The lesser mouse deer (Tragulus javanicus) is one of the most primitive ruminants. Skulls of lesser mouse deer were measured to evaluate their growth. The age was estimated from the eruption and attrition of molars on the mandible. A total of 95 specimens was divided into five age groups, and the growth pattern in each sex was established for some measurements. The relative growth coefficients were calculated for the head and body length and several parts of the cranium and mandible against the profile length. Sexual dimorphism could not be found on the skull in any age group except for measurements related to the upper canine. However, females showed a higher value in head and body length in the oldest group. This feature was consistent with an interpretation of mouse deer society as monogamous. Growth was more rapid in length than in width. Various statistical analyses showed that the visceral cranium related to masticatory facility was much better developed than the neurocranium. However, observations on the shape of the mandible ramus and of the premolars implied that the masticatory function was not suitable for taking roughage. This speculation agreed with previous reports on the feeding habits and digestive organs of the mouse deer.

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A micromethod which utilizes a protein-dye binding reaction on an acetate membrane and a light microscope with an automatic exposure instrument has been developed for measuring small amounts of protein from limited biological materials. After discoidal cellulose acetate membranes (2.0 mm diameter), which absorbed 0.5 μl of protein solution containing 0.2-2.0 μg of protein, were stained with Coomassie Brilliant Blue G-250, they were examined by a light microscope with an automatic exposure instrument. A linear relation was confirmed between the intensity of dye binding to protein and exposure time under specific optical conditions.

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To characterize the active transport of amino acids across the placenta, uptakes of proline, leucine, and alanine were kinetically examined in placental microvillous membrane vesicles (PMV) prepared from rats in the late gestational period. Uptake rates of these amino acids in PMV showed saturable hyperbolic curves that obeyed Michaelis-Menten kinetics. Proline, leucine, and alanine transport were demonstrated to be carrier mediated systems with sodium-dependent, -independent, and both manner, respectively. In addition, sodium-dependent l-alanine transport showed two different systems, and new sodium-independent alanine transport system (K _m of 1.12 mM) was observed in rat placenta. From these results, rat placenta has carrier mediated amino acid transport systems, and posseses at least three different transport systems for alanine.

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Using polymerase chain reaction, bottle-nosed dolphin (Tursiops truncatus) major histocompatibility complex (DoLA) class Ia chain cDNA was cloned and sequenced. Predicted amino acid sequences of DoLA class Iα chain have cystein residues for intradomain disulfide bond formation and N-linked glycosylation sites, suggesting that DoLA class Iα chain molecules construct α1, α2, and α3 domain structures. Similarity of DoLA class Iα chain cDNA to land mammal MHC class Iα chain cDNA was 90.4% in cattle, 90.2% in horse, 89.4% in sheep, and 87.8% in human. This investigation suggests that DoLA is closely related to land mammals.

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The present study was designed to investigate the role of cytokines in the pathogenesis of Babesia caballi in experimentally infected horses. The expression of cytokine mRNA was determined by using reverse transcription-polymerase chain reaction in two B. caballi-infected horses for 2 weeks after the infection. In one horse, there was up-regulation of interferon-gamma, tumor necrosis factor-alpha (TNF-α) and interleukin-2 mRNAs, while in the second horse, expression of only TNF-α mRNA was up-regulated. No change was observed in interleukin-4 mRNA in both of the horses. To know the relation between nitric oxide (NO) production and pathogenesis, NO production was assayed in three dexamethasone treated-B. caballi-infected horses. Production of NO in all 3 horses increased significantly before death, although the parasitemia level remained very low. Treatment with NO inhibitor resulted in the suppression of NO production and increased parasitemia level in a horse, which died of the infection. The pathological examination showed that the main cause of the death was dyspnoea and pulmonary edema. Histopathologically, diffuse global mesangial proliferative glomerulonephritis was also observed. These results suggested that NO may be a critical effector molecule of immune defense against parasite. TNF-α and NO might be contributing to the pathogenesis in B. caballi infection.

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Five Japanese Black cattle showing the tendency of persistent hemorrhage were diagnosed as δ-storage pool deficiency because of lack of dense bodies in platelets. There was no significant difference in the platelet count, fibrinogen concentration, prothrombin time and activated partial thromboplastin time between the cases and normal control cattle. However, the maximum platelet aggregation rate and the values of myosin in the cytoskeletons during platelet aggregation induced by collagen were significantly lower in the cases compared with those in normal control cattle. The quantities of platelet membrane glycoprotein were in the range of 94-160 kDa and were not different between the cases and control cattle. However, a decrease of thrombospondin in a-granules in platelet cytoplasm were suspected in two of the 5 cases

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The Otsuka Long-Evans Tokushima Fatty (OLETF) rat is an animal model for obese-type non-insulin-dependent diabetes mellitus (NIDDM) in humans. The OLETF rat has poor capacity for proliferation of pancreatic β-cells after partial pancreatectomy, which may be the critical pathogenetic event in NIDDM development. The poor pancreatic β-cell proliferation in this model is characterized by reduction in β-cell mass and decrease in insulin content in the remnant pancreas. Our investigation was designed to identify quantitative trait loci (QTLs) responsible for β-cell mass and plasma insulin levels after partial pancreatectomy by performing a genome-wide scan in an F₂ intercross obtained by mating the OLETF and the Fischer-344 (F344) rats. We have identified a suggestive QTL for the plasma insulin levels, near D20Mgh5 on rat chromosome 20, with a maximum lod score of 3.75 which accounts for 20% of the total variance, while no QTLs were detected for β-cell mass. This chromosome 20 QTL, whose OLETF allele is associated with low plasma insulin levels through acting in an incompletely recessive manner, may affect insulin secretion itself rather than β-cell proliferation.

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Mitochondrial capsule selenoprotein (MCS) has been known as a structural protein of the mitochondrial sheath in spermatozoa. In the present study, a full-length cDNA encoding the MCS was first isolated from the testes of 10-week-old golden hamsters using a RACE (rapid amplification of cDNA ends) technique and its mRNA expression pattern was investigated from the hamster tissues by reverse transcription-polymerase chain reaction (RT-PCR) analysis. Hamster MCS cDNA was 820 bp long, including 24 bp of the 5'-untranslated region (UTR) and 243 bp of the 3'-UTR, and showed identity of 75.6% and 73.9% with mouse and rat MCS. According to the deduced amino acid (aa) sequence analysis, hamster MCS encoded a polypeptide of 184 aa, including a cysteine- and proline-rich domain which is the characteristic sequences of MCS, and contained 2 in-frame UGA codons for selenocysteine. Hamster MCS also shared aa identity of 64.4% with mouse MCS and contained an Arg-Lys-Ser-Thr-rich region in the N-terminus similar to the mitochondrial targeting signal. On the other hand, according to the RT-PCR analysis using the specific primers for hamster MCS, hamster MCS mRNA was expressed in various tissues as well as the testes. This finding indicates that MCS in hamster may have more than just a function of mitochondrial sheath formation of spermatozoa.

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A conserved DNA region among the subspecies of koala, of mitochondrial cytochrome b gene, was employed to analyze the genetic variation among 3 available subspecies of koala. This conserved sequence, 307 bp DNA, was sequenced using polymerase chain reaction and direct DNA sequencing technique. Substitutions in the nucleotide sequences were observed, with which koalas can be divided into 3 DNA haplotypes subspecies, but the molecular data provided inconsistency with current classification of the 3 subspecies of koala.

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Nucleic acid biosynthesis of Angiostrongylus costaricensis was examined with various inhibitors; aminopterin (inhibitor of purine and pyrimidine de novo biosynthetic pathways), 8-azaguanine (specific inhibitor of purine salvage pathway) and PALA (specific inhibitor of pyrimidine de novo biosynthetic pathway) were applied in in vitro culture developing from the third stage larvae to young adult in chemically defined medium. It was suggested that A. costaricensis possessed functional purine and pyrimidine de novo biosynthetic pathways and also that they could utilize exogenous sources of purines and pyrimidines by salvage pathways for their development.

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Intrahepatic pylephlebitis was detected in 17 Japanese beef cattle. Grossly, the intrahepatic vessels in the caudate lobe and/or in the periphery of the other hepatic lobes were thickened and protruded above the lobar surface. The vessel lumina were packed with white to red, waste thread-like contents. A few immature flukes were found in the bile ducts in 3 of the 7 cases with biliary thickening. Foci of hepatic necrosis and hemorrhage were scattered around the thickened vessels in 8 cases. Histologically, the interlobular veins were thickened due to severe intimal hyperplasia with endothelial proliferation and eosinophilic accumulation and medial hypertrophy, accompanied by fibrosis and eosinophilic infiltration in the portal areas. Hepatic tissues with necrosis and hemorrhage were surrounded by eosinophils and histiocytes including a granulomatous reaction. One immature fluke was detected in one of these regions of necrosis. Immunoperoxidase staining revealed that the small fluke, Kupffer cells, and histocytes in the liver of all cases were positively stained with anti-Japanese Fasciola sp. antiserum. Enzyme-linked immunosorbent assay of the sera of 15 cases revealed that all were positive for the anti-Fasciola antibody. On the basis of these findings, the present cases were regarded as an atypical form of fascioliasis, characterized by eosinophilic proliferative pylephlebitis of the liver.

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Changes in reactivity of vascular smooth muscles of male alloxan- induced diabetes-susceptible (ALS) and resistant (ALR) mice aorta were investigated at 2 weeks, 1, 2 and 4 month(s) after the injection of alloxan (45 mg/kg, i.v.). The glucose levels in blood and urine of all the alloxan-treated ALS mice were markedly elevated while those in alloxan-treated ALR and non-treated ALS and ALR mice were not altered. The magnitude of high K⁺ (65.4 mM)-induced contractions were not affected by the treatment of alloxan. Norepinephrine-induced contractions in vascular smooth muscles of ALS mice in a diabetic state for 2 or 4 months were significantly potentiated. The contractile sensitivity to prostaglandin F_2α (PGF_2α) was increased in the 4 month-diabetic state. Responsiveness to 5-HT did not vary in the diabetic mouse. Vasorelaxation induced by nitroprusside was attenuated in 2 weeks, 2 or 4 month-diabetic ALS mice. Similarly the inhibitory effects of levcromakalim were attenuated at 2 and 4 months. The influences of diabetes on the inhibitory effects of forskolin or verapamil were very small or not detected. The effects of the vasomodulators used in this study on the vascular smooth muscles of alloxan-treated ALR mice did not differ from those of untreated ALR mice. The results from using ALS and ALR mice suggest that the vasoreactivities to some vasomodulators are changed especially in the long-term diabetic state and that when diabetes was not induced the dose of alloxan does not have any effect on vascular smooth muscle.

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In rats with genetically hypoplastic kidneys (hpk/hpk) and associated hypogonadism (hgn/hgn), their kidneys contain only one-quarter the number of nephrons that are found in those of normal rats [26]. Not surprisingly, therefore, renal excretive function has been shown to be depressed in hpk/hpk rats [26]. In the study presented here, we have examined the process of the progression of renal failure and the development of renal secondary disease in hpk/hpk rats. The plasma concentrations of urea-nitrogen and creatinine were significantly higher in adult hpk/hpk rats than in normal rats. These values elevated gradually and the degree of renal histological damage also progressed with advancing age in the hpk/hpk rats. In addition, renal anemia appeared at 140 days of age or later in these rats, and hyperplasia of the parathyroid glands was visible macroscopically at 280 days of age. In the hpk/hpk rats plasma levels of calcium and phosphorus were significantly lower and higher than in normal rats, respectively, at 280 days of age. Pathologically, the left femora of hpk/hpk rats exhibited fibrous osteodystrophy at 280 days of age and the calcium content of the right femora (as a percentage of the dry weight of bone) was significantly lower than in normal rats at both 210 and 280 days of age. These results indicate that the reduced nephrogenesis of the hpk/hpk rats causes progressive renal failure, secondarily inducing anemia, hyperparathyroidism, and osteodystrophy.

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To identify the source of contamination of raw bulk milk with Listeria, we attempted to isolate the bacteria from various samples such as cattle-related samples, bulk storage tanks and the environments on three farms. On farms A and B, Listeria monocytogenes was repeatedly isolated from raw milk, while on farm C, it was scarcely isolated from it. On the former farms, Listeria was detected in cattle-related samples and the environments. On the other hand, only one fecal sample on the latter farm was Listeria-positive. Especially, we demonstrated that the bulk tank on farm A was contaminated with L. monocytogenes. Then, L. monocytogenes was controlled by continuously washing the bulk tank on farm A with alkaline detergent.

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The in vitro cytotoxicity of recombinant human tumor necrosis factor-α (rh-TNF-α) and actinomycin D (ACT-D) on canine normal and tumor cells was investigated. rh-TNF-α showed dose-dependent cytotoxic and cell-growth inhibitory effects on cultured canine kidney carcinoma cells (CKCa-1). rh-TNF-α alone produced little cytotoxic effect on canine normal cells. However, combined with ACT-D, it showed moderate cytotoxicity on normal canine cells from the kidney medulla, spleen, heart muscle and lung. When the effects on the spontaneous tumor cells were examined, the combination of rh-TNF-α and ACT-D produced substantial cytotoxic effect on 60% of the tumor cells. All mammary mixed tumors and perianal gland tumors tested were susceptible to this combination. The data indicated the combination of rh-TNF-α and ACT-D have in vitro cytotoxicity on certain canine tumor cells.

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Alveolar and peritoneal macrophages (MPs) of mouse, dog and cat were compared in relation to their scanning electron microscopic features and the lysosomal activities of nonspecific esterase, acid phosphatase, and β-glucuronidase. The long spindle shape of peritoneal MPs differed from the spherical form of alveolar MPs in all species. There was no difference in the morphological findings among the three animals. Murine alveolar and peritoneal MPs were strongly positive for all three enzymes. Canine and feline alveolar and peritoneal MPs were strongly positive for acid phosphatase and β-glucuronidase, but weakly positive for nonspecific esterase. These results strongly suggest that acid phosphatase and β-glucuronidase can be used as markers of the MPs in healthy dogs and cats.

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The image processing procedure has become widely applied as a visual aid in imaging diagnosis. The subtraction image of MRI obtained by digitally subtracting an unenhanced image from a contrast-enhanced image, depicted a discrete distribution of the contrast agent. Subtraction images in the clinical cases were compared. The subtraction image in a case of chondroma demonstrated a relatively uniform distribution of a contrast agent, with a well delineated neoplastic lesion. The subtraction image in a case of squamous cell carcinoma, the contrast agent was distributed heterogeneously in the carcinoma and well permeated into the muscle. Different patterns of subtraction image were obtained in the clinical cases of chondroma and squamous cell carcinoma in this study. The findings suggest the potential usefulness of the subtraction image for diagnosing the degree of malignancy.

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To determine whether indomethacin, a potent inhibitor of prostaglandins endoperoxide synthetase, affects the selective follicle-stimulating hormone (FSH) surge during the period of ovulation, the compound was administered intravenously (iv), concurrent with 10 IU human chorionic gonadotropin (hCG), to diestrous female rats at 16:00 hr. Indomethacin inhibited the number of ovulations in a dose-dependent manner, and treatment with 500 μg indomethacin reduced number of oocytes in the ampullae most effectively without enteric lesions. In the histological observation, oocytes that had began to mature were found not only in unruptured luteinized follicles but also in ovarian interstitium beneath ruptured luteinized follicle. Despite the inhibitory effects of indomethacin on ovulation, peri-ovulatory FSH and progesterone surges occurred in comparable levels and duration to vehicle-treated animals. These results indicate that indomethacin-induced inhibition of prostaglandin synthesis does not affect the selective release of FSH during the peri-ovulatory period.

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Day 9 rat embryos were exposed to 1,4-dihydropyridine calcium channel blockers; nifedipine (NIF), nicardipine (NIC) or nitrendipine (NIT), for 48 hr in the whole embryo culture system. There were dose-dependent growth retardation and abnormalities, predominantly in cardiovascular system. The three compounds exhibited very similar pattern of dysmorphogenic effects, but the potency of these compounds were quantitatively different. The incidences of embryos with the abnormalities were 100%, 100% and 85% following either exposure of NIF, NIC or NIT at concentration of 300, 8 and 15 μM, respectively. This study was to investigate whether these blocker-induced embryotoxicity was due to calcium channel blocking properties themselves in the embryos. Day 9 rat embryos were co-exposed to 1,4-dihydropyridine calcium channel agonist, Bay k 8644 (BAY) and each calcium channel blocker under the same culture condition. The retarded embryonic growth induced by 200 or 300 μM of NIF, 8 μM of NIC and 15 μM of NIT nearly or completely ameliorated when embryos were co-exposed with BAY at one-third or half concentration of each calcium channel blocker. Supplementation of BAY reduced the incidence of abnormalities by NIF-, NIC- and NIT-alone. These results suggested that one of mechanisms for embryotoxicity induced by calcium channel blocker was directly related to channel blocking property of the chemicals.

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Recently, a type-specific ELISA using equine herpesvirus type 1 (EHV-1) and type 4 (EHV-4) glycoprotein Gs (gGs) was developed by Crabb and Studdert [1993]. To investigate the dissemination of EHV-1 and -4 among horses in Japan, we applied their ELISA as suitable for discriminating between EHV-1 and -4 infections serologically. Type-specificity of the ELISA was confirmed by using paired sera of infected horses with either EHV-1 or -4. Application of the ELISA to sera collected before and after the winter season of 1995-1996 from 80 racehorses revealed that 30 horses showed significant antibody responses against EHV-1 and 9 against EHV-4, respectively. The results indicated that this ELISA using paired sera is useful for a diagnosis and an epizootiological study on EHV-1 and -4 infections.

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A male fetus of gestation day 187 was aborted from a Holstein-Friesian cow in an epizootic of the Aino virus (AINOV) in September 1995. Neutralizing antibody titers against AINOV were 1:128, 1:16 and 1:64 in the dam serum, fetal ascites and cerebrospinal fluid, respectively. A 10% brain suspension of the aborted fetus was prepared immediately after autopsy, rinsed three times and sonicated before centrifugation. The supernatant was then inoculated into HmLu-1 cell cultures. A cytopathic effect was noted on post-inoculation day 7. The isolated virus was identified as the AINOV based on the physicochemical properties and cross neutralization test. This is the first report on the isolation of AINOV from an aborted bovine fetus.

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