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Biomedical subjects

T E Bunton

Publications and source records attributed to T E Bunton.

At least 19 recordsLinked to original sources

Cytochrome P450 isoenzyme activities in cultured rat and mouse liver slices.

1. The objective of this study was to determine the basal and inducible activities of several cytochrome P450 (CYP) isozymes and monitor the acinar and hepatocyte morphology in precision cut, cultured rat and mouse liver slices. 2. The slices were cultured up to 96 h in Chee's essential medium supplemented with insulin, transferrin, selenium, DMSO, dexamethasone and epidermal growth factor. A dynamic roller system was used to incubate the slices at 37 degrees C in an atmosphere of 95% O2:5% CO2. 3. Histopathology of the liver slices revealed maintenance of normal hepatic lobular architecture with time in culture. 4. CYP isozyme activities were measured at various times of culture. In rat liver slices, at 72 h, CYP1A1/1A2 activity was induced 4-fold by beta NF and 37-fold by dioxin (TCDD) whereas in mouse liver slices, 1A1/1A2 activity was not inducible by beta NF but was induced 19-fold by TCDD. At 72 h, CYP2A5 (coumarin-7-hydroxylase) activity was not detected in rat liver slices but in mouse liver slices, 2A5 was induced 2-fold by beta NF, 11-fold by phenobarbital (PB) and 3-fold by TCDD. 5. Hydroxylation of testosterone at specific positions was used as an indication of the activities of various P450 isoforms. Testosterone was added to the cultures at 0 and 72 h and the metabolites were measured at 24 and 96 h respectively by hplc analysis. Depending upon the species, the treatment and the time in culture, CYP1A, 2A, 3A, 2B and 2C activities were detectable. 3A activity was highly induced by PB in both rat and mouse liver slices. These results demonstrate that this culture system can be used to assess and compare xenobiotic metabolism in liver slices from rodent species.

Animals

Expression of actin and desmin in experimentally induced hepatic lesions and neoplasms from medaka (Oryzias latipes).

Proliferation of stromal cells is a common occurrence in experimentally induced hepatocarcinogenesis in fish. However, the role of these cells in fish hepatic injury and neoplasia is unknown. To better understand the biology of the cells comprising the hepatic stroma, livers from medaka (Oryzias latipes) experimentally induced by diethylnitrosamine or methylazoxymethanol acetate were labeled with keratin, actin, and desmin antibodies. The distribution of these proteins in hepatocellular carcinoma, cholangiocarcinoma, spindle cell proliferative lesions, and spongiosis hepatis was assessed, including three peritoneal sarcomas for comparison. Ductular epithelial cells in cholangiocarcinoma were positive for keratin, with desmin and actin positive ductular walls. Presumptive perisinusoidal cells in primarily trabecular and schirrous hepatocellular carcinoma were also actin positive. Only one spindle cell proliferative lesion was positive for any of the antibodies (desmin), and this lesion was morphologically distinct from others in the same category. Spongiosis hepatis, the peritoneal sarcomas, and normal perisinusoidal and other stromal cells were negative for these proteins. Since actin and desmin can be alternatively or coexpressed by mammalian perisinusoidal cells in association with hepatic fibrosis and neoplasia, the present studies suggest these proteins may serve as functional markers of hepatic stromal cells in fish as well.

Actins

Exclusion of copper from altered hepatocytes in white perch (Morone americana) with hepatic copper storage.

Iron is excluded from foci of hepatocellular alteration in carcinogenesis of rodents and some fish. Among white perch (Morone americana), there is a condition of hepatic copper storage in which copper-loaded livers are produced naturally. In a group of fish collected from the Chesapeake Bay, Maryland (USA), from September to December 1990, we observed hepatic lesions which excluded copper similar to the phenomenon of iron exclusion, in a white perch with over 3,600 micrograms/g wet weight hepatic copper. The lesions were of two types: one with cells morphologically different from normal hepatocytes and which had diminished to absolute exclusion of copper with the copper specific histochemical stain rubeinic acid, and a second with cells morphologically similar to normal hepatocytes which had only a partial exclusion of copper. Although the exact cause and nature of the lesions was not determined, intrinsic copper toxicity, environmental pollution, or a combination of these factors may have contributed to their development.

Animals

Intermediate filament reactivity in hyperplastic and neoplastic lesions from medaka (Oryzias latipes).

To determine if hyperplastic and neoplastic lesions from medaka showed similar immunoreactivity to intermediate filament antibodies as the tissues of origin, two week old medaka were exposed to 10 or 20 mg/L of methylazoxymethanol acetate for two hours and transferred to clean water for up to six months. Using a streptavidin peroxidase method, paraffin embedded Bouins fixed neoplasms were incubated with cytokeratin, vimentin, or neurofilament antibodies. Like their nonneoplastic cellular counterparts, hepatocellular carcinoma, pancreatic acinar carcinoma and mesenchymal neoplasms including hemangioma and hemangiopericytoma reacted negatively to cytokeratin antibodies. Cholangiocarcinoma, mesothelioma, and proliferative lesions containing biliary epithelial cells reacted positively to cytokeratin antibodies. All neoplasms and proliferative lesions were negative with vimentin and neurofilament antibodies. These data indicate that while some epithelial neoplasms showed cytokeratin reactivity similar to the parent tissues, additional markers are needed to identify mesenchymal tissues and neoplasms.

Adenoma, Liver Cell

The immunocytochemistry of cytokeratin in fish tissues.

An increasing interest in fish species as sentinels of environmental pollution and in carcinogenesis research has led to the identification of diagnostically challenging neoplasms of uncertain cellular origin and the need for additional diagnostic methods. To determine the potential of using commercially available antibodies to intermediate filament proteins on paraffin-embedded fish tissues for immunocytochemistry in tumor diagnosis, the application of three antikeratin antibodies to normal adult tissues from two fish species was assessed. Multiple tissues from 12-14-in. striped bass (Morone saxatilis) and 6-month-old medaka (Oryzias latipes) of both sexes were fixed in Bouin's or formalin fixatives. Formalin-fixed neoplasms from several mammalian species, including cat, dog, hedgehog (Atelerix albiventris, Erinaceus europaeus), rhesus macaque (Macaca mulatta), and sloth bear (Melursus ursinus), were also used as positive controls. Using a strepavidin horseradish peroxidase method on paraffin-embedded tissues, the broad spectrum antibodies AE1/AE3 (Boehringer Mannheim, Indianapolis, IN) and MAK-6 (Triton Biosciences, Alameda, CA), which recognize most of the 19 human cytokeratins, and CAM 5.2 (Becton Dickinson, Mountain View, CA), which recognizes cytokeratins present in human liver, were used as primary antibodies. Epithelia from skin, gills, cornea, bile ducts, renal tubules, gastrointestinal tract, and thymus were strongly positive with AE1/AE3 and MAK-6 in striped bass, but nonepithelial tissues such as bone and muscle were negative. Skin, gills, cornea, and portions of the gastrointestinal tract were strongly positive in medaka with the same antibodies, whereas bile duct, renal, and intestinal epithelia were less so. Tissue digestion improved the intensity of staining, and fixation with Bouin's fixative improved results somewhat compared with formalin fixation.(ABSTRACT TRUNCATED AT 250 WORDS)

Animals

Ultrastructure of hepatic hemangiopericytoma in the medaka (Oryzias latipes).

The histologic and ultrastructural features of hepatic hemangiopericytoma from a medaka (Oryzias latipes) exposed for 48 hr to 400 mg/liter of diethylnitrosamine at 14 days of age are described. The predominant histologic pattern was of spindle-shaped cells forming numerous whorls around central capillaries, vacuolated areas, or necrotic debris. The predominant cell type was a spindle-shaped cell with oval nuclei, elongated cell processes, and abundant organelles converging upon normal appearing capillaries. Occasionally, however, they converged upon cells swollen with cytoplasmic filaments and/or containing large fenestrated or debris-filled cytoplasmic vacuoles. These features were reminiscent of endothelial cells undergoing intracellular canalization seen in angiogenesis or neovascularization. Individual capillaries were also seen in the mass independent of whorls. It was not clear, as is the case in man, if capillary formation was an integral part of the neoplastic process or a reactive response. Although the liver is an unusual location for hemangiopericytoma in man, many of the cellular features in the fish tumor were similar to the human tumor. The ultrastructural characterization of tumor cells in fish carcinogenesis correlated with histologic patterns of growth will expand our understanding of how fish cells respond when transformed, and augment the development and use of aquatic bioassays for carcinogenesis research.

Actin Cytoskeleton

Comparison of three histochemical staining methods for the detection of copper in white perch (Morone americana) with abnormal hepatic copper storage.

A comparison of orcein, rubeanic acid and rhodanine histochemical stains in serial sections of liver from white perch (Morone americana) with abnormal hepatic copper storage is described. Staining reactions correlated well with hepatic copper concentrations although the threshold was the most problematic for rhodanine and rubeanic acid. Orcein was positive at a lower concentration than the two other stains. Hepatocellular granules and macrophage centres increased in size and number with copper concentration and could be related to increased lysosomes, as described for other species.

Animals

Hepatopathology of diethylnitrosamine in the medaka (Oryzias latipes) following short-term exposure.

The effect of short-term exposure (48 hr) of 14-day-old medaka (Oryzias latipes) to diethylnitrosamine (DEN) was tested on 4 groups of fish at 0, 100, 200, and 400 mg/L, with interim sacrifices of up to 6 months post exposure. Many of the lesions seen were similar to those seen in other fish DEN carcinogenicity studies using longer term, adult exposures, including a cytotoxic phase with cystic degeneration of the hepatic parenchyma, cellular vacuolation, globular acidophilic cytoplasmic inclusions, spongiosis hepatis, foci of cellular alteration, adenomas, and cholangiomas. There were also several proliferative lesions of uncertain origin which occurred throughout the study. There appeared to be a direct relationship between exposure level and the incidence and severity of the lesions seen, including aggressiveness and invasiveness of the neoplasms seen. However, there was a possible difference from other studies in the types of neoplasms seen with this protocol, which may be related to age at exposure, exposure level, duration of exposure, or a combination of these factors. Similarities in lesions seen in this study and in rodent and other fish studies is an indication of promise for the use of this model in carcinogenicity testing. Differences indicate the variety of ways the fish liver can respond to toxic injury, and lend flexibility to the model.

Animals

Cholangioma in white perch (Morone americana) from the Chesapeake Bay.

Cholangiomas found in two of 21 wild-caught white perch (Morone americana) from the Chesapeake Bay are described. The two fish were part of a study investigating a condition of abnormal hepatic copper storage in this species. The tumors were superficial, solitary masses consisting of cuboidal to columnar cells in tubuloglandular arrangement. Mild to marked peribiliary inflammation and fibrosis was seen also. Environmental pollution, the condition of abnormal copper storage, peribiliary fibrosis, and/or parasites may have contributed to the development of these tumors.

Adenoma, Bile Duct

Herpesvirus-like infection in black-footed penguins (Spheniscus demersus).

A herpesvirus-like infection is described in the black-footed penguin (Spheniscus demersus). Clinically, the infection was characterized by debilitation and respiratory distress. Histopathological lesions were confined to the respiratory tract and consisted of inflammation and syncytial cell formation with Type A intranuclear inclusions in sinuses, trachea, and mainstem bronchi. Electron microscopy demonstrated polyhedral viral particles 80-140 nm in size consistent with Herpetoviridae. The lesions resembled those seen in infectious laryngotracheitis.

Animals

Abnormal hepatic copper storage in a teleost fish (Morone americana).

Excessive copper storage in livers of feral white perch (Morone americana) from the Chesapeake Bay is described. Age-related, progressive accumulation of hepatic copper in levels often exceeding 1,000 micrograms/g wet weight was associated with peribiliary fibrosis and inflammation, bile duct hyperplasia, prominent, enlarged melanomacrophage centers, and disruption of hepatic architecture in older fish. Levels of zinc were mildly elevated compared to striped bass (Morone saxitilis) and adult rats. Cholangiomas were found in two perch. Rubeanic acid-stained liver had abundant copper-positive cytoplasmic granules in hepatocytes and cells of melanomacrophage centers. Subcellular fractionation showed that 90% of hepatocellular copper was in nuclei/cell debris fractions (which also contain tertiary lysosomes). Using electron probe microanalysis, high copper levels were localized in hepatocellular cytoplasmic bodies. Resolution of hepatic cytosol by gel permeation chromatography indicated that approximately 50% of the cytosolic copper in the white perch was bound to non-specific high molecular weight proteins, with the remaining 50% eluting at a peak where rat metallothionein is located. Ultrastructural examination revealed abundant lysosomes, increased size and number of peroxisomes, and increased density and numbers of mitochondrial matrix granules. This study indicates that white perch may be a model for studying effects of excessive copper accumulation and cellular mechanisms which control copper kinetics.

Animals

Incidental lesions in nonhuman primate placentae.

Twenty-one placentae from uncomplicated pregnancies of rhesus macaques in the third trimester (124-164 days) were examined to determine placental incidental lesions. Placental weight (PW) generally correlated with fetal weight (FW); PW:FW equaled approximately 1:3. Most placentae were bidiscoid (9.5% monodiscoid) with eccentric cord insertion and marginal or circummarginate membrane insertion. Macroscopically, there was in varying degrees, subchorionic, perivillous, and perilobular fibrin deposition, focal infarction, retroplacental hematoma, and calcification. Histologic changes included fibrinoid necrosis of villi, multifocal acute inflammation, excessive cytotrophoblasts, formation of syncytial knots, and intimal proliferation in umbilical vessels. Volume density of fibrinoid necrosis and calcification in terminal villi showed nonstatistically significant increases to term. However, calcification in reference to entire placental tissue increased significantly to term. All lesions seen in the rhesus are also seen incidentally in human placentae, but may become significant in correlation with pregnancy wastage if present to a large degree.

Animals

Clinical and histological features of primary progressive, familial thyroiditis in a colony of borzoi dogs.

A six-year study of clinical data and the morphologic changes involved in lymphocytic thyroiditis in a colony of related borzoi dogs was conducted. Lesions observed included initial degenerative thyroidal parenchymal changes which progressed to subacute inflammation with subsequent fibrosis and end stage thyroid gland disease. This study encompasses three successive generations of this borzoi colony and documents the progression of the histologic changes from the initial thyroid gland degenerative lesions to the end stage parenchymal atrophy. Spontaneous, familial thyroiditis has not been reported previously in the borzoi breed.

Animals

Triamcinolone-induced structural alterations in the development of the lung of the fetal rhesus macaque.

To test the effect of dose and time on fetal lung maturation by exogenous glucocorticosteroids, triamcinolone acetonide was given to time-mated pregnant rhesus macaques (Macaca mulatta). Triamcinolone acetonide (10 mg/kg or 1 mg/kg) was administered intramuscularly during the midpseudoglandular (63 to 65 days' gestational age) or midcanalicular phase (110 to 112 days' gestational age) of lung development. Fetectomies were performed at 90, 120, or 150 days' gestational age. Nontreated control fetuses and postnatal animals from 59 days' gestational age to 31 days' postnatal age were collected for study of normal developmental lung morphology in the rhesus monkey. Lungs from fetuses of 90 days' gestational age and older were fixed by tracheal infusion, and lungs from fetuses less than 90 days' gestational age were fixed by immersion and embedded for high-resolution light microscopy. Body weight, crown-rump length, and fixed lung volume were determined for all fetuses. Morphometric evaluation of the volume percent of parenchyma, the volume percent of air space and the mean linear intercept as an estimate of air space size was also done. Lungs from fetuses treated during the canalicular phase had thinner, longer, less cellular septa, larger air spaces, and increased numbers of alveolar divisions in terminal air spaces as compared to those of controls. Lungs of fetuses treated during the pseudoglandular phase also had some septa which were thinner, less cellular, and longer than those of controls and had a marked increase in air space size. However, most septa were greatly reduced in height and appeared fewer in number. Alveolar divisions were decreased. The volume percent of air space was larger in all treated groups. Fixed lung volume normalized to body size (body weight and crown-rump length) was greater than that for controls for the high-dose canalicular phase treatment group. The same parameter was lower than that of controls for the high-dose, pseudoglandular phase treatment group of 150 days' gestational age. Body weight and crown-rump length for all pseudoglandular phase triamcinolone acetonide-treated fetuses also tended to be lower than those of controls. We concluded that: (1) there is both a time-dependent and a dose-dependent effect on triamcinolone acetonide on fetal lung maturation in rhesus macaques, (2) triamcinolone acetonide accelerates maturation of the interstitial and epithelial component of the developing fetal rhesus lung independent of age, (3) triamcinolone acetonide accelerates alveolarization only at later treatment ages, and (4) triamcinolone acetonide induces retardation of growth of some of the lung septa and body growth only

Animals