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Objective To study the clinical significance of the expression of D-amino acid oxidase(DAO) in hepatocellular carcinoma(HCC) tissues.Methods The gene expression profiles and related clinical data of 214 HCC cases were collected.Univariate and multivariate analyses were conducted to investigate the association between DAO expression levels,clinical traits,the prognosis.Results Univariate analysis indicated that there were a lower level of blood alpha fetoprotein(AFP,P=0.001),a smaller number of nodules(P=0.042),a better TNM stage(P=0.014),a lower metastasis risk(P=0.001) and a better prognosis(P=0.011)in the samples with high DAO expression.Multivariate analysis also indicated a lower AFP level(OR:0.162,95%CI:0.078-0.336) and metastasis risk(OR:0.140,95%CI:0.069-0.284) in the samples with high DAO expression,as well as a better prognosis(OR:0.833,95%CI:0.700-0.992).Conclusion DAO expression was associated with blood AFP levels,metastasis risk and prognosis in HCC,and its high expression was a protective factor for HCC.
ABSTRACT
D-amino acid oxidase (DAAO) is biotechnologically relevant enzyme that is used in various food and pharmaceutical industries. DAAO from the yeast Trigonopsis variabilis is an important agent for use in commercial applications because of its high activity with cephalosporin C and is reasonable resistant to the oxidants O2 and H2O2 byproducts of reaction. In this study, response surface methodology (RSM) in shake flask culture was used to enhance the production of DAAO from T. variabilis by optimization of fermentation media composition. The effects of six factors (DL-alanine, glucose, pH, ZnSO4, (NH4)2SO4 and temperature) were evaluated on DAAO production. Results of Placket-Burman design showed that DL-alanine, pH, glucose and ZnSO4 were significant factors for DAAO production (P<0.05). The optimum values of media components as predicted by the central composite design were inducer (DL-alanine) concentration 3 g/L, pH 7.7, glucose 17 g/L and ZnSO4 34 mg/L. At these optimum values of media composition, maximum production of DAAO was 153 U/g yeast dry weight. Two-fold increase in DAAO production was achieved after optimization of the physical parameters by RSM.
Subject(s)
Biostatistics/methods , D-Amino-Acid Oxidase/analysis , Models, Statistical , Research Design/methods , Yeasts/analysisABSTRACT
NG-nitro-D-arginine (D-NNA) produced pressor responses in rats by acting via chiral inversion intoNG-nitro-L-arginine (L-NNA), an inhibitor of nitro oxide synthase. The present investigation aimed to study the roleof the D-amino acid oxidase (DAAO) in chiral inversion of D-NNA and the relationship between DAAO activitieson various D-amino acids and their inversion rate. Benzoate (400 mg/kg) or creatinine (400 mg/kg), two inhibitorsof DAAO, blocked D-NNA-induced pressor responses in rats. Furthermore, the addition of the pure DAAOsignificantly potentiates L-NNA production rate in kidney homogenates by approximately 2-folds. The in vivo andin vitro results suggested that DAAO plays an essential role in the pressor responses elicited by D-NNA.Moreover, crude DAAO solution from the kidney showed marked selection (the maximal ratio of Kcat/Km wasnearly 15 times) on different D-amino acids that exhibited similar chiral inversion rate in vivo, suggesting that otherenzymes, such as transaminase, are also required for the entire process of D-NNA chiral inversion.
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Objective:To study the expression of D-amino acid oxidase(DAO) in gastric cancer cells and gastric mucosa cells. Methods:The expression of DAO was detected in seven gastric cancer cell lines and a normal gastric mucosa cell line GES-1 by real-time quantitative PCR and in tumor tissues and normal gastric mucous tissues of 46 patients with gastric cancer by RT-PCR.The liver and kidney tissues of nude mouse acted as positive controls.Results:DAO was expressed in the liver and kidney tissues of nude mouse,and DAO expression in kidney tissue was higher than that in liver tissue.DAO was not detected in seven gastric cancer cell lines and the normal gastric mucosa cell line GES-1.Except one tumor tissue sample,DAO was not detected in other gastric cancer tissues and all normal mucosa tissues.Conclusion:DAO was not expressed in gastric cancer cells and gastric mucosa cells.The nutritional treatment of D-Met in place of L-Met could avoid the disadvantage of methionine starvation to important organs,such as liver and kidney.
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Trigonopsis variabilis induced for D-amino acid oxidase and catalase was immobilized by entrapment in Polyacrylamide beads obtained by radiation polymerisation. Permeabilization of the cells was found to be essential for optimal activity of the enzymes in free cells. However, the process of entrapment itself was found to eliminate the permeability barrier of cells immobilized in Polyacrylamide. The two enzymes exhibited a differential response on Polyacrylamide entrapment. Thus, D-amino acid oxidase activity was stabilized to heat inactivation whereas catalase in the same cells showed a destabilization on entrapment in Polyacrylamide. The coimmobilized enzyme preparation showed an operational half life of 7-9 days after which the D-amino acid oxidase activity remained stable at a value 35–40% of that of the initial activity for a study period of 3 weeks. Coimmobilization of MnO2 was not effective in enhancing the operational life of the enzyme preparation.
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A simple method using charcoal treatment was developed for the preparation of apo-D-amino acid oxidase from rat kidney homogenates. This apo-D-amino acid oxidase was used to study the effect of progesterone on the apo- and holo-enzyme. Progesterone inhibited the activity of D-amino acid oxidase, when the apo- enzyme, preincubated with saturating amounts of FAD was used; this effect varied with FAD concentration. Progesterone did not inhibit the activity when added to a mixture of non-preincubated apo-enzyme and FAD; this suggests that progesterone has different effects on apo- and holo D-amino acid oxidase.