Inhibition of post-mortem muscle softening following in situ perfusion of protease inhibitors in tilapia

ABSTRACT:     A method of introducing protease inhibitors into fish muscle through the bulbus arteriosus was developed using an in situ perfusion technique. Perfusion efficiency was initially tested using eosin and [³⁵S]-methionine. Visible fluorescence was observed in the gill, liver, intestine and dorsal muscle of the eosin-treated tilapia, and the occurrence of eosin in the blood vessels of the dorsal muscle was confirmed under a fluorescence stereoscopic microscope with ultraviolet light. The radioactivity of [³⁵S]-methionine was taken into the dorsal muscle and liver at a concentration of 7.8 Bq/g and 70.2 Bq/g, respectively, after perfusion with 1000 Bq/mL solution. Using the perfusion technique with four kinds of protease inhibitors dissolved in physiological saline, the type of proteases implicated in the post-mortem muscle softening in tilapia (867 ± 195 g, n  = 10/protease inhibitor) was investigated. After the perfusion of leupeptin (serine and cysteine protease inhibitor), benzyloxycarbonyl-Val-Ala-Asp-fluoromethylketone (Z-VAD-fmk; caspase inhibitor), chymostatin (serine protease inhibitor) and ο -phenanthroline (metalloprotease inhibitor), the breaking strength of the perfused muscle was measured as a parameter of the meat toughness and compared with that of the control fish, which were perfused with physiological saline only. The reduction of breaking strength during storage was inhibited by the perfusion of leupeptin and Z-VAD-fmk.     

The effects of concentrations of lysine in media on differentiation of 3T3‐L1 preadipocytes

Intramuscular fat content is increased by feeding of low lysine diets in pigs. Reduction in dietary lysine intake results in low plasma lysine concentration and low cytosolic lysine concentration in skeletal muscles. From these observations, we hypothesized that low plasma lysine concentration in pigs fed on low lysine diets reduced supply of lysine from blood circulation to preadipocytes, and this limited supply of lysine might promote adipocyte differentiation in porcine muscles. In order to verify the hypothesis, we investigated the effects of low concentrations of lysine in culture medium on differentiation of 3T3‐L1 preadipocytes. Low concentration of lysine suppressed lipid accumulation and messenger RNA (mRNA) expression and enzyme activity of fatty acid synthase. mRNA expressions of peroxisome proliferator‐activated receptor γ (PPARγ) and CCAAT/enhancer binding protein α (C/EBPα) were lower in cells cultured in low lysine medium. On the other hand, mRNA and protein expressions of C/EBPβ and C/EBPδ were not inhibited by low concentrations of lysine in culture medium. These results indicate that low lysine concentrations in culture medium inhibit differentiation of 3T3‐L1 preadipocytes through inhibiting the mRNA expressions of PPARγ and C/EBPα.     

Application of microparticle diets for Japanese flounder Paralichthys olivaceus larvae

ABSTRACT:   A feeding trial was conducted with two newly developed microparticle diets (MD-Q and MD-V), which differed in the mixing ratios of two types of casein hydrolysates (C700 and C800) as the protein source. The ratio of C700 and C800 were adjusted to 7:3 in MD-Q and 6:4 in MD-V, respectively. Japanese flounder larvae were fed from 7 days after hatching (d.a.h.) to 32 d.a.h. on live food (LF), MD solely, MD + 1/3 LF (either of the MD and one-third quantity of the live food) and 1/3 LF (one-third quantity of live food alone), respectively. They were then switched to Kyowa diet B from 33 d.a.h. until 40 d.a.h. The larvae fed on MD-Q exclusively had a higher survival rate (36.4%) than those fed on MD-V (24.2%) by 22 d.a.h. Unfed larvae could survive up to only 12 d.a.h. Larvae fed on MD-Q + 1/3 LF also had a significantly higher survival rate (34.2%) than those on MD-V + 1/3 LF (16.3%) and 1/3 LF solely (15.4%) at 32 d.a.h. These results suggest the potential of MD-Q as a part replacement for LF from the early developmental stage in the seed production of Japanese flounder.     

Environmental effects on recruitment and productivity of Japanese sardine Sardinops melanostictus and chub mackerel Scomber japonicus with recommendations for management

We compared a wide range of environmental data with measures of recruitment and stock production for Japanese sardine Sardinops melanostictus and chub mackerel Scomber japonicus to examine factors potentially responsible for fishery regimes (periods of high or low recruitment and productivity). Environmental factors fall into two groups based on principal component analyses. The first principal component group was determined by the Pacific Decadal Oscillation Index and was dominated by variables associated with the Southern Oscillation Index and Kuroshio Sverdrup transport. The second was led by the Arctic Oscillation and dominated by variables associated with Kuroshio geostrophic transport. Instantaneous surplus production rates (ISPR) and log recruitment residuals (LNRR) changed within several years of environmental regime shifts and then stabilized due, we hypothesize, to rapid changes in carrying capacity and relaxation of density dependent effects. Like ISPR, LNRR appears more useful than fluctuation in commercial catch data for identifying the onset of fishery regime shifts. The extended Ricker models indicate spawning stock biomass and sea surface temperatures (SST) affect recruitment of sardine while spawning stock biomass, SST and sardine biomass affect recruitment of chub mackerel. Environmental conditions were favorable for sardine during 1969–87 and unfavorable during 1951–67 and after 1988. There were apparent shifts from favorable to unfavorable conditions for chub mackerel during 1976–77 and 1985–88, and from unfavorable to favorable during 1969–70 and 1988–92. Environmental effects on recruitment and surplus production are important but fishing effects are also influential. For example, chub mackerel may have shifted into a new favorable fishery regime in 1992 if fishing mortality had been lower. We suggest that managers consider to shift fishing effort in response to the changing stock productivity, and protect strong year classes by which we may detect new favorable regimes.     

Mechanism of body cavity temperature regulation of chum salmon (Oncorhynchus keta) during homing migration in the North Pacific Ocean

Vertical movement patterns of five chum salmon (Oncorhynchus keta) during homing migration were examined using archival tags. The standard deviation of the depth and ambient and body cavity temperatures during daytime were larger than those during night‐time. Vertical movements through the thermocline with a periodicity of less than 1 h were observed during daytime in addition to the diel vertical movement patterns in the open ocean. During these periods of frequent short‐term vertical movements, the difference between the body cavity temperature and ambient temperature was large while the variance of the body cavity temperature was less than that of the ambient temperature. From the results of a random simulation, the variation of the body cavity temperature was shown to decrease due to these periodic high frequency movements in comparison with random vertical movements. The whole‐body heat‐transfer coefficient k (s^?1), which was estimated by a heat budget model, was 1.48 × 10^?3. The k of chum salmon was larger than that of bigeye tuna (Thunnus obesus) by about one order of magnitude for the cooling of the body. The k of chum salmon did not change like tuna, which are physiologically adapted to conserve body cavity temperature. This indicates that the regulation of body cavity temperature by chum salmon is dependent on the vertical movements only. The maintenance of the body cavity temperature is concluded to be advantageous for their maturation and growth from the relationship between energy input and output during their homing migration.     

Carotenoids in marine invertebrates living along the Kuroshio current coast

Carotenoids of the corals Acropora japonica, A. secale, and A. hyacinthus, the tridacnid clam Tridacna squamosa, the crown-of-thorns starfish Acanthaster planci, and the small sea snail Drupella fragum were investigated. The corals and the tridacnid clam are filter feeders and are associated with symbiotic zooxanthellae. Peridinin and pyrrhoxanthin, which originated from symbiotic zooxanthellae, were found to be major carotenoids in corals and the tridacnid clam. The crown-of-thorns starfish and the sea snail D. fragum are carnivorous and mainly feed on corals. Peridinin-3-acyl esters were major carotenoids in the sea snail D. fragum. On the other hand, ketocarotenoids such as 7,8-didehydroastaxanthin and astaxanthin were major carotenoids in the crown-of-thorns starfish. Carotenoids found in these marine animals closely reflected not only their metabolism but also their food chains.     

Granulomatous Nephritis Consistent with Malakoplakia in a Cynomolgus Monkey

Malakoplakia is a rare form of chronic granulomatous inflammation in mammals, and usually affects the urinary tract in humans. In this report, we present a case of granulomatous nephritis consistent with malakoplakia in a 4-year-old male cynomolgus monkey. Gross examination showed that the kidney was markedly enlarged and adhered to the surrounding organs. Histology showed that there was diffuse interstitial infiltration of histiocytes with abundant foamy eosinophilic cytoplasm resembling von Hansemann cells, PAS-positive granular cytoplasm and occasional PAS- and iron-positive intracellular small inclusion bodies. Electron microscopy showed that these histiocytes contained abundant lysosomes and phagolysosomes but no obvious Michaelis-Gutmann bodies. Based on these findings, a diagnosis of granulomatous nephritis consistent with early malakoplakia was made. This is the first report in a monkey of a renal lesion consistent with malakoplakia.     

Proposal of screening method for intestinal mucus adhesive lactobacilli using the enzymatic activity of glyceraldehyde‐3‐phosphate dehydrogenase (GAPDH)

Adhesion tests are complex, time‐consuming and expensive, while the most important criterion for a probiotic lactobacilli is the ability to adhere to the human intestine. Thirty lactobacilli isolates from human intestinal tissues were measured for cell surface glyceraldehyde‐3‐phosphate dehydrogenase (GAPDH) activity using a microtiter plate screening method. GAPDH activities were detected in 21 out of 30 samples from 12 h cultures and in all samples from 18 h cultures. This suggests GAPDH is universally expressed on the bacterial cell surfaces from many lactobacilli. A statistically significant positive correlation was shown between GAPDH activity and adhesion using the BIACORE adhesion assay (P < 0.01). The new screening method using GAPDH enzymatic activity without an adhesion test may be possible due to the significant positive correlation of GAPDH activity with adhesion of lactobacilli derived from the human intestine.     

Suppression of feline coronavirus replication in vitro by cyclosporin A

ABSTRACT: The feline infectious peritonitis virus (FIPV) is a member of the feline coronavirus family that causes FIP, which is incurable and fatal in cats. Cyclosporin A (CsA), an immunosuppressive agent that targets the nuclear factor pathway of activated T-cells (NF-AT) to bind cellular cyclophilins (CyP), dose-dependently inhibited FIPV replication in vitro. FK506 (an immunosuppressor of the pathway that binds cellular FK506-binding protein (FKBP) but not CyP) did not affect FIPV replication. Neither cell growth nor viability changed in the presence of either CsA or FK506, and these factors did not affect the NF-AT pathway in fcwf-4 cells. Therefore, CsA does not seem to exert inhibitory effects via the NF-AT pathway. In conclusion, CsA inhibited FIPV replication in vitro and further studies are needed to verify the practical value of CsA as an anti-FIPV treatment in vivo.     

不同消毒方式对设施草莓花药愈伤组织诱导的影响

以栽培品种京藏香草莓(Fragaria ananassa Duch.)花药(单核靠边期)为材料,75%乙醇为对照,研究了5种不同组合的消毒方式对草莓花药愈伤组织的诱导影响。结果表明,处理T_1经过Tween-80消毒后,用75%乙醇进行消毒30 s可以有效控制花药污染,同时还能减少花药褐化率;其次是处理T_3,该处理虽能减少污染,但是褐化率略高于T_1;处理T_0(CK)、T_2均有不同程度的污染,处理T4褐化率最高。