Detection of cathepsin L in red cell membranes from fish blood

Cathepsin L, one of the cysteine proteases found in fish muscle, is considered to be the main cause of post-mortem autolysis of fish muscle. We have determined the presence of cathepsin L in the membranes of red blood cells of carp, amberjack, and red sea bream and measured its activity. Immunoblotting of an extract of the red blood cell membranes from these three fish species using human anti-cathepsin L antibody revealed the presence of cathepsin L of different molecular masses. The molecular masses of cathepsin L was estimated to be 120 and 85 kDa in the amberjack and 75 and 70 kDa in the carp. These proteins have higher molecular masses than the mature form of cathepsin L, suggesting that they are precursor forms. In contrast, the protein in the red sea bream was estimated to have a molecular mass of 30 kDa, suggesting that this cathepsin L is a mature form. The specific activity of cathepsin L was highest in the red blood cell membranes of the amberjack, followed by the carp and the red sea bream in descending order.     

Characterisation of starch properties and physical characteristics in buckwheat (Fagopyrum esculentum Moench.) mutant lacking accumulation of ‘granule-bound starch synthase a’

The concentration of amylose, which is synthesised using granule-bound starch synthase, affects the physical properties of food. However, no studies have focused on starch properties and physical characteristics of low-amylose buckwheat. Here, we hypothesised that low-amylose buckwheat would be useful to produce new buckwheat products because low-amylose characteristics change the texture of buckwheat food. In this study, we bred relatively low-amylose buckwheat compared to wild type and investigated the causative genes of the traits, starch properties and physical properties of noodles. In the GBSSa mutant, the amylose concentration was lower than that in the wild type. Compared with the wild type, the mutant exhibited the following traits: Amylose concentration decreased by approximately 2%, setback in the Rapid Visco Analyzer decreased by 30 points and the physical characteristics of noodles in the sensory analysis were soft and sticky. These results suggest that this trait may be useful for changing the texture of foods. In addition, the mutant is promising for producing new foods with physical characteristics that are different from those of the wild type.     

Expressed sequence tag analysis of phyllosomas and hemocytes of Japanese spiny lobster Panulirus japonicus

Two complementary DNA (cDNA) libraries were constructed from phyllosomas and hemocytes of adult Japanese spiny lobster Panulirus japonicus and a total of 2,673 expressed sequence tags (ESTs) were obtained. After assembly and clustering, 450 and 458 unique sequences were found from the phyllosoma and hemocyte cDNA libraries, respectively. Of these, 114 and 220 ESTs showed significant homologies with known genes in the National Centre for Biotechnology Information (NCBI) database. The remaining sequences were of unknown function. Immune-related genes found in this study include lectin, proteinase inhibitor, prophenoloxidase, heat-shock protein, antimicrobial peptide, and a few putative defense-related proteins.     

Simultaneous steroids measurement in dogs with hyperadrenocorticism using a column-switching liquid chromatography-tandem mass spectrometry method

We developed an analytical method using an on-line column-switching liquid chromatography with triple quadrupole mass spectrometry (LC/MS/MS) for quantifying multiple steroids in serum. Using the developed method, we evaluated the serum concentration of nine steroids (cortisol, corticosterone, cortisone, 11-deoxycortisol, 21-deoxycortisol, deoxycorticosterone, progesterone, 17α-OH-progesterone and aldosterone) in dogs with hyperadrenocorticism (HAC). Serum was mixed with stable isotope internal standards and thereafter purified by the automated column-switching system. The limit of detection ranged 2–16 pg/ml for nine steroids. In the baseline samples, five steroids (cortisol, corticosterone, cortisone, 11-deoxycortisol, and 17α-OH-progesterone) were detected in all dogs. The concentrations of cortisone, 11-deoxycortisol, and 17α-OH-progesterone in dogs with HAC (n=19) were significantly higher those in dogs without HAC (n=15, P<0.02). After the adrenocorticotropic hormone stimulation test, six steroids (cortisol, corticosterone, cortisone, 11-deoxycortisol, 17α-OH-progesterone, and deoxycorticosterone) were above the limit of quantification in all dogs. Cortisol, corticosterone, cortisone, and deoxycorticosterone concentrations of dogs with HAC were significantly higher than those of dogs without HAC (P<0.02). In addition, 11-deoxycortisol and 17α-OH-progesterone concentration was higher in dogs with HAC than in dogs without HAC (P=0.044 and P=0.048, respectively). The on-line column-switching LC/MS/MS would be feasible for measuring multiple steroids in dog serum. The results suggest that cortisone, 11-deoxycortisol, and 17α-OH-progesterone would be related to HAC. Further studies are warranted to assess the clinical feasibility of steroid profile in dogs with HAC.     

Arteriovenous shunts resembling patent ductus arteriosus in dogs: 3 cases

Three dogs presented for the evaluation of cardiac murmurs were diagnosed with aberrant arteriovenous shunts. All cases demonstrated the following findings: 1) relatively soft continuous murmur loudest at the left heart base resembling patent ductus arteriosus (PDA); 2) shunt flow signals in the pulmonary artery on echocardiography; and 3) no PDA on selective angiography, but evidence of anomalous shunting vessels from thoracic aorta to pulmonary vasculature. An aberrant arteriovenous shunt should be considered when a continuous murmur of relatively small intensity is heard.     

A generic model of thinning and stand density effects on forest growth, mortality and net increment

? For assessing forest thinning effects at large (i.e. continental) scale, data scarcity and technical limitations prevent the application of localized or individual-based thinning models.

? Here we present a simple general framework to analyze and predict the effects of thinning on growth and mortality, including the following stand density development. The effects are modeled in relative terms so that the model can be parameterized based on any thinning experiment that includes an unthinned control, regardless of site conditions and stand age.

? The model was tested against observed thinning effects on growth and mortality from five temperate and boreal species (all species pooled r ² = 0.51). It predicted a maximum increase in net stem biomass increment of 16% and a reduction in density-related mortality of 75% compared to un-thinned conditions at stand densities of around 70% of the maximum (increment optimal density).

? A sensitivity analysis revealed overlapping ranges of near optimal density (net increment within 95% of optimal) among all tested species, suggesting that one thinning scenario can be used for many species. The simple and general formulation of thinning effects based on only five parameters allows easy integration with a wide range of generic forest growth models.

     

Effects of cooling rate on post-thaw motility and fertility of Japanese pearl oyster <Emphasis Type="Italic">Pinctada fucata martensii</Emphasis> spermatozoa

The post-thaw motility and fertility of Japanese pearl oyster sperm show large variances, even among sperm samples obtained from the same individuals. This study aimed to clarify the factors that cause such differences. Spermatozoa were diluted 50 times with diluent comprising 10 % methanol, 18 % fetal bovine serum, and 72 % seawater, and dispensed into 0.25 ml straws. A total of 59 straws were cooled, one by one, at 11 different heights from the surface of liquid nitrogen (LN) to −50 °C, and then immediately immersed in LN. After thawing the straws, the relationships between the cooling rate and the post-thaw motility and post-thaw fertility of the spermatozoa were examined. Both the post-thaw motility and the post-thaw fertility showed a sharp peak when the straws were cooled at around −20 °C/min. There was a strong correlation between post-thaw motility and fertility (P < 0.001). There was a large difference in the cooling rates and the post-thaw motilities and fertilities of the spermatozoa, even between straws cooled at the same height. These results indicate that the optimum range for the cooling rate of oyster spermatozoa is quite narrow, and the method of cooling straws at a fixed distance from the LN surface is unsuitable for the cryopreservation of Japanese pearl oyster spermatozoa.     

Behavioural analysis of a nonionic detergent in the kraft pulp washing process using cryo-time-of-flight secondary ion mass spectrometry and cryo-scanning electron microscopy

In order to understand the kraft pulp decolouring mechanism on using a nonionic detergent, the pulp washing process and the resulting pulp handsheets were investigated by examining the brightness, kappa number, thioacidolysis product yield, and dewatering efficiency in the pressing sheet making process. The pulp decolouring could be attributed to a decrease in the lignin content and an improvement in the dewatering efficiency. Furthermore, the detergent distribution in the aqueous pulp suspension obtained during the pulp washing process was visualised using cryo-time-of-flight secondary ion mass spectrometry/scanning electron microscopy (cryo-TOF-SIMS/SEM). The detergent was clearly observed at the transverse surface of the pulp fibre cell wall and was also detected in the lumen of the fibres, suggesting the permeation of the detergent into the pulp fibre cell wall. Based on these results, the pulp decolouring mechanism can be proposed as follows: the detergent permeates into the pulp fibre cell wall and promotes the solution-exchange between the inside and the outside parts of the fibre cell wall, finally washing away the chromophoric substances such as lignin and its degradation products owing to the enhanced dewatering efficiency.