Lesional expression of thymus and activation-regulated chemokine in canine atopic dermatitis

Glucocorticoids are reported to bias cytokines to a Th2 phenotype. However, this dogma has been advanced largely from studies utilizing potent glucocorticoid analogs. The current study was conducted to revisit the issue of glucocorticoid modulation of Th1/Th2 cytokine production and evaluate migration inhibitory factor (MIF) mRNA expression in cultured pig splenocytes treated with physiologically relevant concentrations of cortisol (CORT). Dexamethasone (DEX) was included for comparison. In Experiment 1, DEX, at 150 and 300 nM, suppressed concanavalin (ConA)-stimulated IFNgamma at both 12 and 24 h in culture, and IL-10 at 24h (P<0.05). Both 150 and 300 nM CORT suppressed IL-10 at 24 h (P<0.05), but neither concentration affected IFNgamma at 24 h. In Experiment 2, cells were cultured with a broader range of CORT for 48 h following ConA. Parallel cultures with identical treatments also were conducted in separate plates for evaluation of glucocorticoid regulation of MIF mRNA. IFNgamma was reduced by 300 nM DEX at 12, 24, and 48 h (P<0.05), whereas 150 and 300 nM CORT blunted IFNgamma at 24 h (P<0.05), but not 48 h. ConA increased IL-2 (P<0.01), but none of the steroid treatments affected IL-2. At both 12 and 24 h, IL-10 was reduced by 300 nM DEX and by 150 and 300 nM CORT (P<0.05). ConA increased relative abundance of MIF mRNA (P<0.001), but no steroid treatment affected MIF mRNA. In Experiment 3, steroid additions were delayed by 24 h after ConA, and cytokine concentrations evaluated 48 h later. Again, separate cultures were used for determination of effect of treatments on MIF mRNA. None of the steroid treatments affected IFNgamma, but 300 nM DEX reduced IL-10 (P<0.05). All of the CORT treatments (75-300 nM) reduced MIF mRNA (P<0.05), whereas DEX did not affect MIF mRNA in this experiment. The current experiments suggest that both DEX and high physiological concentrations of CORT can suppress both type 1 and type 2-like cytokines in cultured pig splenocytes. But, IL-10 was generally more sensitive to CORT suppression with increased time in culture than was IFNgamma. In addition, MIF mRNA could be suppressed by delayed addition of CORT to porcine splenocytes. Taken together, the data do not support the hypothesis that CORT directs the cytokine milieu toward a type 2 bias in cultured pig splenocytes.     

Proliferation of canine intervertebral disk chondrocytes in three-dimensional alginate microsphere culture

Proliferation of chondrocytes from nucleus pulposus (NP) and anulus fibrosus (AF) was confirmed in three-dimensional culture using alginate microspheres. Cells isolated from NP and AF were incorporated in microspheres and cultured for 14 days. Round mononuclear cells of 20-25 microm in diameter proliferated and formed aggregates. At day 14, alcian blue positive matrix surrounded the proliferating cells. The cells had cytoplasmic vacuoles stained positively by toluidine blue. On electron microscopy, the cells contained proteoglycan vacuoles and lipid droplets in the cytoplasm and synthesized collagen fibrils and electron dense granules surrounding the cell. These features of the cells were characteristic for chondrocytes. This culture system should be useful to further investigate metabolic activities of intervertebral disk chondrocytes.     

A retrospective study on adverse reactions to canine vaccines in Japan

Adverse reactions to vaccines were examined in 311 canine cases reported to the Ministry of Agriculture, Forestry and Fisheries in Japan during the period of 6 years from April of 1994 to March of 2000, and classified according to their clinical symptoms. There were 27 cases of adverse reactions to rabies virus vaccines. Gastrointestinal symptoms were the most frequently observed (26%), followed by respiratory and/or cardiovascular symptoms (22%) and dermatologic symptoms (11%). There were 284 cases of adverse reactions to non-rabies monovalent vaccines and mixed vaccines. Dermatologic symptoms were the most frequently observed (53%), followed by gastrointestinal symptoms (16%) and respiratory and/or cardiovascular symptoms (14%). Of the total 311 cases, 11 (3.5%) died of adverse reactions to vaccines.     

Distribution of blastodermal cells transferred to chick embryos for chimera production using windowed eggs

1. To improve the production of chimeras, the distribution of donor blastodermal cells after transferring into recipient embryos was examined morphologically.

2. Donor blastodermal cells were distributed near the site of injection in the epiblast and in the subgerminal cavity and yolk. Some filled the hole made by the micropipette and were distributed outside the epiblast. Many were buried in yolk. In some cases, more donor blastodermal cells were located in the yolk than in the subgerminal cavity and some were located 800 μm below the under‐surface of the epiblast.

3. It is recommended that injection should be as shallow as possible to increase the proportion of chimeras produced, and that some means is needed to prevent blastodermal cells from escaping from the hole produced by injection.     

Identifying novel polymorphisms in the canine tryptophan hydroxylase 2 gene

Tryptophan hydroxylase (TPH) serves as the rate-limiting enzyme in the biosynthesis of serotonin, and two forms of TPH genes, TPH1 and TPH2, have been reported with specific nucleotide sequences and expression patterns. We determined the sequences of these two genes and identified polymorphisms within their coding regions. We identified five novel single nucleotide polymorphisms in the TPH2 gene, one of which resulted in a non-synonymous mutation and the remaining four in synonymous mutations. Comparison of the genotype frequencies of the polymorphisms among five popular breeds with different behavioral traits revealed statistically significant interbreed differences for three polymorphisms. These results suggest that these polymorphisms would be useful genetic markers when performing behavior studies in dogs.     

Effect of Gaseous Acetic Acid Treatment on Breadmaking Properties of Flour

Eleven different wheat flours of varying protein content were treated with gaseous acetic acid. Each wheat flour exhibited maximum breadmaking properties (bread height and specific volume) at different levels of gaseous acetic acid treatment. There was a relationship (r = +0.7384) between the level of gaseous acetic acid treatment required for maximum breadmaking properties and the protein content of the wheat flour. Wheat flours with higher protein contents were more resistant to the decrease in pH value due to acetic acid treatment.     

Effects of temperature,food availability,and body size on daily growth rate of Japanese anchovy <Emphasis Type="Italic">Engraulis japonicus</Emphasis> larvae in Hiuchi-nada

With data sampled from 2003 to 2006, this study presents the effects of temperature, food availability (C), and body size on the somatic growth rate [^(g)] \hat{g} (mm day⁻¹) and on the size-specific growth rates [^(G)] \hat{G} (day⁻¹) of larval Japanese anchovy Engraulis japonicus caught in Hiuchi-nada, Seto Inland Sea, Japan, a semi-enclosed narrow sea. C was calculated using an individual-based model, which is an encounter model between copepods and anchovy where the anchovy resulted in a successful capture of the copepod. [^(g)] \hat{g} decreased with increasing anchovy length, but increased with increasing C. No significant relationship was found between [^(g)] \hat{g} and temperature. [^(G)] \hat{G} decreased with increasing temperature, but anchovy weight, on the other hand, increased with increasing C. In the literature, the mechanism of growth determination for larvae of pelagic fish species is considered to be primarily determined by sea temperature and secondarily by food availability. However, [^(g)] \hat{g} and [^(G)] \hat{G} were found to be dependent on C rather than temperature. [^(G)] \hat{G} was dependent on anchovy weight rather than C and temperature. For forecasting the number of days required for development from an arbitrary pre-recruitment size to the recruitment size, we address not only temperature but also food availability and the size effect when considering the mechanism of growth determination for larvae.     

Effect of Gaseous Acetic Acid on Dough Rheological and Breadmaking Properties

Breads baked from wheat flours (protein contents 14.1–16.5% at 14.0% mb) that were pretreated with 2–3 mL of gaseous acetic acid per kg of wheat flour, showed maximum bread height and specific volume (cm³/g). Flour-water suspension and the crumb pH values were gradually decreased with increased amounts of acetic acid. Gas generation and dough expansion tests with bread dough showed that the addition of the same amount of acetic acid, which achieved maximum specific volume, also showed the highest rate of gas generation and dough expansion. However, increasing acetic acid decreased these values. Scanning electron microscope (Cryo-SEM) observation showed that the bread dough made from the same acetic acid-treated flour indicated continuum and no cracks in the dough matrix. Evaluation of mixograms showed the decrease of mixing stability with increased acetic acid levels. Viscosity and water binding capacity of flour-water suspensions were sharply increased by the addition of acetic acid at pH 5.0–3.5.     

Breed differences in genotype and allele frequency of catechol O-methyltransferase gene polymorphic regions in dogs

Catechol O-methyltransferase (COMT) inactivates catecholamines and catechol-containing drugs such as L-DOPA. The common genetic polymorphism Val158Met in the human COMT gene is suspected to be associated with "persistence" or risk for schizophrenia. In this study, we attempted to identify the canine COMT gene fragment and to find a similar polymorphism and to reveal its genetic distribution among five representative canine breeds. We found that the amplified gene consisted of 663 bp nucleotides and was 84% homologous with the human COMT gene. The single nucleotide polymorphisms, guanine adenine substitution, were observed at the 39th, 216th and 482nd nucleotides. From the genotyping of the 216th polymorphism among 266 dogs by the polymerase chain reaction-restriction fragment length polymorphism method with restriction enzyme EagI, and that of the 482nd polymorphism with restriction enzyme SfcI, we found inter-breed variations of genotypes as well as of allelic frequencies for both of these polymorphic regions. These results suggest that the identified polymorphisms will be useful tools in elucidating the genetic background of canine behavioral traits.