Biomedical subjects

T Imanari

Publications and source records attributed to T Imanari.

At least 55 records · Page 3Linked to original sources

PubMed1997

Chemical microdetermination of heparin in plasma.

A new, simple, and highly sensitive method for the determination of heparin has been established. Heparin was first converted into unsaturated disaccharides through the action of heparin lyases I, II, and III. A major trisulfated unsaturated disaccharide product results, consistent with structural analysis of a number of pharmaceutical heparins using one- and two-dimensional 1H NMR spectroscopy. This disaccharide was analyzed by HPLC using fluorometric postcolumn derivatization. The correlation between the amount of this trisulfated unsaturated disaccharide and anticoagulant activity of heparin as measured by anti-IIa was determined. The analysis of these pharmaceutical heparins showed a linear correlation between both HPLC and bioassay. This HPLC method was then applied to a pharmacokinetic study of heparin intravenously administered to rabbits.

Animals↗

PubMed1997

Structural differences and the presence of unsubstituted amino groups in heparan sulphates from different tissues and species.

This study presents a comparison of heparan sulphate chains isolated from various porcine and bovine tissues. 1H-NMR spectroscopy (500 MHz) was applied for structural and compositional studies on intact heparan sulphate chains. After enzymic digestion of heparan sulphate using heparin lyase I (EC 4.2.2.7) II and III (EC 4.2.2.8), the compositions of unsaturated disaccharides obtained were determined by analytical capillary electrophoresis. Correlations between the N-sulphated glucosamine residues and O-sulphation and between iduronic acid content and total sulphation were discovered using the data obtained by NMR and disaccharide analysis. Heparan sulphate chains could be classified into two groups based on the sulphation degree and the iduronic acid content. Heparan sulphate chains with a high degree of sulphation possessed also a significant number of iduronic acid residues and were isolated exclusively from porcine brain, liver and kidney medulla. The presence and amount of N-unsubstituted glucosamine residues (GlcNp) was established in all of the heparan sulphates examined. The structural context in which this residue occurs was demonstrated to be: high sulphation domain --> 4)-beta-D-GlcAp-(1 --> 4)-alpha-D-GlcNp-(1 --> 4)-beta-D-GlcAp-(1 --> low sulphation domain (where GlcNp is 2-amino-2-deoxyglucopyranose, and GlcAp is glucopyranosyluronic acid), based on the isolation and characterization of a novel, heparin lyase III-derived, GlcNp containing tetrasaccharide and hexasaccharide. The results presented suggest that structural differences may play a role in important biological events controlled by heparan sulphate in different tissues.

Amino Sugars↗

PubMed1997

Measurement of ascorbate and dehydroascorbate contents in biological fluids.

Instabilities of ascorbate and dehydroascorbate throughout sample processing are clearly a significant aspect of quantifying of them. Contents of ascorbate in biological fluids decrease with measurable oxidation occurring within minutes to hours. Similarly, dehydroascorbate disappears with chemical or enzymatic degradation within minutes. The half-life of dehydroascorbate in human heparinized plasma was approximately 2 min. These results indicated that the amount of dehydroascorbate present in sample solutions is a function of both the oxidation of ascorbate and the degradation of dehydroascorbate during the processing of biological fluids. To quantify ascorbate and dehydroascorbate concentrations in biological fluids including circulating blood plasma and urine, we established a high-performance liquid chromatographic method, which requires no pretreatment of sample solutions.

Ascorbic Acid↗

PubMed1997

Structural diversity of dermatan sulphate in porcine dermis.

The structural diversity of dermatan sulphate (DS), the major glycosaminoglycan component of mammalian skin, was investigated by examining different layers of porcine dermal tissue using 1H-NMR and disaccharide compositional analysis by HPLC. Structural reporter signals were assigned using one-dimensional (1D) 1H-NMR differential transient NOE and 1D totally correlated spectroscopy (TOCSY) spectra, measured at different probe temperatures. The results of these studies on 12 sliced layers (average thickness of 250 microns) of skin show that the content of glucuronic acid in DS decreases when moving from the outer surface of the skin to the inside, while the degree of sulfation of the C-2 hydroxy group of iduronate and the C-4 and C-6 hydroxy groups of N-acetylgalactosamine increases with depth. These results suggest that the utility of analysis of DS from various depths in porcine skin clearly show the origin of each sample, and might be useful for the quality control of these biological materials in clinical use.

Animals↗

PubMed1997

[Activation of skin cells by inorganic compounds].

It has been known that the stimulation of skin cells by physiologically active substances is accompanied by the quantitative and qualitative alterations of the glycosaminoglycans (GAGs) in skin. This phenomenon make it possible to evaluate the activity of test substances for the stimulation of skin cells. The purpose of our study is to elucidate the effectiveness of inorganic compounds for the stimulation of skin cells. (1) The cultured skin cells including keratinocytes and fibroblasts, (2) the cultured model skin constituted of collagen gel, fibroblasts and keratinocytes, and (3) the mouse damaged skin were used in the present study. The results obtained from these studies demonstrate that inorganic substances commonly have a possibility to activate cyclic AMP-dependent protein kinase (A-kinase), and the A-kinase activation results in an increase in GAGs. These findings brought the scientific basis for the effectiveness of the Japanese traditional balneotherapy for damaged skin.

Animals↗

PubMed1996

Simultaneous determination of dermatan sulfate and oversulfated dermatan sulfate in plasma by high-performance liquid chromatography with postcolumn fluorescence derivatization.

A chemical method for the determination of dermatan sulfate (DS) and oversulfated dermatan sulfate has been developed and applied to the pharmacokinetic study of these polysaccharides in experimental animals. The analytical procedure includes a simple preparation step of administered DS and oversulfated DS from blood plasma, HPLC for the separation and detection of DS and oversulfated DS using an Asahipak NH2P-50 column, fluorometric reaction of the polysaccharides with guanidine in a strong alkaline medium. DS and oversulfated DS were extracted from plasma by treating it with proteinase to remove plasma proteins and recovered with endogenous plasma glycosaminoglycans by ethanol precipitation. Finally, DS and oversulfated DS were analyzed by fluorometric HPLC. The detection limits of DS and oversulfated DS were 10 and 20 ng, respectively. Furthermore, we demonstrated that artificial oversulfation of DS increased its biological half-life after intravenous administration to rats.

Animals↗

PubMed1996

High-performance liquid chromatographic analysis of glycosaminoglycan-derived oligosaccharides.

High-performance liquid chromatography of glycosaminoglycan (GAG)-derived oligosaccharides has been employed for the structural analysis and measurement of hyaluronan, chondroitin sulphate, dermatan sulphate, keratan sulphate, heparan sulphate and heparin. Recent developments in the separation and detection of unsaturated disaccharides and oligosaccharides derived from GAGs by enzymatic or chemical degradation are reviewed.

Carbohydrate Sequence↗

PubMed1996

Asymptomatic colorectal cancer detected by screening.

PURPOSE: Colorectal cancer screening has become prevalent. To discuss the efficacy of screening, we studied the characteristic of asymptomatic colorectal cancer detected by screening. METHODS: This is a retrospective review of patients with colorectal cancer treated at our institution. During the past 20 years, 96 of 1,046 cases of colorectal cancer were asymptomatic and detected by screening. Sixty-one of these cases were detected in the recent five years. The initial screening procedures were fecal occult blood test in 51 cases, sigmoidoscopy or colonoscopy in 18, barium enema in 9, and other tests in 18. RESULTS: Thirteen lesions (14 percent) were smaller than 1.0 cm and 32 (33 percent) were 1-2 cm in size. There were 34 Tis, 21 T1, and 8 T2 tumors. Of the 55 Tis or T1 lesions, 14 showed nonpolypoid growth (5 flat-elevated, 7 flat-elevated with depression, 1 flat, 1 depressed), and 12 of these were detected on endoscopy. Thirty-four cases were TNM Stage 0, 25 were Stage I, 16 were Stage II, 12 were Stage III, and 9 were Stage IV. Sixty-one percent of those detected by screening were in either Stage 0 or Stage I compared with 16 percent in the symptomatic group. Cumulative five-year disease-free survival rates were 100 percent for both Stage 0 and Stage I, 94 percent for Stage II, and 52 percent for Stage III. Overall cumulative five-year survival rate was 87 percent for those detected by screening, compared with 57 percent in symptomatic patients. CONCLUSIONS: Asymptomatic cancers detected by screening were at a less advanced stage. In particular, many nonpolypoid early cancers were detected by endoscopic screening.

Aged↗

PubMed1996

Compositional analysis of hyaluronan, chondroitin sulfate and dermatan sulfate: HPLC of disaccharides produced from the glycosaminoglycans by solvolysis.

An ion-exchange high-performance liquid chromatography procedure was developed for analysis of mixtures of N-acetyldermosine, N-acetylchondrosine and N-acetylhyalobiuronic acid produced quantitatively by heating dermatan sulfate, chondroitin sulfate and hyaluronan in dimethyl sulfoxide (DMSO) containing 0.1% (v/v) H20 at 80 degrees C for 48 h. These disaccharides were eluted from a TSK gel SAX column using 0.1 M acetic acid containing KCl, and detected fluorometrically by post column derivatization. The eluate was mixed with 2-cyanoacetamide solution and alkaline solution, and heated at 110 degrees C for 4 min. The resultant compounds were detected fluorometrically (Ex. 335 nm and Em. 390 nm). The usefulness and practicality of the present method were verified by applications to the determination of glycosaminoglycans in tissues.

Animals↗

PubMed1995

Determination of a depolymerized holothurian glycosaminoglycan in plasma after intravenous administration by postcolumn HPLC.

Depolymerized holothurian glycosaminoglycan (DHG) produced artificially from sea cucumber has an anticoagulant and antithrombotic activity. In the present study, we attempted to determine the plasma level of DHG using a chemical procedure. A general method for determination of chondroitin sulfates by forming unsaturated disaccharides with chondroitinase digestion was difficult to apply to DHG, because it was resistant to any chondroitinase digestion. We therefore developed a highly sensitive postcolumn HPLC method for determination of intact DHG. DHG was applied to Asahipak NH2P-50, an amino-bonded column, eluted by alkaline solution and then detected with arginine under a strong alkaline condition as a postcolumn reagent. The limit of detection for DHG was 10 ng. The present method was applicable to the determination of DHG in plasma after intravenous administration.

Animals↗

PubMed1995

[Continuous intraarterial infusion of 5-fluorouracil plus leucovorin for liver metastases from colorectal cancer].

Between September 1990 and August 1994, 11 patients (pts) with liver metastases (mets) from colorectal cancer were treated with continuous hepatic arterial infusion chemotherapy of 5-fluorouracil (FU) plus leucovorin (LV). Eight pts had non-resectable liver mets (H3: 7, H2: 1), and 3 had residual small mets after resection of major mets. Drugs were administered via an extracorporeal infusion device connected to the hepatic arterial infusion port. 5-FU and LV were given through a 5- to 7-day continuous infusion at 500-750 mg/body/day and 30 mg/body/day, respectively, with a 3- to 4-week rest period. In the recent 6 pts, cisplatin was administered as a 2-hour infusion at 25 mg/body, one or two times simultaneously. Grade 2 toxicity was noted in two pts (18%). One was stomatitis and another was uncontrolled ascites in an advanced cirrhotic pt. The response rate in the 9 evaluable pts was 67% with 6 PR and no CR. The duration of the response was 5 to 9 months. One- and two-year survival rates were 75% and 22%, respectively. These results were superior to those of the intermittent bolus injection of 5-FU plus MMC (or epirubicin) in 40 pts from 1977 to 1994. These results suggest that continuous 5-FU plus LV arterial infusion is an effective regimen in pts with liver metastases from colorectal cancer. However, the infusion with an extracorporeal device limits the pts' quality of life. Further investigation is needed for a schedule that can be practiced for a longer period.

Aged↗

PubMed1994

Time-course experiment involving glycosaminoglycan synthesis by normal human dermal fibroblasts cultured in the presence of dibutyryl cyclic AMP and retinoic acid.

Glycosaminoglycan (GAG) synthesis by cultured normal human dermal fibroblasts was examined. Hyaluronic acid (HA) synthesis reached a maximum on day 3 (0.3 microgram/ml medium) and then decreased to a low level (0.15 microgram/ml medium). The amounts of dermatan sulfate (DS) and chondroitin sulfate (ChS) synthesized by the cells increased with increasing cell numbers during the initial stage to attain constant levels (0.092 microgram DS/ml, 0.026 microgram ChS/ml medium) after the cells reached confluence. We also tested the effects of dibutyryl cyclic AMP (dbcAMP) and retinoic acid (RA) on GAG synthesis by the cells. The synthesis of HA and ChS by cells was stimulated, when the cells were cultured in medium containing dbcAMP (0.1 mM), whereas DS synthesis was scarcely affected. However, addition of RA (5 mM) suppressed GAG synthesis by the cells.

Bucladesine↗

PubMed1994

Analytical studies on hyaluronic acid synthesis by normal human epidermal keratinocytes cultured in a serum-free medium.

Hyaluronic acid (HA) synthesized by normal human epidermal keratinocytes cultured in a serum-free medium was monitored by a highly sensitive HPLC method, which was established by us for the simultaneous determination of HA, chondroitin sulfate (ChS) and dermatan sulfate (DS) as their unsaturated disaccharides. The major glycosaminoglycan (GAG) in the medium of the keratinocytes was HA, and the ability of the cells to synthesize HA increased relatively with an increase in cell numbers during the logarithmic phase and reached a maximum level after the cells became confluent. HA synthesis by the keratinocytes was inhibited by the addition of calcium chloride to the culture medium, and was strongly stimulated by the addition of retinoic acid (RA), respectively. It was shown that the ability of the cells to synthesize HA exists in the spinous cell stage. Furthermore, we found that HA synthesis by the cells was slightly increased by the addition of dibutyryl cyclic AMP (dbcAMP). Our results indicate that the measurement of time-course levels of HA in the culture medium is useful for the screening of active substances for proliferation and differentiation of the keratinocytes.

Bucladesine↗

PubMed1994

[5-Fluorouracil plus low-dose leucovorin in the treatment of advanced colorectal cancer].

From September 1989 to September 1992, 17 patients (pts) with non-curative or recurrent colorectal cancer were treated with 5-fluorouracil (FU) plus leucovorin (LV) systemic therapy. The sites of evaluable metastases were liver (10), peritoneum (4), lung (3) and others (6). The LV dosage was 30 mg/body (low-dose method) and the FU dosage was 500-750 mg/body. Both drugs were administered either 5 days/every 4 weeks or one day/every 1 to 2 weeks via protracted or bolus infusion. The regimen was generally well tolerated, although 9 pts (53%) experienced toxicity which required dose reductions. Overall response rate was 29% (5 PR). No CR was observed. Improvement in tumor-related symptoms was noted temporarily in 7 out of 10 pts. Serum CEA level decreased in 13 out of 14 pts. All the response sites were liver (5), with simultaneous lung (1) or peritoneum (1) metastasis. The response duration was 3 to 10 months (mean 6.2 months). Second-line therapy after progression of diseases, such as combination with CDDP, was attempted in some of the PR pts, but no re-response was observed. We conclude that FU plus low-dose LV regimen is an effective therapy for advanced colorectal cancer, but further attempts should be made to increase response rate, prolongation of response duration and effective second-line therapy after progression.

Antineoplastic Combined Chemotherapy Protocols↗

PubMed1993

Analysis of chondroitin sulfate/dermatan sulfate chains in rat peritoneal resident macrophages.

Chondroitin sulfate/dermatan sulfate (CS/DS) hybrid polymers obtained from rat peritoneal resident macrophages were analyzed by the enzymatic method; CS/DS chains were digested by chondroitinase, and the resultant unsaturated disaccharides were determined by HPLC with fluorometric detection. CS/DS in the resident macrophages are constituted of Di-4S, Di-6S, Di-diSE, Di-diSB and Di-triS.

Animals↗

PubMed1993

Structural diversity of mammalian hepatic dermatan sulfates.

Dermatan sulfate-proteoglycans (DS-PGs) were extracted from rabbit, rat and bovine defatted livers by magnesium chloride extraction and DEAE-cellulose chromatography, and then submitted successively to Asahipak GS-520 gel filtration chromatography, Asahipak ES-502N anion exchange chromatography, and cellulose acetate membrane electrophoresis. The disaccharide composition of the glycosaminoglycan chains was determined by differential digestion by chondroitinase ABC, AC, ACII and/or B followed by HPLC for analysis of the resulting unsaturated disaccharides. The hepatic dermatan sulfate chains contained disulfated disaccharide units; Di-diSB and Di-diSE. The hepatic DS-PGs were divided into two groups; Di-diSE-poor DS-PGs and Di-diSE-rich DS-PGs. The iduronic acid content of Di-diSE-poor dermatan sulfate chains was higher than that of Di-diSE-rich ones.

Animals↗

PubMed1993

Separation and characterization of dermatan sulfate in normal human urine.

Dermatan sulfate excreted in normal human urine was isolated and characterized by TLC and cellulose acetate strip electrophoresis after cetylpyridinium chloride precipitation and pronase digestion. In these separation methods, dermatan sulfate and chondroitin sulfate were extracted and then monitored by sensitive HPLC methods with post column fluorometric derivatization coupled with chondroitinase ABC, ACII and B digestion. From the results, we demonstrated that human urinary dermatan sulfate contains iduronic acid as its major uronic acid (80-90% of total uronic acid), and is composed mainly of repeated mono-sulfated disaccharide units [Di-4S (structure shown in Fig. 1), 89%] and small numbers of di-sulfated disaccharide units (Di-diSB, 7% and Di-diSE, 1%).

Chondroitin Sulfates↗

PubMed1993

Structural analysis of a low-sulfated chondroitin sulfate chain in human urinary trypsin inhibitor.

The low-sulfated chondroitin 4-sulfate(LSC) chain from human urinary trypsin inhibitor was purified and the structure was characterized. After hyaluronidase SD digestion of LSC, an oligosaccharide which contains the linkage region could be obtained. The structure of oligosaccharide was analyzed by HPLC and 500 MHz 1H-NMR spectroscopy. The analytical results revealed that 4-O-sulfo GalNAc residues were located in the neighborhood of the linkage region.

Carbohydrate Conformation↗