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Biomedical subjects

M Sugimoto

Publications and source records attributed to M Sugimoto.

At least 199 records · Page 11Linked to original sources
PubMed1995

Establishment by the rat lymph node method of epitope-defined monoclonal antibodies recognizing the six different alpha chains of human type IV collagen.

A group of rat monoclonal antibodies recognizing the six different alpha chains of human type IV collagen have been established by our novel method. The method is designated the rat lymph node method in which enlarged medial iliac lymph nodes of a rat injected with an antigen emulsion via hind footpads are used as a source of B cells for cell fusion to produce hybridomas. The immunogens used were synthetic peptides having non-consensus amino acid sequences near the carboxyl termini of type IV collagen alpha chains. Hybridomas were screened both by ELISA with synthetic peptides and by indirect immunofluorescence with cryostat sections of human kidneys. Because the epitopes of all antibodies were determined by multipin-peptide scanning, they were confirmed to be isoform-specific. They are useful for identification of alpha chains of type IV collagen at the protein level in normal and abnormal conditions. The combined use of synthetic peptides as immunogens, the rat lymph node method as making monoclonal antibodies, and the multipin-peptide scanning as epitope mapping is found to be a strong tool for identification of peptides and proteins whose amino acid sequences are known or have been deduced.

Amino Acid Sequence↗
PubMed1995

Shear stress-induced platelet aggregation in various types of von Willebrand disease.

We here examined the shear stress-induced platelet aggregation (SIPA) in various types of von Willebrand Disease (vWD), using a novel cone-and-plate viscometer recently developed by Ikeda et al. [1]. Under experimental conditions of high shear stress (90-108 dyne/cm2) known to induce von Willebrand factor (vWF)-dependent platelet aggregation in normal individuals, a variety of SIPA patterns, from a normal to total defect, were observed in 6 patients with type I vWD, while SIPA was totally absent in all 7 type IIA patients examined. One patient with Normandy type vWD exhibited a normal SIPA pattern under these experimental conditions. These findings, together with the vWF parameters of each patient, suggested that the presence of higher molecular weight multimers of plasma vWF, as well as an appropriate level of vWF antigen, is essential for SIPA to occur under high shear stress. Interestingly, vWF-dependent SIPA in 3 patients with type IIB vWD was found to be initiated by a low shear stress force, which did not induce such platelet aggregation in normal individuals. This might explain the intravessel platelet clumping that occurs, resulting in the intermittent thrombocytopenia observed in this type of the disease.

Adolescent↗
PubMed1995

Intraarterial infusion of dibutyryl cyclic adenosine monophosphate plus mitomycin C for unresectable hepatocellular carcinoma: long-term survival and response to tumor growth inhibition.

RATIONALE AND OBJECTIVES: Dibutyryl cyclic adenosine 3',5'-monophosphate (dBcAMP) has the capacity to promote morphologic differentiation and to inhibit tumor growth in vitro, but it has not been well researched in the clinical setting. In this study we examined the effects of intraarterial infusion of dBcAMP plus mitomycin C (MMC) on long-term survival and growth inhibition of tumors. METHODS: Thirty-one previously untreated patients with unresectable hepatocellular carcinoma (HCC) received intraarterial infusion of dBcAMP plus MMC. According to the International Union Against Cancer staging system, three patients had stage T3, eight had stage T4a, and 20 had stage T4b cancer. Growth inhibition was defined as no computed tomography (CT) scan evidence of increase in tumor diameter for at least 6 months after treatment. RESULTS: In all 31 patients with HCC, the cumulative survival rate was 34% at 1 year, 14% at 3 years, and 9% at 5 years. The median survival was 5.0 months, with the longest survival period being 92 months. Among 21 patients in whom a tumor response could be evaluated on the basis of follow-up CT studies, two had complete regression of their primary tumors. Overall, the response rate was 43% (9 of 21). Among the 12 stage T4b patients who had HCC that included the main trunk and its major branches of the portal vein, portal thrombi had disappeared in four (33%). Among eight patients who survived more than 1 year and were evaluated for tumor response using follow-up CT scan studies, six (75%) had growth inhibition of tumor. CONCLUSION: Chemotherapy combined with dBcAMP and MMC showed a favorable response in approximately one third of 31 patients who had unresectable HCC. Because of our results, combined therapy should be strongly considered in the treatment of patients with HCC including occlusion of the main portal vein.

Antibiotics, Antineoplastic↗
PubMed1995

Differential expression of two basement membrane collagen genes, COL4A6 and COL4A5, demonstrated by immunofluorescence staining using peptide-specific monoclonal antibodies.

Genes for the human alpha 5(IV) and alpha 6(IV) collagen chains have a unique arrangement in that they are colocalized on chromosome Xq22 in a head-to-head fashion and appear to share a common bidirectional promoter. In addition we reported a novel observation that the COL4A6 gene is transcribed from two alternative promoters in a tissue-specific manner (Sugimoto, M., T. Oohashi, and Y. Ninomiya. 1994. Proc. Natl. Acad. Sci. USA. 91:11679-11683). To know whether the translation products of both genes are colocalized in various tissues, we raised alpha 5(IV) and alpha 6(IV) chain-specific rat monoclonal antibodies against synthetic peptides reflecting sequences near the carboxy terminus of each noncollagenous (NC)1 domain. By Western blotting alpha 6(IV) chain-specific antibody recognized 27-kD monomers and associated dimers of the human type IV collagen NC1 domain, which is the first demonstration of the presence in tissues of the alpha 6(IV) polypeptide as predicted from its cDNA sequence. Immunofluorescence studies using anti-alpha 6(IV) antibody demonstrated that in human adult kidney the alpha 6(IV) chain was never detected in the glomerular basement membrane, whereas the basement membranes of the Bowman's capsules and distal tubules were positive. The staining pattern of the glomerular basement membrane was quite different from that obtained with the anti-alpha 5(IV) peptide antibody. The alpha 5(IV) and alpha 6(IV) chains were colocalized in the basement membrane in the skin, smooth muscle cells, and adipocytes; however, little if any reaction was seen in basement membranes of cardiac muscles and hepatic sinusoidal endothelial cells. Thus, both genes are expressed in a tissue-specific manner, perhaps due to the unique function of the bidirectional promoter for both genes, which is presumably different from that for COL4A1 and COL4A2.

Adipocytes↗
PubMed1995

Determination of amounts of polyamines excreted in urine: demonstration of N1,N8-diacetylspermidine and N1,N12-diacetylspermine as components commonly occurring in normal human urine.

An analytical system developed for fractionating free and monoacetylated polyamines [Hiramatsu, K. et al. (1994) J. Biochem. 115, 584-589] was proved useful also in detecting diacetylpolyamines, namely N1,N8-diacetylspermidine (diAcSpd) and N1,N12-diacetylspermine (diAcSpm). Detection limits were 0.9 and 0.6 pmol (S/N = 5) for diAcSpd and diAcSpm, respectively. Analytical recovery and within-run variation were also satisfactory. Human urine samples were found to contain diAcSpd and diAcSpm. These polyamines were identified on the basis of the following observations: (i) their retention times were coincident with those of authentic samples; (ii) they were deacetylated to N8-acetylspermidine and monoacetyl- and free spermine, respectively, by acetylpolyamine amidohydrolase; and (iii) they were practically inert to direct oxidation by bacterial polyamine oxidase as were authentic samples. The amounts of eleven polyamine species including diAcSpd and diAcSpm in urine samples from 52 healthy persons were determined. Mean values for the major polyamine components were consistent with those reported by others. Although the amounts of diAcSpd and diAcSpm were very small, comprising only 1.4 and 0.46% of total polyamines, respectively, these two compounds were found to be always present in healthy human urine as regular constituents. Moreover, variation in their content among individuals was small, suggesting that excretion of these components in urine is strictly regulated.

Biogenic Polyamines↗
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

Changes in adrenergic innervation to chromatophores during prolonged background adaptation in the medaka, Oryzias latipes.

The pattern of adrenergic innervation to scale chromatophores of the wild-type medaka, Oryzias latipes, was examined by autoradiography with 3H-norepinephrine and found for the first time to be changed reversibly during prolonged background adaptation. In scales of the medaka, which was adapted to a black background for 10-15 days, a great number of melanophores and dense networks of varicose fibers were observed: many fibers built up a radial plexus around each melanophore. However, the dense distribution of varicose fibers disappeared with a decrease in the number of melanophores during long-term adaptation to a white background. As to the changes in the innervation pattern to amelanotic melanophores of the medaka, orange-red variety, a similar result was obtained. Although the increase in the number of leucophores was observed in the medaka adapted to a white background, no exact plexuses of labeled fibers were confirmed around leucophores. From these results, it is concluded that the density of chromatic nerve fibers changes in parallel with the variation of the number of melanophores during prolonged background adaptation.

Acclimatization↗
PubMed1995

Transforming growth factor-beta 1 coated beta-tricalcium phosphate pellets stimulate healing of experimental bone defects of rat calvariae.

OBJECTIVE: TGF-beta 1-coated beta-TCP pellets were grafted in experimental defects of rat calvariae to study the effects on new bone formation. MATERIALS AND METHODS: The grafted sites were evaluated by light microscopy using hematoxylin-eosin (H-E) staining for histology and detection of alkaline phosphatase (ALPase) and tartrate-resistant acid phosphatase (TRACPase) activities to demonstrate osteoblastic and osteoclastic cells. Confocal laser scanning microscopy (CLSM) was performed for morphometry of newly formed bone. RESULTS: H-E sections showed more new bone formation in sites grafted with TGF-beta 1-coated beta-TCP pellets (experimental sites) than those with beta-TCP pellets only (control sites). TRACPase-positive and ALPase-positive cells at experimental sites were more frequent than at control sites. The bone formation rate calculated by computerized CLSM pixel image analysis showed more new bone formation at the experimental sites than at control sites (3.4 +/- 0.8% vs 9.3 +/- 1.7% on week 2 and 11.8 +/- 2.1% vs 39.8 +/- 10.9% on week 4). CONCLUSION: TGF-beta 1-coated beta-TCP pellets promote new bone formation and may be a useful modality in synthetic bone grafting.

Acid Phosphatase↗
PubMed1995

Chemotactic heterogeneity of eosinophils in idiopathic pulmonary eosinophilia.

Heterogeneity in the chemotactic response of eosinophils to 5 T cell line eosinophilic chemotactic factors (ECFs) was assessed in 5 patients with idiopathic pulmonary eosinophilia. Eosinophils from 2 patients responded to all 5 ECFs (group 1), whereas eosinophils from the other 3 patients responded to ECF-PI 5, PI 6, PI 7 and PI 8 but failed to respond to ECF-PI 9 (group 2). It was further found that group 1 showed an elevated level of lactate dehydrogenase and a positive tuberculin reaction, whereas group 2 showed neither. The effects of steroid therapy on the chemotactic responses of eosinophils were also examined. In group 1, the chemotactic response of eosinophils to ECF-PI 9 was significantly diminished after therapy; in contrast it was elevated in group 2. This change was accompanied by resolution of both clinical symptoms and pulmonary infiltration of eosinophils. These findings suggest that pulmonary eosinophilia can be divided into two types on the basis of eosinophil chemotactic response and laboratory findings. The heterogeneous responses of eosinophils to ECFs may provide a useful marker for classification of pulmonary eosinophilia and evaluation of therapy.

Adult↗
PubMed1995

A small-volume constant-flow air sampling device for thermal desorption GC analysis.

A glass syringe with disposable membrane filters was tested as a constant-flow air sampling device. The principle of this sampling device is straightforward: when the syringe is inverted vertically, the syringe plunger falls at a constant speed if an appropriate flow restrictor is connected at the syringe inlet. In case of a 100 ml syringe with two Teflon membrane filters of 0.50 microns in pore size and 3 mm in diameter, the flow rate was 3 ml/min. The flow rate was constant for a 15-minute sampling period when two to eight filters were used as a flow restrictor. A sample volume of less than 50 ml is sufficient for the determination of organic solvent vapor at the ppm level, when gas chromatography with thermal desorption method is employed. Therefore, this sampling device is useful for determining the time-weighted average concentrations of organic solvent vapor in a working environment.

Air Pollutants↗
PubMed1995

In vivo proviral burden and viral RNA expression in T cell subsets of patients with human T lymphotropic virus type-1-associated myelopathy/tropical spastic paraparesis.

We used in situ hybridization combined with immunocytochemistry, cell sorting, and the polymerase chain reaction (PCR) to investigate clinical events in three asymptomatic carriers of human T lymphotrophic virus type-1 (HTLV-1) and ten patients with HTLV-1-associated myelopathy/tropical spastic paraparesis (HAM/TSP). The objective was to determine which T cell subset of peripheral blood mononuclear cells (PBMC), CD4 or CD8, were infected by HTLV-1 and the manner in which HTLV-1 proviral DNA was expressed at the level of the single cell. Both CD4-positive and CD8-positive cells of the PBMC from five patients with HAM/TSP were infected with HTLV-1. The proportion of HTLV-1-infected cells was 2.5-40% in the CD4-positive subset and 1.0-65% in the CD8- positive subset, when quantified by PCR using HTLV-1-infected MT2 cells as a positive standard. Proviral DNA of HTLV-1 was expressed in both CD4-positive cells and CD8-positive cells of the PBMC from six patients with HAM/TSP and three asymptomatic HTLV-1 carriers. In patients with HAM/TSP, the proportion of the cells expressing HTLV-1 proviral DNA was 0.02-0.1% in both subsets. In asymptomatic carriers, the expression of HTLV-1 proviral DNA was 0.01-0.02% in the CD4-positive subset and 0.01% in the CD8-positive subset. Therefore, HTLV-1 possessed similar in vivo cellular tropism for both CD4-positive cells and CD8-positive cells and HTLV-1 proviral DNA was expressed in vivo in both circulating T cell subsets.(ABSTRACT TRUNCATED AT 250 WORDS)

Adult↗
PubMed1995

[Glutathione S-transferases (GSTs)].

Glutathione S-transferases (GSTs) are a family of multifunctional detoxifying enzymes that catalyse the conjugation of glutathione with large number of compounds bearing an electrophilic center, including carcinogens, and also bind a variety of nonsubstrate ligands. The transferases are widely distributed in the mammalian species and can be grouped into three classes on the basis of subunit composition: alpha (basic), mu (neutral), and pi (acidic). The liver is an organ possessing abundant GST-alpha. GST-mu is also present in the liver and lymphocytes, but this is absent in approximately 50% of the human population. GST-pi (originally found in the placenta) is widely located in the lung, kidney, GI tract, erythrocytes and cancer cells. The present review describes a new nomenclature of human GST isoenzymes and recent investigations of this enzymes, including ours. Implications of each isoenzyme are also discussed.

Animals↗
PubMed1995

[Shear-induced platelet aggregation (SIPA) monitoring of hemostatic effect during platelet transfusion in a patient with Glanzmann's thrombasthenia: a comparison between SIPA and conventional platelet aggregation].

Shear-induced platelet aggregation (SIPA) in a patient with Glanzmann's thrombasthenia was examined during platelet infusion therapy. Prior to platelet infusion, SIPA measured with the modified cone-and-plate type viscometer, as well as ADP-and collagen-induced platelet aggregation measured with the conventional platelet aggregometer, were absent. The patient's SIPA after multiple infusions of platelet, in parallel with the bleeding time and the clinical hemostatic effect, improved to the normal level, while ADP-and collagen-induced platelet aggregation remained abnormal. These observations imply that SIPA is physiologically more relevant than the conventional agonist-induced platelet aggregometry in this type of the disease.

Adenosine Diphosphate↗
PubMed1995

Differential susceptibility of epidermal keratinocytes and neuroblastoma cells to cytotoxicity of ultraviolet-B light irradiation prevented by the oxygen radical-scavenger ascorbate-2-phosphate but not by ascorbate.

Human or mouse epidermal keratinocytes NHEK or Pam212 was less susceptible to ultraviolet (UV)-B irradiation than mouse neuroblastoma NAs1 cells in culture, undergoing apoptosis-like cell death as shown by cell fragmentation and cell membrane integrity disruption. UV susceptibility was appreciably reduced by the reactive oxygen species (ROS)-scavenger L-ascorbic acid-2-phosphate (Asc2P) endowed with long-lasting functions but not by L-ascorbic acid (Asc) for each cell type. DehydroAsc reduced UV susceptibility of Pam212 or NAs1 established cell lines but not of normal diploid NHEK cells destined to be thereafter submitted to cellular senescence. The susceptibility reduction may not be ascribed to extracellular Asc2P or DehAsc, which was removed by aspirating and/or rinsing upon irradiation after the intracellular channelyzer analysis and dead cell-specific DNA-intercalator ethidium homodimer/fluorometry, respectively. Thus, the three cell types differed in UV susceptibility partly because of their different ROS-scavenging abilities, which may be potently promoted by Asc2P or dehydroAsc but not Asc.

Animals↗
PubMed1994

The effects of intraoperative radiotherapy on bone-healing ability in relation to different doses and postradiotherapy intervals.

PURPOSE: Alterations in bone-healing ability after intraoperative radiotherapy were investigated in relation to different doses and postradiotherapy intervals in an experimental model. METHODS AND MATERIALS: Bilateral osteotomy of the fibulae of 56 rabbits was carried out either immediately, 13 weeks or 39 weeks after unilateral intraoperative radiotherapy with 10-MV X ray using one of three doses, 25, 50, or 100 Gy. The healing of the fibulae was evaluated radiographically and histologically 13 weeks after the osteotomy. RESULTS: Bone-healing ability was irreversibly reduced in all irradiated fibulae and these damages occurred in a dose-dependent manner. The severity of the damage also tended to increase with the time interval between irradiation and osteotomy. This interval-related aggravation was characterized by augmented bone loss at the osteotomy ends and atrophy of the neighboring areas. CONCLUSION: These results serve as useful information concerning the surgical intervention of irradiated bone during and after intraoperative radiotherapy, and also after conventional radiotherapy.

Animals↗
PubMed1994

The genes COL4A5 and COL4A6, coding for basement membrane collagen chains alpha 5(IV) and alpha 6(IV), are located head-to-head in close proximity on human chromosome Xq22 and COL4A6 is transcribed from two alternative promoters.

The genes for the alpha 5(IV) and alpha 6(IV) chains of human basement membrane collagen type IV have been found together on chromosome X at segment q22 and have been reported to be arranged in a head-to-head fashion. Here we report the 5' flanking sequences of COL4A5 and COL4A6 and that COL4A6 is transcribed from two alternative promoters in a tissue-specific fashion. Analysis of the sequence immediately upstream of the transcription start sites revealed some features of housekeeping genes--i.e., the lack of a TATA motif and the presence of CCAAT and CTC boxes. Further analysis revealed that COL4A6 contains two alternative promoters that control the generation of two different transcripts. One transcription start site (from exon 1') is 442 bp away from the transcription start site of COL4A5, while an alternative transcription start site (from exon 1) is located 1050 bp from the first one and drives the expression of a second transcript that encodes an alpha 6(IV) chain with a different signal peptide. Reverse transcription-PCR experiments revealed that the transcript from exon 1' is abundant in placenta, whereas the transcript from exon 1 is more frequently found in kidney and lung. These results provide additional clues to answering the general question of what mechanisms are used to generate unique basement membrane structures in different tissues.

Base Sequence↗