JAPANESE JOURNAL OF VETERINARY RESEARCH

Identification of novel alternative splicing variants within swine Setd8 gene and their high mRNA expression in testis
Yu S, Chen X, Deng Z, Lan X and Pan C
SET domain containing (lysine methyltransferase) 8 (Setd8), a histone modification enzyme, affects cell cycling, chromosome condensation, high efficient repair of DNA double strand breaks and so on. The objective of this study was to identify novel alternative splicing variants of pig Setd8 gene and its mRNA expression. Four 180-day-old male Guanzhong Black (GZB) pigs and six male Landrace piglets (including three 30-day-old and three 7-day-old pigs) were collected to study Setd8 gene. Herein, two novel variants, Setd8a and Setd8b, were found in pig. The entire sequences of Setd8a and Setd8b variants were 1,039 bp and 958 bp, respectively. qRT-PCR results showed that Setd8a and Setd8b were highly expressed in brains and testes of 180-day-old GZB pigs. Moreover, the expressions of the two Setd8 variants were significantly higher in testis than brain of GZB pig (P < 0.05). Further study on testis showed that the mRNA expression of Setd8a variant was significantly lower than Setd8b variant in 30-day-old and 7-day-old pigs (P < 0.05). The mRNA expression of Setd8a variant'was lower than Setd8b variant in GZB pigs (P > 0.05). Moreover, the expressions of the two Setd8 variants were significantly higher along with age enlargement. In conclusion, Setd8a and Setd8b were firstly identified in pigs and both were expressed in pig testis. Setd8b was the major splicing variant of pig Seid8 gene transcript product. Moreover, the expressions of Setd8 variants were time-dependent. All these findings would enrich the study of Setd8 gene in pig testis.
Antibody detection from Middendorf's vole (Microtus middendorffii) against Tula virus captured in Mongolia
Yoshimatsu K, Arai S, Shimizu K, Tsuda Y, Boldgiv B, Boldbaatar B, Sergelen E, Ariunzaya D, Enkhmanda O, Tuvshintugs S, Morikawa S and Arikawa J
Mongolia in 2010 and 2011. A total of 76 voles belonging to the genera Myodes and Microtus were captured. Most of the voles that were seropositive to Tula virus antigen were Middendorf's voles (Microtus middendorffii (6/31)). Two of the 18 Myodes voles were also seropositive to Tula virus antigen. On the other hand, only one vole was seropositive to Puumala virus antigen. The results suggest that Tula virus was maintained in Middendorf's vole. This is the first report of detection of anti-Tula virus antibody in the central part of the Eurasia continent.
Anti-neoplastic effects of topoisomerase inhibitors in canine mammary carcinoma, melanoma, and osteosarcoma cell lines
Ong SM, Yamamoto H, Saeki K, Tanaka Y, Yoshitake R, Nishimura R and Nakagawa T
Numerous topoisomerase inhibitors with proven efficacy have been used extensively to treat various human neoplasms. However, among these, only doxorubicin has been used and studied extensively in veterinary oncology. The current study was performed to evaluate the responsiveness of canine osteosarcoma (cOSA), mammary gland tumour (cMGT), and malignant melanoma (cMM) cell lines to several topoisomerase inhibitors. In addition, the correlation between the sensitivity to treatment and multi-drug resistant (MDR) factors was investigated. cOSA cell lines exhibited higher sensitivity than cMGT and cMM cell lines to all the topoisomerase inhibitors tested in vitro; this was associated with the levels of multi-drug resistance protein 1 (MDR1) gene expression in the cOSA cell lines. Treatment of cOSA (HMPOS) and cMGT cell line (CHMp) xenograft mouse models with etoposide markedly delayed tumour progression in HMPOS xenografts, but failed to elicit lasting anti-tumour effects on CHMp xenograft mice. The present findings suggest that MDR1 represents a molecular signature for prediction of treatment efficacy of topoisomerase inhibitors, especially that of etoposide, which may be a clinically useful anti-tumour agent for cOSA; however, further study is necessary to refine the treatment protocol.
Particle-Induced X-ray Emission Analysis of Zierum Trace and Major Elements in Cattle with Acute Coliform Mastitis
Shimamori T, Noda J, Tsukano K, Sera K, Yokota H, Koiwa M, Suzuki T and Suzuki K
The aim of the present study was to examine the applicability of the direct determination of trace and major element concentrations in serum samples collected from Holstein dairy cattle with acute coliform mastitis (n = 53) compared with a healthy control group (n = 39). Twenty-eight elements (Na, Mg, Al, Si, S, Cl, K, Ca, Ti, V, Cr, Mn, Fe, Ce, Ni, Cu, Zn, Ga, As, Se, Br, Rb, Sr, Y, Zr, Nb, Mo, and Pb) were detected by particle-induced X-ray emission (PIXE). Significant differences were observed in serum K, Fe, Zn, and Br concentrations, but not in those of the remaining twenty-four elements. Furthermore, serum Fe concentrations (0.751 ± 0.583 μg/ml, n = 18) were significantly lower in dairy cattle with a poor prognosis than in those with a good prognosis (0.945 ± 0.393 μg/ml, n = 35, P < 0.05) and healthy controls (1.458 ± 0.391 μg/ml, n = 39, P < 0.01). We proposed a diagnostic cut-off point for serum Fe concentrations of <0.82 μg/ml based on receiver operating characteristic (ROC) curves in order to identify cattle with a poor prognosis. The results of the present study indicated that assessing the elemental composition of serum, particularly iron, is a promising prognostic tool for determining the outcomes of cattle with severe acute coliform mastitis.
Amenorative effects of exogenous gonadotropins on reproductive profiles of replacement gilts with delayed puberty in a farm in Thailand
Am-In N and Roongsitthichai A
This study was to investigate the effect of gonadotropins on reproductive profiles of replacement gilts with delayed puberty. Totally, 136 Landrace X Yorkshire crossbred gilts, were categorized into control (n = 58) and treatment (n = 78) groups. Gonadotropins (400 U eCG plus 200 IU hCG) were administered in treatment group only. The results revealed that gilts in treatment group had higher number of gilts with estrus (92.3 vs 25.9%, P < 0.001), shorter onset to estrus (4.7 ± 0.3 vs 9.0 ± 0.8 d, P < 0.001), higher number of dominant follicles (18.0 ± 0.2 vs 13.2 ± 0.3 follicles, P < 0.001), and higher farrowing rate (87.5 vs 53.3%, P = 0.002) than those in control group. In conclusion, gonadotropins containing 400 IU eCG plus 200 IU hCG could improve reproductive profiles in replacement gilts with delayed puberty.
Effect of β-hydroxy-β-methylbutyrate acid on meat performance traits and selected indicators of humoral immunity in goats
Zabek K, Wojcik R, Milewski S, Malaczewska J, Tanski Z and Siwicki AK
The aim of this study was to determine the effect of β-hydroxy-β-methylbutyrate acid, on parameters of meat performance in goats as well as on selected parameters of non-specific humoral defense. An experiment was performed on 24 Alpine kids divided into two equal groups: I - control and II - experimental. Over a period of 60 days, the animals were fed an HMB-supplemented diet. The following meat performance parameters were determined: body weight, daily gains, growth rate, the dimensions of musculus longissimus dorsi (m.l.d.) sections and fat thickness over the loin "eye". Selected indicators of non-specific humoral immunity were determined in the blood serum of kids: lysozyme activity, ceruloplasmin activity and gamma globulin content. It was found that the kids administered HMB had a significantly higher body weight on days 30 and 60 of the experiment compared to the control group. The kids in this group also had a significantly more favorable musculature development. Simultaneously, a significant impact of HIMB on the examined immunological indices was found. The significance of differences in relation to the control group was confirmed statistically for lysozyme activity and ceruloplasmin activity on days 30 and 60, while the content of gammaglobulins was confirmed statistically on days 15, 30 and 60 of the study. It was also found that the addition of HMB had a stimulating impact on immunity and growth rate as well as on the development of muscles. It is thus justified to administer HMB to early-weaned kids to enhance their rearing parameters.
Regulation of p16 gene expression by histone H3 acetylation in canine lymphoid tumor cell lines
Fujiwara-Igarashi A, Tomiyasu H, Igarashi H, Yu Y, Gogo-Koshino Y, Ohno K and Tsujimoto H
In order to investigate whether suppression of the p16 gene is mediated by histone H3 acetylation in 4 canine lymphoid tumor cell lines, the gene's acetylation status was examined. In 2 canine lymphoid tumor cell lines with low p16 mRNA expression (GL-1 and UL-1), the acetylation level was lower than that in CL-1 cells with high p16 mRNA expression. The expression of the p16 gene in these 2 cell lines was markedly restored after culture in the presence of a histone deacetylase inhibitors trichostatin A, indicating that p16 was inactivated by hypoacetylation. Findings obtained this study will add new insights and lead to the better understanding of the disease pathogenesis and future development of epigenetic therapeutic strategies.
Does the routine handling affect the phenotype of disease model mice?
Ono M, Sasaki H, Nagasaki K, Torigoe D, Ichii O, Sasaki N and Agui T
The three different mouse handling methods, picking up by tails, tunnels, and open hands were performed using the ICGN glomerulonephritis mouse and the severity of symptoms was evaluated. The handling groups exhibited a tendency of more severe symptoms than the non-handling control group. Female mice handled by their tails showed significantly more severe symptoms than the control group. In addition, we subjected the normal laboratory mice, C57BL/6 and BALB/c mice to tail and tunnel handling to assess the stress conditions. The plasma corticosterone level in the tail-handled mice was higher than that in control mice. These results indicate that handling causes stress and may affect the phenotype of disease model mice.
Usefulness of an anesthetic mixture of medetomidine, midazolam, and butorphanol in cotton rats (Sigmodn hispidus)
Nakamura T, Ichii O, Irie T, Hosotani M, Dantsuka A, Nakamura S, Sato S, Sotozaki K, Kouguchi H, Yoshiyasu T, Nagasaki KI and Kon Y
Tne cotton rat (Sigmodon hispidus) is a laboratory rodent used for studying human infectious diseases. However, a lack of suitable anesthetic agents inconveniences the use of cotton rats in surgical manipulation. This study demonstrated that subcutaneous injection of the mixture of medetomidine, midazolam, and butorphanol (0.15, 2.0, and 2.5 mg/kg, respectively), which is a suitable anesthetic agents for mice and rats, produced an anesthetic duration of more than 50 min in cotton rats. We also demonstrated that 0.15 mg/kg of atipamezole, an antagonist of medetomidine, produced a quick recovery from anesthesia in cotton rats. This indicated that the anesthetic mixture of medetomidine, midazolam, and butorphanol, functioned as a useful and effective anesthetic for short-term surgery in cotton rats.
Using Nassarius stolatus as a potential heavy metal biomonitor
Khidkhan K, Imsilp K, Poapolathep A, Poapolathep S, Jermnak U and Tanhan P
Nassarius stolatus was evaluated as the potential heavy metal (Cd, Fe, Mn, Ni and Pb) accumulator in Don Hoi Lot sandbar, Samut Songkhram province, Thailand. This selected species belongs to the Gastropoda, which is widely distributed in the coastal areas from the upper Gulf through the southern part of Thailand. From our findings, the heavy metal accumulations in N. stolatus tissues were Fe > Pb > Mn > Ni > Cd. The retrieved bioaccumulation factor (BAF) indicated that N. Stolatus has high potential to be a biomonitor for the contaminations of Fe and Mn in water and Cd, Ni and Pb in sediment.
Serial measurements of cardiac troponin I (cTnI) in dogs treated with doxorubicin
Surachetpong SD, Teewasutrakul P and Rungsipipat A
The study aimed to evaluate whether cardiac troponin I (cTnI) and pulsed-wave tissue Doppler imaging (TDI) measurements can detect cardiac changes during doxorubicin therapy in dogs with various types of cancers compared to conventional echocardiography. Serial measurements of cTnI and conventional and pulsed-wave TDI echocardiography were performed in 12 dogs diagnosed with various types of cancers at day 0, weeks 3, 6, 9, and 12 prior to each doxorubicin injection. After treatment with doxorubicin, dogs had significantly increased cTnI levels at week 9 (p = 0.027) and 12 (p = 0.027) compared to normal untreated dogs. Dogs had increased cTnI levels during doxorubicin therapy (p = 0.004). Percent left ventricular ejection fraction (LVEF) and fractional shortening (FS) assessed by 2-dimensional and M-mode echocardiography significantly decreased at weeks 9 and 12. Pulsed-wave TDI derived myocardial performance index (MPI) increased significantly at weeks 9 and 12 compared to day 0 (p = 0.028 and 0.040, respectively). In conclusion, dogs treated with doxorubicin had increased cTnI levels. An increase in cTnI levels was detected before echocardiographic value changes. Serum cTnI can be a sensitive marker for detection of cardiotoxicity in dogs treated with doxorubicin.
Cryopreservation of stallion semen: laboratory assessment of sperm injuries after cushioned centrifugation and freezing with conventional and alternative directional freezing methods
Puglisi R, Bornaghi V, Severgnini A, Vanni R, Balduzzi D and Galli A
Fresh 36 ejaculates of 13 stallions were split into two volumes, centrifuged with and without cushion and frozen with Conventional and two prototype, Drum and Directional, methods using 0.5 ml straws for the Conventional and Drum, and 2 ml flat straws for both the Drum and Directional. Cushioned centrifugation increased total motility (61.2 ± 18.6% vs. 57.5 ± 18.6%; P < 0.001) and mean velocity (84.3 ± 15.6% vs. 83.2 ± 13.8%; P < 0.05) when compared to not cushioned centrifugation, estimated after cooling the sperm at 4⁰C for 90 min before freezing. Cushioned centrifugation also increased (P < 0.001) spermatozoa with polarized mitochondrial membranes (46.8 ± 11.4% vs. 43.4 ± 10.6%) and intact plasmatic/acrosomal membranes (41.0 ± 11.2% vs. 38.5 ± 11.3%) of frozen/thawed sperm, with respect to not cushioned centrifugation. However, no effects of the centrifugation were evidenced for classical kinetic parameters. Flat straws had negative effect for almost all the parameters analyzed at thawing (T,) and after 3 hours' incubation at 37⁰C (T₁), while the Drum method with Paillettes did not show appreciable affects. The variability among stallions was relevant (5% to 69% variance for kinetics and membrane status), while the variability among ejaculates was minor (9% to 28%). Factorial analysis identified three relevant, factors with different informational content: Factor 1 represented by membranes status, Factor 2 by kinetics estimated at T₀, and Factor 3 by kinetics estimated at T₁. Cushioned centrifugation had some beneficial effects for the membrane status of the frozen/thawed sperm, while the use of flat straws needs to be improved.
Alleviative effect of some phytochemicals on cyadox-induced oxidative damage in rabbit erythrocytes
Farag MR, Alagawany M, El-Hack MEA and Tufarelli V
This study was carried out to evaluate the effects of different concentrations of cyadox (CYA), cinnamaldehyde (CIN) and resveratrol (RES) on rabbit isolated erythrocytes in two experiments. Experiment 1 evaluated the effects of different concentrations (2.5, 5, 10, 20, 40, 60, 80 and 100) μg/mI of CYA and pM of both CIN and RES separately on isolated erythrocytes in order to make the concentration response curve. Results indicated that CYA caused a significant depletion of reduced glutathione (GSH) content compared to control at 40 μg/ml and the depletion increased by increasing the concentration. On the other hand, both RES and CIN showed a highly significant elevation of GSH content at 40 μM being more effective by increasing concentrations. Experiment 2 investigated the potential benefits of using CIN and RES separately or in combination on CYA induced alterations in isolated rabbit erythrocytes (the used concentrations were based on the results of experiment 1). Results revealed that exposure to CYA caused a significant decrease in superoxide dismutase (SOD) and catalase (CAT) activities and reduced glutathione (GSH) and total protein (TP) contents, CYA also elevated extracellular hemoglobin (Hb) and adenosine triphosphate (ATP), increased the malonaldehyde (MDA) and protein carbonyl (PrC) contents with increasing caspase3 and caspase8 activities suggesting CYA pro-oxidant effect. Both CIN and RES were able to inverse these hazardous effects of CYA. However, CIN was more effective than RES, and their combination showed a positive synergistic effect in protecting cells against oxidative injury caused by CYA.
Gallus gallus coxsackievirus and adenovirus receptor facilitates the binding of fowl adenovirus serotype 1 in chickens
Fujino K, Fujimoto Y, Ujino A, Thanasut K, Taharaguchi M, Taharaguchi S and Takase K
Coxsackievirus and adenovirus receptor (UXADR) is an integral membrane protein that serves as a receptor for coxsackie B viruses and adenovirus types 2 and 5. Previous studies demonstrated that Fowl adenovirus (FAV) can also utilize Homo sapiens CXADR to infect cells. FAV is a double-stranded DNA virus of the family Adenoviridae. FAV causes inclusion body hepatitis and hydropericardium syndrome in chickens. In addition, FAV serotypes 1 and 8 have recently been shown to cause gizzard erosion in chickens. These chicken diseases and growth insufficiency caused by FAV infection result in great economic loss. Thus, identifying and characterizing the viral receptor would further enhance our understanding of the mechanisms underlying virus infection and histocompatibility. Here, in order to determine the FAV receptor in chickens, we investigated the effect of the recently identified Gallus gallus CXADR (ggCXADR) on FAV infection. Overexpression of ggCXADR in CHO cells resulted in increased FAV binding and expression of early FAV genes. However, the propagation of infectious viruses in CHO cells expressing ggCXADR was not detected. These findings provide the basis for further studies aimed at elucidating the infection mechanism of FAV. Further research is required to characterize the additional host factors involved in FAV infection and life cycle.
Methylprednisolone sodium succinate reduces spinal cord swelling but does not affect recovery of dogs with surgically treated thoracolumbar intervertebral disk herniation
Nishida H, Tanaka H, Kitamura M, Inaba T and Nakayama M
The effect of methylprednisolone sodium succinate (MPSS) therapy was studied in 50 dogs with surgically treated Hansen type I thoracolumbar intervertebral disk herniation (TL-IVDH). Administration of MPSS significantly reduced the swelling of the spinal cord. The sensitivity of localization of disk extrusion using myelography in the MPSS group was 92.3%, and in the non-administration group was 83.3%. No significant difference in recovery rate or length of recovery time was found between the two groups. Administration of MPSS reduced spinal cord swelling, but has no effect on recovery in dogs after surgery for TL-IVDH.
The effects of administering lactic acid bacteria sealed in a capsule on the intestinal bacterial flora of cattle
Seyama T, Hirayasu H, Yoshida G, Ohnuma A, Qiu Y, Nakajima C, Kasai K and Suzuki Y
We examined the effects of encapsulated lactic acid bacteria administrated orally to lactating cattle on the intestinal flora. A dose of 3 X 10¹¹ colony forming unit (cfu) of freeze-dried Lactobacillus coryniformis subsp. torquens (JCM1099) encapsulated in an enteric capsule capable of bypassing the rumen was administered for seven days. DNA was extracted from feces 0 and 24 hr after daily administration. Metagenomic analysis showed an increasing trend of the alpha diversity, an index of the species diversity. Furthermore, principal component analysis of intestinal flora revealed that cattle could be differentiated by JCM1099 capsule and suspension administration via principal components 1, 2, and 3. We conclude that administration of encapsulated JCM1099 can alter the intestinal bacterial flora of cattle.
Molecular study of feline hemoplasmas in free- ranging fishing cats (Prionailurus viverrinus) in Thailand
Suksai P, Tangsudjai S, Sariya L, Chamsai T, Sedwisai P, Patumrattanathan S, Prasittichai L, Cutter P, Ratanakorn P and Sangkachai N
Feline hemoplasmas, consisting of Mycoplasma haemofelis (M. haemofelis), Candidatus Mycoplasma haemominutum (Ca. M. haemominutum), and Candidatus Mycoplasma turicensis (Ca. M. turicensis), cause feline infectious anemia and zoonoses. Using multiplex PCR and phylogenetic analysis based on 16S rRNA, 22 blood samples from fishing cats (Prionailurus viverrinus) living in Khao Sam Roi Yot National Park, Thailand were determined positive for M. haemofelis (13.6%)and Ca. M. haemominutum (22.7%). M. haemofelis and Ca. M. haemominutum infection can result severe anemia and asymptomatic, respectively. However, not all positive cases exhibit anemia symptoms. Future study of hemoplasma infection in wild felids is necessary for conservation and the preservation of public health in Thailand.
Genetic diversity of Thoroughbred horse population from Bosnia and Herzegovina based on 17 microsatellite markers
Rukavian D, Hasanbasic D, Ramic J, Zahirovic A, Ajanovic A, Beganovic K, Durmic-Pasic A, Kalamujic B and Pojskic N
The focus of this study was on genetic diversity of TB horse population raised in B&H. Genomic DNA was genotyped by using 17 microsatellite markers. A total of 103 alleles were detected. The average number of alleles per locus was 6.059 and effective number of alleles was 3.293. Means of observed and expected heterozygosity were calculated 0.645 and 0.696, respectively. The average PIC values was 0.649 and inbreeding coefficient was 0.090. Based on all observed parameters, ASB2 locus showed the highest genetic diversity while locus HMS2 was the least diverse. These results suggest that the population of TB horses from B&H is not affected by substantial loss of genetic diversity, indicating the presence of reasonably high level of genetic variability.
Evaluation of weekly bathing in allergic dogs with methicillin-resistant Staphylococcal colonization
Kawarai S, Fujimoto A, Nozawa G, Kanemaki N, Madarame H, Shida T and Kiuchi A
We evaluated the efficacy of weekly bathing in reducing methicillin-resistant Staphylococcus (MRS) colonization in canine allergic dermatitis in a pilot clinical trial. Six dogs with allergic dermatitis controlled by prescription medications were treated with weekly bathing for 1 month. The Canine Atopic Dermatitis Extent and Severity Index version 3 (CADESI-03) and pruritus scores and frequency of mecA-positive Staphylococcus spp. isolated from three body sites between weeks 0 and 4 were compared. There was no significant difference in CADESI-03 scores with bathing, whereas the pruritus scores were significantly reduced (p < 0.05). Furthermore, MRS frequency was decreased in four of the six dogs (p < 0.05). In conclusion, weekly bathing should be considered for reducing MRS colonization in canine allergic dermatitis.
Water intoxication in adult cattle
Kawahara N, Ofuji S, Abe S, Tanaka A, Uematsu M and Ogata Y
Water intoxication is a common disorder in calves and is usually characterized by transient hemoglobinuria. In contrast, the condition is very rare in adult cattle, with few reports on naturally occurring cases. In the present report, four female Japanese Black cattle, aged 16-25 months, showed neurological signs when they drank water following a water outage. Hemoglobinuria was not grossly observed, while severe hyponatremia was revealed by laboratory tests. Autopsy indicated cerebral edema with accumulation of serous fluid in expanded Virchow-Robin spaces. These results indicate the possibility of water intoxication associated with cerebral edema due to severe dilutional hyponatremia in adult cattle.
An unusual case of thoracic ectopia cordis in a Toggenburg Goat and its three-dimensional images constructed with X-ray computed tomography
Lai JM, Wu JT, Yang WC, Chao MH and Nagahata H
A two-day-old female Toggenburg goat with thoracic ectopia cordis (EC) was diagnosed via radiography and computed tomography. The goat was born with EC, defects of the sternum and a supra-umbilical abdominal wall, but without the presence of Cantrell's syndrome. Necropsy and histopathological findings indicated the affected kid had malformation of the heart with an enlarged left ventricle. The findings showed the heart (9 x 5 x 5 cm) stayed outside the thorax, and was covered by a semitransparent membrane. This report is the first to describe a case of thoracic EC in a goat whose sternum was not developed fully and was not connected to the ribs. It is also the first paper to describe three-dimensional images of this condition constructed from computed tomography scans.