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PubMed2004

Hexosamine pathway is responsible for inhibition by diabetes of phenylephrine-induced inotropy.

Hyperglycemia diminishes positive inotropic responses to agonists that activate phospholipase C (PLC) and generate inositol trisphosphate (1,4,5). The mechanisms underlying both the inotropic responses and hyperglycemia's effects on them remain undetermined, but data from isolated cardiomyocytes suggest the involvement of capacitative Ca(2+) entry (CCE), the influx of Ca(2+) through plasma membrane channels activated in response to depletion of endoplasmic or sarcoplasmic reticulum Ca(2+) stores. In neonatal rat cardiomyocytes, hyperglycemia decreased CCE induced by PLC-mediated agonists. The attenuation of CCE was also seen with glucosamine, and the inhibition by hyperglycemia was prevented by azaserine, thereby implicating hexosamine biosynthesis as the responsible metabolic pathway. In the current study, the importance of hexosamine metabolites to hyperglycemia's effects on inotropic responses was examined in isolated perfused rat hearts. The inhibition by hyperglycemia of phenylephrine-induced inotropy was reversed with azaserine and mimicked by glucosamine. An independent inhibitor of CCE, SKF96365, was also effective in blunting inotropy. These treatments did not inhibit inotropy induced by activation of adenylate cyclase through beta-adrenergic receptors. These data thus implicate CCE in responses to PLC-mediated agonists in the intact heart and point to the hexosamine pathway's negative effect on CCE as being central to the inhibition seen with hyperglycemia.

Animals↗
PubMed1981

The effects of manganese, biotin, and choline on hexosamine and hydroxyproline content as related to leg weakness.

Three experiments were conducted with day-old broiler chicks to determine the effects of dietary choline, biotin, and manganese on cartilage hexosamine and hydroxyproline content. The incidence of leg weakness was 75% or higher on all basal diets. The levels of supplementation found to minimize leg weakness were 1540 mg choline chloride/kg diet, .2 mg biotin/kg diet, and 100 mg manganese/kg diet. The levels required to maintain normal epiphyseal cartilage hexosamine content were 385 mg of choline chloride, .05 mg of biotin, and 25 mg of manganese/kg of diet. Substantially higher levels of nutrients were needed to reduce the incidence of leg weakness. Only with manganese supplementation was the incidence of leg weakness significantly reduced at the level of supplementation required for normal hexosamine content. Hydroxyproline content was not affected by varying the levels of supplemental choline, biotin, or manganese.

Animals↗
PubMed1975

Variations in collagen, non-collagenous proteins, and hexosamine in menisci derived from osteoarthritic and rheumatoid arthritic knee joints.

Knee joint menisci from osteoarthritic and rheumatoid knees were analyzed for nitrogen, collagen, non-collagenous proteins (NCP) and hexamine content. Degenerate areas were analyzed separately. The degenerative areas were significantly lower (P less than 0.005) in collagen but NCP and hexosamines were elevated relative to control tissue. The rheumatoid menisci revealed a reduced level of collagen and hexosamine in areas of localized degeneration. However, the collagen and hexosamine content of the remaining tissue was also diminished (P less than 0.025) relative to normal tissue of the same. age.

Adult↗
PubMed1975

[The hexosamines of the umbilical cord vessel walls].

Qualitative and quantitative compositions of hexosamines in arteries and veins of umbilical cord have been determined. The qualitative compositions of hexosamines in umbilical cord vessels are identical, namely they are composed of glucosoamine and galactosoamine. It has been shown that hexosamines occurred mainly as glycosoaminoglycans (GAG). Qualitative and quantitative compositions of GAG have been determined. It has been revealed that qualitative compositions of GAG of umbilical cord vessels are identical and they have been presented as follows: hyaluronic acid, heparane sulphate, chondroitine-6-sulphate, dermatane sulphate. Concerning the GAG, the difference between veins and arteries consist in higher content of hyaluronic acid and lower content of chondroitine-6-sulphate. The high content of hyaluronic acid in vein walls is probably connected with their permability.

Capillary Permeability↗
PubMed1991

Lysis of Neisseria gonorrhoeae initiated by binding of normal human IgM to a hexosamine-containing lipooligosaccharide epitope(s) is augmented by strain-specific, properdin-binding-dependent alternative complement pathway activation.

We studied the specificity of naturally acquired IgM bactericidal for strains of Neisseria gonorrhoeae that varied in sensitivity to the lytic action of normal human serum (NHS) and the relative ability of these strains to deplete the classical (CP) and alternative (ACP) C pathways. Lysis of both highly sensitive and relatively insensitive strains was inhibited by the same gonococcal lipooligosaccharides (LOS), as well as by Salmonella minnesota Re LOS and three hexosamine-containing glycose polymers. A polymer of N-acetylgalactosamine phosphate was the most inhibitory; a polymer of N-acetylglucosamine phosphate only partially inhibited. Neither 3-deoxy-D-manno-octulosonic acid (dOc1A) nor a polymer that contained dOc1A but not hexosamine inhibited NHS lysis. A co-polymer of N-acetylgalactosamine-dOc1A inhibited both bactericidal activity and the binding of IgM to the LOS of a highly serum-sensitive (sers) gonococcal strain. Carboxyl reduction of the dOc1A in this polymer did not affect its inhibitory capacity for gonococcal antibody, but abolished its binding to homologous antibody induced by vaccination. CP activity was not affected by vaccination. CP activity was not affected by absorption of NHS with gonococcal strains, whereas ablation of CP activity markedly but variously diminished lytic activity for highly sers strains. ACP activity was variously depleted by gonococcal strains, and the proportion of bacteria that could be lysed through the ACP varied among strains and among different populations of a given strain. The titer at which a strain was sensitive to NHS lysis was a function of its ACP consumption (p = 0.006), which accounted for 70% of the differences in titer among strains. Analyses of the absorbed sera revealed that the gonococci had variously depleted properdin from NHS as assessed by using an Ag-capture solid-phase RIA. Addition of purified properdin to absorbed sera restored ACP activity to normal levels. Western immunoblots of gonococcal lysates showed that purified properdin bound directly to a 39-kDa outer membrane protein. We conclude that both CP activation by IgM binding to LOS epitopes, one of which contains hexosamine, and ACP activation, which is a function of strain-specific direct binding of properdin, can initiate lysis of sers strains and that ACP activation, also enhances lysis and accounts for variations in sensitivity of sers strains.

Antigens, Bacterial↗
PubMed1988

[Effects of duodenogastric bile reflux on gastric mucosal hexosamine concentrations after selective proximal vagotomy or extended distal gastrectomy for duodenal ulcer].

Effects of duodenogastric reflux (DGR) of bile on hexosamine concentrations in gastric mucosa were studied in 17 healthy controls and 133 patients with duodenal ulcer patients before and after surgery. Total bile acid concentration in gastric juice was measured using enzyme method to estimate DGR. Mucosal hexosamine concentration of the biopsy specimens taken from the gastric corpus and antrum was measured according to Boas's method. The operative procedures included selective proximal vagotomy (SPV) with or without pyloroplasty, and extended distal gastrectomy with Billroth I(BI) or II(BII) anastomosis. The rate of DGR were significantly higher in cases after gastrectomy, especially in BII cases than in cases after SPV. In the early postoperative period after SPV with or without pyloroplasty, DGR was increased significantly. However, the reflux rate was decreased gradually to the preoperative level thereafter, suggesting that normal function of gastric emptying might be recovered with time. The hexosamine concentration of the antral mucosa showed clearly an inverse relationship to the changes in DGR rate. These results suggested that SPV could be the more physiological procedure than gastrectomy from the point of DGR.

Bile Acids and Salts↗
PubMed1977

Hexosamine-containing macromolecules in human colon carcinomas.

Normal, transitional, and carcinoma areas of five colons resected for carcinoma were examined morphologically, histochemically, and biochemically. The transitional area contained a larger amount of non-sulphated acid mucin than the normal mucosa as verified histochemically. Normal mucosa contained mainly sulphated mucin. The hexosamine-containing macromolecules present in different areas were isolated and characterized. They were divided into the following groups: 1) acid glycosaminoglycans, 2) high-molecular-weight glycopeptides, and 3) low-molecular weight glycopeptides. The concentration of the total hexosamine-containing material was in the carcinoma area twice as high as in normal areas. Acid glycosaminoglycans were identified as hyaluronate, heparan sulphate, dermatan sulphate, and chondroitin 4-(6)-sulphate. Their concentraitons were found to increase from normal to transitional and from transitional to carcinoma areas. The high-molecular-weight glycopeptide was composed of fucose, galactose, glucosamine, galactosamine, sialic acid, and variable amounts of sulphate. The sulphation degree of the glycopeptide was higher in normal mucosa than in transitional or carcinoma areas: The low-molecular-weight glycopeptides consisted of about a half of the total hexosamine-containing substances. The concentration of saline-insoluble fraction of the low-molecular-weight glycopeptides was in transitional areas about two times, and in carcinoma areas about four times, higher than in normal mucosa.

Adenocarcinoma↗
PubMed2002

Flux through the hexosamine pathway is a determinant of nuclear factor kappaB- dependent promoter activation.

The hexosamine pathway may mediate some of the toxic effects of glucose. We hypothesized that flux through this pathway might regulate the activity of nuclear factor kappaB (NF-kappaB)-dependent genes in mesangial cells (MCs). In MCs, RT-PCR revealed that high glucose (30 mmol/l) and glucosamine (1 mmol/l) increased mRNA levels for vascular cell adhesion molecule 1 (VCAM-1) and increased the activity of an NF-kappaB enhancer by 1.5- and 2-fold, respectively. Overexpression of glutamine:fructose-6-phosphate amidotransferase (GFAT), the rate-limiting enzyme for flux through the hexosamine pathway, led to a 2.2-fold increase in NF-kappaB enhancer activity; the combination of GFAT overexpression and high glucose increased activity 2.8-fold, and these increases were prevented by 40 micromol/l O-diazoacetyl-L-serine (azaserine) or 6-diazo-5-oxonorleucine. High glucose, glucosamine, and GFAT overexpression increased binding of MC nuclear proteins to NF-kappaB consensus sequences. Immunoblotting revealed that the p65 subunit of NF-kappaB was O-glycosylated in MC cultured in physiologic glucose and that significant enhancement occurred with high glucose and glucosamine. Both glucose and glucosamine dose-dependently increased human VCAM-1 promoter activity. In addition, GFAT overexpression activated the VCAM-1 promoter (2.25-fold), with further augmentation by high glucose and abrogation by inhibitors of GFAT, NF-kappaB, and O-glycosylation. Inactivation of the two NF-kappaB sites in the VCAM-1 promoter abolished its response to high glucose, glucosamine, and GFAT overexpression. These results suggest that increased flux through the hexosamine pathway leads to NF-kappaB-dependent promoter activation in MCs.

Animals↗
PubMed2000

Analysis of N-acetylated hexosamine monosaccharides by ferrocenyl boronation and tandem electrospray ionization mass spectrometry.

A tandem electrospray mass spectrometric (MS(n)) technique for the analysis of N-acetylated hexose carbohydrates using ferrocene boronate (F(c)Bor) derivatization was developed. The biologically important N-acetyl hexosamines can be readily distinguished utilizing this technique. The analysis is made possible by utilizing the inherent electrochemical properties of the electrospray device to produce oxidized, pre-formed single-electron ferrocenyl ions in a non-aqueous solvent system. The electrospray device was modified using a custom built cell consisting of concentric stainless steel tubes, which produced an enhanced signal for the molecular ion of each analyte species. The MS(2) spectra derived from isomeric populations of ferrocenyl boronic esters of these carbohydrates when generated under the same conditions possessed features unique to each sugar allowing easy differentiation between a number of N-acetyl hexosamine isomers.

Acetylation↗
PubMed1979

In vitro hexosamine depletion of intact articular cartilage by E-prostaglandins: prevention by chloroquine.

Short-term incubations of intact canine articular cartilage slices with prostaglandins E1 and E2 caused significant losses of hexosamine from cartilage matrix compared to controls. Chloroquine, an inhibitor of DNA primer, prevented this prostaglandin-induced hexosamine depletion. These data suggest that E-prostaglandins may degrade articular cartilage directly through DNA-dependent RNA synthesis of cathepsin-like proteases. Catabolism of articular cartilage probably involves degradation of existing matrix in addition to inhibition of synthesis.

Animals↗
PubMed1994

A specific fluorometric assay for hexosamines in glycosaminoglycans, based on deaminative cleavage with nitrous acid.

Glycosaminoglycan (GAG) hexosamines were measured after deacetylation (2 h acid hydrolysis), deaminative cleavage by nitrous acid, and coupling of the 2,5-anhydro sugars thus produced with 3,5-diaminobenzoic acid (DABA) to give a fluorescent product. Analyses were performed after the addition of an aliquot of potassium acetate solution to the acid hydrolysates, to adjust the pH. Results on a series of GAGs were compared with an Elson-Morgan (E-M) procedure. Our method is faster, more convenient, 10-20 times more sensitive, and always gave higher figures for hexosamine content. In particular it avoids long, destructive hydrolysis in concentrated strong acid and the complications of the Moggridge-Neuberger effect. Results agreed with calculations and titrimetric data. Using internal standards, our method is tolerant of cetylpyridinium chloride, peptides, and salts. A simple fluorometer easily handled less than 1 microgram GAG per sample in 1.0 ml volume. Results are available in 4-5 h. The method is suitable for automation. A strongly polycationic environment, as in chitosan, markedly slowed nitrous acid deamination, possibly because of Donnan-type exclusion from the domain of the polycation of the cationic nitrosonium species. Whereas all other alpha- and beta-hexosaminides gave yield-hydrolysis time profiles that peaked in approximately 1 h in the DABA method, the E-M profiles, particularly for 2-sulfamato-alpha-hexosaminides, took longer and/or achieved lower optimal yields. All the usually encountered forms of the 2-amino-2-deoxy group (free, acetylated, sulfamato) are readily assayed using the same reagents, with appropriate prefluorometric stages.

Aminobenzoates↗
PubMed1981

Renal medullary hexosamine content following antidiuresis and water-loading in the rat. Effects of antisera against rat urinary and testicular hyaluronidase.

The effect of antiserum raised against rat urinary (renal) hyaluronidase has been examined in rats subjected to antidiuretic stimuli (water-deprivation or vasopressin infusion). Prior administration of antiserum abolishes the reduction in medullary and papillary extractable hexosamine which normally accompanies antidiuresis. Antiserum against rat testicular hyaluronidase was found to be without effect during water-deprivation. Water-loading significantly increased the level of extractable hexosamine. The findings are considered in relation to previous observations on the effects of antisera on renal and urinary composition and collecting duct morphology under identical experimental conditions. It is suggested that a functional relationship exists between the net degradation of medullary mucopolysaccharides by hyaluronidase and the concentrating capacity of the kidney.

Animals↗
PubMed1981

Comparative studies on viscosity and concentration of protein and hexosamine in pure pancreatic juice.

In order to investigate the cause of an increase in viscosity of pancreatic juice in patients with chronic pancreatitis, we performed comparative studies on voscosity and concentration of protein, hexosamine, three hydrolytic enzymes (amylase, chymotrypsinogen and lipase), calcium and gamma-Glutamyltranspeptidase in pure pancreatic juice. Pure pancreatic juice (PPJ) was obtained from seven control subjects, 10 patients with probable chronic pancreatitis and six patients with definite chronic pancreatitis by means of endoscopic retrograde catheterization of the papilla. Specimens were collected in ice-cooled tubes during the initial four-minute period and thereafter at two-minute intervals for 20 minutes after a rapid intravenous injection of secretin (Eisai Secrepan, 1 U/kg of body weight) and then at one-minute intervals for 10 minutes after a rapid intravenous injection of pancreozymin (Boots, 1 U/kg of body weight). It was demonstrated that the viscosity of PPJ was closely related to the concentration of hydrolytic enzyme protein, hexosamine and the presence of mucous substance and protein plugs.

Amylases↗
PubMed1983

Automated analysis of hexosamines by high-performance liquid chromatography with photometric and fluorimetric postcolumn labeling using 2-cyanoacetamide.

Glucosamine and galactosamine were well separated in ca. 60 min on a Hitachi 2617 column (polystyrene sulfonate type, 4 mm X 25 cm) with a borate buffer (pH 7.5) containing sodium chloride. The hexosamines in the eluate were monitored fluorimetrically and photometrically at 331 (excitation)/383 (emission) and 276 nm, respectively, by postcolumn labeling with 2-cyanoacetamide. This simple method allowed the simultaneous, automated determination of 10-500 nmol of glucosamine and galactosamine with high reproducibility. This method was applied successfully to the analysis of hexosamines in some glycoconjugates.

Adult↗
PubMedDate not supplied

Aortic hexosamine, [35S]sulfate uptake, and calcium metabolism related to early arterial degenerative changes induced by lactation and forced weaning in breeder rats.

The relation of lactation and weaning to the development of early arterial lesions in the female breeder rat was investigated. Changes in aortic hexosamine and 35S-uptake, which are indicative of ground substance metabolism were correlated with changes in aortic calcium, phosphorus, and 45Ca-uptake during lactation and weaning. Lactation was associated with reduced aortic hexosamine content and lowered uptake of [S]sulfate. Further dynamic changes in aortic metabolism of these substances occurred following weaning in conjunction with an intense calcium uptake in the aorta of some, but not all, of the post-lactation animals. These latter changes were associated with the beginning aortic calcifications. Histopathologically, early arterial lesions developing during lactation, consist of intimal accumulations of mucopolysaccharide capped over by collagen. With repeated breeding cycles, these early lesions become exacerbated with extensive medial calcification occurring in areas rich in mucopolysaccharide and collagen particularly around degenerating elastic fibers. The hormones associated with reproduction and lactation may have a conditioning effect on these arterial connective tissue alterations and the development of arterial degenerative changes.

Animals↗
PubMed1979

Aortic glycopeptide sialic acid, hexose and hexosamine in a genetically selected (WC-2) strain of atherosclerosis-susceptible pigeon.

The aortic content of glycopeptide sialic acid, hexosamine and hexose was studied in a genetically selected strain of White Carneau pigeons (WC-2) with significantly more severe atherosclerosis than randomly bred White Carneau pigeons (RBWC). Pigeons were fed an atherogenic diet for 3 months and examined to determine differences in content of glycopeptide-sugars between WC-2 and RBWC, changes with the progression of atherosclerosis and the relationship of aortic cholesterol to glycopeptide-sugar content. In animals with mainly normal aorta (cholesterol content of 0.2-0.3 mg/cm2-aorta) sialic acid was significantly lower in WC-2 pigeons. The progression of atherosclerosis was associated with increased aortic glycopeptide sialic acid (r = 0.78; p less than 0.05) in WC-2 pigeons whereas an inverse relationship was suggested in RBWC pigeons. In WC-2, but not RBWC pigeons, significant positive relationships were seen for aortic glycopeptide hexosamine and aortic cholesterol and for aortic glycopeptide hexose and aortic cholesterol. The findings implicate a possible role of aortic glycoproteins in either the initiation or modulation of atherosclerosis of the WC-2 pigeon.

Animals↗
PubMed1976

The last two pathway-specific enzyme activities of hexosamine biosynthesis are present in Blastocladiella emersonii zoospores prior to germination.

Forward direction assays have been developed for the last two pathway-specific enzymes of hexosamine biosynthesis using crude extracts from Blastocladiella emersonii zoospores. The specific enzyme activities measured are substantially higher than those reported with enzyme preparations from other organisms. During the development of one of the assays, another enzyme activity was observed which converts one of the intermediates of the pathway, N-acetylglucosamine-6-phosphate, to N-acetylglucosamine. The finding of these three enzyme activities in zoospore extracts completes the demonstration that all the enzyme activities necessary to synthesize some 2% by weight as chitin early during zoospore germination (de novo cell wall formation) pre-exist in the zoospore. This demonstration is consistent with the conclusion that the hexosamine pathway is regulated at the post-translational level during zoospore germination.

Acetylglucosamine↗
PubMed1993

Development of an HPLC method to estimate hexosamines and its application to determine mucin content in rat and human gastric mucosa.

1. An HPLC analysis of hexosamines and related compounds modified from the Waters Pico-Tag amino acid analysis procedure is described. 2. This method is applicable to determine glucosamine and galactosamine not only in the hydrolyzate of purified glycoconjugate such as gastric mucin and ovalbumin but also in crude materials such as rat gastric mucosal extract and powdered bovine nasal cartilage. 3. Compared with a conventional colorimetric hexosamine determination, sensitivity of this method is almost 100 times higher. 4. By applying this method, a simple but highly sensitive procedure to determine mucin content in a crude stomach specimen of rat and that in a biopsied human gastric mucosa was also developed.

Adult↗