p62/Sequestosome-1: Mapping Sites of Protein-Handling Stress in Canine Cutaneous Mast Cell Tumors.

Canine cutaneous mast cell tumors (MCT) are common, frequently malignant neoplasms that are currently graded histologically for provision of prognostic information. Continuing evidence of subsets of MCT within certain grades (with differing survival times) indicate the need for biomarkers that will facilitate better patient stratification and also provide further information on the biological processes involved in progression. We decided to investigate the expression of p62/sequestosome-1 (p62/SQSTM1), a stress-inducible "hub protein" found in all cell types that shuttles rapidly between the nucleus and cytoplasm and is known to play important roles in protein handling and tumorigenesis. The identity of canine p62/SQSTM1 was confirmed in silico and by validation of a commercial antibody using both Western blotting and functional (pharmaceutical-based) analyses in cell culture. Using immunohistochemistry, 3 patterns of p62 expression were identified based on the predominant intracellular localization, that is, nuclear, mixed (nuclear and cytoplasmic), and cytoplasmic. There was a highly significant association with the 2-tier (Kiupel) grade (P < .0001), with all p62-nuclear immunoreactivity being associated with low grade and most p62-cytoplasmic immunoreactivity (93%) with high grade. Most but not all mixed nuclear-cytoplasmic labeling occurred in low-grade MCT; in other (human) tumor types, this pattern has been interpreted as borderline malignant. These data indicate that there is a shift in protein-handling stress from the nucleus to the cytoplasm in association with increasing malignancy in MCT. Studies to identify the processes and drug-able targets involved in this progression are ongoing.

Masculinization of the distal tubular and external genitalia in female sheep with prenatal androgen exposure.

Prenatal exposure to endogenous or exogenous androgens alters the development of the female reproductive tract. Although lesions in ovaries and external genitalia of androgenized female sheep have been reported, lesions of the tubular genitalia have not. Testosterone propionate (TP) or dihydrotestosterone (DHT) was administered by intramuscular injection twice weekly to 32 ewes from 30 to 90 days of pregnancy. The ewes lambed normally. The reproductive tracts from 24 treated and 13 control postpubertal female offspring were examined at 10 months of age. The ovaries, oviducts, and uteri were grossly and histologically normal in both TP- and DHT-exposed sheep. However, in the DHT-treated sheep, the uterus connected to a misshapen, saccular vagina that opened into the urethra; in the TP-treated sheep, it ended in a blind sac. In both TP- and DHT-treated sheep, the urethra was approximately 5 times longer than that of control sheep, and it resembled a male urethra with bilateral male accessory genital glands. The urethra terminated in a fully developed penis in both TP- and DHT-treated sheep, and a scrotal sac was present (without testes). These results show that prenatal exposure of female sheep to exogenous androgens results in masculinization of the tubular and external genitalia.

Streptococcal infection in dogs: a retrospective study of 393 cases.

Streptococcus spp are opportunistic pathogens that normally reside in the upper respiratory, intestinal, lower urinary, and genital tracts but can cause localized infection or septicemia in dogs of all ages. A retrospective study of streptococcal infection in 393 dogs was conducted to identify the species of Streptococcus isolated, determine demographics of affected dogs, and characterize the disease processes associated with infection. The major streptococcal species isolated were S canis (88 cases, 22.4%), S dysgalactiae ssp equisimilis (13, 3.3%), and S equi ssp zooepidemicus (4, 1.0%). Sex was not a risk factor (P > .30). Fetuses and neonates were more likely to have streptococcal infection than were other age groups (P < .001). Streptococcal septicemia was considered an important cause of abortion and neonatal death and was isolated from all samples submitted for aerobic culture from dogs in that age group. There was a seasonal trend, with dogs more likely to have streptococcal infection in summer months. In dogs for which a disease process was identified, streptococcal infection was associated with dermatitis (29 dogs), pneumonia (24 dogs), adult septicemia (13 dogs), and fetal/neonatal septicemia leading to abortion or neonatal death (16 dogs). Identification of other clinically significant bacterial, viral, fungal, and parasitic organisms was common (267 of 393 dogs, 68%), especially in dogs with dermatitis or pneumonia. Infection with Streptococcus spp should be considered in the differential diagnosis in cases of abortion, septicemia, dermatitis, and pneumonia in dogs. Clinical significance of isolation of streptococcal organisms should be interpreted in context of clinical signs and pathologic findings.

Immunohistochemical expression of c-KIT protein in feline soft tissue fibrosarcomas.

C-KIT is the cellular homolog of the feline sarcoma viral oncogene v-KIT, which encodes the tyrosine kinase receptor protein KIT. Mutations and varied expression of this gene have been demonstrated within multiple neoplasms in people and domestic animals. The purpose of this study was to determine if KIT protein is expressed in feline soft tissue fibrosarcomas (ST FSA) using immunohistochemistry (IHC). The computer database at the Oklahoma Animal Disease Diagnostic Laboratory was searched from January 1, 2006, to December 31, 2007, for any domestic cat with an ST FSA. Routinely stained slides from 46 feline ST FSAs were reviewed and graded based on the scale outlined by Kuntz et al. Immunohistochemistry for KIT protein was performed on one representative section from each cat. There were a total of 12/46 (26%) cats that were immunoreactive for KIT. Immunoreactivity was detected in greater than 80% of the neoplastic cells in 4/46 (9%) cats. Immunoreactivity was detected in less than 10% of the neoplastic cells in 8/46 (17%) cats. Immunoreactivity was characterized by evenly distributed cytoplasmic stippling within the neoplastic spindle-shaped cells and/or multinucleated giant cells. Based on these results, KIT immunoreactivity can be detected within feline ST FSAs using IHC. The results of this study also indicate that KIT immunoreactivity in feline ST FSA does not correlate with the histologic grade (P = .141, X(2) = 2.166), survivability (P = .241, X(2) = 1.373), or whether the neoplasm was a spontaneous or an injection site FSA (P = .074, X(2) = 3.184).

Bovine viral diarrhea virus abortion in goats housed with persistently infected cattle.

Twenty-four border disease virus-seronegative, pregnant, mixed breed goats were experimentally comingled with 3 heifers persistently infected with bovine viral diarrhea virus type 2a (BVDV-2a). Twelve of the 24 exposed does aborted. Twenty-nine fetuses and 16 placentas from affected does were submitted to the Oklahoma Animal Disease Diagnostic Laboratory for a necropsy examination. Infection with BVDV was confirmed with a combination of immunohistochemistry, BVDV-2 polymerase chain reaction, and virus isolation in 19 of the 29 fetuses. On gross examination of the 19 fetuses and placentas in which BVDV-2a infection was confirmed, a mild placentitis (3/19), fetal mummification (1/19), and facial deformities (4/19) were noted. Histologically, placentitis (2/19), myocarditis (4/19), thymic depletion (5/19), choroid plexitis (3/19), encephalitis (2/19), and cerebral gliosis (1/19) were noted. Other causes of abortion in goats, including common bacterial and viral infections, were ruled out with histology, virus isolation, polymerase chain reaction, and aerobic bacteriologic cultures. As supported by the findings in this case, BVDV-2a should be included as a differential for abortion in goats. This is the first report of abortion in goats after exposure to persistently infected cattle.

Distribution of bovine viral diarrhea virus antigen in aborted fetal and neonatal goats by immunohistochemistry.

Bovine viral diarrhea virus (BVDV) infection in goats can result in severe reproductive losses, with abortion rates reaching 80%. Infection with BVDV in aborted goat fetuses and stillborn kids can result in placentitis, encephalitis, myocarditis, and thymic depletion. This study investigates the distribution of viral antigen within the organ systems of aborted goat fetuses, stillborn kids, and nonviable kids infected with BVDV at various stages of gestation using immunohistochemistry (IHC). Virus antigen was detected within the placenta (8/13), thymus (4/9), heart (4/11), and brain (4/15) of affected goats. Uncommonly, BVDV antigen was detected within the skin (1/14), liver (1/13), kidney (1/12), lung (1/11), and trachea (1/3). BVDV antigen was not detected within the spleen (0/9), nasal turbinate (0/2), or thyroid (0/3). The results of this study indicate that placenta, heart, thymus, and brain are the most reliable tissues for BVDV antigen detection using IHC in aborted goat fetuses.

Ovarian and intestinal angiomatosis in a horse.

Dozens of red, raised nodules scattered along the serosal surface of the small intestine and the right and left ovaries were observed as incidental findings on gross examination in a 21-year-old Thoroughbred mare euthanatized for severe lameness. Histologically, these nodules were composed of numerous, variably sized, redundant vascular profiles filled with red blood cells and fibrin thrombi. Based on the presence of multiple nodules composed of benign vascular channels scattered within the small intestine and ovary, a diagnosis of angiomatosis is proposed. To the authors' knowledge, this is the first report of small intestinal and ovarian angiomatosis in a horse.

Advantages of a two-detector system in automated substoichiometric radioisotope dilution analysis.

Automation widens the scope of substoichiometric radioisotope dilution analysis. This is because the very strict requirement of the manual method-chemical separation of exactly equal quantities of the test substance-need no longer be fulfilled: reproducibility of the determination is reached by means of automated operation and activity measurement. The theory given in this paper shows how the choice of suitable chemical reactions is widened and why the reliability and the advantages of isotope dilution analysis are secured by the use of a two-detector system.

Automated determination of traces of mercury in biological materials by substoichiometric radioisotope dilution.

Solid samples (1-2 g) are burned in oxygen in a flask containing radiomercury in dilute hydrochloric acid, in which the non-active mercury to be determined is immediately absorbed. All mercury is subsequently extracted by dithizone in carbon tetrachloride and then re-extracted into dilute hydrochloric acid. This aqueous phase is further analysed automatically (AutoAnalyzer, 20 samples hr ) as previously described. Liquids (up to 100 ml) are analysed in the same way but instead of being burned in oxygen are first oxidized with potassium permanganate in acid medium. Quantities between 2 and 0.00004 ppm Hg were determined in various materials. Results for international biological standards agreed well with values obtained by activation analysis: kale 0.159 ppm Hg (relative standard deviation 2%) and IAEA cereals 0.0435 ppm Hg (+/- 5%). The new method is far more simple and rapid than activation analysis and just as sensitive; it is therefore more suitable for routine work. About 100 samples can be analysed per day.

A new concept of automated radiochemical analysis based on substoichiometric separation.

Tracer methods, such as radioisotope dilution, radiometric analysis, concentration dependent distribution, saturation analysis etc., are compared on a basis of radioactivity-mass balance relationships, and their automation is proposed. The requirements of a chemical separation, which is an integral part of these methods, are discussed. It is shown that automation, in addition to its obvious advantages, enables entirely new procedures to be developed, based on chemical separations which do not give reproducible results when performed normally. Simple commercially available apparatus has been used to verify these concepts by determination of traces of mercury. As little as 0.005 ppm of Hg can be determined, the detection limit being about a tenth of this. In the range 2.4-0.03 ppm, 20 samples/hr can be analysed, for lower amounts the sampling rate is 10 samples/hr.