Hepatic AA amyloidosis in a cat: cytologic and histologic identification of AA amyloid in macrophages

A 3‐year‐old, spayed female, Domestic Shorthair cat presented with anorexia, lethargy, vomiting, probable hemoabdomen, and multiple masses on the right lateral liver lobe. Clinicopathologic and imaging abnormalities included anemia, azotemia, icterus, and hepatomegaly with hypoechoic masses. On cytologic evaluation of a fine‐needle aspiration of a liver mass there was abundant extracellular pink‐ to purple‐colored material between hepatocytes. The amorphous material was stained with direct fast scarlet (DFS), and green birefringent areas were observed under polarized light, confirming the presence of amyloid. A unique finding on the cytologic smear were macrophages containing amorphous and fibrillar amyloid‐like protein. Histopathologic examination using H&E and Congo red staining confirmed amyloid deposits within the space of Disse, along the sinusoids, portal tracts, blood vessel walls, and within the cytoplasm of macrophages. Immunohistochemical staining with anti‐AA amyloid antibodies further confirmed the presence of AA amyloid. To the author's knowledge, this is the first report of the cytologic finding of AA amyloid protein within macrophages and DFS stain detection of amyloid on a cytologic smear.     

A method for visualizing cesium ions adsorbed on wood charcoal using SEM–EDX

A new method based on scanning electron microscopy with energy-dispersive X-ray spectroscopy (SEM–EDX) was developed for visualizing the distribution of cesium ions (Cs⁺) adsorbed by Japanese cedar charcoal. After being soaked in saturated aqueous CsCl solutions, wood charcoal and regular wood samples were embedded in resin, and thin sections of sample were prepared for the Cs⁺ observations. The Cs⁺ ions on charcoal could be seen in EDX mapping images and also in line profiles with enough contrast to identify differences between the various carbonized cell wall regions. On the other hand, Cs⁺ could not be observed in the EDX mapping images of wood samples, despite having been soaked in a saturated aqueous CsCl solution. These results indicate that Cs⁺ adsorbed on charcoal can be visualized using this method, but not Cs⁺ in wood. Additionally, oxygen detected on the charcoal by EDX can be regarded as an indicator of the presence of surface functional groups.     

Identification and characterization of a novel etiological agent of mango malformation disease in Mexico, Fusarium mexicanum sp. nov

The primary objective of this study was to characterize Fusarium spp. associated with the economically devastating mango malformation disease (MMD) in Mexico. In all, 142 Fusarium strains were isolated from symptomatic mango inflorescences and vegetative tissues in eight geographically diverse Mexican states from 2002 through 2007. Initially, all the Mexican isolates were screened for genetic diversity using appolymerase chain reaction and random amplified polymorphic DNA markers and were grouped into seven distinct genotypes. Based on results of these analyses, evolutionary relationships and species limits of the genetically diverse MMD-associated Fusarium spp. were investigated using multilocus DNA sequence data and phylogenetic species recognition. Maximum parsimony analyses of a five-locus data set comprising 5.8 kb of aligned DNA sequence data indicated that at least nine phylogenetically distinct Fusarium spp. within the Gibberella fujikuroi species complex are associated with MMD, including one species within the African clade (Fusarium pseudocircinatum), two species within the Asian clade (F. mangiferae and F. proliferatum), and at least six species within the American clade (F. sterilihyphosum and five undescribed Fusarium spp.). Molecular phylogenetic analyses indicate that a novel genealogically exclusive lineage within the American clade was the predominant MMD associate in Mexico. This new Fusarium sp. caused MMD and could be distinguished from all other known species morphologically by the production of mostly sterile, coiled hyphae which are typically associated with sporodochial conidiophores together with unbranched or sparsely branched aerial conidiophores. Koch's postulates were completed for isolates of the new species on nucellar seedlings of mango cv. Ataulfo. This pathogen is formally described herein as F. mexicanum.     

Molecular typing and antimicrobial resistance of Salmonella enterica subspecies enterica serovar Choleraesuis isolates from diseased pigs in Japan

Salmonella enterica serovar Choleraesuis (S. Choleraesuis) isolates derived from diseased pigs in Japan during 2001 and 2005 were analyzed for biotype, based on H(2)S production and dulcitol fermentation, pulsed-field gel electrophoresis (PFGE) profile, and antimicrobial resistance profile. S. Choleraesuis biotype Choleraesuis (biotype Choleraesuis) was classified into one genotype, while varietas Kunzendorf (var. Kunzendorf) was classified into two genotypes. The isolates of var. Kunzendorf belonging to one genotype were isolated in a limited area of Japan. Variation in the antimicrobial resistance pattern was observed in isolates of both biotypes Choleraesuis and var. Kunzendorf. We have also shown that the PFGE profile was associated with the biotype and isolation region of each isolate.     

Interaction between myostatin and extracellular matrix components

Myostatin, a member of the TGF-β superfamily, is a negative regulator of skeletal muscle mass. We have recently demonstrated that decorin binds to myostatin in vitro , and that immobilized decorin within the collagen matrix prevents myostatin-mediated inhibition of myoblast proliferation. However, little is known about other ECM molecules that bind to myostatin and modulate its activity. Thus, in the present study, we investigated the interaction of several other ECM molecules with myostatin. We here show that fibromodulin, fibronectin and laminin bind to myostatin in the presence of Zn²⁺ with a dissociation constant ( K_D ) of 10⁻¹⁰∼10⁻⁸ mol/L. Fibromodulin shows the highest affinity for myostatin among them. These results suggest that these ECM molecules may modulate myostatin activity like decorin does.     

Energy partitioning in cultured juvenile Pacific bluefin tuna,Thunnus orientalis (Temminck & Schlegel, 1844)

The characteristics of dietary utilization, energy conversion efficiency, metabolic rate and energy partitioning were measured for cultured Pacific bluefin tuna (PBT) juveniles fed on an artificial diet. Thirty‐one juveniles (1.1 ± 0.3 g BW) were stocked into each of two 2500 L tanks to measure oxygen consumption (?O₂), swimming speed, digestibility and growth performance. ?O₂ elevated until 2.5 ± 0.3 times of pre‐feeding level within 1.5 h after feeding, except for the first feeding, and returned to pre‐feeding level within 3 h. Swimming speed fluctuation was corresponded with the ?O₂ fluctuation, and both parameters were stable from 02:00 to 06:00 and also during the whole day for starved fish. These indicate that feeding has strong influence on their metabolic rate. Energy partitioning for faecal, urinary and branchial, heat increment and voluntary activity, standard metabolism, and retained energy were calculated to be 17.2%, 5.9%, 14.9%, 41.3% and 20.7% of total ingested energy, respectively. The results indicate that, unlike other fish, juvenile PBT distribute large amount of energy for maintenance, which allows only a little proportion of ingested energy available for growth.     

Construction of a system for exploring mitotic homologous recombination in the genome of Pyricularia oryzae

To investigate mitotic homologous recombination (HR) in Pyricularia oryzae, we created an HR detection system. The system consists of two non-functional enhanced yellow fluorescent protein (YFP) and blasticidin S deaminase (BSD) fusion genes (YFP::BSD). If mitotic HR occurs between the two non-functional genes in the genome, restoration of the functional YFP::BSD gene can be expected. The expression of the functional YFP::BSD gene can be detected by both YFP fluorescence and resistance against BS. When the P. oryzae genome was transformed simultaneously with two non-functional genes, all six lines of transformants with both genes had some portion of their hyphae exhibiting YFP fluorescence and BS resistance during growth. Up to ca. 10 % of conidia harvested from the mycelium of each of the six lines had YFP fluorescence, suggesting that HR consistently occurs during mycelium growth. To determine whether and how the HR-mediated phenotypic changes occurred at the DNA level, we analyzed the genomic DNA of BS-resistant mycelia by PCR-RFLP and sequencing and were able to confirm the existence of a restored functional YFP::BSD gene and a non-functional recombinant gene. Collectively, these findings demonstrate that we established a successful HR detection system for P. oryzae, which can be used for other plant pathogens, and that mitotic HR actually occurs in P. oryzae and constitute the first experimental evidence for mitotic HR in a fungus.     

Hypoglycemic activity of Gymnema sylvestre extracts on oxidative stress and antioxidant status in diabetic rats

Diabetes mellitus, which is associated with oxidative damage, has a significant impact on health, quality of life, and life expectancy. An ethanol extract of Gymnema sylvestre leaf was examined in vitro and in vivo to investigate the role of antioxidants in diabetic rats. The extract exhibited strong antioxidant activity in the assays, including TBA (56%), SOD-like (92%), and ABTS (54%). Blood glucose levels in the diabetic rats fed G. sylvestre extract decreased to normal levels. The presence of the antihyperglycemic compounds gymnemagenin and gymnemic acids in G. sylvestre extract was detected by LC/MS analysis. Lipid peroxidation levels were decreased by 31.7% in serum, 9.9% in liver, and 9.1% in kidney in the diabetic rats fed the extract. Feeding G. sylvestre extract to the diabetic rats decreased the activity of glutathione peroxidase in cytosolic liver and glutamate pyruvate transaminase in serum to normal levels.     

Flavonoids with potent antioxidant activity found in young green barley leaves

Saponarin, a flavonoid found in young green barley leaves, possesses potent antioxidant activities, which are determined by its inhibition of malonaldehyde (MA) formation from various lipids oxidized by UV light or Fenton's reagent. Lipids used were squalene, ethyl linoleate, ethyl linolenate, ethyl arachidonate, octadecatetraenoic acid (ODTA), eicosapentaenoic acid (EPA), docosahexaenoic acid (DHA), cod liver oil, lecithin I, lecithin II, and blood plasma. The addition of saponarin inhibited the formation of MA from squalene upon UV irradiation at the level of 2 μmol/mL by almost 100%, whereas BHT inhibited its formation by 75% at the same level. Saponarin showed potent antioxidant activity toward fatty acid ethyl esters at levels >100 μg/mL. Saponarin inhibited MA formation in ODTA by 60%, in EPA by 50%, and in DHA by 43% at the level of 15 μmol/mL. Saponarin exhibited strong antioxidant activities with dose-response levels toward cod liver oil and lipoproteins (lecithins I and II), higher than those of α-tocopherol. A mixture of saponarin/lutonarin (4.5:1, w/w) inhibited MA formation appreciably from all lipids tested with dose response. This mixture exhibited highest effect toward cod liver oil (86%), followed by DHA, lecithin II, blood plasma, EPA, and lecithin I. Supplementation of young green barley leaves containing saponarin should be beneficial to health and may prevent diseases caused by oxidative damage such as various cancers, inflammations, and cardiovascular diseases.     

Characterization of spheres derived from canine mammary gland adenocarcinoma cell lines

There is increasing evidence for the presence of cancer stem cells in several solid tumors, and these cancer stem cells have a potential role in tumor initiation, aggression, and recurrence. The stem cell-like properties of spheres derived from canine mammary tumors remain largely elusive. We attempted to induce sphere formation using four cell lines of canine mammary adenocarcinoma, and characterized the spheres derived from a CHMp line in vitro and in vivo. The CHMp-derived spheres showed predominantly CD44⁺CD24⁻ population, higher expression of stem cell-related genes, such as CD133, Notch3 and MDR, and higher resistance to doxorubicin compared with the CHMp-derived adherent cells. Xenograft transplantations in nude mice demonstrated that only 1 × 10⁴ sphere cells were sufficient for tumor formation. Use of the sphere assay on these sphere-derived tumors showed that sphere-forming cells were present in the tumors, and were maintained in serial transplantation. We propose that spheres derived from canine mammary adenocarcinoma cell lines possess a potential characteristic of cancer stem cells. Spheres derived from canine mammary tumors could be a powerful tool with which to investigate novel therapeutic drugs and to elucidate the molecular and cellular mechanisms that underlie tumorigenesis.