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issn:1976-555
1.  An overview of calf diarrhea - infectious etiology, diagnosis, and intervention 
Journal of Veterinary Science  2014;15(1):1-17.
Calf diarrhea is a commonly reported disease in young animals, and still a major cause of productivity and economic loss to cattle producers worldwide. In the report of the 2007 National Animal Health Monitoring System for U.S. dairy, half of the deaths among unweaned calves was attributed to diarrhea. Multiple pathogens are known or postulated to cause or contribute to calf diarrhea development. Other factors including both the environment and management practices influence disease severity or outcomes. The multifactorial nature of calf diarrhea makes this disease hard to control effectively in modern cow-calf operations. The purpose of this review is to provide a better understanding of a) the ecology and pathogenesis of well-known and potential bovine enteric pathogens implicated in calf diarrhea, b) describe diagnostic tests used to detect various enteric pathogens along with their pros and cons, and c) propose improved intervention strategies for treating calf diarrhea.
doi:10.4142/jvs.2014.15.1.1
PMCID: PMC3973752  PMID: 24378583
calf diarrhea; etiology; intervention
2.  Mouse Fyn induces pseudopodium formation in Chinese hamster ovary cells 
Journal of Veterinary Science  2014;15(1):111-115.
Molecular mechanisms underlying the effects of Fyn on cell morphology, pseudopodium movement, and cell migration were investigated. The Fyn gene was subcloned into pEGFP-N1 to produce pEGFP-N1-Fyn. Chinese hamster ovary (CHO) cells were transfected with pEGFP-N1-Fyn. The expression of Fyn mRNA and proteins was monitored by reverse transcription-PCR and Western blotting. Additionally, transfected cells were stained with 4',6-diamidino-2-phenylindole and a series of time-lapse images was taken. Sequences of the recombinant plasmids pMD18-T-Fyn and pEGFP-N1-Fyn were confirmed by sequence identification using National Center for Biotechnology Information in USA, and Fyn expression was detected by RT-PCR and Western blotting. The morphology of CHO cells transfected with the recombinant vector was significantly altered. Fyn expression induced filopodia and lamellipodia formation. Based on these results, we concluded that overexpression of mouse Fyn induces the formation of filopodia and lamellipodia in CHO cells, and promotes cell movement.
doi:10.4142/jvs.2014.15.1.111
PMCID: PMC3973753  PMID: 24378585
filopodia; Fyn; lamellipodia; time-lapse
3.  Photodynamic hyperthermal chemotherapy with indocyanine green: a novel cancer therapy for 16 cases of malignant soft tissue sarcoma 
Journal of Veterinary Science  2014;15(1):117-123.
Sixteen cases of malignant soft tissue sarcoma (STS; 10 canines and six felines) were treated with a novel triple therapy that combined photodynamic therapy, hyperthermia using indocyanine green with a broadband light source, and local chemotherapy after surgical tumor resection. This triple therapy was called photodynamic hyperthermal chemotherapy (PHCT). In all cases, the surgical margin was insufficient. In one feline case, PHCT was performed without surgical resection. PHCT was performed over an interval of 1 to 2 weeks and was repeated three to 21 times. No severe side effects, including severe skin burns, necrosis, or skin suture rupture, were observed in any of the animals. No disease recurrence was observed in seven out of 10 (70.0%) dogs and three out of six (50.0%) cats over the follow-up periods ranging from 238 to 1901 days. These results suggest that PHCT decreases the risk of STS recurrence. PHCT should therefore be considered an adjuvant therapy for treating companion animals with STS in veterinary medicine.
doi:10.4142/jvs.2014.15.1.117
PMCID: PMC3973754  PMID: 24136207
cancer; chemotherapy; hyperthermia; PDT; soft tissue sarcoma
4.  Prostaglandin F receptor expression in intrauterine tissues of pregnant rats 
Journal of Veterinary Science  2014;15(1):125-131.
In this investigation, we studied the expression and localization of rat prostaglandin F (FP) receptor in uterine tissues of rats on gestational Days 10, 15, 18, 20, 21, 21.5 and postpartal Days 1 and 3 using Western blotting analysis, real-time PCR, and immunohistochemistry. A high level of immunoreactivity was observed on gestational Days 20, 21, and 21.5 with the most significant signals found on Day 20. FP receptor protein was expressed starting on gestational Day 15, and a fluctuating unsteady increase was observed until delivery. Uterine FP receptor mRNA levels were low between Days 10 and 18 of gestation (p < 0.05). The transcript level increased significantly on Day 20 and peaked on Day 21.5 just before labor (p < 0.05). There was a positive correlation between FP receptor mRNA expression and serum estradiol levels (rs = 0.78; p < 0.01) along with serum estradiol/progesterone ratios (rs = 0.79; p < 0.01). In summary, we observed an increase FP receptor expression in rat uterus with advancing gestation, a marked elevation of expression at term, and a concominant decrease during the postpartum period. These findings indicate a role for uterine FP receptors in the mediation of uterine contractility at term.
doi:10.4142/jvs.2014.15.1.125
PMCID: PMC3973755  PMID: 24136214
immunohistochemistry; pregnancy; prostaglandin F receptor; rat; uterus
5.  Regulation of matrix metalloproteinase-9 protein expression by 1α,25-(OH)2D3 during osteoclast differentiation 
Journal of Veterinary Science  2014;15(1):133-140.
To investigate 1α,25-(OH)2D3 regulation of matrix metalloproteinase-9 (MMP-9) protein expression during osteoclast formation and differentiation, receptor activator of nuclear factor κB ligand (RANKL) and macrophage colony-stimulating factor (M-CSF) were administered to induce the differentiation of RAW264.7 cells into osteoclasts. The cells were incubated with different concentrations of 1α,25-(OH)2D3 during culturing, and cell proliferation was measured using the methylthiazol tetrazolium method. Osteoclast formation was confirmed using tartrate-resistant acid phosphatase (TRAP) staining and assessing bone lacunar resorption. MMP-9 protein expression levels were measured with Western blotting. We showed that 1α,25-(OH)2D3 inhibited RAW264.7 cell proliferation induced by RANKL and M-CSF, increased the numbers of TRAP-positive osteoclasts and their nuclei, enhanced osteoclast bone resorption, and promoted MMP-9 protein expression in a concentration-dependent manner. These findings indicate that 1α,25-(OH)2D3 administered at a physiological relevant concentration promoted osteoclast formation and could regulate osteoclast bone metabolism by increasing MMP-9 protein expression during osteoclast differentiation.
doi:10.4142/jvs.2014.15.1.133
PMCID: PMC3973756  PMID: 24136216
1α,25-(OH)2D3; bone lacunar resorption; MMP-9; osteoclast; TRAP
6.  Betulinic acid prevents alcohol-induced liver damage by improving the antioxidant system in mice 
Journal of Veterinary Science  2014;15(1):141-148.
Betulinic acid (BA), a pentacyclic lupane-type triterpene, has a wide range of bioactivities. The main objective of this work was to evaluate the hepatoprotective activity of BA and the potential mechanism underlying the ability of this compound to prevent liver damage induced by alcohol in vivo. Mice were given oral doses of BA (0.25, 0.5, and 1.0 mg/kg) daily for 14 days, and induced liver injury by feeding 50% alcohol orally at the dosage of 10 ml/kg after 1 h last administration of BA. BA pretreatment significantly reduced the serum levels of alanine transaminase, aspartate transaminase, total cholesterol, and triacylglycerides in a dose-dependent manner in the mice administered alcohol. Hepatic levels of glutathione, superoxide dismutase, glutathione peroxidase, and catalase were remarkably increased, while malondialdehyde contents and microvesicular steatosis in the liver were decreased by BA in a dose-dependent manner after alcohol-induced liver injury. These findings suggest that the mechanism underlying the hepatoprotective effects of BA might be due to increased antioxidant capacity, mainly through improvement of the tissue redox system, maintenance of the antioxidant system, and decreased lipid peroxidation in the liver.
doi:10.4142/jvs.2014.15.1.141
PMCID: PMC3973757  PMID: 24378582
alcohol; antioxidant capacity; betulinic acid; lipid peroxidation; liver damage
7.  Echocardiographic assessment of coronary artery flow in normal canines and model dogs with myocardial infarction 
Journal of Veterinary Science  2014;15(1):149-155.
This study was conducted to evaluate the usefulness of coronary arterial profiles from normal dogs (11 animals) and canines (six dogs) with experimental myocardial infarction (MI) induced by ligation of the left coronary artery (LCA). Blood velocity of the LCA and right coronary artery (RCA) were evaluated following transthoracic pulsed-wave Doppler echocardiography. The LCA was observed as an infundibular shape, located adjacent to the sinus of Valsalva. The RCA appeared as a tubular structure located 12 o'clock relative to the aorta. In normal dogs, the LCA and RCA mean peak diastolic velocities were 20.84 ± 3.24 and 19.47 ± 2.67 cm/sec, respectively. The LCA and RCA mean diastolic deceleration times were 0.91 ± 0.14 sec and 1.13 ± 0.20 sec, respectively. In dogs with MI, the LCA had significantly (p < 0.01) lower peak velocities (14.82 ± 1.61 cm/sec) than the RCA (31.61 ± 2.34 cm/sec). The RCA had a significantly (p < 0.01) rapid diastolic deceleration time (0.71 ± 0.06 sec) than that found in the LCA (1.02 ± 0.22 sec) of MI dogs. In conclusion, these profiles may serve as a differential factor for evaluating cardiomyopathy in dogs.
doi:10.4142/jvs.2014.15.1.149
PMCID: PMC3973758  PMID: 23820197
coronary artery; dog; echocardiography; myocardial infarction; peak diastolic velocity
8.  Effect of Harderian adenectomy on the statistical analyses of mouse brain imaging using positron emission tomography 
Journal of Veterinary Science  2014;15(1):157-161.
Positron emission tomography (PET) using 2-deoxy-2-[18F] fluoro-D-glucose (FDG) as a radioactive tracer is a useful technique for in vivo brain imaging. However, the anatomical and physiological features of the Harderian gland limit the use of FDG-PET imaging in the mouse brain. The gland shows strong FDG uptake, which in turn results in distorted PET images of the frontal brain region. The purpose of this study was to determine if a simple surgical procedure to remove the Harderian gland prior to PET imaging of mouse brains could reduce or eliminate FDG uptake. Measurement of FDG uptake in unilaterally adenectomized mice showed that the radioactive signal emitted from the intact Harderian gland distorts frontal brain region images. Spatial parametric measurement analysis demonstrated that the presence of the Harderian gland could prevent accurate assessment of brain PET imaging. Bilateral Harderian adenectomy efficiently eliminated unwanted radioactive signal spillover into the frontal brain region beginning on postoperative Day 10. Harderian adenectomy did not cause any post-operative complications during the experimental period. These findings demonstrate the benefits of performing a Harderian adenectomy prior to PET imaging of mouse brains.
doi:10.4142/jvs.2014.15.1.157
PMCID: PMC3973759  PMID: 23820224
adenectomy; Harderian gland; mouse; positron emission tomography; statistical parametric mapping
9.  Development of a multiplex PCR assay to detect Edwardsiella tarda, Streptococcus parauberis, and Streptococcus iniae in olive flounder (Paralichthys olivaceus) 
Journal of Veterinary Science  2014;15(1):163-166.
A multiplex PCR protocol was established to simultaneously detect major bacterial pathogens in olive flounder (Paralichthys olivaceus) including Edwardsiella (E.) tarda, Streptococcus (S.) parauberis, and S. iniae. The PCR assay was able to detect 0.01 ng of E. tarda, 0.1 ng of S. parauberis, and 1 ng of S. iniae genomic DNA. Furthermore, this technique was found to have high specificity when tested with related bacterial species. This method represents a cheaper, faster, and reliable alternative for identifying major bacterial pathogens in olive flounder, the most important farmed fish in Korea.
doi:10.4142/jvs.2014.15.1.163
PMCID: PMC3973760  PMID: 24378589
Edwardsiella tarda; multiplex PCR; olive flounder; Streptococcus iniae; Streptococcus parauberis
10.  Relationship between pregnancy rate and serum progesterone concentration in cases of porcine embryo transfer 
Journal of Veterinary Science  2014;15(1):167-171.
The level of P4 at the time of embryo transfer (ET) is important. P4 concentrations and numbers of corpora lutea for 126 recipients were evaluated. Nuclear transfer embryos were transferred into 126 surrogates. 11 maintained their pregnancy until full-term delivery, 17 miscarried, and implantation failed in 98 animals. P4 levels in the full-term group were significantly different from those of the pigs that aborted or in which implantation failed (p < 0.05). However, the numbers of corpora lutea were not significantly different. These findings indicate that the concentration of progesterone can be an important factor for successful ET in pigs.
doi:10.4142/jvs.2014.15.1.167
PMCID: PMC3973761  PMID: 24378584
corpus luteum; pregnancy; progesterone; somatic cell nuclear transfer
11.  Sequential alterations of glucocorticoid receptors in the hippocampus of STZ-treated type 1 diabetic rats 
Journal of Veterinary Science  2014;15(1):19-26.
Type 1 diabetes is a common metabolic disorder accompanied by increased blood glucose levels along with glucocorticoid and cognitive deficits. The disease is also thought to be associated with environmental changes in brain and constantly induces oxidative stress in patients. Therefore, glucocorticoid-mediated negative feedback mechanisms involving the glucocorticoid receptor (GR) binding site are very important to understand the development of this disease. Many researchers have used streptozotocin (STZ)-treated diabetic animals to study changes in GR expression in the brain. However, few scientists have evaluated the hyperglycemic period following STZ exposure. In the present study, we found GR expression in the hippocampus varied based on the period after STZ administration for up to 4 weeks. We performed immunohistochemistry and Western blotting to validate the sequential alterations of GR expression in the hippocampus of STZ-treated type 1 diabetic rats. GR protein expression increased significantly until week 3 but decreased at week 4 following STZ administration. GR expression after 70 mg/kg STZ administration was highest at 3 weeks post-treatment and decreased thereafter. Although STZ-induced increase in GR expression in diabetic animals has been described, our data indicate that researchers should consider the sequential GR expression changes during the hyperglycemic period following STZ exposure.
doi:10.4142/jvs.2014.15.1.19
PMCID: PMC3973762  PMID: 23820217
glucocorticoid receptor; hippocampus; hyperglycemic period; streptozotocin; type 1 diabetes
12.  Cell proliferation and neuroblast differentiation in the dentate gyrus of high-fat diet-fed mice are increased after rosiglitazone treatment 
Journal of Veterinary Science  2014;15(1):27-33.
In this study, we determined how rosiglitazone (RSG) differentially affected hippocampal neurogenesis in mice fed a low-fat diet (LFD) or high-fat diet (HFD; 60% fat). LFD and HFD were given to the mice for 8 weeks. Four weeks after initiating the LFD and HFD feeding, vehicle or RSG was administered orally once a day to both groups of mice. We measured cell proliferation and neuroblast differentiation in the subgranular zone of the dentate gyrus using Ki67 and doublecortin (DCX), respectively, as markers. In addition, we monitored the effects of RSG on the levels of DCX and brain-derived neurotrophic factor (BDNF) in hippocampal homogenates. At 8 weeks after the LFD feeding, the numbers of Ki67- and DCX-positive cells as well as hippocampal levels of DCX and BDNF were significantly decreased in the RSG-treated group compared to the vehicle-treated animals. In contrast, the numbers of Ki67- and DCX-positive cells along with hippocampal levels of DCX and BDNF in the HFD fed mice were significantly increased in the RSG-treated mice compared to the vehicle-treated group. Our data demonstrate that RSG can modulate the levels of BDNF, which could play a pivotal role in cell proliferation and neuroblast differentiation in the hippocampal dentate gyrus.
doi:10.4142/jvs.2014.15.1.27
PMCID: PMC3973763  PMID: 24136217
brain-derived neurotrophic factor; dentate gyrus; high-fat diet; rosiglitazone
13.  Immunohistochemical evaluation of the goat forestomach during prenatal development 
Journal of Veterinary Science  2014;15(1):35-43.
Here we report the detection and distribution of synaptophysin (SPY), non-neuronal enolase (NNE), glial fibrillary acidic protein (GFAP), vimentin (VIM), neuropeptide Y (NPY), and vasoactive intestinal peptide (VIP) expression in the goat forestomach during prenatal development. A total of 140 embryos and fetuses were examined to evaluate protein expression from the first stage of prenatal life until birth. In all cases, SPY immunoreactivity was detected at 53 days gestation in the lamina propria-submucosa, tunica muscularis, serosa, and myenteric plexuses. Immunoreactivity to NNE was observed at 64 days gestation in the same locations as well as the epithelial layer. Glial cells were found at 64 days as indicated by signals corresponding to GFAP and VIM at 39 days. Positive staining for NPY and VIP was observed at 113, 75, and 95 days in the rumen, reticulum, and omasum, respectively, in the lamina propria-submucosa, tunica muscularis, and myenteric plexuses of each of these gastric compartments. These findings indicate possible preparation of the fetal goat forestomach for postnatal function. Compared to other ruminant species, neuroendocrine cells, glial cells and peptidergic innervations markers were detected earlier compared to sheep but at around the same stage as in deer.
doi:10.4142/jvs.2014.15.1.35
PMCID: PMC3973764  PMID: 24136206
forestomach; goat; immunohistochemistry; prenatal development
14.  Training-induced changes in clotting parameters of athletic horses 
Journal of Veterinary Science  2014;15(1):45-49.
The purpose of this study was to investigate the effects of training on prothrombin time, activated partial thromboplastin time, and fibrinogen (Fb) concentrations in horses to assess potential adaptive response to training. Fifteen clinically healthy horses were enrolled in the present study and equally divided into three groups. Group A completed an intense training program, group B participated in a light training program, and group C included sedentary horses. After 5 weeks, group B was subjected to the same training program completed by group A and renamed group B1. Blood samples were collected by jugular venipuncture from each animal at rest and analyzed within 2 h after sampling. A two-way ANOVA for repeated measures showed a significant effect of training (p < 0.05) on Fb concentrations in group B1 alone during the first week after changing the training program. Our findings demonstrated that Fb is a parameter susceptible to training. Fb plasma levels increase with a more intense training program. However, Fb plasma levels decreased after the first week and returned to basel levels, suggesting that the horses had adapted to the new training program.
doi:10.4142/jvs.2014.15.1.45
PMCID: PMC3973765  PMID: 24136203
activated partial thromboplastin time; fibrinogen; horse; prothrombin time; training
15.  Protective effects of silymarin on fumonisin B1-induced hepatotoxicity in mice 
Journal of Veterinary Science  2014;15(1):51-60.
The present study was conducted to investigate the effect of silymarin on experimental liver toxication induced by Fumonisin B1 (FB1) in BALB/c mice. The mice were divided into six groups (n = 15). Group 1 served as the control. Group 2 was the silymarin control (100 mg/kg by gavage). Groups 3 and 4 were treated with FB1 (Group 3, 1.5 mg/kg FB1, intraperitoneally; and Group 4, 4.5 mg/kg FB1). Group 5 received FB1 (1.5 mg/kg) and silymarin (100 mg/kg), and Group 6 was given a higher dose of FB1 (4.5 mg/kg FB1) with silymarin (100 mg/kg). Silymarin treatment significantly decreased (p < 0.0001) the apoptotic rate. FB1 administration significantly increased (p < 0.0001) proliferating cell nuclear antigen and Ki-67 expression. Furthermore, FB1 elevated the levels of caspase-8 and tumor necrosis factor-alpha mediators while silymarin significantly reduced (p < 0.0001) the expression of these factors. Vascular endothelial growth factor (VEGF) and fibroblast growth factor-2 (FGF-2) expressions were significantly elevated in Group 4 (p < 0.0001). Silymarin administration alleviated increased VEGF and FGF-2 expression levels (p < 0.0001). In conclusion, silymarin ameliorated toxic liver damage caused by FB1 in BALB/c mice.
doi:10.4142/jvs.2014.15.1.51
PMCID: PMC3973766  PMID: 24136215
caspase-8; fumonisin B1; fibroblast growth factor-2; galectin-3; silymarin
16.  Toxic effects of methylmercury, arsanilic acid and danofloxacin on the differentiation of mouse embryonic stem cells into neural cells 
Journal of Veterinary Science  2014;15(1):61-71.
This study was performed to assess the neurotoxic effects of methylmercury, arsanilic acid and danofloxacin by quantification of neural-specific proteins in vitro. Quantitation of the protein markers during 14 days of differentiation indicated that the mouse ESCs were completely differentiated into neural cells by Day 8. The cells were treated with non-cytotoxic concentrations of three chemicals during differentiation. Low levels of exposure to methylmercury decreased the expression of GABAA-R and Nestin during the differentiating stage, and Nestin during the differentiated stage. In contrast, GFAP, Tuj1, and MAP2 expression was affected only by relatively high doses during both stages. Arsanilic acid affected the levels of GABAA-R and GFAP during the differentiated stage while the changes of Nestin and Tuj1 were greater during the differentiating stage. For the neural markers (except Nestin) expressed during both stages, danofloxacin affected protein levels at lower concentrations in the differentiated stage than the differentiating stage. Acetylcholinesterase activity was inhibited by relatively low concentrations of methylmercury and arsanilic acid during the differentiating stage while this activity was inhibited only by more than 40 µM of danofloxacin in the differentiated stage. Our results provide useful information about the different toxicities of chemicals and the impact on neural development.
doi:10.4142/jvs.2014.15.1.61
PMCID: PMC3973767  PMID: 24136205
arsanilic acid; danofloxacin; embryonic stem cell test; methylmercury; neural cell
17.  A simplified one-step nuclear transfer procedure alters the gene expression patterns and developmental potential of cloned porcine embryos 
Journal of Veterinary Science  2014;15(1):73-80.
Various somatic cell nuclear transfer (SCNT) techniques for mammalian species have been developed to adjust species-specific procedures to oocyte-associated differences among species. Species-specific SCNT protocols may result in different expression levels of developmentally important genes that may affect embryonic development and pregnancy. In the present study, porcine oocytes were treated with demecolcine that facilitated enucleation with protruding genetic material. Enucleation and donor cell injection were performed either simultaneously with a single pipette (simplified one-step SCNT; SONT) or separately with different pipettes (conventional two-step SCNT; CTNT) as the control procedure. After blastocysts from both groups were cultured in vitro, the expression levels of developmentally important genes (OCT4, NANOG, EOMES, CDX2, GLUT-1, PolyA, and HSP70) were analyzed by real-time quantitative polymerase chain reaction. Both the developmental rate according to blastocyst stage as well as the expression levels CDX2, EOMES, and HSP70 were elevated with SONT compared to CTNT. The genes with elevated expression are known to influence trophectoderm formation and heat stress-induced arrest. These results showed that our SONT technique improved the development of SCNT porcine embryos, and increased the expression of genes that are important for placental formation and stress-induced arrest.
doi:10.4142/jvs.2014.15.1.73
PMCID: PMC3973768  PMID: 23820223
CDX2; EOMES; HSP70; porcine; somatic cell nuclear transfer
18.  Elucidating the role of ApxI in hemolysis and cellular damage by using a novel apxIA mutant of Actinobacillus pleuropneumoniae serotype 10 
Journal of Veterinary Science  2014;15(1):81-89.
Exotoxins produced by Actinobacillus (A.) pleuropneumoniae (Apx) play major roles in the pathogenesis of pleuropneumonia in swine. This study investigated the role of ApxI in hemolysis and cellular damage using a novel apxIA mutant, ApxIA336, which was developed from the parental strain A. pleuropneumoniae serotype 10 that produces only ApxI in vitro. The genotype of ApxIA336 was confirmed by PCR, Southern blotting, and gene sequencing. Exotoxin preparation derived from ApxIA336 was analyzed for its bioactivity towards porcine erythrocytes and alveolar macrophages. Analysis results indicated that ApxIA336 contained a kanamycin-resistant cassette inserted immediately after 1005 bp of the apxIA gene. Phenotype analysis of ApxIA336 revealed no difference in the growth rate as compared to the parental strain. Meanwhile, ApxI production was abolished in the bacterial culture supernatant, i.e. exotoxin preparation. The inability of ApxIA336 to produce ApxI corresponded to the loss of hemolytic and cytotoxic bioactivity in exotoxin preparation, as demonstrated by hemolysis, lactate dehydrogenase release, mitochondrial activity, and apoptosis assays. Additionally, the virulence of ApxIA336 appeared to be attenuated by 15-fold in BALB/c mice. Collectively, ApxI, but not other components in the exotoxin preparation of A. pleuropneumoniae serotype 10, was responsible for the hemolytic and cytotoxic effects on porcine erythrocytes and alveolar macrophages.
doi:10.4142/jvs.2014.15.1.81
PMCID: PMC3973769  PMID: 23820218
Actinobacillus pleuropneumoniae; ApxI; attenuation; cytotoxicity; mutant
19.  Development and clinical evaluation of a rapid diagnostic kit for feline leukemia virus infection 
Journal of Veterinary Science  2014;15(1):91-97.
Feline leukemia virus (FeLV) causes a range of neoplastic and degenerative diseases in cats. To obtain a more sensitive and convenient diagnosis of the disease, we prepared monoclonal antibodies specific for the FeLV p27 to develop a rapid diagnostic test with enhanced sensitivity and specificity. Among these antibodies, we identified two clones (hybridomas 8F8B5 and 8G7D1) that specifically bound to FeLV and were very suitable for a diagnostic kit. The affinity constants for 8F8B5 and 8G7D1 were 0.35 × 109 and 0.86 × 109, respectively. To investigate the diagnostic abilities of the rapid kit using these antibodies, we performed several clinical studies. Assessment of analytical sensitivity revealed that the detection threshold of the rapid diagnostic test was 2 ng/mL for recombinant p27 and 12.5 × 104 IU/mL for FeLV. When evaluating 252 cat sera samples, the kit was found to have a kappa value of 0.88 compared to polymerase chain reaction (PCR), indicating a significant correlation between data from the rapid diagnostic test and PCR. Sensitivity and specificity of the kit were 95.2% (20/21) and 98.5% (257/261), respectively. Our results demonstrated that the rapid diagnostic test would be a suitable diagnostic tool for the rapid detection of FeLV infection in cats.
doi:10.4142/jvs.2014.15.1.91
PMCID: PMC3973770  PMID: 24136209
anti-FeLV p27 monoclonal antibody; feline leukemia virus; polymerase chain reaction; rapid diagnostic test; recombinant FeLV p27
20.  Positive effects of porcine IL-2 and IL-4 on virus-specific immune responses induced by the porcine reproductive and respiratory syndrome virus (PRRSV) ORF5 DNA vaccine in swine 
Journal of Veterinary Science  2014;15(1):99-109.
The purpose of this study was to investigate the effects of porcine interleukin (IL)-2 and IL-4 genes on enhancing the immunogenicity of a porcine reproductive and respiratory syndrome virus ORF5 DNA vaccine in piglets. Eukaryotic expression plasmids pcDNA-ORF5, pcDNA-IL-2, and pcDNA-IL-4 were constructed and then expressed in Marc-145 cells. The effects of these genes were detected using an indirect immunofluorescent assay and reverse transcription polymerase chain reaction (RT-PCR). Characteristic fluorescence was observed at different times after pcDNA-ORF5 was expressed in the Marc-145 cells, and PCR products corresponding to ORF5, IL-2, and IL-4 genes were detected at 48 h. Based on these data, healthy piglets were injected intramuscularly with different combinations of the purified plasmids: pcDNA-ORF5 alone, pcDNA-ORF5 + pcDNA-IL-2, pcDNA-ORF5 + pcDNA-IL-4, and pcDNA-ORF5 + pcDNAIL-4 + pcDNA-IL-2. The ensuing humoral immune responses, percentages of CD4+ and CD8+ T lymphocytes, proliferation indices, and interferon-γ expression were analyzed. Results revealed that the piglets co-immunized with pcDNA-ORF5 + pcDNA-IL-4 + pcDNA-IL-2 plasmids developed significantly higher antibody titers and neutralizing antibody levels, had significantly increased levels of specific T lymphocyte proliferation, elevated percentages of CD4+ and CD8+ T lymphocytes, and significantly higher IFN-γ production than the other inoculated pigs (p < 0.05).
doi:10.4142/jvs.2014.15.1.99
PMCID: PMC3973771  PMID: 24136204
IL-2; IL-4; immunogenicity; ORF5 DNA vaccine; porcine reproductive and respiratory syndrome virus
21.  Time-dependent changes of calbindin D-28K and parvalbumin immunoreactivity in the hippocampus of rats with streptozotocin-induced type 1 diabetes 
Journal of Veterinary Science  2013;14(4):373-380.
The hippocampus is affected by various stimuli that include hyperglycemia, depression, and ischemia. Calcium-binding proteins (CaBPs) have protective roles in the response to such stimuli. However, little is known about the expression of CaBPs under diabetic conditions. This study was conducted to examine alterations in the physiological parameters with type 1 diabetes induced with streptozotocin (STZ) as well as time-dependent changes in the expression of two CaBPs changes of were being evaluated. Rats treated with STZ (70 mg/kg) had high blood glucose levels (>21.4 mmol/L) along with increased food intake and water consumption volumes compared to the sham controls. In contrast, body weight of the animals treated with STZ was significantly reduced compared to the sham group. CB-specific immunoreactivity was generally increased in the hippocampal CA1 region and granule cell layer of the dentate gyrus (DG) 2 weeks after STZ treatment, but decreased thereafter in these regions. In contrast, the number of PV-immunoreactive neurons and fibers was unchanged in the hippocampus and DG 2 weeks after STZ treatment. However, this number subsequently decreased over time. These results suggest that CB and PV expression is lowest 3 weeks after STZ administration, and these deficits lead to disturbances in calcium homeostasis.
doi:10.4142/jvs.2013.14.4.373
PMCID: PMC3885729  PMID: 23628656
calbindin D-28K; calcium binding protein; hippocampus; parvalbumin; type I diabetes
22.  Expression of E-cadherin in pig kidney 
Journal of Veterinary Science  2013;14(4):381-386.
E-cadherin is a cell adhesion molecule that plays an important role in maintaining renal epithelial polarity and integrity. The purpose of this study was to determine the exact cellular localization of E-cadherin in pig kidney. Kidney tissues from pigs were processed for light and electron microscopy immunocytochemistry, and immunoblot analysis. E-cadhedrin bands of the same size were detected by immunoblot of samples from rat and pig kidneys. In pig kidney, strong E-cadherin expression was observed in the basolateral plasma membrane of the tubular epithelial cells. E-cadherin immunolabeling was not detected in glomeruli or blood vessels of pig kidney. Double-labeling results demonstrated that E-cadherin was expressed in the calbindin D28k-positive distal convoluted tubule and H+-ATPase-positive collecting duct, but not in the aquaporin 1-positive, N-cadherin-positive proximal tubule. In contrast to rat, E-cadherin immunoreactivity was not expressed at detectable levels in the Tamm-Horsfall protein-positive thick ascending limb of pig kidney. Immunoelectron microscopy confirmed that E-cadherin was localized in both the lateral membranes and basal infoldings of the collecting duct. These results suggest that E-cadherin may be a critical adhesion molecule in the distal convoluted tubule and collecting duct cells of pig kidney.
doi:10.4142/jvs.2013.14.4.381
PMCID: PMC3885730  PMID: 23820247
E-cadherin; kidney; pig
23.  Chemopreventive and metabolic effects of inulin on colon cancer development 
Journal of Veterinary Science  2013;14(4):387-393.
Prebiotics modulate microbial composition and ensure a healthy gastrointestinal tract environment that can prevent colon cancer development. These natural dietary compounds are therefore potential chemopreventive agents. Thirty Sprague-Dawley rats (4 months old) were experimentally treated with procarcinogen dimethylhydrazine to induce colon cancer development. The rats were randomly assigned to three groups: a control group (CG), a group treated with dimethylhydrazine (DMH), and a group given DMH and inulin, a prebiotic (DMH+PRE). The effects of inulin on the activities of bacterial glycolytic enzymes, short-chain fatty acids, coliform and lactobacilli counts, cytokine levels, and cyclooxygenase-2 (COX-2) and transcription nuclear factor kappa beta (NFκB) immunoreactivity were measured. Inulin significantly decreased coliform counts (p < 0.01), increased lactobacilli counts (p < 0.001), and decreased the activity of β-glucuronidase (p < 0.01). Butyric and propionic concentrations were decreased in the DMH group. Inulin increased its concentration that had been reduced by DMH. Inulin decreased the numbers of COX-2- and NFκB-positive cells in the tunica mucosae and tela submucosae of the colon. The expression of IL-2, TNFα, and IL-10 was also diminished. This 28-week study showed that dietary intake of inulin prevents preneoplastic changes and inflammation that promote colon cancer development.
doi:10.4142/jvs.2013.14.4.387
PMCID: PMC3885731  PMID: 23820222
chemoprevention; colon cancer; prebiotic; Sprague-Dawley rats
24.  Effect of chronic lead intoxication on the distribution and elimination of amoxicillin in goats 
Journal of Veterinary Science  2013;14(4):395-403.
A study of amoxicillin pharmacokinetics was conducted in healthy goats and goats with chronic lead intoxication. The intoxicated goats had increased serum concentrations of liver enzymes (alanine aminotransferase and γ-glutamyl transferase), blood urea nitrogen, and reactivated δ-aminolevulinic acid dehydratase compared to the controls. Following intravenous amoxicillin (10 mg/kg bw) in control and lead-intoxicated goats, elimination half-lives were 4.14 and 1.26 h, respectively. The volumes of distribution based on the terminal phase were 1.19 and 0.38 L/kg, respectively, and those at steady-state were 0.54 and 0.18 L/kg, respectively. After intramuscular (IM) amoxicillin (10 mg/kg bw) in lead-intoxicated goats and control animals, the absorption, distribution, and elimination of the drug were more rapid in lead-intoxicated goats than the controls. Peak serum concentrations of 21.89 and 12.19 µg/mL were achieved at 1 h and 2 h, respectively, in lead-intoxicated and control goats. Amoxicillin bioavailability in the lead-intoxicated goats decreased 20% compared to the controls. After amoxicillin, more of the drug was excreted in the urine from lead-intoxicated goats than the controls. Our results suggested that lead intoxication in goats increases the rate of amoxicillin absorption after IM administration and distribution and elimination. Thus, lead intoxication may impair the therapeutic effectiveness of amoxicillin.
doi:10.4142/jvs.2013.14.4.395
PMCID: PMC3885732  PMID: 23820209
amoxicillin; bioavailability; disposition; lead intoxication; pharmacokinetics
25.  Inhibitory effects of osteoprotegerin on osteoclast formation and function under serum-free conditions 
Journal of Veterinary Science  2013;14(4):405-412.
The purpose of this study was to determine whether osteoprotegerin (OPG) could affect osteoclat differentiation and activation under serum-free conditions. Both duck embryo bone marrow cells and RAW264.7 cells were incubated with macrophage colony stimulatory factor (M-CSF) and receptor activator for nuclear factor κB ligand (RANKL) in serum-free medium to promote osteoclastogenesis. During cultivation, 0, 10, 20, 50, and 100 ng/mL OPG were added to various groups of cells. Osteoclast differentiation and activation were monitored via tartrate-resistant acid phosphatase (TRAP) staining, filamentous-actin rings analysis, and a bone resorption assay. Furthermore, the expression osteoclast-related genes, such as TRAP and receptor activator for nuclear factor κB (RANK), that was influenced by OPG in RAW264.7 cells was examined using real-time polymerase chain reaction. In summary, findings from the present study suggested that M-CSF with RANKL can promote osteoclast differentiation and activation, and enhance the expression of TRAP and RANK mRNA in osteoclasts. In contrast, OPG inhibited these activities under serum-free conditions.
doi:10.4142/jvs.2013.14.4.405
PMCID: PMC3885733  PMID: 23820214
activation; differentiation; osteoclast; osteoprotegerin; serum-free

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