Correlation between sphingomyelin and the membrane stability of mammalian erythrocytes.

Lipid compositions of mammalian erythrocyte membranes are different among species. Therefore, the information on hemolysis from mammalian erythrocytes is useful to understand membrane properties of human erythrocytes. In this work, pressure-induced hemolysis and hypotonic one were examined using erythrocytes of human, sheep, cow, cat, dog, pig, horse, rat, and mouse. Pressure-induced hemolysis was suppressed by membrane sphingomyelin, whereas hypotonic hemolysis decreased upon increment of cell diameter. Mass spectra of erythrocyte membrane lipids demonstrated that sphingomyelin with an acyl chain 24:1 was associated with the suppression of pressure-induced hemolysis. In cow erythrocytes, pressure-induced hemolysis was greatly suppressed and the detachment of cytoskeletal proteins from the membrane under hypotonic conditions was also inhibited. Taken together, these results suggest that sphingomyelin with 24:1 fatty acid plays an important role in the stability of the erythrocyte membrane, perhaps via cholesterol.

African pygmy hedgehog adenovirus: Virus replication, virus-induced cytopathogenesis and activation of mitogen-activated protein kinase signaling pathways in infected MDCK cells.

We examined several aspects of African hedgehog adenovirus (AhAdv-1) that was isolated from an African pygmy hedgehog, including: replication kinetics of, virus-induced cytopathic effect (CPE), activation status of mitogen-activated protein kinase (MAPK) signaling pathways, and possible roles of these signaling pathways in virus replication and virus-induced CPE in MDCK cells. AhAdv-1 efficiently replicated and induced CPE in infected cells and caused accumulation of cleaved caspase-3 at 24 h post-infection (p.i.), suggesting apoptosis induction. Analysis of several intracellular signal transduction pathways, which are involved in apoptosis, showed activation of p38 MAPK, Akt and ERK1/2 pathways at 3 h p.i., and upregulation of phosphorylated SAPK/JNK at 24 h p.i. Although p38 MAPK inhibitor and SAPK/JNK inhibitor suppressed activation of the respective pathways in infected cells, they did not inhibit virus-induced CPE. Treatment of infected cells with inhibitor of the Akt pathway, the p38 pathway, the SAPK/JNK pathway or the ERK pathway revealed that inhibitors of p38 pathway inhibited viral replication by real-time PCR and TCID50 assay in infected MDCK cells, suggesting that AhAdv-1 uses p38 pathway for multiplication in infected cells.

A novel glucokinase activator TMG-123 causes long-lasting hypoglycemia and impairs spermatogenesis irreversibly in rats.

The importance of glucose is well known as an energy source in testes. In order to evaluate the effects of long-lasting hypoglycemia on testes, a novel glucokinase activator, TMG-123, was dosed to rats at 5, 20 and 100 mg/kg for 13 weeks. As a result, plasma glucose levels decreased for several hours with increasing doses over the dose range of 5 to 100 mg/kg. No toxicological findings attributable to the test article were observed in clinical observation, measurements of body weight and food consumption, necropsy, and organ weight measurement. Histopathology showed scattered degeneration of seminiferous tubules in testes, and exfoliation of germ cells related to the degeneration of seminiferous tubules was observed in the lumen of both epididymides in the same animals at the end of the dosing period. Similar histopathological findings were noted at the end of the recovery period. In addition, a fertility study was conducted at the same doses for 13 weeks for males and 5 weeks for females. Sperm analysis showed decreases in the sperm concentration and the motility index and an increase in the incidences of sperm malformations. However, there were no abnormalities in the copulation or fertility rate. These results suggest that long-lasting hypoglycemia in rats is harmful to spermatogenesis and the testicular damage does not recover.

Complete Genome Sequence of an Adenovirus-1 Isolate from an African Pygmy Hedgehog (Atelerix albiventris) Exhibiting Respiratory Symptoms in Japan.

This study reports the complete genome sequence of an African pygmy hedgehog adenovirus-1 isolate from an African pygmy hedgehog which displayed respiratory symptoms that included nasal discharge, sniffling, coughing, and respiratory distress. The viral genome is 31,764 bp long and shows four deletion sites compared to that of skunk adenovirus-1.

Phylogenetic analysis of novel posaviruses detected in feces of Japanese pigs with posaviruses and posa-like viruses of vertebrates and invertebrates.

Posaviruses and posa-like viruses are unclassified viruses with sequence similarity to viruses of the order Picornavirales. They have been reported in various vertebrates and invertebrates. We identified 11 posavirus-like sequences in porcine feces and performed phylogenic analysis. Previously reported Japanese posaviruses and those identified in this study clustered with posavirus 1, 4, and 7 and husavirus 1, while five viruses branched into three independent lineages, tentatively named posavirus 10, 11, and 12. Interestingly, posaviruses, except for posavirus 8 and 9, husaviruses, and rasaviruses, formed a cluster consisting of viruses only from pigs, humans, and rats, while posavirus 8 and 9, fisavirus, and basaviruses clustered with posa-like viruses from invertebrates.

Transrectal guidance of the ovaries reduces operative time during bovine laparoscopic ovariectomy.

The main objective of this study was to evaluate the effects of transrectal guidance of the ovaries by an assistant on operative time during bovine laparoscopic ovariectomy. Twenty four clinically healthy Holstein dairy cows were divided randomly into two groups. In the transrectal guidance group, an assistant grasped the ovaries via the transrectal route and pulled them to a position where they could be visualized with a camera. On the other hand, the control group was operated without guidance. The time required to remove both ovaries in the guidance group was shorter than that in the control group (P<0.01). We concluded that laparoscopic ovariectomy with transrectal guidance of the ovaries can substantially shorten operative time, thereby greatly contributing to animal welfare and to reducing the burden on the operator.

A nasal osteoma with an acute course in a Japanese Black heifer.

A 14-month-old Japanese Black heifer presented with unilateral epistaxis and mild swelling of the right face. Radiography revealed a mass with increased radiopacity on the right side of the nasal bridge, extending to the left side. Intranasal endoscopy confirmed a large tumor-like structure protruding into the nasal cavity. Following euthanasia, cranial computed tomography (CT) was performed, revealing a tumor 24.3 × 17.5 × 14.8 cm in size. The tumor occupied the entire right nasal cavity and the frontal and sphenoid sinuses. Histopathological examination revealed that the tumor consisted of well-differentiated trabecular bones and loose connective tissue. Based on these findings, a diagnosis of osteoma was established. This report describes a case of osteoma with an acute course in a Japanese Black heifer.

Mycobacterium avium subsp. hominissuis menigoencephalitis in a cat.

A 33-month old, neutered female Abyssinian cat died. The cat had sudden onset of widespread neurologic signs about half a year after birth. Mycobacterium avium subsp. hominissuis (MAH) group was isolated and identified from the brain of a cat affected with pyogranulomatous meningoencephalitis. The central nervous system (CNS) was involved in the disseminated MAH infection. MAH infection should be considered in cats with neurologic signs in regard to zoonotic aspects. Comparatively, this is a first case of MAH infection observed in the brain in either humans or animals.

Antibodies reacting to carbonic anhydrase isozymes (I and II) and albumin in sera from dogs.

IgGs to carbonic anhydrase isozymes (CA-I and CA-II) and albumin were identified in dog serum. IgG titers were determined in the sera of asymptomatic dogs, and in dogs with atopic dermatitis, diarrhea and/or vomiting, diabetes and/or pancreatitis, kidney disease, hepatic disease, and thyroid gland disease, using ELISA. Low titres of IgG-reactive CA-I, CA-II, BSA, and CSA were found in the sera of healthy beagles. Compared with healthy beagles, there was a significant difference in the titers of antibodies against CA-I in asymptomatic dogs, dogs with diabetes and/or pancreatitis, or thyroid gland disease, or hepatic disease. Compared with healthy beagles, there was a significant difference in the antibody titer of anti-CA-II IgG in asymptomatic dogs and in those with hepatic disease. There was a significant difference in the antibody titer of anti-BSA IgG between healthy beagles and dogs with hepatic disease.

Agglutination of human erythrocytes by the interaction of Zn(2+)ion with histidine-651 on the extracellular domain of band 3.

Clustering of band 3, chloride/bicarbonate exchanger, has been reported in Zn(2+)-treated human erythrocytes. However, the agglutination of human erythrocytes is also induced by the interaction of Zn(2+)ion with histidine on band 3. Identification of histidine that interacts with Zn(2+)ion remains to be determined. The Zn(2+)-induced agglutination of human erythrocytes was unaffected by chymotrypsin cleavage of the small loop region containing His-547 in the extracellular domain of band 3. On the other hand, papain digestion of the large loop region containing His-651 in band 3 inhibited such Zn(2+)-induced agglutination. Moreover, Zn(2+)-induced erythrocyte agglutination was inhibited by the peptide (ARGWVIHPLG) containing His-651, but not by the peptide such as ARGWVIRPLG, which His-651 was substituted by arginine. Among 10 kinds of animal erythrocytes tested, interestingly, no agglutination by Zn(2+)ions was observed in cow cells only that the forth amino acid in the upstream from His-669 on the large loop of cow band 3 is aspartate (Asp-665) instead of glycine. As expected, the agglutination of human erythrocytes by Zn(2+) ions was inhibited in the presence of aspartate. These data indicate that the interaction of Zn(2+) ion with His-651 residue of band 3 plays an important role in the Zn(2+)-induced agglutination of human erythrocytes.

Involvement of the 3' Untranslated Region in Encapsidation of the Hepatitis C Virus.

Although information regarding morphogenesis of the hepatitis C virus (HCV) is accumulating, the mechanism(s) by which the HCV genome encapsidated remains unknown. In the present study, in cell cultures producing HCV, the molecular ratios of 3' end- to 5' end-regions of the viral RNA population in the culture medium were markedly higher than those in the cells, and the ratio was highest in the virion-rich fraction. The interaction of the 3' untranslated region (UTR) with Core in vitro was stronger than that of the interaction of other stable RNA structure elements across the HCV genome. A foreign gene flanked by the 3' UTR was encapsidated by supplying both viral NS3-NS5B proteins and Core-NS2 in trans. Mutations within the conserved stem-loops of the 3' UTR were observed to dramatically diminish packaging efficiency, suggesting that the conserved apical motifs of the 3´ X region are important for HCV genome packaging. This study provides evidence of selective packaging of the HCV genome into viral particles and identified that the 3' UTR acts as a cis-acting element for encapsidation.

Analysis of L-type amino acid transporter in canine hepatocellular carcinoma.

p. 527, lines 5 to 1 from the bottom in the right column. While Kadosawa et al. reported the LAT1 expression in the canine mammary gland tumor and melanoma in the veterinary clinic [9-11], little information is available concerning LAT expression in hepatocellular carcinoma cells (HCCs) of dog; should have been While Fukumoto et al. reported the LAT1 expression in the canine mammary gland tumor and melanoma in the veterinary clinic [9-11], little information is available concerning LAT expression in hepatocellular carcinoma cells (HCCs) of dog.

Suspected panosteitis in a crossbred calf.

A male crossbred calf developed a limp and pain upon deep pressure on the right hind limb and the right forelimb. The radiographic findings of affected limbs and pathological findings of bone biopsy were similar to those observed in canine panosteitis. This is the first case of suspected panosteitis reported in cattle.

Panostéite suspectée chez un veau de race croisée. Un veau mâle de race croisée a développé une boiterie et de la douleur à l'application d'une pression profonde sur la jambe arrière droite et la jambe avant droite. Les résultats de la radiographie des membres touchés et les résultats pathologiques d'une biopsie osseuse étaient semblables à ceux observés dans la panostéite canine. Il s'agit du premier cas de panostéite suspectée chez le bétail.(Traduit par Isabelle Vallières).

Analysis of L-type amino acid transporter in canine hepatocellular carcinoma.

Analysis of L-type amino acid transport expression of hepatocellular carcinoma cells (HCCs) of the dog was performed. The leucine transport activity of canine HCCs was 0.628 ± 0.018 nmol/mg protein/min. The inhibitor of LAT 2-aminobicyclo[2.2.1]heptane-2-carboxylic acid (BCH) reduced 90% of the activity at 1 mM. The deduced amino acid sequences of canine LAT2, LAT3 and LAT4 were well conserved in mammalians, exhibiting 89, 88 and 77% homology, respectively. RT-PCR revealed distinct LAT1 expression compared with normal hepatocytes. Western blotting analysis confirmed the potent LAT1 expression in canine HCCs but not hepatocytes, and real-time RT-PCR analysis indicated that canine HCCs possessed 28 times higher LAT1 expression than hepatocytes. These results indicated that the leucine transport activity of canine HCCs was due to LAT1.

Evidence of LAT1 expression in canine caput epididymis.

L-type amino acid transporter 1 (LAT1), the first isotype of amino acid transport system L, transports aromatic and branched amino acids pivotal for fundamental cellular activities such cellular growth and proliferation. LAT1 expression was high only in the brain in contrast to its limited distribution and low level of expression in normal tissues. We found potent LAT1 expression in canine caput epididymis by quantitative RT-PCR and Western blotting analysis. Immnuno-histochemical examination revealed observable LAT1 in microvillous epithelial cells.

Urinary carbonic anhydrase VI as a biomarker for kidney disease in pigs.

This study investigated whether carbonic anhydrase (CA)-VI has utility as a biomarker in swine kidney disease. Serum chemistry, histopathology, immunohistochemical staining and enzyme-linked immunosorbent assay (ELISA) analyses were performed. In the kidney of normal healthy pigs, CA-VI was localized in the epithelial cells of the renal distal straight tubules. CA-VI levels were 16 ± 35 ng/g wet tissue and 50 ± 66 ng/mL in normal pig kidney and urine, respectively, and 136 ± 173 ng/mL in the urine of pigs with kidney disease. CA-VI urinary concentration was not correlated with urinary urea nitrogen (UUN), urinary creatinine (Cre), or urinary albumin levels in pigs with kidney disease. However, UUN and Cre levels were positively correlated in the urine of pigs with kidney disease. These data suggest that urinary CA-VI may represent a biomarker for kidney disease in pigs, particularly for disorders affecting distal straight tubules.

Detection of a pneumonia virus of mice (PVM) in an African hedgehog (Atelerix arbiventris) with suspected wobbly hedgehog syndrome (WHS).

A pneumonia virus of mice (PVM) from an African hedgehog (Atelerix arbiventris) with suspected wobbly hedgehog syndrome (WHS) was detected and genetically characterized. The affected hedgehog had a nonsuppurative encephalitis with vacuolization of the white matter, and the brain samples yielded RNA reads highly homogeneous to PVM strain 15 (96.5% of full genomic sequence homology by analysis of next generation sequencing). PVM antigen was also detected in the brain and the lungs immunohistochemically. A PVM was strongly suggested as a causative agent of encephalitis of a hedgehog with suspected WHS. This is a first report of PVM infection in hedgehogs.

Distribution, molecular structure and functional analysis of carnitine transporter (SLC22A5) in canine lens epithelial cells.

While carnitine has been reported to have an anti-oxidative role on the ocular surface, there has been no report on the existence of a carnitine transporter (SLC22A5) in the lens. Therefore, we investigated the carnitine transport activity of canine lens epithelial cells (LEC) and determined the molecular structure of canine SLC22A5. The carnitine transport activity was 7.16 ± 0.48 pmol/mg protein/30 min. Butyrobetaine, the analogue of carnitine, reduced 30% of the activity at 50 µM. A coding sequence of canine carnitine transporter was 1694 bp long and was predicted to encode 557 amino acid polypeptides. The deduced amino acid sequence of canine carnitine transporter showed >80% similarity to that of mouse and human. Western blot analysis detected the band at 60 kDa in the membrane of lens epithelial cells. The high content of carnitine in the lens is possibly transported from aqueous humor by SLC22A5.

Detection of enterovirus genome sequence from diarrheal feces of goat.

Goat diarrheal feces were subjected to metagenome analysis by the next-generation sequencing. Nucleotide sequences with homology to enteroviruses were obtained. Primers for RT-PCR were designed based on the nucleotide sequence of these sequences at the 5'-untranslated region, and we determined 563 bp nucleotide sequences that showed homology to bovine-like and ovine enteroviruses (77-87 %). We named the virus detected in this study goat enterovirus G1 (GEV-G1). In the phylogenetic analysis, GEV-G1 belonged to a cluster containing ovine enteroviruses. To our knowledge, this is the first report on nucleotide sequences of an enterovirus infecting Japanese goats.