A mixture of Persistent Organic Pollutants (POPs) and Azoxymethane (AOM) show potential synergistic effects on intestinal tumorigenesis in the A/J Min/+ mouse model.

A multitude of cancer types, including breast, testicular, liver and colorectal cancer, have associations with exposure to Persistent Organic Pollutants (POPs). The present study aimed to investigate whether a mixture of POPs could affect intestinal tumorigenesis in the A/J Min/+ mouse, a model for human colorectal cancer (CRC). Pollutants were selected for their presence in Scandinavian food products and the mixture was designed based on defined human estimated daily intake levels. Mice were exposed through the diet, at control, low and high mixture concentrations, for 10 weeks. In a separate experiment, mice also received one subcutaneous injection of Azoxymethane (AOM) to explore whether this carcinogenic compound influenced the effect of the POPs. Intestinal tumorigenesis was examined by surface microscopy and histopathology. Moderate and dose-dependent increases in tumorigenesis were observed after dietary POP exposure. The AOM treatment alone stimulated the growth of colonic lesions, but did not increase the formation of new lesions. Combined AOM treatment and POP exposure demonstrated a synergistic effect on lesion formation in the colon, and to a lesser extent in the small intestine. This synergy was also evident by an increased number of malignant colonic tumors (carcinomas). In conclusion, the study shows that a mixture of POPs interacted synergistically with a known carcinogen (AOM), causing increased intestinal tumorigenesis in the A/J Min/+ mouse model.

Providing high-quality research training for veterinary pathologists in Europe.

Despite their key role in a wide range of fields relating to animal and public health, there is currently a lack of veterinary pathologists in Europe. In 1999, to help address the problem, the European College of Veterinary Pathologists (ECVP) and the European Society of Veterinary Pathology (ESVP) established a joint Education Committee. In this Special Article, Professor Anja Kipar and colleagues, all members of the committee, describe the ECVP/ESVP Summer Schools in Veterinary Pathology programme, which aims to provide high-quality research training for veterinary pathologists from all over Europe and beyond.

Detection of PrP(Sc) in rectal biopsy and necropsy samples from sheep with experimental scrapie.

Scrapie diagnosis is based on the demonstration of disease-associated prion protein (PrP(Sc)) in brain or, in the live animal, in readily accessible peripheral lymphoid tissue. Lymphatic tissues present at the rectoanal line were readily obtained from sheep without the need for anaesthesia. The presence of PrP(Sc) in such tissue was investigated in sheep infected orally with scrapie-infected brain material. The methods used consisted of immunohistochemistry and histoblotting on biopsy and post-mortem material. PrP(Sc) was detected in animals with PrP genotypes associated with high susceptibility to scrapie from 10 months after infection, i.e., from about the time of appearance of early clinical signs. In the rectal mucosa, PrP(Sc) was found in lymphoid follicles and in cells scattered in the lamina propria, often near and sometimes in the crypt epithelium. By Western blotting, PrP(Sc) was detected in rectal biopsy samples of sheep with the PrP genotype VRQ/VRQ, after electrophoresis of material equivalent to 8 mg of tissue. This study indicated that rectal biopsy samples should prove useful for the diagnosis of scrapie in sheep.

Ontogeny, distribution and structure of aggregated lymphoid follicles in the large intestine of sheep.

Aggregates of lymphocytes were demonstrated from 70 days gestation (term 150 days in sheep) in the proximal colon and rectum. Immunoperoxidase staining for 5'-bromo-2'-deoxyuridine incorporation and IgM, indicated that the lymphocyte population of lymphoid follicles in fetal sheep colon was actively dividing and surface IgM positive. Enzyme histochemistry for 5'-nucleotidase showed that the lymphocytes developed in a meshwork of positive reticular cells, suggestive of developing follicles. Follicle aggregates were distributed in a characteristic pattern in lambs, with major accumulations in the ascending colon and in the rectum. In adult sheep a partial atrophy of follicle aggregates was observed. The microscopic structure of large intestinal aggregates showed similarities to the jejunal Peyer's patch (PP), with broad follicles containing a prominent corona and wide interfollicular areas in older lambs. The apparent deep penetration of crypts into the lymphoid tissue proper, which is a frequently reported phenomenon for colon follicles, was dependent on the contractile state of the mucosa, as judged from its absence in specimens where the intestinal wall had been stretched before fixation.

The intestinal habitat for organized lymphoid tissues in ruminants; comparative aspects of structure, function and development.

Unlike the Peyer's patches of rats and mice, which are considered to be secondary lymphoid organs, the ileal Peyer's patch of sheep is thought to be responsible for the primary generation of B cells, like the bursa of Fabricius of birds. The ileal Peyer's patch of sheep shows prenatal maturation, antigen-independent lymphopoiesis, a rate of lymphocyte production larger than that of the thymus, and involution at a young age. Follicles contain few T cells and have an IgM+, relatively immature B lymphocyte population, as judged by B-cell differentiation markers. The follicle-associated epithelium of the ileal Peyer's patch is of a special type that sheds carbonic anhydrase-rich, 50-nanometer membrane-bounded particles (carbonic anhydrase-reactive particles; CAP) into the intercellular spaces. The CAP filter into the follicle centre and are taken up by lymphocytes. They represent the epithelial (bursa-like) element in an otherwise mesenchymal stroma of reticular cells embedding the follicle lymphocytes. Transepithelial transport of macromolecules, with the formation of multivesicular body-like cytoplasmic vacuoles, appears to be the basis for CAP formation. The jejunal Peyer's patches are devoid of CAP, persist in the adult animal, contain M cells with clusters of B cells in the follicle-associated epithelium, and have many CD4+ lymphocytes in the follicles and in the interfollicular areas. Aggregates of lymphoid follicles in the large intestine resemble the jejunal Peyer's patches with respect to their lymphocyte population and the ileal Peyer's patch with respect to their follicle-associated epithelium.

Morphological changes in the testis after long-term valproate treatment in male Wistar rats.

Uncertainty exists about the effect of antiepileptic drugs on gonadal function. In females, long-term valproate treatment has been shown to induce endocrine disturbances and an increased number of ovarian cysts. The aim of the present study was to investigate whether valproate can also induce morphological changes in the testis of male animals. In addition, possible morphological changes in the liver, heart, lungs, lymphatic nodes, pancreas, kidney or brain were studied. The carcinogenic implications were evaluated by the measurement of p53. Male Wistar rats were fed perorally with valproate mixture 200 mg kg(-1)(n= 15) or 400 mg kg(-1)(n= 20), or control solution (n= 15) twice daily for 90 days. Serum concentrations measured 4-6 hours after the last dose were 105 and 404 micromol l(-1)in low- and high-dose valproate treated animals respectively. There was a highly significant, 51% decrease (P< 0.001) in testicular weight in the high-dose treated valproate rats with no changes in the other groups. There was widespread testicular atrophy with histologically verified spermatogenic arrest in 15/20 of the high-dose valproate treated animals. No changes in the testis were seen in the low-dose valproate treated rats, nor in the control rats. There were no morphological changes in the other investigated organs. None of the groups showed over-expression of p53. In conclusion, a dose-dependent effect of chronic valproate treatment was found on testicular morphology in rats. Caution must be taken before these results can be applied to humans.

Diabetes mellitus and bilateral cataracts in a kitten.

An 18-week-old male domestic long-hair kitten was presented with a history of polyuria and polydipsia for several weeks. The general condition was unremarkable, but the kitten was considerably smaller than expected for the age and showed cataracts in both eyes. Serum glucose concentrations were persistently elevated and based on clinical findings and an elevated serum fructosamine concentration, a diagnosis of diabetes mellitus was established. Diabetes mellitus is not commonly diagnosed in young kittens, nor are cataracts recognised as a frequent feature of this disease in cats. The cataracts progressed in spite of the insulin therapy and the kitten was euthanised 10 weeks after referral. Histopathological examination of the pancreas revealed few and small islets of Langerhans compared to the examination of pancreas from a healthy kitten of the same age. Histopathological changes in the eyes included cataracts affecting both cortex and nucleus.

Malignant catarrhal fever caused by ovine herpesvirus-2 in pigs in Norway.

This paper describes the first cases of malignant catarrhal fever (MCF) in pigs in which the diagnosis was verified aetiologically by polymerase chain reaction (PCR) and DNA analysis and by the demonstration of antibodies. Three pigs on two separate premises showed clinical signs, gross pathological and histopathological lesions which were in many respects similar to those of MCF in ruminants. The pigs were housed adjacent to sheep and DNA of ovine herpesvirus-2 (OHV-2) was detected by PCR in tissues of all the pigs. In addition, antibody to alcelaphine herpesvirus-1 was detected in the serum of the pigs and in five in-contact sheep. It is concluded that the disease described is MCF of pigs caused by OHV-2.

Pancreatic insulin-secreting carcinoma in a dog: fructosamine for determining persistent hypoglycaemia.

A five-year-old intact male rottweiler was presented with a history of episodic weakness and mild-generalised seizures. A tentative diagnosis of an insulin-secreting tumour in the pancreas was made based on fasting hypoglycaemia with concomitant hyperinsulinaemia and a subnormal fructosamine value. The diagnosis was confirmed by exploratory coeliotomy, intravenous infusion of methylene blue, histopathology and immuno-histochemical analysis of suspected neoplastic tissue. Fructosamine assays are traditionally used for monitoring the metabolic status of diabetics where a single elevated measurement reflects persistent hyperglycaemia. This report suggests that a single low measurement of fructosamine may indicate persistent hypoglycaemia and may be helpful, in conjunction with an insulin measurement, in the diagnosis of insulin-secreting tumours.

Adult Dirofilaria repens in a subcutaneous granuloma on the chest of a dog.

A 10-year-old, 9 kg, intact male crossbred dog was treated for nasal mites with milbemycin oxime using a dose of 1 mg/kg bodyweight orally, three times at 10-day intervals. One month after the initiation of this treatment a subcutaneous nodule developed on the sternum of the dog. The nodule was removed and found to contain a single, 82 mm long, thread-like nematode. Several exotic parasites were suspected as possibilities because the dog had been imported to Norway from South Africa. Microfilariae were not detected in the blood and heartworm antigen tests were negative. The worm was identified morphologically as an adult, female Dirofilaria repens. This is the first report of D repens from Norway. The case is of interest because of the differential diagnostic problem it posed and because infestation was recognised following treatment of another parasitic condition with a broad-spectrum, antiparasitic drug. In addition, the case provides a reminder of the necessity to be aware of geographical differences in disease occurrence which can produce unexpected disease in non-endemic areas as a consequence of increased international travel with pets.

Diagnosis and clinical course of canine oral papillomavirus infection.

A six-month-old intact male rottweiler presented with papillomatous growths protruding from the oral mucous membranes. A tentative diagnosis of canine oral papillomavirus (COPV) infection was made based on the gross appearance of the numerous lesions and the young age of the patient. Two warts from the oral mucosa were removed surgically for further diagnostic investigations. The viral aetiology of the diseases was confirmed by histopathological and electron microscopic findings, and by the identification of specific COPV DNA is removed oral papillomatous tissue. The patient was followed clinically and complete regression of the oral lesions occurred after four weeks. Neither the route of transmission nor the source of infection was found. Immunodeficiency as a contributing aetiological factor to the development of COPV-induced lesions as discussed.

Potassium-dependent p-nitrophenyl phosphatase and carbonic anhydrase reactivities suggest that lymphoid follicles in the large intestine of lambs are lined with a uniform type of epithelial cell distinct from the absorptive epithelium.

The epithelium covering the large intestinal lymphoid follicles in fetal and postnatal lambs was examined for potassium-dependent p-nitrophenyl-phosphatase (K(+)-NPPase), carbonic anhydrase, magnesium-dependent adenosine triphosphatase (Mg(2+)-ATPase) and acid phosphatase. Reactivities for these enzymes indicated a homogenous population of cells in the follicle-associated epithelium (FAE), distinct from the absorptive epithelium. There were essentially no differences in the enzyme reactivities of the large intestinal FAE between fetuses in late gestation and postnatal lambs. The FAE showed a weak reaction for K(+)-NPPase and a variable staining for Mg(2+)-ATPase and acid phosphatase. In contrast, the adjacent absorptive epithelium demonstrated strong reactions for these enzymes. Carbonic anhydrase gave a strong reaction at the luminal and apparent basolateral cell borders of the large intestinal FAE. This distribution of reactivity for carbonic anhydrase resembled that found in the ileal FAE. In absorptive epithelial cells, only the luminal cell border reacted strongly for carbonic anhydrase. Serial sections of large intestinal tissue showed a variation in the basolateral staining of FAE from one section to the next, a finding which suggested that the reaction may be associated with transcytosis. The lymphoid follicles and domes of the large intestine showed a variable granular pattern of carbonic anhydrase staining, which also suggested a dependence on epithelial transcytosis.

Scarcity of gamma delta T cells in intestinal epithelia containing coccidia despite general increase of epithelial lymphocytes.

Lymphocytes of intestinal epithelia were examined in lambs experimentally infected with coccidia. Fourteen conventional, coccidia-free 3-week-old lambs of the Dala breed were given either 250,000 (eight lambs in experiment 1) or 100,000 (six lambs in experiment 2) sporulated oocysts of Eimeria ovinoidalis (> 99.9% pure) via a stomach tube. Nine lambs (five in experiment 1 and four in experiment 2) of the same age served as uninoculated controls. The infected lambs developed diarrhea and dehydration between days 10 and 13 after infection. Intestinal specimens were collected 10-20 days after inoculation. A heavy multifocal infection of the epithelium comprising schizonts, gamonts, and oocysts resulted, with changes being most pronounced in the large intestine and the terminal part of ileum. Lesions included villous atrophy, villous fusions, and crypt hyperplasia. Computer-assisted morphometric analysis was applied to immunoperoxidase-stained sections for quantification of intraepithelial T cell subsets and included the distal jejunum of lambs of experiment 1. A significant increase in the epithelial area of the distal jejunum occupied by CD8+ lymphocytes was found in the infected lambs as compared with uninfected controls (18.3% versus 3.7%). The increased presence of CD8+ cells may be of significance in the intestinal immune response to ovine coccidiosis. A moderate, overall increase was also found for the gamma delta lymphocytes in the jejunal epithelium. Examination of the ileum of infected lambs of both experiments showed a scarcity of gamma delta cells (0.4% of epithelial area) in stretches of epithelium infected by coccida as compared with adjacent noninfected epithelium (4.0% of epithelial area).(ABSTRACT TRUNCATED AT 250 WORDS)

Distribution of lymphocyte subsets in the large intestinal lymphoid follicles of lambs.

The phenotypes of lymphocytes in the large intestinal patches (LIP) of lambs were examined by flow cytometry and immunohistology, using a panel of monoclonal antibodies (mAb), and compared to those found in the jejunal (JPP) and ileal Peyer's patches (IPP). T-cell markers were detected on 25% of the LIP and JPP lymphocytes by cytofluorometry, and nearly all T cells expressed the CD4 molecule. In contrast, T cells were scarce in the IPP (less than 1%). The B-cell marker p220 was expressed by 74% of the LIP lymphocytes, whereas surface immunoglobulin-positive cells comprised 50-60% of the lymphocyte population. The adhesion molecule CD2 was expressed by a larger proportion of cells from the LIP and JPP than from the IPP, whereas the adhesion molecule CD44 was detected on more IPP lymphocytes. Major histocompatibility complex (MHC) class I antigens were expressed by nearly all lymphocytes from the LIP, JPP and IPP. The LIP contained 70-80% cells with MHC class II expression, whereas the majority of IPP cells (greater than 95%) were MHC class II positive. Immunohistology showed many CD4+ T lymphocytes in the follicles of the LIP and JPP, but none in the IPP follicles. CD8+ lymphocytes were found in the interfollicular areas and were absent from the follicles. The interfollicular areas of the rectal patch contained about 15% tau delta T cells. In contrast, the JPP, IPP and the colon patch at the beginning of spiral colon contained less than 3% tau delta T cells.