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PubMed1976

The heterogeneity of dermatan sulfate and heparan sulfate in rat liver and a shift in the glycosaminoglycan contents in carbon tetrachloride-damaged liver.

The glycosaminoglycan of rat liver can be separated into five distinct fractions; a hyaluronic acid fraction, a heparan sulfate fraction with a molar ratio of sulfate to hexosamine (S/HexN) around 0.7, a heparan sulfate fraction with a S/HexN ratio around 1.4, a dermatan sulfate fraction with a S/HexN ratio near unity, and a dermatan sulfate fraction with a S/HexN ratio around 1.3. Enzymatic analysis of the two dermatan sulfate fractions indicates that they differ significantly in that the high sulfated fraction contains relatively more N-acetylgalactosamine 4,6-bissulfate units (about 26% of the total hexosamine). In experimental injury produced by carbon tetrachloride, the low sulfated fraction increases as much as 9-fold on a dry weight basis, bearing no linear relationship to the amount of the high sulfated fraction which increases only 2-fold. A significant shift is also observed in the levels of the two heparan sulfate fractions. In this case, however, the high sulfated fraction shows a much more pronounced increase than does the low sulfated fraction. On the basis of these observations, it is suggested that for each of the dermatan sulfate and heparan sulfate classes there are at least two pools, distinguished by sulfation degree and perhaps by turnover rate and physiological function.

Animals↗
PubMed1976

Stimulatory effect of bleomycin on the synthesis of acidic glycosaminoglycans in cultured fibroblasts derived from rat carrageenin granuloma.

Cultured fibroblasts derived from rat carrageenin granuloma were treated with bleomycin and the synthesis of hexosamine-containing substances was compared with that in control cells. Four day treatment with o.1 mug bleomycin/ml resulted in a significant increase of the production of these macromolecules by the cells, though DNA synthesis was remarkably inhibited at this dose of bleomycin. The stimulatory effect could be seen as early as the second day of bleomycin treatment, and was enhanced with increasing treatment time. Further fractionation of the hexosamine-containing substances revealed that synthesis of acidic glycosaminoglycans was more sensitive to bleomycin than that of glycoproteins, i.e., acidic glycosaminoglycans increased by 80% and glycoproteins by 53% after four day treatment with 0.1 mug bleomycin/ml. The increased components of acidic glycosaminoglycans included not only hyaluronic acid but also sulphated glycosaminoglycans. Collagen synthesis was increased by 23% by the same dose of bleomycin. N-Acetyl-beta-glucosaminidase, one of the degradation enzymes for acidic glycosaminoglycans released into the cultured medium, was decreased significantly by bleomycin.

Acetylglucosaminidase↗
PubMed1985

The proteoglycans of the canine intervertebral disc.

The high-buoyant-density proteoglycans of the nucleus pulposus and annulus fibrosus of the beagle intervertebral disc have been isolated by CsCl density gradient ultracentrifugation. The sulphated proteoglycans were labelled in vivo with 35SO4, 24 h and 60 days prior to killing. The hydrodynamic size and aggregation of the 24 h, 60 day and resident (from hexuronic acid and hexosamine analysis) proteoglycan subunit populations were determined by Sepharose CL-2B chromatography in the presence or absence of excess hyaluronic acid. The hydrodynamic size of the keratan sulphate-proteoglycan core protein complexes were also determined by Sepharose CL-2B chromatography after chondroitinase ABC digestion of proteoglycans. When initially synthesised (24 h) or after 60 days, the percentage aggregation and hydrodynamic size of the proteoglycans derived from the annulus fibrosus were larger than those present in the nucleus pulposus. Hexosamine, hexuronic and protein determination of the high-buoyant-density fractions showed that the proteoglycans of the nucleus pulposus were richer in chondroitin sulphate than those in the annulus. However there was no difference in Mr of the chondroitin sulphate and keratan sulphate attached to the proteoglycans of the two disc regions, nor were differences detected by HPLC between the proportions of chondroitin 4-sulphate and chondroitin 6-sulphate present in these high-density fractions. In contrast, the low-buoyant-density (1.54 greater than p greater than 1.45) proteoglycan fractions and tissue residues remaining after 4 M GuHCl extraction were found to contain dermatan sulphate, suggesting the presence of a third proteoglycan species possibly associated with the collagen of the fibrocartilagenous matrix.

Animals↗
PubMed1977

Phosphonoglycoprotein from Metridium senile--heterogeneity of glycoproteins containing aminoethylphosphonic acid.

1. After separation by SDS gel-chromatography, analysis of AEP-containing glycoproteins from M. senile, indicated 66% amino acids with 220 AEP res./1000 res. and 30% carbohydrate for high mol. wt (greater than 10(7) forms and 80% amino acids with 25-50 AEP res./1000 res. and 10% carbohydrate for low mol. wt (2-4 x 10(4) forms. 2. Uronic acids, sulfate, lipid, and sialic acids were absent. 3. Mild base digestion released AEP-hexosamine containing oligosaccharides and destroyed ser-thr residues in the high mol. wt components. 4. Phosphonoglycoproteins appear to be acidic connective tissue components with AEP linked to hexosamine containing oligosaccharide side chains.

Amino Acids↗
PubMed2007

Ascorbic acid increases healing of excision wounds of mice whole body exposed to different doses of gamma-radiation.

Because of the practical importance of acute radiation exposure associated with combined injuries, it is imperative to investigate the efficacy of cost-effective nutritional factors in the reconstruction of irradiated wounds. Therefore, effect of pretreatment of ascorbic acid was studied on the healing of excised wounds in mice exposed to 2, 4, 6 and 8 Gy whole body gamma-radiation. A full-thickness wound was created on the dorsum of the irradiated mice and the progression of wound contraction was monitored by capturing video images of the wound at various varying days after irradiation. Irradiation caused a dose dependent delay in wound contraction and wound healing time, while ascorbic acid pretreatment resulted in a significant acceleration in the rate of wound contraction and a decrease in the mean wound healing time. To understand the mechanism of healing, collagen, hexosamine, DNA, nitrite and nitrate contents were measured in the granulation tissue of wounded mice treated with ascorbic acid before exposure to 6 Gy gamma-radiation. Ascorbic acid treatment prior to irradiation enhanced the synthesis of collagen, hexosamine, DNA, nitrite and nitrate contents. The histological assessment of wound biopsy revealed an improved collagen deposition, and increase in fibroblast and vascular densities. The present study demonstrates that ascorbic acid pretreatment has a beneficial effect on the irradiated wound and could be a substantial therapeutic strategy to accelerate wound repair in irradiated wounds and in the cases of combined injury situations.

Animals↗
PubMed2005

Mosquito glucosamine-6-phosphate N-acetyltransferase: cDNA, gene structure and enzyme kinetics.

Mosquito midgut epithelial cells secrete digestive enzymes as well as components of the peritrophic matrix in response to blood-feeding. The peritrophic matrix is composed of proteins, glycoproteins and chitin fibrils in a proteoglycan matrix and may function to protect the midgut epithelium from mechanical damage and insult from pathogens and toxins. Chitin biosynthesis takes place via the hexosamine pathway converting fructose-6-phosphate to UDP-N-acetylglucosamine, which is then polymerized to chitin by chitin synthase. Glucosamine-6-phosphate N-acetyltransferase (GNA) is one of the hexosamine pathway enzymes and catalyzes the transfer of the acetyl group from acetyl-CoA to the primary amine of glucosamine-6-phosphate. We cloned and sequenced the GNA cDNA, gene (AeGna) and its putative promoter regions from Aedes aegypti. AeGna consists of five exons and four introns and lacks a TATA box near the transcription start site. The AeGna cDNA is 1.3 kb in length and the predicted protein is approximately 23.6 kDa. The amino acid sequence of AeGna has high homology to its orthologues. AeGna mRNA is constitutively expressed in all developmental stages and blood-feeding causes no obvious effect on levels of AeGna transcript in the midgut. The Km value of recombinant GNA for glucosamine-6-phosphate was 330 microM and the Km for acetyl-CoA was 500 microM.

Acetyltransferases↗
PubMed2005

Effect of procyanidolic oligomers of Vitis vinifera on the biosynthesis and excretion of corneal glycosaminoglycans.

Glycosaminoglycans (GAG) as hyaluronan and proteoglycans (PG) as dermatan sulfate (DSPG) and keratan sulfate (KSPG) play important role in the structure and physiological functions of cornea. Several corneal pathologies are associated with qualitative and quantitative alterations of GAG and/or PG biosynthesis. For these reasons we investigated the effect of procyanidolic oligomers (PCO) on the biosynthesis and excretion of GAG-s in bovine corneal explant cultures. This was accomplished by the determination of their chemical components, hexoses, hexosamines and uronic acids as well as the incorporation of (35)S-sulfate in the presence and without PCO in the culture medium. In presence of 1 mg/ml PCO total hexoses, hexosamines and uronic acids increased at 5 and 24 h of incubation, hexoses less than the two other components. Stimulation of biosynthesis concerned essentially the uronic acid containing GAG-s, DS and hyaluronan. (35)S-sulfate incorporation decreased in presence of PCO, showing a decreased sulfation of GAG-s. The most striking effect was however the strongly increased excretion of neosynthesized GAG-s in the culture medium. This effect could be beneficial in decreasing corneal hydration in inflammatory conditions, together with the previously demonstrated protection of corneal macromolecules against proteolytic degradation.

Animals↗
PubMedDate not supplied

Salvage pathways in glycosphingolipid metabolism.

In this review, the focus is on the role of salvage pathways in glycosphingolipid, particularly, ganglioside metabolism. Ganglioside de novo biosynthesis, that begins with the formation of ceramide and continues with the sequential glycosylation steps producing the oligosaccharide moieties, is briefly outlined in its enzymological and cell-topological aspects. Neo-synthesized gangliosides are delivered to the plasma membrane, where their oligosaccharide chains protrude toward the cell exterior. The metabolic fate of gangliosides after internalization via endocytosis is then described, illustrating: (a) the direct recycling of gangliosides to the plasma membrane through vesicles gemmated from sorting endosomes; (b) the sorting through endosomal vesicles to the Golgi apparatus where additional glycosylations may take place; and (c) the channelling to the endosomal/lysosomal system, where complete degradation occurs with formation of the individual sugar (glucose, galactose, hexosamine, sialic acid) and lipid (ceramide, sphingosine, fatty acid) components of gangliosides. The in vivo and in vitro evidence concerning the metabolic recycling of these components is examined in detail. The notion arises that these salvage pathways, leading to the formation of gangliosides and other glycosphingolipids, sphingomyelin, glycoproteins and glycosaminoglycans, represent an important saving of energy in the cell economy and constitute a relevant event in overall ganglioside (or glycosphingolipid, in general) turnover, covering from 50% to 90% of it, depending on the cell line and stage of cell life. Sialic acid is the moiety most actively recycled for metabolic purposes, followed by sphingosine, hexosamine, galactose and fatty acid. Finally, the importance of salvage processes in controlling the active concentrations of ceramide and sphingosine, known to carry peculiar bioregulatory/signalling properties, is discussed.

Animals↗
PubMed1985

Biological characterization of human bone tumors. IV. Combined biochemical and histological analyses of different osteosarcomas.

A new technique was applied to the study of human osteosarcoma. Ten slices of 10 micron were cut serially from 2 X 2 X 6 mm shock frozen blocks of human osteosarcoma for chemical analysis. Before and after each series of 10 slices, one slice of 10 micron was separated for morphological analysis. Four different types of osteosarcoma were investigated: Case 1 was an atypical osteoblastic osteosarcoma, case 2 a small cell sclerosing osteosarcoma, case 3 a well-differentiated parosteal osteosarcoma grade I, and case 4 a highly malignant anaplastic osteosarcoma. Alkaline phosphatase, acid phosphatase, beta-glucuronidase and proteolytic activities were analysed as well as matrix collagen and hexosamine, phosphorus (Pi and Po), protein, DNA, and water content. In accordance with the morphology, the obtained data illustrate the great heterogeneity of osteosarcomas. Although case 1, 2 and 3 all represent calcifying types of the tumor, characteristic differences exist with regard to the matrix and the degree of calcification. In contrast to these three, case 4 presents a noncalcified type of osteosarcoma whose matrix contains relatively high amounts of hexosamine and low amounts of collagen, whereas DNA and water contents are high. The data from the analysis of osteosarcoma were compared with previous results from the calf epiphyseal growth plate in order to define differences and similarities between the formation of tumor bone and the physiological formation of hard tissue.

Acid Phosphatase↗
PubMed2000

The wound healing properties of Channa striatus-cetrimide cream-wound contraction and glycosaminoglycan measurement.

Haruan has been proved to influence the different phases of wound healing process. The current research focuses on the effects of haruan on the different constituents of extracellular matrix of healing wounds in normal and diabetic rats. Anaesthetized normal and streptozotocin induced diabetic rats were provided with excision wounds at the back and then animals were divided into four groups as: group 1, wounds treated with cetrimide+haruan cream; group 2, wounds treated with haruan cream; group 3, wounds treated with cetrimide (commercial) cream; and group 4, wounds untreated and served as control. Animals were sacrificed after 3, 6, 9 and 12 days. These wounds were used to determine the hexosamine, protein, uronic acid and glycosaminoglycan contents and the wound contraction. The results suggested a marked increase (P<0.05) in the uronic acid, hexosamine and dermatan sulfate contents on day 3 of group 1 when compared with groups 2-4. Wound contraction of group 1 was also markedly enhanced of group 1 (P<0.01) when compared with groups 2- 4. On the basis of these results, we conclude that haruan enhances the synthesis of different glycosaminoglycans in healing wounds, which are the first component of extracellular matrix to be synthesized during the wound healing process. The enhanced levels of glycosaminoglycans may help in the formation of a resistant scar and enhanced wound contraction represents the positive influence of haruan on the fibroplastic phase of wound healing.

Animals↗
PubMed2002

Hypolipidemic effect of glycosaminoglycans from the sea cucumber Metriatyla scabra in rats fed a cholesterol-supplemented diet.

The purpose of this study was to determine the hypolipidemic effects of the glycosaminoglycans (GAGs) extracted from a sea cucumber, Metriatyla scabra. Using DEAE column chromatography, two major peaks containing GAGs were obtained: peak 1 (P-1) contained mainly GAGs (as hexuronic acid and hexosamine), whereas P-2 contained mostly free glycan (as fucose) with little hexuronic acid or hexosamine. Therefore, we used only the P-1 fraction (with molecular weights in the range 200-500 kDa) for evaluation of hypolipidemic effects. The lyophilized GAGs were administered orally to male Wistar rats at 5, 10, 20, and 50 mg/kg body weight for six consecutive weeks, during which the rats were fed ad libitum a basal laboratory diet with or without 1% cholesterol. The results show that the 1% cholesterol diet significantly increased plasma total cholesterol, LDL-cholesterol, and atherogenic index. Cholesterol supplementation also significantly increased hepatic TG, cholesterol, phospholipid, and liver weight. When rats fed the 1% cholesterol diet were supplemented orally with the sea cucumber GAGs, plasma levels of total cholesterol, LDL-cholesterol, and atherogenic index were significantly decreased, while HDL-cholesterol was significantly increased, although these effects of the GAGs were only dose-dependent at doses lower than 20 mg/kg b.w. Similarly, the GAGs significantly prevented the increase (p < 0.05) in hepatic contents of triglyceride, cholesterol, and phospholipid. Thus, the present study demonstrates that the sea cucumber GAGs have the potential of being used for reducing the risk of atherosclerosis and hyperlipoproteinemia.

Animals↗
PubMed1998

The salubrious effect of tamoxifen [correction of Tamaxifen] on serum marker enzymes, glycoproteins, and lysosomal enzymes level in breast cancer woman.

Tumour markers correlate strongly with prognosis based on tumour burden and surgical resectability. If chemotherapy is extremely effective in certain stage of the disease, the sensitive marker may be of great use in monitoring disease response and drug treatment. Hence, this study was launched to evaluate the changes in tumour marker enzymes like lactate dehydrogenase (LDH), glutamate oxaloacetate transaminase (SGOT), glutamate pyruvate transaminase (SGPT), alkaline phosphatase, and acid phosphatase in before and after 3 and 6 months tamoxifen treated breast cancer patients. In addition, the changes in serum glycoproteins viz., hexose, hexosamine, and sialic acid and lysosomal enzymes such as N-acetyl-beta-D-glucosaminidase, beta-D-galactosidase, and beta-D-glucuronidase were analysed in these patients. These values were compared with their age matched healthy control subjects. At 6 months evaluation, the tamoxifen treated postmenopausal breast cancer women showed a statistically significant decreased (p < 0.001, 0.05 respectively) levels of LDH, SGOT, SGPT, alkaline and acid phosphatases than their baseline values. Similarly, the levels of hexose, hexosamine, and sialic acid and N-acetyl-beta-D-glucosaminidase, beta-D-galactosidase, and beta-D-glucuronidase were decreased significantly (p < 0.001) in tamoxifen received postmenopausal women. The result of this study suggested that tamoxifen potentially retard the metastasis of breast cancer as well as the bone demineralisation in postmenopausal breast cancer women. Thus, tamoxifen may also have its antitumour activity through its beneficial effects on tumour marker enzymes and serum proteins in breast cancer women.

Adult↗
PubMed1999

New insights on the specificity of heparin and heparan sulfate lyases from Flavobacterium heparinum revealed by the use of synthetic derivatives of K5 polysaccharide from E. coli and 2-O-desulfated heparin.

The capsular polysaccharide from E. Coli, strain K5 composed of ...-->4)beta-D-GlcA(1-->4)alpha-D-GlcNAc(1-->4)beta-D-GlcA (1-->..., chemically modified K5 polysaccharides, bearing sulfates at C-2 and C-6 of the hexosamine moiety and at the C-2 of the glucuronic acid residues as well as 2-O desulfated heparin were used as substrates to study the specificity of heparitinases I and II and heparinase from Flavobacterium heparinum. The natural K5 polysaccharide was susceptible only to heparitinase I forming deltaU-GlcNAc. N-deacetylated, N-sulfated K5 became susceptible to both heparitinases I and II producing deltaU-GlcNS. The K5 polysaccharides containing sulfate at the C-2 and C-6 positions of the hexosamine moiety and C-2 position of the glucuronic acid residues were susceptible only to heparitinase II producing deltaU-GlcNS,6S and deltaU,2S-GlcNS,6S respectively. These combined results led to the conclusion that the sulfate at C-6 position of the glucosamine is impeditive for the action of heparitinase I and that heparitinase II requires at least a C-2 or a C-6 sulfate in the glucosamine residues of the substrate for its activity. Iduronic acid-2-O-desulfated heparin was susceptible only to heparitinase II producing deltaU-GlcNS,6S. All the modified K5 polysaccharides as well as the desulfated heparin were not substrates for heparinase. This led to the conclusion that heparitinase II acts upon linkages containing non-sulfated iduronic acid residues and that heparinase requires C-2 sulfated iduronic acid residues for its activity.

Bacterial Capsules↗
PubMed1981

Mitogenicity and binding properties of beta-galactoside-binding lectin from chick-embryo kidney.

THe beta-galactoside-binding lectin binds to glucosamine, mannosamine and galactosamine in addition to beta-galactoside, as determined by the inhibition of haemagglutination. Haemagglutination is further extended to examine the interaction of the binding sites for hexosamines and beta-galactosides, indicating that the binding of hexosamine and beta-galactoside is competitive. The lectin also shows strong mitogenic activity toward lymphocytes from mouse lymph node, as determined by the stimulation of thymidine incorporation.

Animals↗
PubMed1999

Ecabet sodium, a novel locally-acting anti-ulcer agent, protects the integrity of the gastric mucosal gel layer from pepsin-induced disruption in the rat.

BACKGROUND: Ecabet sodium, a novel non-systemic anti-ulcer agent, possesses high affinity to gastric adherent mucus, which plays an important role in the protection of the gastric epithelium against acid and pepsin. AIM: To assess the effect of ecabet on pepsin-induced degradation of the structure of the mucus gel layer. METHODS: Everted sacs of rat stomach were incubated in HCl solution containing pepsin with or without ecabet. Pepsin-induced release of the cleaved peptides and hexosamine from the sacs was determined. Changes in the molecular size of glycoproteins in the adherent mucus (using gel filtration methods) and in the morphology of the epithelium (using both light and scanning electron microscopy) were also examined. RESULTS: Ecabet reduced the pepsin-induced release of peptides and hexosamine, depending on its content in the adherent mucus. Pepsin treatment partially lowered the molecular weight of native glycoproteins in the adherent mucus, caused exfoliation of the epithelial cells, and degraded the network-like ultrastructure of the mucus layer, giving it a lumpy, globular appearance. Ecabet prevented both the pepsin-induced molecular size shift in mucus glycoproteins, and morphological alteration of the epithelium, including ultrastructural derangement of the mucus gel layer. CONCLUSION: Ecabet protects the polymeric structure of mucus glycoproteins from proteolytic degradation by pepsin, and thus maintains integrity of the gastric mucus gel layer.

Abietanes↗
PubMed2002

Glucosamine:fructose-6-phosphate aminotransferase: gene characterization, chitin biosynthesis and peritrophic matrix formation in Aedes aegypti.

Glucosamine:fructose-6-phosphate aminotransferase (GFAT) catalyses the formation of glucosamine 6-phosphate and is the first and rate-limiting enzyme of the hexosamine biosynthetic pathway. The final product of the hexosamine pathway, UDP-N-acetyl glucosamine, is an active precursor of numerous macromolecules containing amino sugars, including chitin in fungi and arthropods. Chitin is one of the essential components of insect cuticle and peritrophic matrix. The peritrophic matrix is produced in the midgut of mosquitoes in response to bloodfeeding, and may affect vector competence by serving as a physical barrier to pathogens. It is hypothesized that GFAT plays a regulatory role in biosynthesis of chitin and peritrophic matrix formation in insects. We cloned and sequenced the GFAT gene (AeGfat-1) and its 5' regulatory region from Aedes aegypti. There is no intron in AeGfat-1 and there are two potential transcription start sites. AeGfat-1 cDNA is 3.4 kb in length and its putative translation product is 75.4 kDa. The amino acid sequence of GFAT is highly conserved in lower and higher eukaryotes, as well as in bacteria. AeGfat-1 message is constitutively expressed but is gradually up-regulated in the midgut after bloodfeeding. The putative regulatory region of the gene contains the ecdysone response element, E74, and Broad complex motifs, similar to what is found in the glutamine synthetase gene in Ae. aegypti. Results suggest that Ae. aegypti GFAT-1 may have a regulatory role in chitin biosynthesis and peritrophic matrix formation, and probably is under the regulation of ecdysteroids.

5' Untranslated Regions↗
PubMed1998

Glucosamine induces translocation of protein kinase C isoenzymes in mesangial cells.

Activation of protein kinase C (PKC) has been implicated in the high glucose-induced stimulation of matrix protein production in mesangial cells. Since we have found (Kolm-Litty et al., 1998) that glucosamine, similar to the PKC activator phorbol myristate acetate (PMA), mimicks high glucose-induced TGF-beta1 overexpression and subsequent matrix overproduction, the action of these agents on the translocation of PKC isoenzymes was studied in cultured mesangial cells. Exposure to 12 mM glucosamine resulted in rapid and specific translocation of PKC-isoenzymes in mesangial cells i.e. glucosamine caused an increased and sustained translocation of PKC-alpha, -beta and -epsilon while PKC-zeta was essentially unaffected. Comparison with PMA-induced translocation exhibited distinct differences. Exposure to high glucose concentrations of mesangial cells induced translocation of PKC-beta and down-regulation of PKC-epsilon while PKC-alpha and -zeta were essentially unaltered. Presence of azaserine an inhibitor of glutamine: fructose-6-phosphate amidotransferase, the key enzyme of the hexosamine pathway, attenuated the high glucose-induced effects on the membrane fraction of PKC-beta. Our results indicate that i) glucosamine is a potent stimulator of PKC-translocation exhibiting an isoenzyme specific translocation kinetic which is different from PMA-induced PKC-isoenzyme translocation ii) the hexosamine pathway may be possibly involved in the high glucose-induced activation of PKC.

Animals↗
PubMed1999

In vivo metabolism of aloemannan.

The metabolism of fluoresceinyl isothiocyanate labeled aloemannan (FITC-AM) was examined by p.o. and i.v. administration in mice at a dose of 120 mg/kg. Analysis of FITC-AM in urine and feces showed that FITC-AM (MW 500 KD) was metabolized into smaller molecules that mainly accumulated in the kidneys. AM was catabolized by the human intestinal microflora to catabolites 1 and 2 with molecular weights of 30 and 10 KD, respectively. Hydrolysis of AM showed hexosamine peaks on HPAE. The findings suggest that the immunomodulation of AM may come from not only neutral polysaccharides but also contaminated hexosamine in AM.

Adjuvants, Immunologic↗