Cellular injury and lipid peroxidation induced by hexavalent chromium in isolated rat hepatocytes

In order to elucidate the relationship between cellular injury and lipid peroxidation induced by hexavalent chromium (CrVI), isolated rat hepatocytes treated with any one of scavengers of active oxygen species, antioxidants or antichromium agent were incubated with K2Cr2O7 as CrVI (1 mM Cr). After the incubation, the development of lipid peroxidation was determined as thiobarbituric acid (TBA)-reacting materials in total lipid extracts from the incubated hepatocytes. Cellular injury was observed as a leakage of lactate dehydrogenase (LDH) from hepatocytes into incubation medium. The contents of reduced glutathione (GSH) in hepatocytes were also assessed. Results obtained were as follows: (1) CrVI facilitated lipid peroxidation in isolated hepatocytes after 20 min of incubation. On the other hand, the cellular injury induced by CrVI was barely observed even after 60 min of incubation. (2) The CrVI-induced lipid peroxidation was inhibited by catalase and mannitol as scavengers of active oxygen species, or N,N'-diphenyl-p-phenylenediamine and alpha-tocopherol as antioxidants. However the cytotoxicity of CrVI could not be prevented by these chemicals. (3) CrVI depleted the contents of intracellular GSH and diminished the activities of glutathione reductase (GR) and glutathione-S-transferase (GST) except glutathione peroxidase. (4) The scavengers of active oxygen species and the antioxidants could not prevent the depletion of intracellular GSH induced by CrVI. (6) Ascorbic acid, antichromium agent, prevented all of the lipid peroxidation, the cellular injury and intracellular GSH depletion induced by CrVI.(ABSTRACT TRUNCATED AT 250 WORDS)     

Catalog of Micro-Tom tomato responses to common fungal, bacterial, and viral pathogens

Lycopersicon esculentum cultivar Micro-Tom is a miniature tomato with many advantages for studies of the molecular biology and physiology of plants. To evaluate the suitability of Micro-Tom as a host plant for the study of pathogenesis, Micro-Tom plants were inoculated with 16 well-known fungal, bacterial, and viral pathogens of tomato. Athelia rolfsii, Botryotinia fuckeliana, Oidium sp., Phytophthora infestans, and Sclerotinia sclerotiorum caused typical symptoms and sporulated abundantly on Micro-Tom. Micro-Tom was resistant to Alternaria alternata, Corynespora cassiicola, and Fusarium oxysporum. When Micro-Tom was inoculated with 17 isolates of Ralstonia solanacearum, many isolates induced wilt symptoms. Agrobacterium tumefaciens also was pathogenic, causing crown galls on stem tissue after needle prick inoculation. In Micro-Tom sprayed with Pseudomonas syringae pv. tomato, P. s. pv. tabaci, or P. s. pv. glycinea, bacterial populations did not increase, and yellow lesions appeared only on leaves sprayed with P. s. pv. tomato. Tomato mosaic virus, Tomato aspermy virus, and Cucumber mosaic virus systemically infected Micro-Tom, which developed symptoms characteristic of other cultivars of tomato after infection with the respective virus. These results indicated that Micro-Tom was generally susceptible to most of the important tomato pathogens and developed typical symptoms, whereas certain pathogens were restricted by either hypersensitive resistance or nonhost resistance on Micro-Tom. Therefore, an assortment of Micro-Tom–pathogen systems should provide excellent models for studying the mechanism of susceptible and resistant interactions between plants and pathogens.     

Identification of the core motif of the BRCA2 C-terminal RAD51-binding domain by comparing canine and human BRCA2

Mammary tumors are the most common tumors in women and non-spayed female dogs. One of the reasons for mammary tumors is mutations of the tumor suppressor gene, BRCA2. BRCA2 participates in homologous recombination repair by interacting with the RAD51 recombinase. BRCA2 has two RAD51-binding domains, consisting of BRC repeats and the C-terminal RAD51-binding domain, respectively. Although several studies have addressed the function of the C-terminal RAD51-binding domain of human BRCA2, the amino acid sequences required for the RAD51-interaction activity remain unclear. In this study, the C-terminal RAD51-binding domains of canine and human BRCA2 were compared; the canine domain displayed a weaker interaction with RAD51. This difference was attributed to the C-terminal portion of the domain via a comparison between canine and human domains. Furthermore, peptides shorter than those previously reported displayed RAD51-interacting activity, and a core motif of this domain consisting of 25 amino acids was identified. Since a mutation (S3323N) was reported in the core motif of this domain, the effect of this mutation was evaluated. The mutant exhibited similar RAD51-binding activity as that of the wild-type protein, suggesting that the mutation was functionally neutral. These data suggested that the C-terminal portion of the BRCA2 C-terminal RAD51-binding domain influenced its RAD51-interaction activity, and a minimum core motif of 25 amino acids was identified in this domain. These data may help clarify BRCA2 function, as well as the tumorigenic effects of BRCA2 mutation.     

Development of a Simple Method for Evaluation of Water Absorption Rate and Capacity of Rice Flour Samples

A simple method employing a commercially available canister was developed for the determination of mode of water absorption of rice flour samples. The samples prepared by four different grinding methods were used to analyze water absorption. The total amount of water in a flour sample was described by using an exponential model. Capacity and rate of water absorption of the samples were determined, and the relationship to baking quality of partially substituted rice bread was investigated. The water absorption was highly dependent on the method of grinding. Flours produced by wet jet‐milling of the grains, which absorbed a small amount of water at high speed, were most suitable for rice bread. The method was applicable to other food powders, provided that flour particles do not stick together or swell immediately upon contact with water.     

Lactobacillus rhamnosus GG SpaC pilin subunit binds to the carbohydrate moieties of intestinal glycoconjugates

Lactobacillus rhamnosus GG (LGG) is a well‐established probiotic strain. The beneficial properties of this strain are partially dependent on its prolonged residence in the gastrointestinal tract, and are likely influenced by its adhesion to the intestinal mucosa. The pilin SpaC subunit, located within the Spa pili structure, is the most well studied LGG adhesion factor. However, the binding epitopes of SpaC remain largely unknown. The aim of this study was to evaluate the binding properties of SpaC to the carbohydrate moieties of intestinal glycoconjugates using a recombinant SpaC protein. In a competitive enzyme‐linked immunosorbent assay, SpaC binding was markedly reduced by addition of purified mucin and the mucin oligosaccharide fraction. Histochemical staining revealed that the binding of SpaC was drastically reduced by periodic acid treatment. Moreover, in the surface plasmon resonance‐based Biacore assay, SpaC bound strongly to the carbohydrate moieties containing β‐galactoside at the non‐reducing terminus of glycolipids. We here provide the first demonstration that SpaC binds to the oligosaccharide chains of mucins, and that the carbohydrate moieties containing β‐galactoside at the non‐reducing termini of glycoconjugates play a crucial role in this binding. Our results demonstrate the importance of carbohydrates of SpaC for mucus interactions.     

Canine fetal echocardiography: correlations for the analysis of cardiac dimensions

The aim of this study was to develop regression models for correlation of canine fetal heart development with body size to characterize normal development or suggest cardiac anomalies. Twenty clinically healthy pregnant bitches, either brachycephalic and non-brachycephalic, were examined ultrasonographically. Transabdominal fetal echocardiography was conducted every 4 days from the beginning of cardiac chambers differentiation until parturition. Ten cardiac parameters were measured: length, width and diameter of the heart; heart area; left and right ventricular dimensions; left and right atrial dimensions; and aortic and pulmonary artery diameter. Femoral length, biparietal diameter and abdominal cross-sectional area were also recorded. Regression equations were developed for each parameter of fetal body size, and linear and logarithmic models were compared. The model with the highest correlation coefficient was chosen to produce equations to calculate relative dimensions based on the correlations. Only the left-ventricular chamber differed between the two racial groups. Biparietal diameter was the independent parameter that produced the highest correlation coefficient for the most fetal cardiac dimensions, although good correlations were also observed using femoral length and abdominal cross-sectional area. Heart width and heart diameter were used as surrogates of cardiac development, as these measurements showed the best statistical correlation. Quantitative evaluation of fetal cardiac structures can be used to monitor normal and abnormal cardiac development.     

Preventive effects of CIDR-based protocols on premature ovulation before timed-AI in Ovsynch in cycling beef cows

Ovsynch is a program developed to synchronize ovulation for timed breeding. In this paper, the authors investigate whether controlled internal drug release (CIDR)-based protocols prevent premature ovulation before timed-artificial insemination (AI) when Ovsynch is started a few days before luteolysis in cycling beef cows. Nine beef cows at 16 days after oestrus were treated with (1) Ovsynch, i.e. gonadotropin releasing hormone (GnRH) analogue on day 0, prostaglandin (PG) F(2alpha) analogue on day 7 and GnRH analogue on day 9 with timed-AI on day 10, (n=3); (2) Ovsynch+CIDR (Ovsynch protocol plus a CIDR for 7 days from day 0, n=3), or (3) oestradiol benzoate (OB)+CIDR+GnRH (OB on day 0 in lieu of the first GnRH treatment, followed by the Ovsynch+CIDR protocol, n=3). In the Ovsynch group (1) plasma progesterone concentrations fell below 0.5 ng/mL earlier (day 5) than in both CIDR-treated groups (2) and (3), where this occurred on day 8. Plasma oestradiol-17beta concentrations peaked on day 8 in the Ovsynch group and on day 9 in both CIDR-treated groups. The dominant follicle ovulated on day 10 in the Ovsynch group and on day 11 in both CIDR-treated groups. Thus, both CIDR-based protocols prevented premature ovulation before timed-AI in Ovsynch when the protocol was started a few days before luteolysis. This reflects the fact that progesterone levels remained high until the beef cattle were treated with PGF(2alpha).     

Assessment of rabbit spleen size using ultrasonography

Assessment of spleen size using the ultrasonography has become a standard practice in human. However, the assessment is not established method in experimental animals. To establish the index to assess the spleen size using ultrasonography, we measured the cross-section image of rabbit spleen during endotoxin shock. The image of the cross-section was appeared as triangle, and the height of the triangular image was defined as the spleen index. This spleen index showed strong correlation with the spleen weight. In conclusion, this method is suitable for observation of changes in rabbit spleen size and may reduce the number of rabbit in the longitudinal studies.     

Hemorrhagic necrotizing splenitis in a slaughter pig infected with Arcanobacterium species

A 6-month-old barrow presented with lethargy, inappetence and dysstasia. At necropsy, multiple coalescing hemorrhagic foci were detected in the margins of the spleen. Gram-positive bacilli were isolated from the spleen, kidney, muscle and liver. Comparative 16S rDNA gene sequencing analysis of the isolates (TO16177) revealed that they would be the same species of unpublished Arcanobacterium species strain HJ57-14E (accession no. gi 18873551) (99.7% similarity based on a comparison of 675 bp). Histologic examination of the splenic tissue sections revealed extensive necrosis and inflammation, and gram-positive bacilli were discernible. Multifocal necrosis was also detected in the liver. Immunohistochemically, the isolates were cross-reacted with polyclonal antibodies against Arcanobacterium pyogenes and Actinomyces naeslundii, and the reaction was strong for the latter. Similar reactions were found in the suppurative lesions of the tonsil, and occasionally in the spleen and lymph nodes. The present results indicate that the unpublished Arcanobacterium species induced multiple organ failure accompanied by acute hemorrhagic necrotizing splenitis in this growing-finishing pig.