Effects Of N-acetylglucosamine Research Articles

Anti-inflammatory activities of two new deoxygenated N-acetyl glucosamines in lipopolysaccharide-activated mouse macrophage RAW264.7 cells

BackgroundGlucosamine and N-acetyl-glucosamine (NAG) are amino sugars found in human extracellular matrix with previously described anti-inflammatory effects. Despite mixed results from clinical studies, these molecules have been used extensively in supplements. ObjectiveWe investigated the anti-inflammatory properties of two synthesized derivatives of N-acetyl-glucosamine (NAG), bi-deoxy-N-acetyl-glucosamine (BNAG) 1 and 2. MethodsUsing mouse macrophage RAW 264.7 cells with lipopolysaccharide (LPS) to induce inflammation, the effects of NAG, BNAG 1, and BNAG 2 on the expression of IL-6, IL-1β, inducible nitric oxide synthase (iNOS) and COX-2 were studied using ELISA, Western blot and quantitative RT-PCR. Cell toxicity and nitric oxide (NO) production were evaluated using WST-1 assay and the Griess reagent, respectively. ResultsAmong the three tested compounds, BNAG1 shows the highest inhibition of iNOS, IL-6, TNF α and IL-1β expression and NO production. All three tested compounds show slight inhibition on cell proliferation of RAW 264.7 cells, except that BNAG1 displays a remarkable toxicity at the tested maximum dose of 5 mM. ConclusionBNAG 1 and 2 exhibit notable anti-inflammatory effects compared to the parent NAG molecule.

The NAG Sensor NagC Regulates LEE Gene Expression and Contributes to Gut Colonization by Escherichia coli O157:H7.

Enterohemorrhagic Escherichia coli (EHEC) O157:H7 are human pathogens responsible for bloody diarrhea and renal failures. EHEC employ a type 3 secretion system to attach directly to the human colonic epithelium. This structure is encoded by the locus of enterocyte effacement (LEE) whose expression is regulated in response to specific nutrients. In this study, we show that the mucin-derived sugars N-acetylglucosamine (NAG) and N-acetylneuraminic acid (NANA) inhibit EHEC adhesion to epithelial cells through down-regulation of LEE expression. The effect of NAG and NANA is dependent on NagC, a transcriptional repressor of the NAG catabolism in E. coli. We show that NagC is an activator of the LEE1 operon and a critical regulator for the colonization of mice intestine by EHEC. Finally, we demonstrate that NAG and NANA as well as the metabolic activity of Bacteroides thetaiotaomicron affect the in vivo fitness of EHEC in a NagC-dependent manner. This study highlights the role of NagC in coordinating metabolism and LEE expression in EHEC and in promoting EHEC colonization in vivo.

Effects of N-acetyl glucosamine and chondroitin sulfate supplementation on knee pain and self-reported knee function in middle-aged and older Japanese adults: a randomized, double-blind, placebo-controlled trial.

There was no evidence of satisfying the standard to decide the efficacies of glucosamine and chondroitin in middle-aged and older Japanese adults with knee pain and/or stiffness. To investigate the effects of 24week oral N-acetyl glucosamine and chondroitin sulfate supplementation on knee pain, self-reported knee function, physical activity, and physical performance. We randomly assigned 11 men and 39 women (aged 52-87years) to receive 100mg of N-acetyl glucosamine and 180mg of chondroitin sulfate daily (Glu/Cho group) or a placebo (control, C group) for 24weeks. The primary outcomes were a 100mm visual analog pain scale (VAS) and the Japanese Knee Osteoarthritis Measure (JKOM) score. The secondary outcomes were physical activity and physical performance. We observed a significant group×time interaction on the JKOM score. According to the post hoc test, it significantly decreased (i.e., improved knee function) from the 4- to 12-week follow-up in the Glu/Cho group and the Glu/Cho group score was significantly lower than the C group at the 12-week follow-up. We found a significant interaction on household physical activity. There was no significant interaction on VAS or physical performance tests. The results of the present study were consistent with previous studies mainly conducted in European and American countries. These results suggest that consumption of N-acetyl glucosamine and chondroitin sulfate for 12weeks or longer has a positive effect on self-reported knee function and household physical activity in middle-aged and older Japanese adults with knee pain and/or stiffness.

Neuroprotective effects of N-acetylglucosamine against hydrogen peroxide-induced apoptosis in human neuronal SK-N-SH cells by inhibiting the activation of caspase-3, PARP, and p38

Neuroprotective effects of N-acetylglucosamine (GlcNAc), a monosaccharide derivative of glucose, against H2O2-induced neurotoxicity and its underlying mechanism in human SK-N-SH neuroblastoma cells were investigated. Pretreatment of GlcNAc prior to exposure of cells to H2O2 stress significantly reduced the H2O2-mediated neuronal cell death and apoptosis. The GlcNAc dose-dependently decreased the level of intracellular reactive oxygen species (ROS) in H2O2-treated cells and also effectively inhibited H2O2-induced apoptotic features such as DNA fragmentation, caspase-3, and poly ADP-ribose polymerase (PARP) cleavages, and p38 phosphorylation. These results suggested that GlcNAc might potentially serve as agents for prevention of neurodegenerative diseases caused by oxidative stresses and this effect may be associated with the suppression of caspase-3, PARP, and p38 activation.

N-acetyl glucosamine improves intestinal mucosal barrier function in rat

Our study investigated the effect of N-acetylglucosamine (GlcNAc) on the intestinal mucosal barrier function in rats. Rats were randomly assigned into normal control group, diarrhea-predominant irritable bowel syndrome (IBS-D) group and GlcNAc group. IBS-D was introduced into the IBS-D group without any treatment. The GlcNAc group were treated with GlcNAc. Microvilli and tight junctions of intestinal epithelial cells were detected. The D-lactic acid level and diamine oxidase (DAO) activity in the serum were determined. Compared with normal rats, microvilli were sparsely distributed on the intestinal epithelial cells, the tight junction gap also widened, and D-lactic acid level and DAO activity were significantly higher in the IBS-D group. After GlcNAc treatment, the microscopic structure of the intestinal mucosa became largely normal, and the level of D-lactic acid and the DAO activity were lowered. In conclusion, GlcNAc can effectively improve the intestinal mucosal barrier dysfunction, perhaps through enhancing the cellular metabolism.

Establishment of An Advanced Chemically Defined Medium for Early Embryos Derived from In Vitro Matured and Fertilized Bovine Oocytes

A chemically defined medium would be useful for analyzing promoters or inhibitors in in vitro culture (IVC) of bovine embryos. However, an IVC system for bovine embryos in a chemically defined medium has not been fully established. The present study was carried out to establish an advanced chemically defined medium for bovine embryos that supports a high rate of embryo development to the blastocyst stage. In the first experiment, we examined the effects of addition of Medium RD (RPMI1640 and Dulbecco's MEM, 1:1 v/v) to mKSOM/aa on developmental competence. The addition of 10% RD to mKSOM/aa with BSA improved the rate of development to the blastocyst stage; however, 10% RD-mKSOM/aa with PVP, which is a chemically defined medium, caused a reduction in the percentage of hatching blastocysts. In the second experiment, embryos were cultured in the chemically defined medium of 10% RD-mKSOM/aa containing 11.7, 23.4, 46.8, 70.2 or 96.8 µM inositol. Inositol at the concentration of 70.2 µM improved the rate of development to the hatching blastocyst stage. In the third experiment, the optimal RD concentration in the IVC medium was evaluated. Embryos were cultured in the chemically defined medium supplemented with 10, 20 or 30% (v/v) RD. The rate of development to the blastocyst stage was highest with 20% RD. In the fourth experiment, the effects of N-acetylglucosamine (GlcNAc) as an IVC medium supplement on developmental competence were examined. The rate of development to the blastocyst stage with 1.0 mM GlcNAc was significantly higher than that without GlcNAc, but the rate of development with 1.2 mM GlcNAc was not different from that without GlcNAc. We also evaluated the ability of blastocysts produced in RD-mKSOM/aa to develop to normal calves after being transferred into recipients. Ten of the 16 recipients became pregnant, with 9 delivering normal calves. These results indicate that 20% RD-mKSOM/aa containing 70.2 µM myo-inositol and 1 mM GlcNAc is useful as a chemically defined medium for IVC of bovine embryos.

Effects of N-acetylglucosamine on tyrosinase activity and melanogenesis in B16 melanoma cells.

Objective To study the effects of N-acetylglucosamine on the activity of mushroom tyrosinase in vitro as well as on tyrosinase activity and melanogenesis in B16 melanoma cells. Methods Various concentrations (1.5 - 50 mmol/L) of N-acetylglucosamine were used to incubate with mushroom tyrosinase for 10 minutes following by the measurement of dopa oxidase activity of tyrosinase. B16 melanoma cells were cultured and treated with six concentrations (1.5 - 100 mmol/L) of N-acetylglu-cosamine for 72 hours; then, the cell proliferation of, melanogenesis and expression of tyrosinase protein and activity of tyrosinase in cultured B16 cells were measured by methyl thiazolyl tetrazolium (MTT) assay, sodium hydroxide (NaOH)-lysis method, immunocytochemical method and dopa oxidation assay, respectively. Results N-acetylglucosamine of 1.5, 3, 6.25, 25 and 50 mmol/L significantly inhibited the activity of mush-room tyrosinase compared with phosphate buffered saline (0.7380 ± 0.0254, 0.7293 ± 0.0382, 0.7247 ± 0.0389, 0.7233 ± 0.0186, 0.7043 ± 0.0166 vs 0.8183 ± 0.0326, P < 0.05 or 0.01). After treatment with N-acetylglucosamine of 25, 50 and 100 mmol/L, a significant suppression was observed in cell proliferation (ahsorbance: 0.5410 ± 0.0496, 0.4480 ± 0.0246 and 0.1273 ± 0.0137 vs 0.6523 ± 0.0569) of and melano-genesis (absorbance at 460 nm: 0.0070 ± 0.0008, 0.0049 ± 0.0012 and 0.0015 ± 0.0014 vs 0.0096 ± 0.0014) in B16 melanoma cells. Also, decreased activity (absorbance at 460 nm: 0.1003 ± 0.0404 and 0.0130 ± 0.0053 vs 0.2283 ± 0.0691 ) and protein expression ( 13.2700 ± 0.9741 and 8.5667 ± 2.0345 vs 17.4703 ± 2.0583 ) of tyrosinase were noted in B16 cells treated with N-acetylglucosamine of 50 and 100 mmol/L. Conclusions These studies show that N-acetylglucosamine inhibits tyrosinase activity and melanogenesis in murine B16 melanoma cells. Hence, N-acetylglucosamine may serve as a skin lightening agent in the future. Key words: Tumor cells; cultured; Monophenol monooxygenase; Melanins; N-acetylglucosamine

P-175: Effect of N-acetyl glucosamine and β glycerophosphate supplementation in cryopreservation media on post thaw sperm functional characteristics

The cryosurvival rates of human spermatozoa remain poor despite of modern cryopreservation methods for human semen. This is due to factors such as membrane damage and sub lethal damage at chromatin level. The goal of the present study was to investigate the supplementation of N-acetyl-D-glucosamine (NAG), a major structural element in cell walls and coats, and β-glycerophosphate (β-GP) in cryoprotective agent (CPA) on post thaw sperm characteristics.

Synergistic Effect of N-Acetylglucosamine and Retinoids on Hyaluronan Production in Human Keratinocytes

Hyaluronan (HA) is well known to reside in the extracellular matrix as a water-sorbed macromolecule. The aims of this study were twofold: to investigate the regulation of HA synthesis in keratinocytes, and to develop a method to modulate this regulatory process. We found that N-acetylglucosamine (NAG) increased the production of HA by cultured keratinocytes dose dependently, but had no effect on the production by skin fibroblasts. The effect of NAG in keratinocytes was found to be specific for HA production, as there was no change in sulfated glycosaminoglycan formation. The copresence of NAG with either of two retinoids, retinoic acid (RA) or retinol, exerted a synergistic effect on HA production. To investigate whether human HA synthase (HAS) genes were regulated by NAG or retinoids, total RNA extracted from cells treated with these agents was subjected to Northern blot analysis. We observed that RA and retinol markedly induced the expression of HA synthase-3 (HAS3) mRNA. Moreover, β-carotene, a provitamin A, influenced HA production and HAS3 gene expression in a manner similar to the retinoids. Conversely, NAG had no effect on the expression of HAS3 transcripts. Pretreatment of cells with RA stimulated the activity of membrane-associated HAS, whereas pretreatment with NAG did not. These results suggest that HA production is regulated by at least two pathways: one involving the regulation of HAS gene expression, and the other independent of such a regulatory effect. Taken together, our findings suggest that NAG is a new modulator of HA synthesis.

Distribution of chitinases in the entomopathogen Metarhizium anisopliae and effect of N-acetylglucosamine in protein secretion.

For a long time, fungi have been characterized by their ability to secrete enzymes, mostly hydrolytic in function, and thus are defined as extracellular degraders. Chitin and chitinolytic enzymes are gaining importance for their biotechnological applications. Particularly, chitinases are used in agriculture to control plant pathogens. Metarhizium anisopliae produces an extracellular chitinase when grown on a medium containing chitin, indicating that synthesis is subject to induction by the substrate. Various sugar combinations were investigated for induction and repression of chitinase. N-acetylglucosamine (GlcNAc) shows a special dual regulation on chitinase production. M. anisopliae has at least two distinct, cell-bound, chitinolytic enzymes when cultured with GlcNAc as one of the carbon sources, and we suggest that this carbohydrate has an important role in protein secretion.

Non-identity of human plasma lysozyme and 4-methylumbelliferyl-tetra-N-acetyl-β- d-chitotetraoside hydrolase

1. 1. Using 4-methylumbelliferyl-tetra-N-acetyl-β- d-chitotetraoside (MU-TACT) as substrate, it is possible to measure the activity of purified lysozyme and to demonstrate lysozyme activity in the urine of patients with acute monocytic leukemia, characterized by massive lysozymuria. 2. 2. Notwithstancling this observation, we present evidence that in normal human plasma another acid endoglucosaminidase is hydrolyzing the substrate. 3. 3. The following data support the hypothesis of the existence of a separate hydrolase: (a) Thermoinactivation is different for MU-TACT hydrolase and lysozyme. (b) In plasma and many other biological samples, the concentration of lysozyme is too low to be measured with the artificial substrate and there is no correlation between MU-TACT hydrolase and lysozyme. (c) Serum of lysozyme deficient rabbits has normal MU-TACT hydrolase activity. (d) On Sephadex G-200 and DEAE cellulose chromatography, lysozyme and MU-TACT hydrolase are eluted separately. (e) Immunoremoval of lysozyme from human plasma does not affect the activity towards MU-TACT. (f) The effect of N-acetylglucosamine and N-acetylmuramic acid on the activity of lysozyme and MU-TACT hydrolase is different.

Effects of N-acetylglucosamine and platelet inhibitors on the synergistic interaction of platelets and aggregating agents in the presence of wheat germ agglutinin.

Low concentrations of wheat germ agglutinin (4 micrograms/ml) have been shown to act synergistically to induce platelet aggregation with epinephrine, collagen, arachidonate and ionophore A23187. Aggregation ceased on the addition of the haptenic sugar N-acetylglucosamine at any time following the onset of aggregation with these agonists and a small degree of disaggregation was observed during the reversible first wave with the biphasic aggregating agents epinephrine and ADP. Cyclooxygenase inhibitors such as indomethacin and aspirin blocked the second wave of aggregation with the biphasic aggregating agents epinephrine and ADP but a synergistic response continued to be shown with the first wave in the presence of these inhibitors. Release of [14C]serotonin and the mobilization of [3H]arachidonate by epinephrine and collagen were markedly stimulated in the presence of wheat germ agglutinin but there was no increase of either radiolabel in the case of ADP. Platelet shape change, but not aggregation, occurred with low levels of wheat germ agglutinin and the synergistic response with ADP, collagen or ionophore A23187 occurred without further shape change. Wheat germ agglutinin did not affect the basal or stimulated levels of cyclic AMP. The membrane fluidity of platelets was not affected by the lectin or by thrombin as shown by the lack of change in fluorescence polarization with diphenylhexatriene. It is suggested that the binding of wheat germ agglutinin to the platelet surface induces platelet activation by mechanisms similar to those of other agonists and that it may affect the distribution of membrane-bound Ca2+ by a reversible perturbation of the platelet membrane.

Effects of N-acetyl glucosamine on platelet aggregation

N-acetyl glucosamine (NAG) alone did not aggregate platelets but in low concentrations shortened the delay phase of the Staphylococcus aureus induced platelet aggregation in a concentration dependent manner, which suggests that NAG may be one of the components of the S. aureus cell wall responsible for platelet aggregation. NAG, on the other hand, inhibited collagen induced platelet aggregation and the secondary platelet aggregation induced by epinephrine and ADP. This inhibition appears to be due to impurities that occur at higher levels in some NAG lots than in others. NAG has also been shown to decrease the length of time before the release of ATP from platelets when S. aureus is the aggregating agent.

The effects of glucosamine on platelet aggregation

The effects of glucosamine on platelet aggregation induced by a variety of agents were investigated. When compared with the effects of N-acetyl glucosamine (NAG), an acetylated derivative of glucosamine (1), it became apparent that the N-acetyl group was important in stimulating Staphylococcus aureus induced platelet aggregation. The absence of the N-acetyl group from NAG changes it to glucosamine which is an inhibitor rather than a stimulator of S. aureus induced platelet aggregation and release. Glucosamine also inhibits collagen, epinephrine, and ADP induced platelet aggregation and release.

An examination of the distribution of hen- and goose-type lysozymes in anseriformes

1. 1. Enzymic tests, based on the effect of N-acetylglucosamine and on tryptic digestion, have been developed to distinguish hen- and goose-type lysozyme activities in egg whites. 2. 2. The lytic activity of the egg whites of 25 Anseriforme species has been measured and grouped into types by the enzymic tests. 3. 3. The simultaneous occurrence of both types of lysozyme in an egg was frequently observed; 6 species were mainly or completely goose-type; 3 species contained mainly hen-type enzyme.

Interaction of alcohols with lysozyme. I. Studies on circular dichroism.

The circular dichroic (CD) spectra of hen egg-white lysozyme [EC 3.2.1.17] in the presence of ethylene glycol, methanol, ethanol, propanol, and sucrose were measured. These reagents enhanced the CD bands in the aromatic absorption region in a non-denaturing concentration range. By analyzing the increase in the CD extremum at 294 mμ with increasing the reagent concentration, it was found that one mole of the respective reagent interacts with one mole of lysozyme. All the association constants for the interaction with the reagents studied were smaller than that for the interaction with N-acetylglucosamine. The effect of N-acetylglucosamine or ethanol on the CD spectrum of lysozyme of which Trp-62 is oxidized was also studied. It was suggested that Trp-62 has a large contribution to the enhancement of the CD extremum by the interaction of N-acetylglucosamine or ethanol.

The Effect of N-Acetylglucosamine and Glucosamine on Carbohydrate Metabolism in Rat Liver Slices

The Effect of N-Acetylglucosamine and Glucosamine on Carbohydrate Metabolism in Rat Liver Slices