Plaque assay of equine influenza virus

ESK cells, a stable cell line derived from a swine embryo kidney, were found to be a good medium for plaque formation of the Prague and Miami strains of equine influenza virus. Factors influencing the plaque formation were investigated and a plaque assay for these viruses was worked out. The method is not only simple enough for routine use, but also is as sensitive as the egg inoculation method. The method was readily adapted for a neutralization test.     

Promotion of lateral female strobili production inPinus densiflora by cytokinin application at a specific stage

BAP (N ⁶-benzylaminopurine) was sprayed 5, 10 and 15 times at four different periods to promote flowering of a grafted Japanese red pine (Pinus densiflora) clone. Four stages were I, August 4 to September 1; II, September 6 to October 4; III, October 5 to November 14 and IV, November 30 to December 28, in 1994. Concentration of BAP was 250 mg/l. In May, 1995, no apical female strobili were observed at the top of the new shoots in any treatment. However, BAP application at stage III (floral differentiation period) produced lateral female and bisexual strobili at the lower part of new shoots. The numbers of lateral female and bisexual strobili were 87 out of 147 to 234 out of 162 new shoots. Frequency of BAP application did not affect the numbers. Marked decrease in number of male strobili induced by BAP application at stages I and II was observed. In contrast, the number of new shoots was increased by the BAP application at the above stages. Treatment at stage III and IV did not affect significantly the number of male strobili and new shoots. These results indicate that BAP application at stage III specifically induces the development of lateral female and bisexual strobili in Japanese red pine.     

Effects of medetomidine and xylazine on intraocular pressure and pupil size in healthy Beagle dogs

ObjectiveTo determine the effects of intramuscular (IM) administration of medetomidine and xylazine on intraocular pressure (IOP) and pupil size in normal dogs.Study designProspective, randomized, experimental, crossover trial.AnimalsFive healthy, purpose-bred Beagle dogs.MethodsEach dog was administered 11 IM injections of, respectively: physiological saline; medetomidine at doses of 5, 10, 20, 40 and 80 μg kg⁻¹, and xylazine at doses of 0.5, 1.0, 2.0, 4.0 and 8.0 mg kg⁻¹. Injections were administered at least 1 week apart. IOP and pupil size were measured at baseline (before treatment) and at 0.25, 0.50, 0.75, 1, 2, 3, 4, 5, 6, 7, 8 and 24 hours post-injection.ResultsA significant decrease in IOP was observed at 6 hours after 80 μg kg⁻¹ medetomidine compared with values at 0.25 and 0.50 hours, although there were no significant changes in IOP from baseline. In dogs treated with 8.0 mg kg⁻¹ xylazine, significant reductions in IOP were observed at 4 and 5 hours compared with that at 0.25 hours after administration. In dogs treated with 5, 10, 20 and 40 μg kg⁻¹ medetomidine and 0.5, 1.0 and 2.0 mg kg⁻¹ xylazine, there were no significant changes in IOP. Pupil size did not change significantly after any of the medetomidine or xylazine treatments compared with the baseline value.Conclusions and clinical relevanceLow or moderate doses of medetomidine or xylazine did not induce significant changes in IOP or pupil size. In contrast, high doses of medetomidine or xylazine induced significant changes up to 8 hours after treatment, but values remained within the normal canine physiological range. The results of this study suggest a lack of significant change in IOP and pupil size in healthy dogs administered low or moderate doses of xylazine or medetomidine.     

Morphology of thyroid follicular cells of methimazole-treated rats

Thyroids of rats treated with methimazole (MMI) were analysed histologically, immunohistochemically and electron microscopically. Thyroids of MMI-treated rats displayed greater hypertrophy and were heavier than those of control rats. Thyroids of MMI-treated rats were composed of variant or small follicles and contained more columnar cells than those of control rats. Strong staining of thyroid granules by azocarmine G was evident in the apical regions of follicular cells from MMI-treated rats. Anti-thyroglobulin positivity was detected in the apical regions of follicular epithelial cells, especially those of MMI-treated rats. After application of a post-embedding method to detect thyroglobulin, colloidal gold particles were often observed in apical vesicles of MMI-treated rats. Our study suggests that apical vesicles may serve as markers of hyperthyroidism. When the functional state of the thyroid gland is known from histological analysis, use of the Azan stain could provide useful additional information.     

Measurement of porcine luteinizing hormone concentration in blood by time-resolved fluoroimmunoassay

We validated a time-resolved fluoroimmunoassay (TR-FIA) using the lanthanide element europium as a non-radiological tracer for measuring luteinizing hormone (LH) in porcine peripheral blood. The dose-response curve of the reference standard ranged from 0.2048 to 50 ng/ml. Good parallelism was noted between the LH standard and plasma sample. The profile of LH throughout the estrous cycle, assessed by daily blood sampling, was consistent with the previous findings obtained by radioimmunoassays (RIAs). Moreover, the secretory patterns of pulsatile LH in the follicular phase and the preovulatory LH surge were also similar to those obtained in previous RIAs. We conclude that TR-FIA can be used to measure LH levels in porcine blood, with practical and convenient applications.     

Cell proliferation activity of proliferating bile duct after bile duct ligation in rats

The cell proliferation activity of proliferating bile ducts produced by bile duct ligation (BDL) in rats was examined histologically, immunohistochemically, and ultrastructurally. Proliferating bile ducts, which were similar to normal bile ducts, increased with time after BDL. The cell proliferation activity of proliferating bile ducts, measured using proliferating-cell nuclear antigen and 5-bromo-2'-deoxyuridine antibodies, tended to be high at 1 and 3 days after BDL and decreased progressively at 2 to 4 weeks after BDL. On the other hand, alpha-smooth muscle actin-positive myofibroblast-like cells increased continuously after BDL. These findings indicate that there is a negative correlation between the cell proliferation activity of proliferating bile ducts and that of myofibroblast-like cells.     

Involvement of nitric oxide generation in hypersensitive cell death induced by elicitin in tobacco cell suspension culture

Recent studies suggest that nitric oxide (NO), an important signaling and defense molecule in mammals, plays a key role in activating disease resistance in plants. We characterized NO production by tobacco Bright Yellow-2 cells pharmacologically after treatment with INF1, the major elicitin secreted by the late blight pathogen Phytophthora infestans, prepared from Escherichia coli. NO production rapidly occurred within 1h and reached a maximum level 3–6h after the addition of INF1. Carboxy-PTIO, a NO-specific scavenger, abolished INF1-induced NO production in a dose-dependent manner. Pretreatment of protein synthesis inhibitor cycloheximide and protein kinase inhibitor K252a blocked NO production 3–12h after INF1 treatment, indicating that NO production requires de novo protein synthesis and protein phosphorylation. In an investigation of the relations between NO generation and several defense responses induced by INF1, carboxy-PTIO completely suppressed activation of a 41-kDa protein kinase and cell death by INF1. Carboxy-PTIO also suppressed the induction of hypersensitive-related (hsr) genes HSR515 and HSR203J, the expression of which is strongly correlated with the hypersensitive response in plants. The results suggest that NO plays a crucial role in the induction of hypersensitive cell death.     

A case of gastrinoma in a Shih-Tzu dog

A five-year-old male Shih-Tzu dog presented with severe vomiting and weight loss. The clinical signs were successfully improved by an eight-day treatment with an H(2)-receptor antagonist, gastrointestinal protectant and antibiotics. Ten days later, however, recurrence of vomiting was seen despite continuous medical treatment. Based on clinical signs and the results of various diagnostic tests including CBC, biochemical analysis, contrast radiography, and endoscopy, a duodenal or pancreatic neoplasm was suspected and exploratory laparotomy was conducted. Some swollen pancreatic regions were found, and biopsy of the pancreas indicated the diagnosis of a gastrin-secreting tumor. Consequently, based on a high serum gastrin level as well as clinical signs and immunohistological findings, we diagnosed the disease as canine gastrinoma, a rare tumor of the pancreas.     

Immune responses against measles virus in cynomolgus monkeys

Measles virus (MV) induces profound suppression of the immune response during and for weeks after acute infection. On the other hand, virus-specific immune responses that mediate viral clearance and confer long-lasting immunity are efficiently generated. To investigate this paradox, we studied the immune responses to MV using a monkey model of acute measles. Cynomolgus monkeys were experimentally infected with wild-type MV (MV-HL) and showed marked leukopenia associated with a steady reduction in CD4+ T cell numbers for 18 days post-inoculation. Transient expression of interferon and IL-6 were observed in the serum between 4 and 6 days post-inoculation, and IL-10 levels increased after 11 days post-inoculation. Interestingly, IL-8 showed a three-peak increase that correlated with an increase in neutrophils. A non-human primate model of measles allows the early immune response against MV to be studied in more detail.