Ethanol Does Not Promote MeIQx-initiated Rat Colon Carcinogenesis Based on Evidence from Analysis of a Colon Cancer Surrogate Marker

Epidemiological studies suggest that alcohol consumption increases the risk of developing colorectal cancer. However, the data are confounded by numerous cosegregating variables. To cast further light on the relationships between alcohol intake and colon cancer development, 21-day-old male F344/DuCrj rats were fed 200 ppm 2-amino-3,8-dimethylimidazo[4,5-f]quinoxaline (MeIQx) in their diet for 8 weeks and doses of 0, 0.1, 0.3, 1, 3, 10 and 20% of ethanol in their drinking water ad libitum for 16 weeks thereafter. The rats were sacrificed after 24 weeks of experiment, and aberrant crypt foci (ACF), surrogate lesions for colon cancer, were examined under a light microscope at low magnification. Ethanol was found not to affect the ACF formation at any dose compared with the initiated-controls. Furthermore, ethanol did not alter colon epithelial cell proliferation. These data, obtained by analysis of a colon cancer surrogate marker lesion, indicate that ethanol lacks promotion activity for MeIQx-initiated rat colon carcinogenesis.     

In vitro effect of recombinant human granulocyte colony-stimulating factor on canine neutrophil apoptosis

Apoptosis is essential in eliminating neutrophils (polymorphonuclear leukocytes: PMNs) in animals. The suppression of PMN apoptosis is believed to be beneficial in eradicating pathogens and is implicated in the pathogenesis of human inflammatory diseases. In the present study, canine PMNs were stimulated with recombinant human granulocyte colony-stimulating factor (rhG-CSF) to investigate the in vitro effect on the apoptosis of canine PMNs. Apoptotic cell rates were assessed by flow cytometry in relation to the ability of PMNs to produce reactive oxygen species (ROS). Canine PMN apoptosis was markedly suppressed by rhG-CSF treatment, in association with the retention of the PMN ability to produce ROS. The addition of cycloheximide abolished this suppression by rhG-CSF. Moreover, canine PMNs, which were stimulated by rhG-CSF, expressed high levels of anti-apoptotic mcl-1 gene mRNA, as quantified by real-time polymerase chain reaction method. The results suggest that PMNs, stimulated by G-CSF, could work effectively over a longer period to eliminate pathogens, and that the prolongation of the PMN life-span might occasionally aggravate tissue injuries in dogs. In addition, the suppression of PMN apoptosis seems to be mediated by the induction of anti-apoptotic mcl-1 gene expression.     

Dysfunction of erythropoietin-producing interstitial cells in the kidneys of ICR-derived glomerulonephritis (ICGN) mice

Anemia is a major secondary symptom in chronic renal disorder (CRD), but the precise cause of insufficient production of erythropoietin (EPO) remains unclear owing to the controversial localization of EPO-producing cells in the kidneys. The ICR-derived glomerulonephritis (ICGN) mouse, a new hereditary nephrotic mouse, is an appropriate model of anemia associated with CRD. By using an amplified in situ hybridization technique, we detected and counted the renal EPO-producing cells under both normoxic and hypoxic conditions. The expression levels of renal EPO mRNA were quantified and oxygen gradients were also assessed immunohistochemically. Amplified in situ hybridization clarified that EPO-producing cells were peritubular interstitial cells in the middle region of renal cortex in both ICR and ICGN mice. Hypoxia (7% O2) induced low oxygen tension in proximal tubular epithelial cells of renal cortex, and increased the expression of EPO mRNA and the number of EPO-producing cells in both ICR and ICGN mice. However, hypoxia did not increase the serum EPO levels in ICGN mice. The ICGN mouse is a good model for anemia associated with CRD, and the suppression of EPO protein production in the renal EPO-producing cells is considered to be a potential cause of anemia associated with CRD.     

Detection of DNA of 'Candidatus Mycoplasma haemominutum' and Spiroplasma sp. in unfed ticks collected from vegetation in Japan

DNA fragments of 'Candidatus Mycoplasma haemominutum', a feline heamobartonella pathogen, were detected from unfed Ixodes ovatus collected from vegetation in Hokkaido, Fukushima and Yamaguchi Prefectures, and unfed Haemaphysalis flava in Yamaguchi Prefecture. This finding suggests that ixodid tick is a possible vector of 'C. Mycoplasma haemominutum'. Spiroplasma DNA was also detected from unfed I. ovatus in Hokkaido, Fukushima and Yamaguchi Prefectures. The analysis of nucleotides sequence suggested that this Spiroplasma was distinct from registered species.     

Effect of amino acid substitutions in the epitope regions of pyolysin from Arcanobacterium pyogenes

Pyolysin (PLO), secreted by Arcanobacterium pyogenes, is a novel member of the thiol-activated cytolysin (TACY, cholesterol-dependent cytolysin) family of bacterial toxins. Recently, we demonstrated that the epitopes of monoclonal antibodies (mAbs) S, H, C, and G lie in the regions of amino acids regions 55-73, 123-166, 482-506, and 482-506 of PLO, respectively, by the reaction of mAbs with truncated PLOs. In this study, we substituted the amino acids in these epitope regions of PLO by site-directed mutagenesis and examined the effect of these amino acid substitutions. Mutants I70S/R71A/L73S, Y131S/P132S, and L163S/P164S for mAbs H or S completely lost the hemolytic activity of the proteins, but these mutants still bound to erythrocyte membranes. Mutants L495S/W497S and W500S/W501S for mAbs C and G also completely lost their hemolytic activity, but still bound to erythrocyte membranes. In the undecapeptide region of PLO, the cysteine residue required for thiol activation is replaced with alanine. Therefore, we substituted Ala-492 of the undecapeptide region for Cys. The hemolytic activity of this mutant A492C decreased by adding hydrogen peroxide or storing at 4 degrees C, and the decreased hemolytic activity was restored by adding L-cysteine.     

Structural gene and strain specificity of a novel cysteine protease produced by Staphylococcus aureus isolated from a diseased chicken

Recently whole genome sequencing of Staphylococcus aureus has revealed the genes encoding cysteine proteases such as staphopain and SspB. In this study, we cloned and sequenced the structural gene (ScpA) encoding a cysteine (thiol) protease of S. aureus strain CH-91 from a chicken with dermatitis using polymerase chain reaction (PCR) and inverse PCR methods. The sequence information revealed a coding sequence (CDS) of 1200 nucleotides encoding the ScpA preproenzyme of 399 amino acids with a molecular mass of 45,071 Da. The deduced amino acid sequence of the ScpA differed at many positions from those of staphopain and SspB with identities of 64 and 42%, respectively. In the Southern blot analysis with a total DNA of S. aureus strain CH-91, the ScpA probe hybridized with a single 7.7 kb XbaI fragment or 2.8 and 0.8 kb EcoRI fragments, whereas the staphopain and SspB probes did not hybridize with these DNA fragments. These results suggest that this ScpA gene is a single-copy gene and is a novel gene, which is not found in the published whole genome sequences of S. aureus. In immunoblot, PCR, and Southern blot assays, the ScpA or its gene was detected in high protease-producing strains from chickens, but was not recognized in bovine and porcine strains or low protease-producing avian strains. These results indicate that the ScpA of CH-91 type may be specific to the high protease-producing strains of S. aureus from chickens, namely, there is a strain specificity of the ScpA.     

Foot Rot of Ulluco Caused by Pythium aphanidermatum

Severe rot of stem bases caused by Pythium aphanidermatum was found on ulluco (Ullucus tuberosus) grown in Kagawa Prefecture, Japan, in September 1999. The name “foot rot of ulluco” is proposed for this new disease. Received 14 November 2001/ Accepted in revised form 7 January 2002     

Dynamic left ventricular outflow tract obstruction secondary to hypovolemia in a German Shepard dog with splenic hemangiosarcoma

Dynamic left ventricular outflow tract obstruction (DLVOTO) is a common condition in cats and humans. In this case report, a dog is described with DLVOTO secondary to severe intra-abdominal hemorrhage caused by a hemangiosarcoma. The dog was a 9-year-old, 35.7-kg, spayed female German Shepard dog that presented with a history of tachypnea and collapse. A Levine II/VI systolic murmur was present at the heart base. Abdominal ultrasonography revealed a splenic mass and a large amount of ascites. Echocardiography showed a reduced left ventricular diameter and an increased aortic velocity caused by systolic anterior motion (SAM) of the mitral valve apparatus. The heart murmur and the SAM were resolved after treatment including a splenectomy and a blood transfusion.     

Critical limits of the degree of compactness and soil penetration resistance for the soybean crop in N Brazil

In agricultural fields soil compaction is a major cause of physical degradation. Degree of compactness (DC) is a useful parameter for characterizing compaction and the response of crops for different soils. The objectives of this study were: (1) to identify the critical DC and PR values for soybean [Glycine max (L.) Merrill] using plant growth variables and (2) to verify the relationship between DC and PR, and assess which parameter is recommended for the evaluation of soil compaction. The study was conducted in a greenhouse in a completely randomized factorial design of 4 textures × 5 compaction levels for sandy loam and sandy clay loam soils, and 3 compaction levels for the clayey and very clayey soils. Soil samples were collected from the surface of a Xantic Kandiudox from the NE region of the State of Pará, Brazil. The DC was calculated from the maximum bulk density obtained by the Proctor test, and the PR curve was determined in undisturbed samples equilibrated in different matric potentials. The growth and development of the soybean was favored in the DC range of 80 to 85%, regardless of soil texture. The critical degree of compactness for the growth of soybean was around 98% regardless of soil texture, while the critical values for penetration resistance at field capacity varied according to soil texture and bulk density and were 28.2, 5.6, 3.5, and 5.2 MPa for the sandy loam, sand clay loam, clayey and very clayey soils, respectively. The root length was the plant growth variable most susceptible to soil compaction. Change in soil penetration resistance was poorly related with change in degree of compactness showing that one parameter cannot be replaced by the other. Because PR is quickly determined in field and have a direct relationship with plant growth, for the soils evaluated in this study we recommend the use of soil PR to assess the state of soil compaction.     

茶叶β-葡萄糖苷酶亲和层析纯化与性质研究

以化学合成的β-葡萄糖苷酶的抑制剂β-葡萄糖脒为配体,通过N-糖苷键交联到支持物上,使糖脒N-糖苷键一侧的氨基保持带正电荷的活化状态,合成了一种β-葡萄糖苷酶的亲和层析树脂。这种合成的亲和层析树脂在弱酸条件下非常稳定,能选择性地吸附β-葡萄糖苷酶,并能多次重复使用。利用这种亲和层析树脂,在pH6和pH5条件下,从薮北种茶树叶片中纯化出两种分子量分别为63和75kDa的β-葡萄糖苷酶单体酶。以对硝基苯酚β-葡萄糖苷为底物,研究了两种酶的性质。TLC分析结果表明,这两种β-葡萄糖苷酶都能水解(Z)-3-己烯醇、苯甲醇和苯乙腈的β-D-葡萄糖苷。