Quantitative measurement of influenza virus replication using consecutive bronchoalveolar lavage in the lower respiratory tract of a ferret model

The ferret is an established animal model of influenza virus infection. Although viral replication in the upper respiratory tract is usually measured with consecutively collected nasal washes, daily evaluation of viral replication in the lung is limited because a large numbers of ferrets need to be sacrificed at consecutive time points. To overcome this limitation, we performed a virus quantification assay using bronchoalveolar lavage (BAL) fluid. This non-invasive BAL technique allows consecutive quantification of virus replication in the lungs of living ferrets. Our method can be used for the longitudinal evaluation of virus tropism in the lower respiratory tract.     

Reactive oxygen scavenging effect of enzymatic extracts from Sargassum thunbergii

The free radical scavenging activity of water soluble natural antioxidants from Sargassum thunbergii, which is a brown marine alga, was evaluated by examining the radical scavenging activities of the extracts of hydrolyzates from S. thunbergii on hydroxyl, 1,1-diphenyl-2-picrylhydrazyl (DPPH), and alkyl radicals. A spin-trapping electron spin resonance (ESR) spectrometer was employed, and the results were compared for their ESR signal intensity. S. thunbergii was enzymatically hydrolyzed to prepare water soluble extracts by five carbohydrases (AMG, Celluclast, Termamyl, Ultraflo, and Viscozyme) and proteases (Alcalase, Flavorzyme, Kojizyme, Neutrase, and Protamex). The scavenging activity of the radicals increased with increased concentrations of the extracts. The scavenging results were higher for hydroxyl and alkyl radicals and lower for DPPH radical as compared with vitamin C as a reference. The hydrogen peroxide scavenging activity of the extracts was also investigated; the Alcalase extract showed the highest scavenging activity among the extracts prepared with the five proteases and five carbohydrates. In addition, the DNA damage was determined by using the comet assay with alkaline electrophoresis and was quantified by measuring the tail length. The preventive effect of Alcalase extract from S. thunbergii against DNA damage increased with increments of concentration of the enzymatic extracts.     

Reevaluation of the Phosphorus Requirement of Juvenile Olive Flounder Pavalichfhys olivaceus and the Bioavailability of Various Inorganic Phosphorus Sources

A 6‐wk feeding trial was conducted to reevaluate the phosphorus (P) requirement of juvenile olive flounder and the bioavailability of various inorganic phosphorus sources (IPS). Eight experimental diets were prepared such that all diets contained the same amount of calories, nitrogen, and calcium. Each diet included 0.33% total phosphorus (TP) and 0.60 % total calcium supplied by the basal diet. The eight experimental diets were: the basal diet without P supplementation (BD), three diets consisting of the BD supplemented with NaH,PO₄.2H₂O (NaP_0.45 NaP_0.57 or NaP_1.14) to supply 0.45, 0.57 or 1.14% TP, and four diets consisting of the BD supplemented with K₂HPO₄ (KP_0.57), Ca(H₂PO₄)_;H₂O (Cap_0.57), CaH₂PO_4;2H₂O (CaHP_0.57) or flounder bone meal (FBP_0.57) to supply 0.57% TP. Fish (N = 480)averaging 4.02 ± 0.03 g (Mean ± SD) were distributed randomly into 24 aquaria (20 fish per aquarium), and were fed one of the eight experimental diets in triplicate groups. The weight gain (WG), specific growth rate (SGR), protein efficiency ratio (PER) and feed efficiency (FE) of fish fed the NaP_0.57 diet were significantly higher than those of fish fed the BD, NaP_1.14, KP_0.57, CaHP_0.57 and FBP_0.57 diets (P > 0.05). There was no significant difference in WG, SGR, PER, and FER among fish fed the Nap_0.45, NaP_0.57 and Cap_0.57, diets. Whole body P retention (WBP_ret) in fish fed the Nap_0.57 diet was significantly higher than in fish fed the BD, NaP_1.14, KP_0.57 CaHP_0.57 and FBP_0.57diets (P > 0.05). There was no significant difference in WBP_ret among fish fed the NaP_0.47, NaP_0.57, and CaP_0.57, diets. The ability of the fish to digest the phosphorus in the NaP_0.45, Nap_0.57, NaP_1.14, and CaP_0.57 diets was significantly better than that of fish fed the other diets (P > 0.05). These results indicated that the dietary P requirement for juvenile olive flounder could be 0.45457%. Also, NaH,PO_; 2H₂O and Ca(H₂PO₄)_;H₂O appeared to have a better bioavailability than the other P sources in juvenile olive flounder.     

Isolation and identification of phlorotannins from Ecklonia stolonifera with antioxidant and hepatoprotective properties in tacrine-treated HepG2 cells

Four kinds of phlorotannins having antioxidant activity were isolated from the ethyl acetate fraction of Ecklonia stolonifera ethanolic extract. The structures of the phlorotannins were determined on the basis of spectroscopic evidence, including 1D and 2D nuclear magnetic resonance. The isolated phlorotannins showed potential radical-scavenging activities against 2,2-diphenyl-1-picrylhydrazyl and suppressed the intracellular reactive oxygen species in tacrine-treated HepG2 cells. Among them, eckol and 2-phloroeckol showed hepatoprotective activity in tacrine-treated HepG2 cells; however, phlorofucofuroeckol B and 6,6'-bieckol did not show the activity, even though having high antioxidant activity. Both eckol and 2-phloroeckol inhibited the expression of Fas-mediated cell-death proteins, including tBid, caspase-3, and poly(ADP-ribose) polymerase, and suppressed the release of cytochrome c from mitochondria to cytosol in a dose-dependent manner in tacrine-treated HepG2 cells. These results suggest that eckol and 2-phloroeckol are the principal hepatoprotective constituents of the ethyl acetate fraction of E. stolonifera ethanolic extract.     

Attenuating Effects of Dieckol on High-Fat Diet-Induced Nonalcoholic Fatty Liver Disease by Decreasing the NLRP3 Inflammasome and Pyroptosis

Nonalcoholic fatty liver disease (NAFLD), which promotes serious health problems, is related to the increase in the nucleotide-binding oligomerization domain-like receptor family, pyrin domain containing 3 (NLRP3) inflammasome and pyroptosis by a high-fat diet (HFD). Whether dieckol (DK), a component of Ecklonia cava extracts (ECE), attenuated NAFLD in an HFD-induced NAFLD animal model was evaluated. The expression of high mobility group box 1/Toll-like receptor 4/nuclear factor-κB, which initiated the NLRP3 inflammasome, was increased in the liver of HFD-fed animals and significantly decreased with ECE or DK administration. The expression of NLRP3/ASC/caspase-1, which are components of the NLRP3 inflammasome, and the number of pyroptotic cells were increased by HFD and decreased with ECE or DK administration. The accumulation of triglycerides and free fatty acids in the liver was increased by HFD and decreased with ECE or DK administration. The histological NAFLD score was increased by HFD and decreased with ECE or DK administration. The expression of lipogenic genes (FASN, SREBP-2, PPARγ, and FABP4) increased and that of lipolytic genes (PPARα, CPT1A, ATGL, and HSL) was decreased by HFD and attenuated with ECE or DK administration. In conclusion, ECE or DK attenuated NAFLD by decreasing the NLRP3 inflammasome and pyroptosis.     

Hematological adverse effects and pharmacokinetics of ribavirin in pigs following intramuscular administration

Ribavirin (RBV) is a synthetic guanosine analog that is used as a drug against various viral diseases in humans. The in vitro antiviral effects of ribavirin against porcine viruses were demonstrated in several studies. The purposes of this study were to evaluate the adverse effects and pharmacokinetics of ribavirin following its intramuscular (IM) injection in pigs. Ribavirin was formulated as a double‐oil emulsion (RBV‐DOE) and gel (RBV‐Gel), which were injected into the pigs as single‐dose IM injections. After injection of RBV, all of the pigs were monitored. The collected serum and whole blood samples were analyzed by liquid chromatography–tandem mass spectrometry and complete blood count analysis, respectively. All of the ribavirin‐treated pigs showed significant decreases in body weight compared to the control groups. Severe clinical signs including dyspnea, anorexia, weakness, and depression were present in ribavirin‐treated pigs until 5 days postinjection (dpi). The ribavirin‐treated groups showed significant decrease in the number of red blood cells and hemoglobin concentration until 8 dpi. The mean half‐life of the RBV‐DOE and RBV‐Gel was 27.949 ± 2.783 h and 37.374 ± 3.502 h, respectively. The mean peak serum concentration (C_max) and area under the serum concentration–time curve from time zero to infinity (AUC_inf) of RBV‐DOE were 8340.000 ± 2562.577 ng/mL and 16 0095.430 ± 61 253.400 h·ng/mL, respectively. The C_max and AUC_inf of RBV‐Gel were 15 300.000 ± 3764.306 ng/mL and 207526.260 ± 63656.390 h·ng/mL, respectively. The results of this study provided the index of side effect and pharmacokinetics of ribavirin in pigs, which should be considered before clinical application.     

Indigenous parasitoids as effective natural enemies of <Emphasis Type="Italic">Phyllocnistis citrella</Emphasis> (Lepidoptera: Gracillariidae) in Korea

This study is aimed to investigate the occurrence of Phyllocnistis citrella Stainton (citrus leafminer; CLM), which has recently caused increasing damage to citrus farms in southern South Korea, and its natural enemies, which can act as biological control agents. In the present study, CLM monitoring using sex pheromone traps were conducted in the major citrus farms in the Jeju region in South Korea. Also a survey of parasites was carried out to detect the effective control agent for CLM. During the investigation from 2011 to 2015, a relatively large number of adult CLM (2,386,990) were observed in the sex pheromone traps. The mean annual occurrence of CLM was 477,398, which showed an increasing trend with the increase in temperature. A survey of parasites was conducted in November 2015. Six parasites were found, of which, Quadrastichus sp. and Sympiesis sp. showed the highest rates of parasitism. The mean rate of parasitism during the survey period was 3.97 %. We discovered four new species of parasites of CLM in South Korea.     

Ishophloroglucin A,Isolated from Ishige okamurae,Alleviates Dexamethasone-Induced Muscle Atrophy through Muscle Protein Metabolism In Vivo

The in vitro capacity of Ishige okamurae extract (IO) to improve impaired muscle function has been previously examined. However, the mechanism underlying IO-mediated muscle protein metabolism and the role of its component, Ishophloroglucin A (IPA), in mice with dexamethasone (Dexa)-induced muscle atrophy remains unknown. In the present study, we evaluated the effect of IO and IPA supplementation on Dexa-induced muscle atrophy by assessing muscle protein metabolism in gastrocnemius and soleus muscles of mice. IO and IPA supplementation improved the Dexa-induced decrease in muscle weight and width, leading to enhanced grip strength. In addition, IO and IPA supplementation regulated impaired protein synthesis (PI3K and Akt) or degradation (muscle-specific ubiquitin ligase muscle RING finger and atrogin-1) by modulating mRNA levels in gastrocnemius and soleus muscles. Additionally, IO and IPA upregulated mRNA levels associated with muscle growth activation (transient receptor potential vanilloid type 4 and adenosine A1 receptor) or inhibition (myostatin and sirtuin 1) in gastrocnemius and soleus muscle tissues of Dexa-induced mice. Collectively, these results suggest that IO and IO-derived IPA can regulate muscle growth through muscle protein metabolism in Dexa-induced muscle atrophy.     

Effects of irradiated phytic acid on antioxidation and color stability in meat models

Lipid oxidation and color stability of meats treated with irradiated phytic acid were investigated during storage for 2 weeks at 4 degrees C. The phytic acid in deionized distilled water (DDW) was degraded by irradiation at 10 and 20 kGy, and the irradiated phytic acid showed a strong antiradical activity. For measuring the antioxidant effects of irradiated phytic acid in food models, beef and pork were prepared with DDW (control), irradiated (10 and 20 kGy) or non-irradiated phytic acid, and ascorbic acid as a model system. Irradiated phytic acid significantly inhibited the lipid oxidation in meats compared to the control and ascorbic acid treated samples during storage (P < 0.05). The redness of the meats treated with phytic acid had a higher value than did the control and ascorbic acid treated samples, but a significant difference was not observed in the samples treated with phytic acid regardless of irradiation treatment. Irradiated phytic acid was also effective in inhibiting the loss of heme iron and metmyoglobin formation during storage. Results indicated that irradiation might be helpful for improving the antioxidant activity of phytic acid in meats.     

Cytotoxic and antitumor activities of thiosulfinates from Allium tuberosum L

In this study we isolated crude thiosulfinates from Allium tuberosum L. using CH 2Cl 2 and then with silica gel column chromatography purified S-methyl methanthiosulfinate and S-methyl 2-propene-1-thiosulfinate from the crude thiosulfinates. Subsequently, in vitro cytotoxicities against human cancer cells and in vivo antitumor activities of the thiosulfinates were investigated. Their cytotoxicities were strong in human cancer cells, in the order of S-methyl 2-propene-1-thiosulfinate, crude thiosulfinates, and S-methyl methanthiosulfinate. When thiosulfinates were administered consecutively for 7 days at 10, 30, and 50 mg/kg ip, in mice, we found significant increases in the life spans of mice that had been inoculated with Sacorma-180 tumor cells. The crude thiosulfinates also induced apoptosis in MCF-7 cancer cells. These results suggest that thiosulfinates from Allium tuberosum L. inhibit the proliferation of cancer cells via apoptosis and have antitumor activities.