Gross anatomical observations of bovine thoracic duct pathways and the lymph-venous junctions revealed that 37% of these ducts connected to the left venous angle at one location, whereas the other terminal connected to other areas, such as the left internal jugular vein, the left subclavian vein and the right venous angle, at more than one location. The thoracic duct pathways were classified according to Adachi's classification as types III, VI and IX. The frequencies of types VI, IX and III were 76%, 15%, and 9%, respectively and 48% of cattle had more than one ring formation in the thoracic duct pathway. These findings demonstrate many anatomical variations in bovine thoracic duct pathways and lymph-venous junctions.
Glucose is essential for the development of the fetus. We address here the quantitative expression and immunohistochemical localization of glucose transporter (GLUT1 and GLUT3) in the rabbit placenta during successful pregnancy. Blood glucose level showed a significant decrease at the gestation period in comparison with non-pregnancy. Maternal serum glucose was gradually increased according to fetal development. Quantitative RT-PCR results showed that expression of GLUT1 was significantly increased from day 13 to day 18, while GLUT3 mRNA level was significantly decreased during the same periods. Western blot analysis demonstrated that GLUT1 protein did not change significantly in the placenta during pregnancy when compared to non-pregnant uteri. Immunohistochemistry indicated that distribution of GLUT1 was observed mainly to the surface of the outer trophoblasts, whereas GLUT3 mainly localized to the basal site of the inner trophoblasts and fetal blood vessels. These results suggest that glucose is transported through GLUT1 from the maternal blood stream for use as a placental fuel and for further transport through GLUT3 to the fetal circulation, thus signifying the distinct anatomical localization of GLUT1 and GLUT3 in the rabbit placenta during successful pregnancy.
To understand effects of Bisphenol-A (BPA) exposure on the reproductive organ across generations, we analyzed morphology of the uterus and ovary, and the methylation pattern of HOXA10 gene of the 2nd generation. Pregnant mice (F0) were treated with sc injection of BPA in sesame oil at various doses of 0-1,000 mg/kg Bwt on days 12-16 of gestation. Their offspring (F1) were bred by foster mice, and the offspring (F2) from F1 mice were prepared. That is, F1 mice experienced in utero BPA exposure during the developmental period of reproductive organs, while F2 mice did not at all. Using these F2 mice, the present study was carried out. Comparing to the control, the body weights in BPA exposure groups were significantly increased. Correlating with the increase of body weight, the relative weights of the ovary and uterus in each group were decreased. The histological analysis revealed expansion or emphraxis of the uterine lumen and partial loss of the uterine epithelium. Unmethylation of HOXA10 gene in the uterus was observed in the intron region. The present study suggested that BPA exposure to F0 mice could affect reproductive organ of F2 mice who were not exposed to BPA.
Sugars in the glycocalyx play an important role in the attachment of infectious agents to the respiratory mucosa. We examined the histochemistry of 23 lectins to survey the sugar expression in the glycocalyx of the respiratory mucosa of the Pacific white-sided dolphin, Lagenorhynchus obliquidens. The ciliated and basal cells were positive for all of the lectins studied. SBA, WFA, GSL-II, STL, S-WGA, and PNA staining in the cytoplasm showed different intensities between basal cells and ciliated cells. These results suggest that multiple terminal glycosylation occurs on ciliated and basal cells, such as GalNAc, GlcNAc, NeuNAc, galactose, glucose/mannose, oligosaccharide, and fucose, and that sugar residue expression changes during cell differentiation. The Pacific white-sided dolphin respiratory mucosa might express multiple sugar residues in the glycocalyx, to prevent the attachment and colonisation of infectious agents.
A new selective medium containing cephem antibiotics was developed for isolation of methicillin-resistant Staphylococcus aureus (MRSA). MRSA colonies on a medium containing ceftazidime (CAZ) were most easily identifiable and a medium containing cefoperazone (CPZ) was superior in suppressing the growth of other bacteria. With the medium containing a couple of CAZ and CPZ, MRSA and methicillin-resistant coagulase-negative staphylococci (MRCNS) were detected from 2 and 1 of 15 chicken meat samples respectively. The MRSA and MRCNS recovery test showed that the medium was effective for MRSA isolation, suppressing the growth of other bacteria efficiently. These results suggested that the medium containing a couple of CAZ and CPZ was useful for MRSA detection from foods and animals.
Tibia fractures are common in small animal practice. Over the past decade, improvements to animal internal fracture fixation have been developed. TGF-β1 has been shown to be crucial in the development, induction and repair of bone. In present study, we investigate the effect of local application of a graft demineralized bone matrix (DBM) along with TGF-β1 in a model of open osteotomy induced experimentally in dogs. Tibia fracture was brought about by using an open osteotomy model in young male dogs. Fracture repair was evaluated by a histological and biochemical analysis. Collagen content, proteolytic activity and urokinase-type plasminogen activator (uPA) expression were analyzed at the end of the study. Radiographic analysis, alkaline phosphatase and hematological evaluation were performed weekly. At the fifth week, there was an improvement and restoration of bone architecture in animals treated with a graft containing TGF-β1 (5 ng/ml) compared with the control and graft groups, as was evidenced by the presence of an early formation of wide callus and bone regeneration. In addition, local application of TGF-β1 led to an increase in collagen and proteolytic activity. More immunopositive osteoclast and mesenchymal cells were found in bone tissue from animals treated with TGF-β1 as compared with the control group. No changes in alkaline phosphatase, hematological and clinical parameters were observed. This study shows that the combined use of DBM along with TGF-β1 is able to improve and accelerate the bone repair.
A previous serosurvey of Japanese encephalitis virus (JEV) among dogs suggested that dogs are well suited for use as sentinels for assessing the risk of JEV transmission to humans. To examine the clinical symptoms and duration of anti-JEV antibodies in dogs, three dogs were experimentally challenged with JEV. All JEV-infected dogs did not show any clinical signs or abnormal blood tests, except for C-reactive protein. Virus-neutralization titers rapidly increased and were maintained until 70 days postinfection, and neither the virus nor the viral genome was detected in blood. Thus, since dogs live in close proximity to humans as companion animals, they are well suited for use as sentinels for surveying the human risk of JEV infection.
1,5-anhydro-D-glucitol (1,5AG) is a pyranoid polyol compound found in human circulating blood. Myo-inositol (MI) is a stereoisomer of inositol and serves as a precursor of inositol phospholipids. 1,5AG and MI are filtered by the glomerulus and almost completely reabsorbed through the renal tubules. However, under hyperglycemic conditions, reabsorption through the renal tubules is competitively inhibited because the structures of 1,5AG and MI resemble that of glucose. In this study, we investigated the kinetics of serum and urine 1,5AG and MI levels in healthy dogs. We demonstrated that 1,5AG and MI exist in canine serum and urine by gas chromatography-mass spectrometry. Under continuous hyperglycemic conditions, the serum 1,5AG concentration in healthy dogs decreased while the serum MI concentration remained unchanged. Urinary excretion of 1,5AG and MI increased significantly after blood glucose concentrations reached 200 to 220 mg/dl. A significant negative correlation was observed between serum 1,5AG and glucose concentrations during hyperglycemia. However, no significant correlation was observed between serum MI and glucose concentrations. In this study, we demonstrated that serum and urine 1,5AG and MI levels were changed by blood glucose concentrations. The serum 1,5AG concentration was decreased by continuous hyperglycemia. However, the serum MI concentration does not reflect hyperglycemia.
High-mobility group box 1 (HMGB1), a nonhistone chromosomal protein, has recently been suggested as a late mediator of the inflammatory cascade. Blood HMGB1 levels are increased in a number of human diseases, and HMGB1 has been suggested to be a useful marker for disease severity and prognosis. The objective of this study was to assess the clinical usefulness of HMGB1 in dogs. Plasma HMGB1 levels, as well as C-reactive protein (CRP), a typical canine inflammatory marker, were measured in dogs with various diseases, especially systemic inflammatory response syndrome (SIRS), and dogs that had undergone surgery. HMGB1 gradually increased and attained a maximum level 72 hr after surgery, whereas CRP increased rapidly, peaking at 24 hr. Although both HMGB1 and CRP levels were significantly increased in dogs with various diseases compared with the control dogs, no correlation was found between the HMGB1 and CRP values. HMGB1 levels in the SIRS group were significantly elevated compared with those in the non-SIRS group. However, the increase in HMGB1 levels above the reference range was not indicative of SIRS. Instead, the presence of increased HMGB1 and CRP levels above the reference ranges significantly affects the poor outcome of SIRS. The present study indicates that HMGB1 is a novel canine inflammatory marker and is distinct from CRP. However, the additional clinical value of HMGB1 measurement remains unclear, and further studies are warranted.
Gastric motility is affected by several pathological conditions which may induce upper gastrointestinal clinical symptoms. The pathogenesis of canine gastric motility disorders is poorly understood because of methodological limitations. This study aimed at establishing a simple method for evaluating postprandial gastric motility in dogs. Gastric motility was ultrasonographically assessed in 7 healthy beagles using a technique previously described in humans. The motility index (MI), an indicator of gastric antral motility, was calculated by measuring the area of the gastric antrum in both a contracted and relaxed phase and by counting the number of contractions. The MI was measured every 30 min for 3 hr after feeding and compared with gastric emptying as assessed by a 13C-octanoic acid breath test. The MI at 30 min had the lowest variability in the 7 dogs (mean SD, 9.77 ± 0.42; coefficient of variance, 4.25%), and a significant correlation was observed with gastric emptying coefficient (R2=0.8126, P=0.005) and half-emptying time (R2=0.654, P=0.027). When atropine was administered, a significant decrease in the MI at 30 min was observed compared with the control (9.77 ± 0.42 vs. 5.19 ± 0.22, P=0.0003). In conclusion, evaluation of the MI at 30 min is suitable for assessing gastric motility and enables us to assess gastric motility simply in a short time. By using this method, further studies for the pathogenesis of canine gastric motility disorders are warranted.
A 2-month-old intact female Miniature Pinscher puppy presented with footpad swelling and crusted pustules of ear pinnae. The dog had been vaccinated with a polyvalent canine vaccine 5 days prior to the onset of clinical signs. With the history of recent vaccination, the clinical presentation and the histopathological observations were suggestive of ischemic dermatopathy. Treatment involved oral prednisolone, azathioprine, and other immune modulating drugs, which did not work. Chlorambucil plus cyclosporine therapy was initiated for vigorous immune suppression after rush therapy using intravenous immunoglobulin. Clinical signs again gradually improved with no relapse or side effects, even at a 4-month follow-up. The case report is suggested ischemic dermatopathy refractory to conventional therapy and suggests effective approaches to long-term management of the disease.
Otitis externa in 27 toy poodles and 40 miniature dachshunds was treated using a video otoscope. A distinct concavity (external tympanic concavity) was observed at the junction between the ventral part of the external surface of the tympanum and the ear canal to which a considerable amount of hair and debris had adhered. All hair and debris adhering to the external tympanic concavity were removed, and systemic antibiotic and antifungal agents were administered, after which all of the dogs recovered. The pattern of hair growth observed in the external tympanic concavity could be characterized according to the breed of dog. All of the toy poodles presented with curly hairs, while the miniature dachshunds had upright or flat-lying hairs.
The purpose of this study was to evaluate whether dityrosine and advanced oxidation protein products (AOPP) were useful as biomarkers for monitoring the development of acetaminophen-induced liver injury. Dityrosine immunoexpression in the liver along with plasma AOPP concentration was examined up to 24 hr post-acetaminophen injection in rats. The histopathological changes in the liver appeared 3 hr after acetaminophen injection and became exacerbated with time. The immunohistological expression of dityrosine was also first detected in the damaged hepatocytes 3 hr after the injection and became more accentuated at 6, 12 and 24 hr in accompanying with the elevation of plasma AOPP concentration. These results suggested that dityrosine and AOPP expressions might be useful biomarkers for monitoring the development of acetaminophen-induced liver injury.
We report an atypical mycobacterial infection in an Indian flap-shelled turtle, Lissemys punctata punctata, that died in an aquarium in Japan. At necropsy, the turtle showed multiple white nodules on the capsular surface and parenchyma of various organs such as the liver, spleen, intestine, and lung. Histologically, granulomatous inflammation surrounding a central zone of necrosis was observed. Sections stained by the Ziehl-Neelsen method revealed numerous acid-fast bacilli in the cytoplasm of macrophages and in the central area of necrosis. The organisms were identified as a mycobacterial species by PCR and nucleotide sequence analysis and revealed 98-100% homology to M. ulcerans. This is, to our knowledge, the first report of mycobacteriosis due to M. ulcerans in a turtle.
A lymphoplasmacytic lymphoma was diagnosed in a 12- year-old domestic cat that had a primary cutaneous mass involving the stomach, liver, kidneys, heart, abdominal wall, diaphragm, bone marrow and several lymph nodes. Histopathologically, the most characteristic feature of this tumor was the heterogeneity of cell components, such as small lymphocytes, well-differentiated plasma cells and plasmacytoid transformed lymphocytes. Amyloid was deposited in the skin, stomach, and several lymph nodes. Immunohistochemically, neoplastic small lymphocytes were positive for CD20, and well-differentiated plasma cells and plasmacytoid transformed lymphocytes were positive for λ-Ig light chains and MUM1/IRF-4. These results emphasize the importance of lymphoplasmacytic lymphoma as a differential diagnosis of extramedullary cutaneous plasmacytoma in cats.
Acidic and osmotic treatments are part of hurdle systems to control pathogens such as Salmonella in food. In the current study, Salmonella enterica isolates previously shown to differ in their ability to form biofilms were grown in diluted tryptic soy broth (TSB) (1:5 dilution in distilled water) and subsequently exposed to phosphate-buffered saline (PBS) adjusted to pH 3.0 with HCl, PBS adjusted to pH 3.9 with acetic acid or rice vinegar diluted 1:15 with distilled water (pH 3.9). Cells grown in diluted TSB were also exposed to distilled water, pH 7.6, containing 5 M NaCl. No differences in survival upon exposure to PBS adjusted to pH 3.0 with HCl or distilled water containing high salt were observed between the isolates; however, exposure to acetic acid and rice vinegar resulted in lower survival levels of isolates previously shown to be poor biofilm formers. The numbers (log10 cfu/ml) of surviving cells after exposure for 36 hr to acetic acid and rice vinegar were 4.43 ± 0.24 vs. 2.27 ± 0.87 (P<0.05) and 5.19 ± 0.12 vs. 2.33 ± 0.93 (P<0.05) for isolates with a high vs. low biofilm-forming ability. The survival data could be fitted with the Weibull model. The data suggest that the ability of Salmonella strains to survive in the presence of acetic acid and rice vinegar parallels their ability to form biofilms. Thus, Salmonella with a high biofilm-formation capability might be more difficult to kill with acetic acid found in foods or cleaning solutions.
To reveal the antimicrobial susceptibilities of Escherichia coli isolates from wild mice, 81 E. coli isolates were obtained from 109 voles (Clethrionomys spp.), 52 large Japanese field mice (Apodemus speciosus) and 19 small Japanese field mice (A. argenteus) captured in a forest of a natural park in Hokkaido, Japan. Seventy-eight of the 81 E. coli isolates were susceptible to all 10 antimicrobial agents tested. One E. coli isolate was resistant to ampicillin, dihydrostreptomycin, kanamycin, chloramphenicol and oxytetracycline. Two isolates were resistant to oxytetracycline. A low prevalence of antimicrobial resistance was maintained among wild mice that inhabited the forest.
In the present study, the effect of 4-day fasting on steroid hormone metabolism in the liver and secretion of LH was examined in cows. Six non pregnant, dry Holstein cows were used. The estrous cycle was synchronized in all cows using CIDR-Ovsynch. Cows were allocated to a control group (n=3) and a fasting group (n=3). In the fasting group, cows were fasted for four days from day -4 to day -1 (day 0=day of 2nd GnRH injection) but otherwise were fed ad libitum. The experiment was repeated in a crossover design after an interval of about one month. The peripheral progesterone (P4) concentration in the fasting group was significantly higher than in the control group on day -1 and 0. The peripheral estradiol-17β concentration in the fasting group was also significantly higher than in the control group on day -1 and 0. The portal vein P4 concentration in the fasting group was significantly higher than in the control group. On day 0, there was no difference in LH secretion between groups. The mean percentages of lipid droplets in liver cells in the fasting group were significantly higher than in the control group on day 0. These results suggest that short-term fasting leads to reduced hepatic steroid hormone metabolism by accumulation of fat in the liver, which causes high peripheral steroid hormone concentrations.
The captive breeding program of the Okinawa rail started in 2008. For successful captive breeding, information related to reproduction, such as age at sexual maturity, testicular cycles and ovulatory cycles, is essential to predict when reproduction is possible and when certain reproductive behaviors are most likely to occur. We made gross and histological observations of the reproductive organs of Okinawa rails to gain understanding of sexual maturity, the testicular cycle and the ovulatory cycle. We found that the weight of the testis was smallest in December and largest in March. Changes in the diameter of the seminiferous tubule showed the same pattern. Mature sperm were observed from March to June. The heaviest ovary was observed in April. A single peak of reproduction, from March to April, was observed in males and females. Our observations suggested that the Okinawa rail is a seasonal breeder. Establishing suitable breeding pairs will be critical to ensure success of the Okinawa rail captive breeding program. Our results suggested that pairing must be started before March. If supportive breeding is used, semen should be collected from March to June and artificial insemination conducted in April.
The objective of this study was to investigate the cellular immunolocalization of inhibin a and inhibin/activin (βA and βB) subunits in the muskrat testes and scented glands during the breeding season. Inhibin α and inhibin/activin (βA and βB) subunits were expressed in Sertoli cells and Leydig cells of testes and glandular cells of scented glands, respectively. Also, positive signals of inhibin α and inhibin/activin (βA and βB) subunits by Western blotting were both observed in testicular and scented glandular tissues. These results suggested that the testes and scented glands of the muskrats had the ability to synthesize inhibins and activins and that activins and inhibins might play an important role in testicular and scented glandular function in muskrats.
A six-month-old Japanese Black bull was found to have no left testis in the scrotum. A fist-sized mass was palpated per rectum. Two months later, hCG was injected and blood samples were collected before and after injection. No testosterone response to hCG was observed. On the cut surface of the excised mass, most of the mass was composed of homogeneous adipose-like tissue. The rest of the surface was composed of a well-circumscribed testicular parenchyma-like tissue (18 × 16 × 15 mm). Histology revealed diffusely distributed mature adipocytes and septa of fibrous connective tissue. Neither germ cells nor spermatozoa were observed in the seminiferous tubules. A diagnosis was made of fibrolipoma and aspermatogenesis of the left cryptorchid testis.
Cytochrome P450 (CYP) 1B1 is involved in the metabolic activation of various procarcinogens, and some CYP1B1 genetic variants alter CYP1B1-dependent procarcinogen metabolism. Cynomolgus and rhesus macaques are frequently used in toxicity tests due to their evolutionary closeness to humans. In this study, we attempted to identify CYP1B1 genetic variants in 13 cynomolgus and 4 rhesus macaques. A total of 17 genetic variants were identified, including 8 non-synonymous genetic variants, indicating that, similar to humans, CYP1B1 is polymorphic in macaques. These CYP1B1 genetic variants could be the basis for understanding potential inter-animal differences in macaque CYP1B1-dependent metabolism of promutagens.
In this study, we carried out an experimental infection in pigs using a foot-and-mouth disease virus isolated from the 2010 epidemic in Japan to analyze the clinical manifestation, antibody response and virus shedding patterns in pigs. We found that the virus was virulent in pigs, producing a synchronous disease in the inoculated pigs and efficient spread to direct contact pigs. These results are useful for epidemiologically investigating the 2010 epidemic in Japan and improving the measures for controlling possible future FMD outbreaks in Japan or elsewhere.
We evaluated loop-mediated isothermal amplification (LAMP) as a means of detecting equine herpesvirus type 1 (EHV-1) DNA directly from nasal swabs. To increase the sensitivity, we added a step in which the samples were heat-treated to the original LAMP procedure. The detection limit of the LAMP assay with heat treatment was 10 times more sensitive than the original LAMP assay even when the DNA extraction step was omitted. In addition, the LAMP assay with heat treatment was more sensitive than the original LAMP assay and the polymerase chain reaction using clinical samples. The LAMP assay with heat treatment is easy to perform and so should be applicable to the diagnosis of EHV-1 infections in clinical laboratories.
Borna disease virus (BDV) is a non-segmented, negative-strand RNA virus that is characterized by nuclear replication and persistent infection. A unique feature of BDV is that it releases only a small number of infectious particles from infected cells. Although these characteristics might make it difficult to obtain a large amount of recombinant viruses in a reverse genetics system, the mechanism underlying the budding or assembly of BDV particle has remained largely unknown. In this study, as a first step toward understanding the virion formation of BDV, we investigated the intracellular distribution and mobility of the fluorescent marker fusion envelope glycoprotein (G) of BDV in living cells. Expression analysis revealed that fusion proteins seem to cleave into functional subunits and localize in the endoplasmic reticulum (ER)/Golgi apparatus, as well as the authentic BDV G. Furthermore, we demonstrated using fluorescence recovery after photobleaching analysis that BDV G fluorescence shows rapid recovery in both the ER/Golgi and plasma membrane regions, indicating that BDV G fusion protein may be a useful tool to investigate not only the maturation of BDV G but also the budding and assembly of BDV particles in living cells.