A case of dedifferentiated liposarcoma discovered due to an intrascrotal calcified ossification.

Dedifferentiated liposarcoma is a rare cancer with a poor prognosis. A 52-year-old man presented with a chief complaint of a mass in his left scrotum. He came with suspected testicular tumor, but all the measured tumor markers were negative. Imaging test showed approximately 2 cm diameter mass accompanied by calcification with some substantial components between the testis and epididymis. Left high testicular resection was performed. The tumor had no continuity between the testis and epididymis, and the spermatic cord transection was negative. Pathological findings showed well differentiated fatty component and a dedifferentiated component around the trabecular bone-like tissue. We observed dedifferentiated dysmorphic cells mixed with fatty droplets of unequal size. Immunostaining led to the diagnosis of dedifferentiated liposarcoma. No additional postoperative therapy was performed. The possibility of dedifferentiated liposarcoma should be kept in mind even if mass is confined to the scrotum and consisted of calcification. In the case of an intrascrotal calcified mass with malignant perspective, radical surgery is highly recommended.

Dynamics of D-amino acid oxidase ain kidney epithelial cells under amino acid starvation.

D-amino acid oxidase (DAO) is a flavoenzyme catalyzing the oxidation of D-amino acid (AA)s. In the kidney, its expression is detected in proximal tubules, and DAO is considered to play a role in the conversion of D-form AAs to α-keto acids. LLC-PK1 cells, a pig renal proximal tubule cell line, were used to elucidate the regulation of DAO protein synthesis and degradation. In this study, we showed that trypsinization of LLC-PK1 cells in culture system rapidly reduced the intracellular DAO protein level to ∼33.9% of that before treatment, even within 30 min. Furthermore, we observed that the DAO protein level was decreased when LLC-PK1 cells were subjected to AA starvation. To determine the degradation pathway, we treated the cells with chloroquine and MG132. DAO degradation was found to be inhibited by chloroquine, but not by MG132 treatment. We next examined whether or not DAO was degraded by autophagy. We found that AA starvation led to an increased accumulation of LC3-II, suggesting that DAO protein is degraded by autophagy due to AA starvation conditions. Furthermore, treatment with cycloheximide inhibited DAO protein degradation. Taken together, DAO protein is degraded by autophagy under starvation. The present study revealed the potential dynamics of DAO correlated with renal pathophysiology.

P219L substitution in human D-amino acid oxidase impacts the ligand binding and catalytic efficiency.

Human D-amino acid oxidase (DAO) is a flavoenzyme that is implicated in neurodegenerative diseases. We investigated the impact of replacement of proline with leucine at Position 219 (P219L) in the active site lid of human DAO on the structural and enzymatic properties, because porcine DAO contains leucine at the corresponding position. The turnover numbers (kcat) of P219L were unchanged, but its Km values decreased compared with wild-type, leading to an increase in the catalytic efficiency (kcat/Km). Moreover, benzoate inhibits P219L with lower Ki value (0.7-0.9 µM) compared with wild-type (1.2-2.0 µM). Crystal structure of P219L in complex with flavin adenine dinucleotide (FAD) and benzoate at 2.25 Å resolution displayed conformational changes of the active site and lid. The distances between the H-bond-forming atoms of arginine 283 and benzoate and the relative position between the aromatic rings of tyrosine 224 and benzoate were changed in the P219L complex. Taken together, the P219L substitution leads to an increase in the catalytic efficiency and binding affinity for substrates/inhibitors due to these structural changes. Furthermore, an acetic acid was located near the adenine ring of FAD in the P219L complex. This study provides new insights into the structure-function relationship of human DAO.

Age- and gender-dependent D-amino acid oxidase activity in mouse brain and peripheral tissues: implication for aging and neurodegeneration.

D-amino acid oxidase (DAO) is a flavoenzyme, catalysing oxidative deamination of D-amino acids to produce corresponding α-keto acids, ammonia and hydrogen peroxide. In our search for DAO activity among various tissues, we developed a sensitive assay based on hydrogen peroxide production involving enzyme-coupled colorimetric assay with peroxidase. We first optimized buffer components to extract DAO protein from mouse tissues. Here we show that DAO activity was detected in kidney, cerebellum, medulla oblongata, midbrain and spinal cord, but not in liver. In addition, we observed that DAO activity and expression were decreased in thoracic and lumbar regions of spinal cord in aged mice when compared with young mice, indicating that decreased DAO is involved in motoneuron degeneration during senescence. We also found gender difference in DAO activity in the kidney, suggesting that DAO activity is influenced by sexual dimorphism. We newly detected DAO activity in the epididymis, although undetected in testis. Furthermore, DAO activity was significantly higher in the caput region than corpus and cauda regions of epididymis, indicating that D-amino acids present in the testis are eliminated in epididymis. Taken together, age- and gender-dependent DAO activity in each organ may underlie the human pathophysiology regulated by D-amino acid metabolism.