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Administration of the plasticizer di (2-ethylhexyl) phthalate alters glycoconjugate profile.

This study was undertaken to evaluate the level of glycoproteins and sialic acid in rats fed di (2-ethylhexyl)-phthalate (DEHP) in the diet for 24 weeks. Protein-bound hexose, hexosamine and sialic acid were increased in plasma and liver of rats treated with DEHP, whereas the erythrocyte membrane showed a reduction following DEHP administration. Evaluation of glycoproteins is a useful indicator of the carcinogenic process. It is suggested that profound alterations in membrane components observed in the present study may be related to the carcinogenic potential of DEHP.

Animals↗

Characteristics of proteoglycans of buffalo ovarian follicular fluid during maturation of follicles.

The proteoglycans (PGs) and glycosaminoglycans (GAGs) of buffalo ovarian follicular fluid (FF) have been studied in small (2-4.9 mm), medium (5-9.9 mm) and large (> or = 10 mm) follicles. GAGs in different categories of follicles were isolated, assayed and analysed. On the basis of hexosamine analysis, glucosamine accounted for all the free GAGs in FF of small and medium follicles. No free GAG was found in large follicles. The concentration of GAGs in the form of PGs decreased significantly with follicular maturation. Qualitative analysis of GAGs from PGs showed higher galactosamine than glucosamine. The ratios of GalNH2:GluNH2 and neutral sugars were highest in small follicles followed by medium and large follicles. On the other hand, the percentage of sialic acid in GAGs was highest in large follicles followed by medium and small follicles. The fractionation of PGs by gel filtration indicated the presence of two types of PGs in buffalo ovarian FF. Difference in distribution of two types of PGs in small and large follicles was also noted.

Animals↗

Proteoglycan and collagen of "achondroplastic" (cn/cn) neonatal mouse cartilage.

Proteoglycans and collagen were analyzed in epiphyseal cartilage of 5 day old mice homozygous for the recessive gene, achondroplasia, and their normal littermates. No significant differences were observed in either hydroxyproline or hexosamine content. Extractable proteoglycans from control and mutant tissue were found to contain a similar distribution of aggregate and monomer. The component glycosaminoglycans were similar in size and composition. Collagen from both was identified as type II collagen. In addition, the collagen content of hind limb bones of 28 day dwarf mice was found to be similar to control amounts. These findings suggest that dwarfism in the cn/cn mouse is not associated with a defect in any of the major matrix constituents.

Achondroplasia↗

Alaskan Malamute chondrodysplasia III. Connective tissue of bone.

The dwarf Alaskan Malamute was compared in these studies with normal Alaskan Malamutes of the same age with regard to collagen and mucopolysaccharide components of the bone. The hydroxyproline concentration of bone segments was normal in most instances, whereas, the hexosamine concentration was increased (P less than .05) in distal segments. Alterations in amounts of collagen soluble in neutral salt, dilute acid and 5 M guanidine hydrochloride were observed and these were related to changes in subunits found following chromatography on carboxymethyl (CM) cellulose. The presence of an increased keratosulfate fraction in dwarf Alaskan Malamutes was consistent with an apparent delayed bone ossification process. These changes in the connective tissue moieties may account for the gross morphology with respect to the bowed legs. The primary cause for these changes has not yet been determined but alterations in factors related to processes of normal development in bone are suggested. The present observations tend to support the hypothesis that the changes in the bone are secondary to some other primary metabolic defect.

Animals↗

Rat intestinal brush border membrane peptidases. I. Solubilization, purification, and physicochemical properties of two different forms of the enzyme.

Two brush border peptidases have been isolated from the particulate fraction of the rat intestinal mucosa and purified to homogeneity as judged by polyacrylamide gel electrophoresis, starch gel electrophoresis, isoelectric focusing, and double immunodiffusion. For convenience, the peptidases have been designated peptidase F (fast) and S (slow) on the basis of their anodic mobilities. The isoelectric point of peptidase F was 4.76 and of peptidase S, 5.10. Both enzymes are glycoproteins. The amino acid compositions of the two peptidases are similar. The same carbohydrates are found in both enzymes, but there are differences in the molar concentrations of individual sugars. Peptidase S has greater concentrations of mannose and galactose and of hexosamines than peptidase F, while sialic acid is slightly greater in peptidase F. Carbohydrate accounted for approximately 19% and 23% of the weight of peptidases F and S, respectively. Estimates of the molecular weights of both enzymes by gel filtration gave values of 280,000. Electrophoresis of the enzymes under denaturing conditions on sodium dodecyl sulfate polyacrylamide gels indicated that each enzyme is a dimer consisting of two subunits of equal molecular weight, 140,000.

Amino Acids↗

[The effects of teprenone on the ulcer healing rate and the quality of healing in the treatment of active gastric ulcers].

UNLABELLED: One hundred and six patients with active gastric ulcer proven by endoscopy were randomly allocated to receive cimetidine 800 mg at bedtime + teprenone 50 mg thrice daily; or cimetidine 800 mg at bedtime alone in a trial comparing the ulcer healing rate and the quality of healing at 4 or 8 weeks. At 4 weeks 72.4% (42/58) of those on cimetidine + teprenone and 52.1% (25/48) on cimetidine alone had ulcers healed in comparison of 93.1% (54/58) and 89.6% (43/48) respectively at 8 weeks. The difference was significant at 4 weeks, indicating the ulcer heal more rapidly in the combination therapy group. The stage S2 achievement rates were 34.5% (20/58) and 10.4% (5/48) at 4 weeks (P < 0.05), 50.0% (29/58) and 20.8% (10/48) at 8 weeks (P < 0.05) respectively, indicating the quality of ulcer healing was much better in the combination therapy group. The gastric mucosal hexosamine levels increase significantly after treatment (17.79 +/- 2.00 micrograms/mg) as compared to that before treatment (14.27 +/- 2.47 micrograms/mg) in the combination therapy group, indicating that teprenone stimulates macromolecular glycoprotein synthesis in gastric mucosa. This is thought to be a significant factor in the healing process of gastric ulceration. CONCLUSION: It is reasonable to administer teprenone, an agent increasing mucosal resistance, concurrently with H2 blockers in the treatment of active gastric ulcers.

Anti-Ulcer Agents↗

Effects of administration of testosterone on seminal vesicle activity in the catfish Clarias batrachus during preparatory phase: a study correlating changes in testis and serum sex steroid levels.

In the catfish Clarias batrachus administration of testosterone (0.25, 0.5 and 1.0 microgram/g BW for 30 days) during the preparatory phase increased significantly gonadosomatic index (GSI), seminal vesicle-somatic index (SV-SI) and concentrations of proteins, fructose, hexosamines, and sialic acid in the SV and testis in a dose-dependent manner. These stimulatory effects could be correlated positively with the increase of serum testosterone level. Serum estradiol-17 beta was also significantly high in the testosterone groups.

Analysis of Variance↗

Mucolipidosis IV. Clinical, ultrastructural, histochemical, and chemical studies of a case, including a brain biopsy.

A 7-year-old Ashkenazi Jewish boy with normal early development started to regress at 8 months of age and made no further developmental progress. Corneal clouding was noted at age 10 months. Corneal and conjunctival biopsy at 14 months, cerebral biopsy at 24 months, and fibroblast cultures at 32 months showed lysosomal inclusions, suggesting the storage of lipid-like and mucopolysaccharide-like material. In the brain, dense fluorescent inclusions resembled those in ceroid-lipofuscinosis. Total ganglioside content of white matter was raised, but the pattern was normal. The level of nonlipid hexosamine in the brain was normal. The cornea and conjunctiva contained electronlucent vacuoles resembling those in the mucopolysaccharidoses. Cornea, brain, and lymphocytes contained concentric membranous lamellar structures reminiscent of those in the gangliosidoses. The clinical picture and ultrastructural findings support the impression that this case belongs to a new variant of the mucolipidoses, mucolipidosis IV.

Cerebral Cortex↗

Sandhoff's disease (GM2 gangliosidosis type 2). Histopathology and ultrastructure of the eye.

Sandhoff's disease (GM2 gangliosidosis type 2) was diagnosed in an infant in whom a progressive neurologic disorder and cherry-red foveal spots developed. At autopsy, ultrastructural examination of the retina and optic nerve disclosed abundant pleomorphic storage cytosomes in all neurons of the retina, including the inner segments of the photoreceptor cells, and in glial cells of the optic nerve. Electron microscopy of the cornea showed, within the keratocytes, distended clear lysosomes that contained some fibrillogranular material and an occasional collection of lamellae. We discuss the pathogenesis of the clinical and pathologic ocular findings with regard to the inherited absence of the enzymes hexosaminidase A and B and an accumulation of the substrates, GM2 ganglioside and asialo GM2, in the nervous system (including retina and optic nerve) and globoside and other hexosamine-containing substances in the viscera (including cornea).

Adolescent↗

Correlations among serum protein-bound carbohydrates, serum glycoproteins, lymphocyte reactivity, and tumors burden in cancer patients.

Levels of glycoprotein-associated carbohydrates (neutral hexoses, hexosamine, sialic acid and fucose) were determined in the serum of patients with either local, regional or metastatic cancer, patients clinically cured of cancer, and controls (smokers and nonsmokers). Total protein-bound carbohydrates were compared with levels of 17 normal serum glycoproteins, carcinoembryonic antigen (CEA), and with lymphocyte reactivity to phytohemagglutin (PHA). Tumor burden was directly related to protein-bound carbohydrate levels in patient groups. Levels of bound carbohydrates reflect the sum of all the changes in serum glycoproteins, but primarily changes in the acute-phase proteins (alpha 1-acid glycoprotein, alpha 1-antitrypsin, haptoglobin, ceruloplasmin) found in the alpha-globulin fraction of serum. Increases in protein-bound carbohydrates in tumor-bearers were not related to increases in CEA. Increased levels of the acute-phase proteins occurred in individuals with depressed in vitro lymphocyte reactivity to PHA. A significant positive correlation was found between lymphocyte reactivity and level of alpha 2HS-glycoprotein. The results suggest that serum protein-bound carbohydrates or glycoproteins may be of adjunctive value is assessing tumor burden and immune reactivity in cancer patients.

Carbohydrates↗

Measurement of sialyl transferase activity and serum glycoproteins in malignant and beningn (hyperparathyroid) hypercalcemia.

Serum sialyl tranferase activity (STA) and protein bound carbohydrate were measured in hypercalcemic patients with surgically documented hyperparathyroidism (n = 15) or with cancer without skeletal metastases (n = 10). Precipitable protein-bound sialic acid and hexosamine, but not neutral hexoses or fucose were significantly (P less than 0.025) increased 40% and 21% respectively, in the perchlorate treated sera of cancer patients. Measurements of some specific serum glycoproteins by radial immunodiffusion assay were also different (P less than 0.05). Mean seromucoid protein was almost twice as great in the group with cancer (P less than 0.005), but four of these values overlapped those from the patients with hyperparathyroidism. The mean STA was 21.1 (range, 9.0-46.8) activity units in the patients with the cancer and 9.2 (range 1.0-17.8) in the patients with hyperparathyroidism (P less than 0.005). Six of the patients with cancer had values above the upper limit of the group with hyperparathyroidism. A discriminant function generated from these data correctly classified all the patients with hyperparathyroidism and 70% of those with cancer. Measurements of seromucoid and STA deserve further consideration as a way to discriminate some cases of hypercalcemia.

Adult↗

Carbohydrate components of lipovitellin of the sand crab Emerita asiatica.

Lipovitellin II (Lv II), the major yolk protein of the anomuran crab Emerita asiatica, was purified using heparin-sepharose affinity column chromatography. The purified Lv II was a glycoprotein as it was stainable with periodic acid-Schiff's reagent. Quantitative analysis of sugars showed the presence of fucose, mannose, galactosamine, N-linked oligosaccharides, as well as O-linked oligosaccharides containing N-acetyl hexosamine as the terminal residue. The amount of N-linked oligosaccharides is higher than that of the O-linked oligosaccharides. Biogel P-4 column chromatographic separation of the radiolabeled oligosaccharides of Lv II showed the presence of five different O-linked oligosaccharides and four different N-linked oligosaccharide species. HPTLC separation of the neoglycolipids prepared from the O-linked oligosaccharides also showed the presence of five different O-linked oligosaccharide species. N-linked oligosaccharides contain significant quantities of mannose. Unisil column chromatographic purification in conjunction with HPTLC separation revealed three neutral glycolipid species such as monoglycosylceramide, diglycosylceramide, and triglycosylceramide in the Lv II. The functional significance of these carbohydrate components of the major yolk protein during embryogenesis of the sand crab is discussed.

Animals↗

Secretory granule proteoglycans of mast cells and natural killer cells.

Proteoglycan research on cells that participate in immune responses has progressed from the early novel finding that heparin proteoglycans are present in the secretory granules of the connective tissue mast cell to the more recent findings that mucosal mast cells and natural killer (NK) cells possess chondroitin sulphate proteoglycans in their granules. Characterization studies of these intracellular proteoglycans have revealed that they all possess peptide cores which are very resistant to proteolytic degradation. Their glycosaminoglycans, however, differ in such parameters as the type of hexosamine, location of sulphation degree of sulphation, or extent of epimerization of the uronic acid. Amino acid compositional analyses of heparin proteoglycans from rat connective tissue mast cells and chondroitin sulphate E proteoglycans from mouse mucosal mast cells indicate that their peptide cores are homologous to, but possibly distinct from one another. It is not yet known if these differences reflect a species variation, are due to different post-translational proteolytic processing, or are the result of expression of distinct genes coding for different peptide cores. The proteoglycans of mast cells and natural killer cells are packaged in the granules with cationic proteins. In mast cells these proteins have been shown to be serine proteases, and when bound to the acidic proteoglycans their enzymic activity is inhibited. Since the type of glycosaminoglycan linked to the proteoglycan has been found to be a characteristic of that cell, the structure of the cell-associated proteoglycan has become one of the markers used to distinguish cells phenotypically. By following the expression of different proteoglycans during differentiation, the relationship of the two subclasses of mast cells has been determined.

Animals↗

Metabolic and ultrastructural changes in articular cartilage of rats fed dietary supplements of omega-3 fatty acids.

A "marginally deficient" essential fatty acid state was produced in male Sprague-Dawley rats by dietary supplementation with omega 3 fatty acids. Animals fed diets containing the highest amounts of these fatty acids (10% menhaden fish oil) demonstrated a 70% maximum decrease in the linoleic and arachidonic acid content of articular cartilage, a 30-40% decrease in cartilage hexosamine content, with little effect on hydroxyproline levels, and a 32% inhibition of proteoglycan synthesis. Histologic analysis revealed an occasional surface irregularity and localized depletion of Safranin O and toluidine blue staining of articular cartilage on the femoral heads from animals taking the higher doses. Electron microscopic analysis revealed a marked decrease in "dark-staining" chondrocytes relative to "light-staining" cells in all animals fed menhaden fish oil. The cartilaginous changes noted in this study reflect a causal relationship between chondrocyte metabolism and an altered unsaturated fatty acid content. The observed responses of chondrocytes to omega 3 fatty acids may be similar to those commonly associated with the development of early osteoarthrosis. It is not known whether similar changes are induced in other species, including humans, but these observations suggest that some caution must be taken in the long-term administration of menhaden fish oil or other omega 3 fatty acid-containing preparations in rheumatoid arthritis patients.

Animals↗

Avian thrombocyte thrombospondin.

A high molecular weight glycoprotein (450,000) was obtained from thrombin-treated duck thrombocytes by barium citrate adsorption technique followed by heparin-agarose affinity chromatography. Amino acid composition (high number of acidic amino acids and cystine) as well as carbohydrate contents (1.3 per cent hexosamine, 0.9 per cent sialic acid and 1.5 per cent hexose) showed similarity to mammalian platelet thrombospondin.

Amino Acids↗

Thiamine and benfotiamine prevent increased apoptosis in endothelial cells and pericytes cultured in high glucose.

BACKGROUND: High glucose induces pathological alterations in small and large vessels, possibly through increased formation of AGE, activation of aldose reductase and protein kinase C, and increased flux through the hexosamine pathway. We showed previously that thiamine and benfotiamine correct delayed replication and increase lactate production in endothelial cells subjected to high glucose. We now aim at verifying the effects of thiamine and benfotiamine on cell cycle, apoptosis, and expression of adhesion molecules in endothelial cells and pericytes, under high ambient glucose. METHODS: Human umbilical vein endothelial cells and bovine retinal pericytes were cultured in normal (5.6 mmol/L) or high (28 mmol/L) glucose, with or without thiamine or benfotiamine, 50 or 100 micro mol/L. Apoptosis was determined by two separate ELISA methods, measuring DNA fragmentation and caspase-3 activity, respectively. Cell cycle and integrin subunits alpha3, alpha5, and beta1 concentration were measured by flow cytometry. RESULTS: Apoptosis was increased in high glucose after 3 days of culture, both in endothelium and pericytes. Thiamine and benfotiamine reversed such effects. Neither cell cycle traversal nor integrin concentrations were modified in these experimental conditions. CONCLUSIONS: Thiamine and benfotiamine correct increased apoptosis due to high glucose in cultured vascular cells. Further elucidations of the mechanisms through which they work could help set the basis for clinical use of this vitamin in the prevention and/or treatment of diabetic microangiopathy.

Animals↗

Electrophoresis-assisted open-tubular liquid chromatography/mass spectrometry for the analysis of lipooligosaccharide expressed by Campylobacter jejuni.

Lipooligosaccharide (LOS) is the major component of the external membrane of Campylobacter jejuni. LOS contains a hydrophobic moiety, lipid A, and a hydrophilic moiety, oligosaccharide. Due to the unique mimicry of human ganglioside structures and potential involvement in the induction of the autoimmune polyneuropathies, Guillain-Barré and Miller Fisher syndromes, the structural characterization of C. jejuni LOS has received much attention. We have been using capillary zone electrophoresis-mass spectrometry to analyze O-deacylated LOS from C. jejuni. In an attempt to optimize the separation conditions, the effect of methanol on the separation of LOS was investigated. It was found that methanol resulted in stronger adsorption of LOS onto the wall of fused-silica capillary. In this paper, we applied this adsorption to perform electrophoresis-assisted open-tubular liquid chromatography electrospray mass spectrometry for the analysis of O-deacylated LOS mixtures from C. jejuni. The analytical potential of the proposed strategy for the analysis of O-deacylated LOS glycoforms from five bacterial colonies is demonstrated. Online tandem mass spectrometry is shown to provide a powerful tool for characterization of variations in the hexosamine backbone, phosphorylation of the lipid A, and sialic acid sequence information.

Campylobacter jejuni↗

Changes in gallbladder bile composition and crystal detection time in morbidly obese subjects after bariatric surgery.

The aim of the present study was to elucidate the mechanisms of development of cholesterol crystals and gallstones during weight reduction in obese subjects. Twenty-five morbidly obese, gallstone-free subjects underwent vertical-banded gastroplasty. Gallbladder bile was collected at the time of the operation via needle aspiration and 1.1-7.3 months after the operation via ultrasound-guided transhepatic puncture of the gallbladder. The mean weight loss was 17 kg. Two patients developed gallstones and 10 patients displayed cholesterol crystals in their bile. In patients with a follow-up time of less than 2 months (n = 13), cholesterol saturation increased from 90% to 114% but tended to decrease in the patients with a follow-up time of more than 2 months. The extraction of the concanavalin-A-binding fraction from gallbladder bile obtained after weight reduction in 7 patients prolonged crystallization detection time from 6 to 10 days. The hexosamine concentration, a marker for mucin, was increased by about 100% in bile obtained in 6 of 7 patients after weight reduction. In conclusion, the results indicate that crystallization-promoting compounds (mucin) are of great importance in the development of cholesterol crystals and gallstones in obese subjects during weight reduction, probably because of defective gallbladder emptying.

Adult↗