The role of cell-mediated cytotoxicity in acute GVHD after MHC-matched allogeneic bone marrow transplantation in mice.

The role of cell-mediated cytotoxicity in the complex pathophysiology of graft-versus-host disease (GVHD) has remained poorly defined for several decades. We transplanted T cells from Fas-ligand (FasL)-defective and perforin-deficient mutant donor mice into lethally irradiated MHC-matched allogeneic recipient mice to characterize the role of cell-mediated cytotoxicity in GVHD. Although recipients of allogeneic FasL-defective donor T cells underwent severe GVHD-associated cachexia, they exhibited only minimal signs of hepatic and cutaneous GVHD pathology. Recipients of perforin-deficient allogeneic donor T cells developed signs of acute GVHD, but the time of onset was significantly delayed. These findings demonstrate that Fas-mediated anti-recipient cytotoxicity may be critical for the development of hepatic and cutaneous GVHD, but is not required for GVHD-associated cachexia. In addition, perforin-mediated anti-recipient cytotoxicity appears to play an important role in the kinetics of GVHD pathophysiology, but is not required for GVHD-associated tissue damage.

The influence of DL-methionine on the metabolism of S-adenosylethionine in rats chronically treated with DL-ethionine.

The concentration of S-adenosylethionine in the liver of ethionine-fed rats was increased gradually during the process of carcinogenesis. This increase may have been due to the decreased capacity of the treated rats to acetylate ethionine sulfoxide. Ethionine sulfoxide is considered as the main reserve pool of ethionine for the synthesis of S-adenosylethionine. When the ethionine diet was supplemented by DL-methionine (0.3 to 0.9%), the increase in the concentration of S-adenosylethionine during the period of observation (28 to 150 days) was lower and the acetylation of ethionine sulfoxide was significantly higher. The concentration of the total S-adenosyl compounds in the liver of rats on a diet supplemented with DL-methionine was increased over the concentration of S-adenosylethionine in rats fed ethionine alone, and the S-adenosylethionine portion of this fraction was only about 30% lower. The supplementation of the diet with methionine restored the diurnal oscillation of adenosine 5'-triphosphate in the liver, which had been absent in rats ingesting only ethionine.

The effect of cupric acetate on ethionine metabolism.

The addition of cupric acetate, a potent inhibitor of ethionine carcinogenesis, to a diet containing ethionine increased the ethionine toxicity. The concentration of S-adenosylethionine in liver was found to be significantly higher when compared to animals fed only ethionine in the diet. Ethionine forms a complex(es) with cupric acetate that is insoluble at a pH higher than 4; however, this complex can be solubilized at a low pH. Ethionine, if administered p.o. in the form of this complex, was absorbed from the intestinal lumen in the same order of magnitude as when administered alone; however, as the body weight increased over 200 g, the portion of absorbed ethionine decreased. The absorption of ethionine bound in the complex was completed within 16 hr compared to 2 hr for free ethionine. This time delay was accompanied by a shift in the concentration maximum of ethionine metabolities in the liver form 8 to 24 hr. When ethionine was administered alone, it was metabolized in the intestinal lumen as demonstrated by the analysis of the soluble intestinal contents; the presence of cupric acetate inhibited this process. The chromatographic analysis of ethionine metabolites in urine of rats treated by the complex revealed an increased excretion of ethionine sulfoxide and other ethionine metabolities at the expense of N-acetylethionine sulfoxide. The increased concentration of S-adenosylethionine in the liver in chronic experiments may be, at least partly, a result of a diminished capacity of the rat to detoxify (acetylate) ethionine sulfoxide, which is considered the main reserve pool of ethionine for the maintenance of a high level of S-adenosylethionine.

Failure of pure 1-naphthylamine to induce bladder tumors in dogs.

1-Naphthylamine was administered p.o. to beagles for approximately 9 years at a daily dose of 15 mg/kg (5 days a week). At autopsy, no tumors or other pathological changes were observed in the bladders of any of these animals. With the possible exception of the excessive accumulation of lipofuscin in the hepatocytes of these dogs, no test compound-related pathological changes in other tissues of the body were observed. Analysis of the urine revealed the presence of small amounts of N-oxidation products (1-nitrosonaphthalene plus N-hydroxy-1-naphthylamine), ranging from 2.85 to 125 microgram, following a dose of 1-naphthylamine. The theoretical significance of these findings is discussed.

Prostate cancer progression, metastasis, and gene expression in transgenic mice.

We previously reported that a transgenic mouse line containing the fetal globin promoter linked to the SV40 T antigen (T Ag) viral oncogene (Ggamma/T-15) resulted in prostate tumors. In this study, we further explored tumor origin, frequency, invasiveness, androgen sensitivity, and gene expression pattern. T Ag was detected in adult but not fetal and neonatal prostates, suggesting a role for androgens in tumor progression. However, castration shortly after prostate morphogenesis did not prevent tumor development, suggesting an androgen-independent phenotype. Tumors originated within ventral or dorsal prostate lobes and involved intraepithelial neoplasia, rapid growth in the pelvic region, and metastasis to lymph nodes and distant sites. In addition, the primary cancers could be propagated in nude mice or nontransgenic mice. Seventy-five percent of hemizygous and 100% of homozygous transgenic males developed prostate tumors, suggesting a T Ag dosage effect. Biochemical characterization of advanced tumors revealed markers of both neuroendocrine and epithelial phenotypes; markers of terminal differentiation are lost early in tumorigenesis. Tumor suppressor genes (p53 and Rb), normally bound to T Ag, were up-regulated; bcl-2 proto-oncogene, which prevents apoptosis, was slightly up-regulated. Myc, a stimulus to cell cycle progression, was unchanged. We propose the Ggamma/T-15 transgenic line as a model of highly aggressive androgen-independent metastatic prostate carcinoma with features similar to end-stage prostate cancer in humans.

Topical fluorouracil. II. Postoperative administration in an animal model of glaucoma filtering surgery.

Unilateral posterior lip sclerectomies were performed in ten owl monkeys. Five milligrams of fluorouracil was injected subconjunctivally in each operated eye immediately after surgery. Three drops (approximately 2.4 mg/drop) of fluorouracil were instilled ten minutes apart in each operated eye twice daily on postoperative days 1 through 7 and once daily on postoperative days 8 through 15, 17, 19, and 21. One monkey died on the seventh postoperative day; its death could not be attributed to systemic fluorouracil toxicity. All of the operated eyes had filtering blebs after the full course of fluorouracil, but seven also had corneal epithelial defects. By the seventh postoperative week, two of the operated eyes manifested moderately severe corneal opacification. Ten weeks postoperatively, the electroretinographic a- and b-wave amplitudes averaged 17% and 12% less, respectively, in the seven operated eyes without clinically significant corneal opacification than in the unoperated fellow eyes. Only two eyes had blebs after the 12th postoperative week. Histopathologic examination was performed on five eyes, of which only two revealed patent sclerostomies. Although topical fluorouracil appears to delay bleb scarring, the corneal findings suggest that it may be more toxic than subconjunctival fluorouracil.

Macrophage inhibitory factor-A3 derived from Mycobacterium avium serovar 2 inhibits candidacidal activity of murine peritoneal macrophages.

Macrophage inhibitory factor-A3 (MIF-A3), a fraction derived from Mycobacterium avium serovar 2 inhibited candidacidal activity in macrophages from C57BL/6, C57BL/10, C3H/HeJ and A/J strains of mice. Inhibition of candidacidal activity was demonstrated at MIF-A3 concentrations ranging from 100-400 micrograms/ml in macrophages without additional stimulators (exception C3H/HeJ macrophages) and in macrophages additionally stimulated with 200 U/ml interferon-gamma, 100 ng/ml phorbol myristate acetate and 0.4 ng/ml E. coli lipopolysaccharide from all mouse strains tested. The decreased candidacidal effect produced by MIF-A3 was dose-dependent and appeared greatest in macrophages treated with phorbol myristate acetate and lipopolysaccharide. This effect was neutralized by the addition of goat anti-MIF-A3 antiserum. Macrophages from the Bcgs mouse strains (C57BL/6 and C57Bl/100 were more sensitive to the effect(s) of MIF-A3 than macrophages from the Bcgr mouse strains (C3H/HeJ and A/J).

Histopathologic study of herpes virus-induced retinitis in athymic BALB/c mice: evidence for an immunopathogenic process.

In order to determine whether antiviral immunity is pathogenic in mouse eyes, HSV-1 was injected into the anterior chamber of one eye of adult athymic BALB/c mice. The eyes of these T cell deficient mice were examined clinically and histopathologically for ocular disease. The anterior segment of injected eyes developed progressive inflammatory reactions that eventually destroyed the ciliary body and then progressed to the posterior compartment where partial necrosis occurred, but only in the inner layers of the retina. A milder form of the same process developed between 7 and 10 days in the contralateral eye. Uninoculated eyes displayed little evidence of choroiditis, hemorrhage, massive necrosis, or disintegration of the architecture of the retina. Since these are features that are found in contralateral retinas of euthymic BALB/c mice infected in one eye via the anterior chamber route, it is concluded that acute retinitis found in contralateral eyes of immunocompetent mice has an immunopathogenic basis. However since euthymic mice develop anterior chamber associated immune deviation (ACAID) (and therefore do not display virus-specific delayed hypersensitivity), the identity of the relevant immune effector remains unknown. Based on these observations and our previous ocular findings following intracameral inoculation of HSV-2, we suggest that in susceptible mice, herpes simplex viruses can induce several pathogenetically distinct forms of retinitis, some of which are mediated by virus-specific immune effector cells.

Feline immunodeficiency virus infection in a population of pet cats from southeastern Florida.

Blood samples from 95 randomly selected pet cats that were brought to veterinarians in southeastern Florida were tested for antibodies to feline immunodeficiency virus (FIV). Virus-specific antibodies (indicative of virus infection) were found in 8 of the 95 (8.4%) cats tested. All of the virus-infected cats were males (statistically significant, P less than or equal to 0.016) and were at least 1 year of age. The 3 most severely ill cats infected with FIV were also infected with feline leukemia virus.

Identification of immunoglobulin light chains in canine extramedullary plasmacytomas by thioflavine T and immunohistochemistry.

Extramedullary plasmacytomas were studied in 29 dogs. The site at which tumors occurred and the age and sex of the dogs were similar to those in previous reports. The skin of the digits, chin, ear, and lip represented the most common (17/29) tumor sites. Males and females were equally represented, and tumors occurred in middle-aged to old dogs (mean age, 9.0 years). A breed predilection was seen in the Cocker Spaniel (n = 7; 24%); Cocker Spaniels represented only 4% (210/4,725) of the submissions during the same period. Tumors were stained with immunohistochemical markers (lambda light chain, K light chain) and thioflavine T. Immunoreactivity was limited to either lambda or K light chains, consistent with a monoclonal plasma cell population. The majority of tumors expressed lambda light chains, consistent with previously reported canine plasma cell dyscrasias. Thioflavine T cytoplasmic fluorescence was seen in the majority (18/29) of plasmacytomas and with inflammatory plasma cells present in control specimens. Other round cell neoplasms (lymphosarcoma, histiocytoma, and mastocytoma) were negative with thioflavine T, indicating that positive staining with thioflavine T was specific for plasma cells (neoplastic and inflammatory). This study confirms by immunohistochemistry that canine extramedullary plasmacytomas disproportionately express lambda light chains and establishes thioflavine T staining as a rapid histochemical method for diagnosis of these tumors.

Four cases of neoplasia in captive wild birds.

Four cases of malignant neoplasia in captive wild birds are described: an adenocarcinoma of the adrenal gland in a Mountain duck (Tadorna tadornoides), a malignant melanoma in the thoracic cavity of a Combed duck (Sarkidiornis melanotos), a hepatocellular carcinoma with pulmonary metastasis in an Asian Purple Swamphen (Porphyrio porphyrio), and an undifferentiated carcinoma in the abdomen with metastasis to skeletal muscle in a White-Breasted Waterhen (Amaurornis phoenicurus). The tumors were diagnosed during a 1-year period and represented an incidence of neoplasia of 3.1%. These appear to be the first documented cases of a malignant adrenal gland tumor and a non-ocular melanoma in the order Anseriformes. The hepatocellular carcinoma failed to react with an immunoperoxidase stain for alphafetoprotein.

Metastatic malignant melanoma in a mandarin duck (Aix galericulata).

A biopsy taken from a mass on the dorsal surface of the bill of an adult female mandarin duck (Aix galericulata) was diagnosed as a malignant melanoma by light microscopy. Two months later, the tumor had enlarged considerably; the duck developed severe dyspnea and was euthanatized. At necropsy, there were metastases to lymphoid tissues in the lower regions of the neck. To our knowledge, this is the first report of a malignant melanoma in a mandarin duck.

Neoplasia in ferrets: eleven cases with a review.

Records from a veterinary diagnostic laboratory in south Florida, U.S.A. were reviewed for cases of neoplasia in pet ferrets. Twelve ferret tumours were received over a four-year period; one case, a ferret with lymphocytic leukaemia and multi-organ involvement, had been reported previously. The other eleven tumours were: two chordomas of the tail, two sebaceous adenomas of the skin, a sebaceous epithelioma of the skin, a cutaneous mastocytoma, a malignant fibrous histiocytoma from the eyelid, a malignant mesenchymoma and an undifferentiated sarcoma from the dorsal abdominal cavity, a leiomyosarcoma found unattached in the abdominal cavity and an interstitial cell tumour of the testicle. A review of the literature yielded reports of 83 other tumours in domestic ferrets, black-footed ferrets and European polecats. Of the 95 ferret tumours, 46 were considered malignant. Tumours occurred in all organ systems except the respiratory tract and central nervous system. Affected ferrets ranged in age from 209 days to 12 years. The most frequently occurring tumours were ovarian stromal tumours (24 of 95), haemangiomas/haemangio-sarcomas (15 of 95). This information indicates that, contrary to previous opinion, ferrets appear to be subject to a similar incidence and variety of tumours as other animals.

Metastasizing extra-adrenal paraganglioma with neurological signs in four dogs.

Extra-adrenal paragangliomas associated with vertebral pain and clinical neurological abnormalities as a result of metastasis to the vertebral column were diagnosed in four dogs of different breeds by light microscopy. All were males (two intact and two neutered) aged 8 years. Metastatic neoplasms occurred as extradural masses with associated bone lysis at vertebrae C4 (2 cases), T12-L1 (1 case) and L4 (1 case). The neoplastic cells exhibited similar morphology with little variation between cases. All neoplasms showed cytoplasmic granules after staining with the Churukian-Schenk modification of the Pascual argyrophil stain for neurosecretory granules or for membrane bound electron-dense granules (dense-core granules). On immunohistochemical examination the neoplastic cells gave positive results for neuron-specific enolase and negative results for chromogranin and epithelial membrane antigen. Multiple organ metastasis and metastasis to bone have been reported previously, but these cases were unusual due to the involvement of the spine as an apparent predilection site for metastasis, and the sex (male) and age of the animals affected.

Salivary gland oncocytes in African hedgehogs (Atelerix albiventris) mimicking cytomegalic inclusion disease.

The salivary glands from three African hedgehogs contained multiple foci of cytomegalic cells, which occasionally had a mild to moderate infiltrate of lymphocytes at the periphery. The cytomegalic cells were 35 to 40 microns in diameter with abundant acidophilic granular to hyalin cytoplasm. The nuclei were enlarged with clumped marginalized chromatin and a large, (6 to 8 microns in diameter) central, brightly eosinophilic nucleolus that had the appearance of an inclusion body by light microscopy. Histochemically most of the cytomegalic cells contained cytoplasmic metachromatic granules with Feyrter's thionine inclusion stain. Scattered cells at the periphery of the cytomegalic foci contained periodic acid-Schiff-positive cytoplasmic granules. Ultrastructurally the cytomegalic cells contained numerous tightly-packed, often bizarre, enlarged mitochondria that completely filled the cytoplasm. The nucleus consisted of a dense central core of chromatin associated with the nucleolus and the remaining chromatin was clumped and marginalized. Nuclear and cytoplasmic virions consistent with cytomegalovirus were not present. Histochemical stains of the nucleus for heavy metals were negative. The ultrastructural and histochemical findings of the cytomegalic cells were consistent with oncocytes. Previous reports in the literature of similar cells in the salivary glands of insectivores appear to have been erroneously described as cytomegalovirus infections.

Metastasis of a myxoid leiomyosarcoma via the renal and hepatic portal circulation in a sarus crane (Grus antigone).

A 12-year-old female sarus crane (Grus antigone) developed a recurrent proliferative lesion in the subcutaneous tissue of the tarsometatarsus, which failed to respond to medical and surgical therapy. The crane was killed and microscopic examination of the tissues taken at necropsy revealed a myxomatous, poorly-differentiated sarcoma with metastasis to the liver and kidney. Immunohistochemical staining for muscle actin, smooth muscle myosin and vimentin were positive, indicating that the primary and metastatic tumours were leiomyosarcomas. Location of the metastatic lesions in only the portal venous system of the liver and veins of the kidney indicated that the route of metastasis was the portal circulation via the ischiatic vein, caudal mesenteric vein and both the renal-portal shunt and hepatic portal vein. This is the first report of metastatic subcutaneous leiomyosarcoma in an avian species.

Hematologic, plasma protein, and biochemical profiles of brown pelicans (Pelecanus occidentalis).

OBJECTIVE: To establish hematologic and biochemical reference values for the brown pelican (Pelecanus occidentalis). ANIMALS: 31 captive, healthy, but permanently disabled pelicans and 35 wild-caught, healthy pelicans from a rehabilitation facility on the east coast of Florida. PROCEDURES: Samples of venous blood were collected from each pelican, and hematologic, plasma biochemical, and electrophoretic protein analyses were performed. Student t-tests were used to compare blood values between captive versus wild-caught, adult male versus adult female, and adult versus juvenile pelicans. RESULTS: Hematologic and electrophoretic values were similar between male and female, adult and juvenile, and captive and wild-caught pelicans. Significant sex differences existed for plasma calcium and triglyceride concentrations. Plasma concentrations of calcium, cholesterol, and CO2 content differed between captive and wild-caught adults. No significant differences were found between wild-caught adult and juvenile pelicans. CONCLUSIONS AND CLINICAL RELEVANCE: Our plasma biochemical results are similar to those of other brown pelicans and confamilial species. Additional studies on seabirds are encouraged, as age, sex, reproductive status, feeding habits, and captivity are important variables for health assessment in this and other aquatic species.

Cardiomyopathy in stranded pygmy and dwarf sperm whales.

Necropsy and histologic examinations were performed in 23 pygmy sperm whales (Kogia breviceps) and 6 dwarf sperm whales (Kogia simus) that had been stranded singly or in cow-calf pairs along the southeastern coastline of the United States. At necropsy, the gross findings in the adult whales included pale, flabby right ventricles. Microscopically, lesions in the hearts of the whales were characterized by moderate to extensive myocellular degeneration, atrophy, and fibrosis. Similar changes were not seen in 5 of 6 sexually immature whales or in the whale calves. Hepatic changes were consistent with heart failure. The cause of the myocardial lesions was not determined. The systemic effects of failing myocardium probably were a major reason for the stranding of the adult whales.

Protothecosis in two cats.

Protothecosis was diagnosed in 2 cats from Florida with nodular skin lesions. Microscopically, lesions were composed of macrophages and multinucleated giant cells arranged in solid sheets in the dermis and subcutis. In both cats, characteristic algal endospores identified as Prototheca wickerhamii were observed within the cytoplasm of infiltrating cells.

Gastric extramedullary plasmacytoma in a dog.

A 10-year-old mixed-breed dog was examined because of a 6-week history of daily vomiting and sporadic diarrhea. On gastroscopy, a crateriform mass was observed on the greater curvature of the stomach. Partial gastrectomy and lymphadenectomy of a large mesenteric lymph node was performed. Gastric plasmacytoma with lymph node metastasis was diagnosed by histologic and immunoperoxidase methods, and chemotherapy was initiated with doxorubicin hydrochloride and diphenhydramine hydrochloride. The dog remains clinically normal 30 months after initial diagnosis. Although gastric plasmacytomas are rare in dogs, long-term survival appears to be better with this disease than with other types of gastric neoplasia.