Screening of Human CYP1A2 and CYP3A4 Inhibitors from Seaweed In Silico and In Vitro

Phenolic compounds and carotenoids are potential inhibitors of cytochrome P450s. Sixteen known compounds, phenolic compounds and carotenoids from seaweed were examined for potential inhibitory capacity against CYP1A2 and CYP3A4 in silico and in vitro. Morin, quercetin, and fucoxanthin inhibited the enzyme activity of CYP1A2 and CYP3A4 in a dose-dependent manner. The IC₅₀ values of morin, quercetin, and fucoxanthin were 41.8, 22.5, and 30.3 μM for CYP1A2 and 86.6, 16.1, and 24.4 μM for CYP3A4, respectively. Siphonaxanthin and hesperidin did not show any significant effect on CYP1A2, but they slightly inhibited CYP3A4 activity at high concentrations. In silico modeling of CYP’s binding site revealed that the potential inhibitors bound in the cavity located above the distal surface of the heme prosthetic group through the 2a or 2f channel of CYPs. This study presents an approach for quickly predicting CYP inhibitory activity and shows the potential interactions of compounds and CYPs through in silico modeling.     

Glutathione content of in vivo and in vitro matured canine oocytes collected from different reproductive stages

Glutathione (GSH) concentrations of oocytes are considered as an important marker of the cytoplasmic maturation. The present study was designed to compare GSH concentrations of in vivo and in vitro matured canine oocytes. In vivo matured oocytes were collected 72 hr after ovulation by flushing fallopian tubes after laparotomy. Ovaries were collected from bitches with different reproductive stages, and collected oocytes were divided into 2 groups according to the size viz. < 120 microm and > 120 microm in diameter and cultured for 72 hr in Tissue Culture Medium-199 supplemented with 10% FBS, 2.2 mg/ml sodium bicarbonate, 2.0 microg/ml estrogen, 0.5 microg/ml FSH, 0.03 IU/ml hCG, and 1% penicillin-streptomycin solution in the presence or absence of 50 microM beta-mercaptoethanol. GSH concentrations were determined by the dithionitrobenzoic acid-glutathione disulfide (DTNB-GSSG) reductase recycling assay. GSH concentrations of immature canine oocytes were 2.9 and 3.8, 3.5 and 6.8, and 3.1 and 6.5 pM/oocyte for < 120 microm and > 120 microm in diameter oocyte groups at anestrous, follicular and luteal stage, respectively (P<0.05). In vivo matured oocytes had significantly higher GSH concentrations compared with in vitro matured oocytes. The GSH content was 19.2 pM/oocyte for in vivo matured oocytes, while 4.1 to 8.1 and 5.7 to 13.2 pM/oocyte for in vitro matured oocytes cultured in the absence or presence of beta-mercaptoethanol, respectively (P<0.05). Presence of beta-mercaptoethanol increased GSH synthesis in canine oocytes cultured in vitro, and oocytes collected from follicular and luteal stage was superior to anestrus oocytes.     

Disseminated transmissible venereal tumor in a dog.

Transmissible venereal tumor (TVT) is a well-documented transplantable tumor in dogs, with no breed or sex predilection and a low metastatic rate. In this report, a 2-year-old intact female Mastiff that had numerous, rapidly growing masses throughout the subcutis mainly at the dorsal body plane, the caudal half of the ventral abdomen, and around the vulva was euthanized due to poor prognosis. Neoplastic nodules similar to those seen in the subcutis were also noted in the lung, anterior mediastinum, liver, spleen, kidney, and superficial and deep lymph nodes in both abdominal and thoracic cavities. The neoplastic nodules from the subcutis as well as metastatic foci revealed similar cytologic and histologic features, which were consistent with canine TVT. By immunohistochemical staining, the neoplastic cells were positive for lysozyme and vimentin but were negative for cytokeratin, desmin, CD3, and CD79a. The diagnosis of the TVT was further supported by the identification and analysis of long interspersed nuclear elements (LINE) from paraffin-embedded tumor tissue. This case is a rare example of TVT with multiorgan metastasis. In this case, the polymerase chain reaction technique was useful in differential diagnosis of canine round cell tumors because this technique can be applied in retrospective as well as future study.     

Development and evaluation of an indirect enzyme-linked immunosorbent assay for detection of foot-and-mouth disease virus nonstructural protein antibody using a chemically synthesized 2B peptide as antigen.

Forty peptides were synthesized corresponding to hydrophilic clusters of amino acids within the sequences of foot-and-mouth disease virus (FMDV) nonstructural proteins (NSP). Six peptides were studied in more detail and the most promising, a 2B peptide, was evaluated in enzyme-linked immunosorbent assay (ELISA) using sera from naive, vaccinated, and vaccinated-and-challenged cattle as well as bovine sera from field outbreaks. The performance of the new NSP peptide ELISA was compared to that of 4 commercial NSP ELISA kits. Antibody to 2B was detectable from the end of the first week to the second week after infection in most of the nonvaccinated animals and by the second to third week in vaccinated-and-challenged animals. The sensitivity of the 2B peptide ELISA was comparable to the 3ABC Ceditest (Ceditest FMDV-NS, Cedi Diagnostics B.V.; Chung et al., 2002). With some modification and further validation, this 2B test could be useful as a screening or conformational NSP test in postvaccination surveillance for FMD.     

Guided bone regeneration with beta-tricalcium phosphate and poly L-lactide-co-glycolide-co-epsilon-caprolactone membrane in partial defects of canine humerus

This study was performed to evaluate the effect of beta-tricalcium phosphate and poly L-lactide-co-glycolide-co-epsilon-caprolactone (TCP/PLGC) membrane in the repair of partial bone defects in canine proximal humerus. Three adult mixed-breed dogs were used during the experimental period. The length of the defect was quarter of the full length of humerus, and width of the defect was quarter of middle diameter of the lateral aspect of humerus. The humeri of each dog were divided into treatment (TCP/PLGC) and control groups. The defect was covered with TCP/PLGC membrane in treatment group. To evaluate regeneration of the bone, computerized tomography (CT) and histopathologic examination were performed. The radiopaque lines were appeared at the original defect sites in TCP/PLGC group but below the original site in control at 4th week. Radiopacity and thickness of the defect sites, and radiopaque lines were more increased at 8th week than those of 4th week. Histopathologic findings revealed fibrous connective tissue migration into the defect and the migration inhibited the structure of new cortex to be placed in the original level in control whereas new cortex growth was found in the level of original line in TCP/PLGC group. However, the new cortical bone in the TCP/PLGC group was thinner and less organized than the adjacent intact cortex, and the amount of new cancellous bones were also scanty. The result suggested that TCP/PLGC membrane is a good guided bone regeneration material to restore the original morphology of humerus in partial defect.     

Nutrient-flow wick culture system for potted plant production: System characteristics and plant growth

To compliment the current subirrigation systems used for production of potted plants, a nutrient-flow wick culture (NFW) system was developed and compared with other subirrigation systems, such as an ebb and flow culture (EBB) system and a nutrient-stagnant wick culture (NSW) system in relation to their system characteristics and plant growth. Kalanchoe (Kalanchoe blossfeldiana cv. New Alter) was cultivated in a 6 cm pot for 10 weeks in each subirrigation system. The water-absorption pattern of the medium, water content of the medium, water loss, algal growth, salt-buildup and plant growth under various culture systems were observed. The water contents of medium under the NFW and EBB systems showed fluctuations from 30 to 40% and from 50 to 60% (by volume), respectively, whereas the water content under the NSW system gradually increased to over 40% without fluctuation. Relative to other systems, the water loss in the NFW system was 50–70% due to the reduction in the evaporation from the surfaces of the trough and medium. Algae appeared in the NSW system because the nutrient solution was always stagnant in the trough, while it was not observed under the NFW system. The dissolved oxygen in the nutrient solution was the highest during the irrigation period and the salinity in the medium was the lowest in the NFW system. With regard to system characteristics, the NFW system was simple, water-saving and efficient. In addition, the growth of kalanchoes in the NFW system was similar to those in the NSW and EBB systems at an irrigation frequency of five times a day.     

Compensatory growth and body composition of juvenile black rockfish <Emphasis Type="Italic">Sebastes schlegeli</Emphasis> following feed deprivation

ABSTRACT:   Compensatory growth, feeding rate, feed efficiency and chemical composition of juvenile black rockfish (mean weight 1.43 g) were investigated for 35 days after a 14-day feed deprivation treatment under four feeding conditions: one group continuously fed (control) and the other three groups fasted for 5 days (F5), 10 days (F10) and 14 days (F14). All fasted fish were re-fed from day 15. Only F5 achieved the same body weight as the control, indicating that complete compensation occurred in F5. The specific growth rate (SGR) of F5 was the highest at day 21 and then decreased thereafter, showing higher values than the control at days 21, 28 and 42. In contrast, although SGRs of F10 and F14 were higher than that of the control during the whole refeeding period except day 21, they did not catch up the control in body mass, indicating that only partial compensation occurred in F10 and F14. The feeding rate (FR) of all groups except F14 changed in a pattern similar to SGR (Spearman's rank correlation, r _s > 0.9), suggesting that SGR varied depending on FR. Similar feeding efficiencies (FEs) were found in the four groups and they did not vary significantly during the whole refeeding period, suggesting that FE was not the factor affecting SGR. At day 14, the ratios of lipid to lean body mass in F10 and F14 were lower than those in the control and F5, and there was no difference between the control and F5. At day 49, however, only F14 showed a lower value than the other three groups, and there was no difference among the three groups. These results indicate that juvenile black rockfish fasted for 5–14 days can exhibit compensatory growth after refeeding, but timing and degree vary depending on the duration of feed deprivation.