by high-performance liquid chromatography using 4-(5,6-dimethoxy-2-phthalimidinyl)phenylsulfonyl chloride as a pre-column fluorescent labelling reagent.

A fluorescent labelling reagent, 4-(5,6-dimethoxy-2-phthalimidinyl)phenylsulfonyl chloride, was designed for the determination of amines by precolumn HPLC and was applied to the simultaneous determination of hydroxyproline and proline in serum. The reagent reacted with hydroxyproline and proline at 30 degrees C for 10 min to produce the fluorescent derivatives, which were separated on a reversed-phase column by gradient elution with phosphate buffer (1 mmol l-1, pH 7) and acetonitrile and detected by fluorescence measurement at 315 nm (excitation) and 385 nm (emission). The detection limits (signal-to-noise ratio = 3) for both hydroxyproline and proline were 10 fmol per injection. The within-day (n = 10) and day-to-day (n = 5) relative standard deviations using human sera were less than 2.16% and 2.75%, respectively, for hydroxyproline and less than 2.30% and 3.25%, respectively, for proline. The concentrations of free hydroxyproline and proline in normal human sera (n = 13) were 5.6-18.0 and 137.6-252.6 mumol l-1, respectively. The proposed method was also applied to the determination of hydroxyproline and proline in sera from patients with chronic renal failure. The mean concentrations of hydroxyproline and proline in chronic renal failure were about 2.6 and 1.6 times higher, respectively, than those in normal human sera.

Chromatography, High Pressure Liquid↗
PubMed1995

Expression of the choline acetyltransferase gene depends on protein kinase A activity.

Choline acetyltransferase activity is barely detectable in a mutant pheochromocytoma PC12 cell line, A123.7, which is deficient in protein kinase A activity. Northern blot and polymerase chain reaction analyses showed that this mutant cell line has dramatically reduced levels of choline acetyltransferase mRNA, which correlates with the low level of enzyme activity. Transient transfection analysis was used to assess the functionality, in these cells, of an enhancer element and a cholinergic-specific repressor element derived from the human choline acetyltransferase gene. The results show that the enhancer element is inactive in the protein kinase A-deficient cell line. Cotransfection experiments with plasmids expressing the catalytic subunit of protein kinase A support this conclusion. These data indicate that protein kinase A regulates expression of the choline acetyltransferase gene at the transcriptional level by controlling the activity of an enhancer element.

Animals↗
PubMed1995

8-Hydroxyguanine (7,8-dihydro-8-oxoguanine) in hot spots of the c-Ha-ras gene: effects of sequence contexts on mutation spectra.

We previously reported that 8-hydroxyguanine (7,8-dihydro-8-oxoguanine) at the second position of codon 12 of the c-Ha-ras gene induces many types of mutations in NIH3T3 cells. In this study we incorporated the modified base into the first and second positions of codon 12 in the coding strand and into the first position of codon 61 in the non-coding strand of the gene using a new 8-hydroxyguanine phosphoramidite as a building block during oligonucleotide synthesis. The ras genes with 8-hydroxyguanine were transfected into NIH3T3 cells and the mutations induced were analyzed. 8-Hydroxyguanine residues at the first positions of codons 12 and 61 induced mutations to T at the modified sites almost exclusively. On the other hand, the DNA lesion at the second position of codon 12 induced a G-->A transition in addition to a G-->T transversion, confirming our previous results. Mutations in 5'-flanking sites were observed with 8-hydroxyguanine at the second position of codon 12 or the first position of codon 61. These results indicate that 8-hydroxyguanine in mammalian cells mainly induces a G-->T transversion at the modified site, but that other types of mutations are also elicited.

3T3 Cells↗
PubMed1995

Structural analysis of the L-methionine gamma-lyase gene from Pseudomonas putida.

The gene encoding L-methionine gamma-lyase from Pseudomonas putida was cloned and the primary structure of the enzyme was deduced from its nucleotide sequence. The L-methionine gamma-lyase gene was expressed in Escherichia coli. The amino acid sequences of BrCN-digested peptides agreed with the corresponding parts of the L-methionine gamma-lyase sequence determined from the gene structure. The polypeptide is composed of 398 amino acid residues with a calculated molecular weight of 42,626, corresponding to the subunit of the homotetrameric enzyme. The deduced amino acid sequence of L-methionine gamma-lyase only showed extensive homology with other well known alpha,gamma-elimination and/or gamma-replacement pyridoxal 5'-phosphate-dependent enzymes, such as cystathionine gamma-lyase, cystathionine gamma-synthase, and O-acetylhomoserine O-acetylserine sulfhydrylase, that participate in the biosynthesis of sulfur amino acids. However, the deduced essential cysteine residue of L-methionine gamma-lyase was not conserved in these enzymes. We confirmed the presence of a part of an open reading frame in the 3'-flanking region of the L-methionine gamma-lyase gene, which showed high homology with the N-terminal region of pyruvate dehydrogenase (lipoamide) from E. coli, suggesting that it participates in the degradative pathway for L-methionine together with L-methionine gamma-lyase.

Amino Acid Sequence↗
PubMed1995

The effects of endothelin-1 on flap necrosis.

In reconstructive surgery, flap necrosis is a major obstacle. The vasospasm in microvascular circulation can be one of the causes of flap necrosis. The vascular endothelium has an important role in the regulation of vascular tone by producing vasodilating and vasoconstricting factors. Endothelin-1, a 21-amino acid polypeptide isolated from the vascular endothelium, is reported to be one of the most potent vasoconstrictors. The purpose of this study was to determine the significance of endothelin-1 on flap necrosis using a rat skin flap. First, we introduced into the experiment endothelin-1 and ETA receptor antagonists, FR139317, then we tried to determine the time course changes of endothelin-1 levels in the flap. Endothelin-1 antagonist had no effect on survival length, although endothelin-1 injection clearly reduced survival length. The combined treatment of antagonist and injection suppressed the effect of endothelin-1 in reducing survival length. Endothelin-1 levels in the flap base significantly increased from 6 hours, and its levels in the flap periphery presented no remarkable changes. These results may suggest that endothelin-1, especially that produced in the flap base, exerts a major effect on flap microcirculation.

Animals↗
PubMed1995

Immunohistochemical features of HLA-DR antigen expression and lymphoid infiltrates in gastric carcinoma after low-dose interleukin-2 and mitomycin C.

We immunohistochemically evaluated lymphoid cell infiltration and HLA-DR antigen expression in gastric tumor tissue obtained from advanced gastric cancer patients 1 day after the completion of the treatment with mitomycin C (MMC) 12 mg/m2 i.v. on day 1 and recombinant interleukin-2 (IL-2) i.v. every 12 h from day 4 through day 8. Then the results were compared with those in 11 patients pretreated with MMC alone, 5 treated with IL-2 alone, and 24 untreated patients. Widespread lymphoid infiltration was observed in 17% of untreated tumors, 27% of MMC-pretreated tumors, and 40% of tumors treated with IL-2 alone. However, 71% of carcinomas pretreated with MMC plus IL-2 exhibited widespread infiltration. The frequency of cases with high-grade infiltration of CD4+ cells was significantly higher in either group of patients treated with MMC alone or MMC plus IL-2. Because the CD8+ cell infiltration was not significantly altered, the ratio of CD4+ to CD8+ cells estimated as being > 1 was more frequently noted in patients given MMC alone or MMC plus IL-2, as compared with untreated control. Furthermore, 86% of tumors pretreated with MMC plus IL-2 exhibited positive HLA-DR antigen expression, whereas 29% of untreated carcinomas did so. MMC or IL-2 alone did not significantly increase HLA-DR expression. These results indicate that the combination of low-dose of IL-2 with MMC enhances the intensity of lymphoid cell infiltration in tumors, with the predominance of CD4+ cells, and HLA-DR antigen expression on tumor cells in patients with advanced gastric carcinoma.

Aged↗
PubMed1995

Prolongation of intraventricular conduction time associated with fatal [correction of fetal] impairment of defibrillation efficiency during treatment with class I antiarrhythmic agents.

To test whether fatal deterioration of defibrillation efficiency during antiarrhythmic therapy can be prevented by avoiding extreme decrease in ventricular prevented by avoiding extreme decrease in ventricular conduction or toxic plasma drug levels, we determined the defibrillation threshold (DFT) before and during infusion of incremental doses of disopyramide (n = 8), mexiletine (n = 9), or flecainide (n = 9) in anesthetized dogs. Disopyramide did not alter DFT [from 4.4 +/- 1.5 to 4.4 +/- 1.6 J (3.1 +/- 1.2 micrograms/ml)]. Mexiletine tended to increase DFT [from 4.6 +/- 1.2 to 6.1 +/- 2.0 J (1.8 +/- 0.6 micrograms/ml); p < 0.05], and defibrillation eventually was unsuccessful in 3 of the 9 dogs. Although the plasma mexiletine level before refractory fibrillation was far beyond the human therapeutic range, prolongation of intraventricular conduction time (CT) was moderate (16 +/- 3%). Flecainide increased DFT from 4.2 +/- 1.3 to 6.1 +/- 1.5 J at a plasma level of 1.04 +/- 0.37 micrograms/ml (p < 0.0005). In 3 of 5 dogs that developed refractory fibrillation, plasma flecainide level before terminal ventricular fibrillation (VF) was not toxic, but prolongation of CT in the 5 dogs was remarkable (30 +/- 9%). Thus, VF resistant to defibrillation is not necessarily associated with both toxic plasma drug level and remarkably decreased conduction. Reliability of these valuables as indicators of fatally deteriorated defibrillation efficiency may vary among antiarrhythmic agents.

Analysis of Variance↗
PubMed1995

Effect of isoproterenol on facilitation of electrical defibrillation by E-4031.

To determine whether isoproterenol could reverse enhancement of electrical defibrillation effectiveness by class III antiarrhythmic agents, we measured the internal defibrillation threshold (DFT) in 12 anesthetized dogs during infusion of (a) saline (baseline), (b) isoproterenol, (c) isoproterenol + E4031 (a new class III antiarrhythmic agent), and (d) E4031 alone. The isoproterenol infusion was adjusted so that heart rate (HR) was at least 30 beats/min greater than baseline. E4031 was given as a 40-micrograms/kg bolus at the beginning of the third stage of the study, followed by constant infusion at 2 micrograms/kg/min. Eight dogs completed the study. Although the energy-based DFT was not affected by isoproterenol (from 6.1 +/- 1.5 to 6.0 +/- 1.7 J), it was decreased to 3.7 +/- 1.6 J in the third stage by infusion of E4031 and isoproterenol (p < 0.01 vs. baseline and vs. isoproterenol). After the discontinuation of isoproterenol in the fourth stage, i.e., during infusion of E4031 alone, DFT was 3.4 +/- 1.6 J (p < 0.01 vs. baseline and vs. isoproterenol). Therefore, isoproterenol did not antagonize the effect of E4031 on the DFT, suggesting the possible clinical usefulness of class III agents for facilitating defibrillation even in the presence of augmented sympathetic activity.

Animals↗
PubMed1995

Sheep red blood cell instillation at palatine tonsil effectively induces specific IgA class antibody in saliva in rabbits.

We have shown that the palatine tonsil effectively incorporates exogenous foreign substances instilled at its surface. It is not clear whether antigen-specific IgA can be induced by the instillation. Sheep red blood cells (SRBC) were instilled at the palatine tonsil every three days as the antigen, and the agglutination titer of specific IgA in saliva was examined. Nasal or intragastric administration, which have been shown to induce specific antibody in saliva, were done as control experiments. Anti-SRBC antibody in saliva from the tonsillar instillation group was detected in the second week, and the agglutination titer reached a maximum in the 6th week after instillation. The maximum titers in the tonsillar instillation group and nasal administration group were 16 (P < 0.01, n = 7) and 4 times (P < 0.01, n = 7) higher, respectively, than that in the intragastric administration group. In the tonsillar instillation group, the number of specific antibody- producing cells per 10(5) lymphocytes was the highest in the parotid glands compared with the lymphoid tissues such as the retropharyngeal lymph nodes, nasal mucosa, mesenteric lymph nodes, Peyer's patches, cervical lymph nodes, palatine tonsil and spleen. In the nasal administration group, the number of lymphocytes was the highest in the nasal mucosa. The results indicate that tonsillar instillation was more effective than nasal administration in inducing specific iIgA in saliva.

Administration, Intranasal↗
PubMed1995

Comparison of incorporation and extension of nucleotides in vitro opposite 8-hydroxyguanine (7,8-dihydro-8-oxoguanine) in hot spots of the c-Ha-ras gene.

DNA templates with 8-hydroxyguanine (7,8-dihydro-8-oxoguanine, oh8Gua) at a site corresponding to the first or second position of codon 12 of the c-Ha-ras gene were prepared, and the nucleotides inserted opposite the modified base were compared. The Klenow fragment (KF) of Escherichia coli DNA polymerase I inserted C opposite oh8Gua at both positions. Taq DNA polymerase incorporated C and A opposite oh8Gua, and the ratio of C to A was higher at the first position than at the second position. DNA polymerase alpha (pol alpha) inserted A and C at the first position, and A at the second position of codon 12, indicating that the ratio of C to A was higher at the first position. Moreover, we studied the extensions of bases paired with oh8Gua by DNA polymerases with or without 3'-5' exonuclease activity. G and T opposite oh8Gua were removed, and subsequently C was inserted by KF. We found that an oh8Gua:A pair was recognized by the exonuclease activity of the enzyme and that A was partially substituted by C. On the other hand, pol alpha extended only C and A opposite oh8Gua. No difference was observed with oh8Gua at the two positions. These results indicate that the ratio of nucleotides incorporated opposite oh8Gua depends on the sequence context, while there is no particular difference in the extension of base pairs involving oh8Gua by DNA polymerases.

Animals↗
PubMed1995

Familial hyperparathyroidism associated with jaw fibroma: case report and literature review.

A 53-year-old female suffering from renal stones and hypercalcaemia was diagnosed as having primary hyperparathyroidism caused by hyperplasia of the parathyroid glands. She underwent total parathyroidectomy and implantation of parathyroid tissue. After one year, she underwent surgery for a jaw tumour. The pathological findings indicated it to be a cementifying fibroma. Jackson et al. (1990) reported the familial association of hyperparathyroidism with jaw tumours, and they suggested that this condition represents a new clinical syndrome. We believe that our case belongs to this syndrome.

Female↗
PubMed1995

B cell mitogenic activity of Toxocara canis adult worm antigen.

The effect of Toxocara (T.) canis antigen (TcAg) on lymphocytes was studied in vitro using normal murine spleen cells and human peripheral blood lymphocytes. TcAg prepared from adult worms stimulated murine spleen cells to proliferate at concentrations of 1-125 micrograms/ml. The responder cells TcAg are B cells, because the response was depleted by the treatment of spleen cells with anti-immunoglobulin (Ig) antibody and complement and after separation on a nylon wool column. This response was not due to the contamination of lipopolysaccharide (LPS), because TcAg could stimulate C3H/HeJ spleen cells which are low responders to LPS. Not only the proliferative response but also polyclonal IgG and IgE production were stimulated with TcAg. TcAg also stimulated macrophages to produce interleukin-1 and could stimulate human B cells. These results suggest that TcAg is a potent B cell mitogen and this activity may be relevant to the alteration of immunological functions in hosts infected with T. canis.

Animals↗
PubMed1995

Experimental osteodystrophy of chronic renal failure induced by aluminum- and ferric-nitrilotriacetate in Wistar rats.

The aluminum (AI) and iron (Fe) chelate complexes of nitrilotriacetate (NTA) cause renal insufficiency when they are administered intraperitoneally to rats. Their effects on bone metabolism were studied in 4 week old Wistar rats. Daily intraperitoneal administration of AI-NTA (3mg AI/kg for 11 weeks) induced osteomalacia, impaired bone growth, decreased bone mineral density, lower serum PTH levels than normal as well as renal insufficiency. Al staining showed diffuse deposition in the trabecula and a strong linear band of aluminum deposited at the mineralization front and along the cement line. The osteoid seen markedly within the trabecula was probably the decalcified portion of the bone, the calcium apatite of which was defectively fabricated because of diffuse Al deposition in the trabecula. Al deposition along the cement line would make it much more susceptible to external shear stress than normal. Although daily intraperitoneal administration of Fe-NTA (6 mg Fe/kg for 11 weeks) caused impaired bone growth, decreased bone mineral content and renal insufficiency, the osteoid volume did not increase. Fe staining showed that Fe was deposited diffusely in the cytoplasm of osteoblasts. The results of this study demonstrated that during renal insufficiency, different minerals exhibit different modes of action on bone metabolism, and that Al-NTA is useful for experimental animal models of Al-induced osteomalacia in renal insufficiency.

Animals↗
PubMed1995

Laparoscopic mesenterioadhesiotomy and Tenckhoff catheter placement in patients with predisposing abdominal surgery.

Peritoneoscopic surgery has been performed widely for a variety of abdominal surgical diseases. We describe here a safe and reliable technique of laparoscopic-assisted mesenterioadhesiotomy and peritoneal Tenckhoff catheter placement in patients who have previously undergone abdominal surgery. Five patients suffering from end-stage renal failure previously underwent single and/or polyabdominal surgery. The surgical procedures included hysterectomy, ovarian resection, appendectomy, and transabdominal right nephrectomy. Under general endotrachial anesthesia, a laparoscope was placed down through a direct cut made using a trocar. After CO2 gas insufflation, another one or two trocars were put in place for surgical procedures. To avoid intestinal injury, mesenterioadhesiotomy was performed carefully using a high-frequency hook electrode, forceps, and scissors forceps, and the Tenckhoff catheter was subsequently inserted with forceps directly into Douglas' fossa. Peritoneal equilibration tests performed 30-70 days after the initiation of continuous ambulatory peritoneal dialysis (CAPD) treatment revealed moderate to good peritoneal effectiveness. This procedure permits the surgeon to perform safe and exact catheter placement into Douglas' fossa even when there is a possibility that peritoneal and mesenterial adhesion are present. We believe that this technique of catheter placement may extend the indication for CAPD treatment in patients with predisposing lower abdominal surgery.

Aged↗
PubMed1995

Immunohistochemical demonstration of fibronectin in the most superficial layer of normal rabbit articular cartilage.

OBJECTIVE: To locate fibronectin ultrastructurally in the most superficial layer of normal articular cartilage of rabbits, in order to clarify its role in joint physiology. METHODS: Articular cartilage was obtained from the femoral condyle of seven normal adult rabbits and prepared by a method that included tannic acid fixation. Polyclonal antibodies against rabbit fibronectin were used in an immunohistochemical electron microscopic study, without any enzymic digestion but with a pre-embedding method for the transmission electron microscopy. RESULTS: The cartilage surface was successfully preserved by tannic acid fixation. The most superficial layer in electron photomicrographs was approximately 200-300 nm thick, cell free, and appeared to have two parallel components: the more superficial lamina and the deeper lamina. Gold labelled fibronectin lined this layer in immunohistochemical electron photomicrographs. CONCLUSIONS: Fibronectin covering the surface of the articular cartilage may have a role in joint lubrication and protection of the cartilage by binding with the collagenous matrix and hyaluronic acid in synovial fluid. Chondroitin sulphates may act as a charge barrier in close relationship with the collagen fibrils in the deeper lamina. Significant alteration in these functions may be one of the first causal steps leading to destruction of the articular cartilage.

Animals↗
PubMed1995

Tolbutamide inhibits glucagon-induced phosphorylation of 6PF-2-K/Fru-2,6-P2ase in rat hepatocytes.

In previous studies, we demonstrated that tolbutamide inhibits a phosphorylation of hepatic 6-phosphofructo-2-kinase (6PF-2-K)/fructose-2,6-bisphosphatase (Fru-2,6-P2ase) catalyzed by the adenosine 3',5'-cyclic monophosphate-dependent protein kinase in a reconstruction system using the purified enzyme from the rat liver. In the current study, to assess a role of tolbutamide on hepatic 6PF-2-K/Fru-2,6-P2ase physiologically, we used intact rat hepatocytes and examined effects of tolbutamide on a phosphorylation of the bifunctional enzyme in the presence of glucagon. Glucagon induced a rapid phosphorylation of hepatic 6PF-2-K/Fru-2,6-P2ase accompanied by an inhibition of 6PF-2-K activity and a stimulation of Fru-2,6-P2ase activity in a dose-dependent manner. Tolbutamide inhibited glucagon-induced phosphorylation of the bifunctional enzyme protein in a dose-dependent manner. By adding 2 mM tolbutamide, reduced activity of 6PF-2-K and increased activity of Fru-2,6-P2ase in the presence of 10(-9) M glucagon were partially restored. The present results suggest the possibility that tolbutamide modulates the activity of hepatic 6PF-2-K/Fru-2,6-P2ase through inhibiting a phosphorylation of the enzyme protein. The counterregulatory influence of tolbutamide on the effect of glucagon suggests a possible mechanism for the extrapancreatic effect of sulfonylurea drugs.

Animals↗
PubMed1995

Staphylococcus aureus stimulates neutrophil recruitment by stimulating interleukin-8 production in dog trachea.

The present study examined whether neutrophil recruitment in dog airways by Staphylococcus aureus is mediated by interleukin-8 (IL-8). S. aureus culture supernatant was superfused into an isolated tracheal segment in six dogs, and neutrophil recruitment and IL-8 concentrations were measured in the superfusate. Dog IL-8 was cloned, expressed in Escherichia coli, purified by chromatography, and shown to be biologically active. With the use of an enzyme-linked immunoabsorbent assay (ELISA) for the measurement of dog IL-8, we showed that S. aureus supernatant induced neutrophil recruitment and increased IL-8 concentration in the superfusate in a time-dependent manner. The chemotactic activity present in the superfusate 6 h after superfusion with S. aureus was inhibited by an anti-IL-8 antibody. S. aureus supernatant also stimulated IL-8 production and gene expression by cultured canine tracheal epithelial cells. These results provide evidence that IL-8 plays a major role in S. aureus-induced neutrophil recruitment in the airways by stimulating IL-8 production in airway cells.

Amino Acid Sequence↗