Histogenesis of hyperplasia and carcinomas of the liver arising around central veins in mice ingesting chlorinated hydrocarbons.

The development of hyperplastic and neoplastic lesions of parenchymal cells of the liver adjacent to central veins was observed in C3H mice ingesting the chlorinated hydrocarbons, dieldrin or aldrin, in the diet. Lesions could be followed from pericentral hyperplasia to areas of hyperplasia, nodules of hyperplasia, small hepatocellular carcinomas, and large well-developed carcinomas, occasionally with metastases. Sometimes pericentral hyperplasia was diffuse throughout most or all of one lobe of the liver. These hyperplastic cells collided to become one large nodule and also one large carcinoma. The carcinomas were well-differentiated or moderately well-differentiated and grew on transplantation to isologous hosts. Histologically, the hyperplastic cells adjacent to central veins were increased in size, frequently with double nuclei. Carcinoma cells varied in size and shape and were huge with large nuclei, prominent nucleoli, and eosinophilic cytoplasm. Similar hepatocellular carcinomas were seen previously with carbon tetrachloride, another organochlorine chemical.     

Carcinomas of the liver in Osborne-Mendel rats ingesting methoxychlor.

Osborne-Mendel female and male rats ingested 0, 10, 25, 100, 200, 500, or 2000 ppm technical methoxychlor for periods up to two years. Male and female rats developed significant incidences of hepatocellular carcinomas of the liver. The carcinomas varied from well differentiated to undifferentiated. Carcinomas of the ovary were also significantly increased in treated female rats.     

Metabolism of organochlorine pesticide heptachlor and its metabolite heptachlor epoxide by white rot fungi, belonging to genus Phlebia

White rot fungi of the genus Phlebia have demonstrated a high capacity to degrade organic pollutants, including polychlorinated dibenzo-p-dioxins and polychlorinated biphenyls. In this study, we evaluated the ability of 18 white rot fungi species of genus Phlebia to degrade heptachlor and heptachlor epoxide, and described the metabolic pathways by selected white rot fungi. Phlebia tremellosa, Phlebia brevispora and Phlebia acanthocystis removed about 71%, 74% and 90% of heptachlor, respectively, after 14 days of incubation. A large amount of heptachlor epoxide and a small amount of 1-hydroxychlordene and 1-hydroxy-2,3-epoxychlordene were detected as metabolic products of heptachlor from most fungal cultures. The screening of heptachlor epoxide-degrading fungi revealed that several fungi are capable of degrading heptachlor epoxide, which is a recalcitrant metabolite of heptachlor. Phlebia acanthocystis, P. brevispora, Phlebia lindtneri and Phlebia aurea removed about 16%, 16%, 22% and 25% of heptachlor epoxide, respectively, after 14 days of incubation. Heptachlor diol and 1-hydroxy-2,3-epoxychlordene were produced in these fungal cultures as metabolites, suggesting that the hydrolysis and hydroxylation reaction occur in the epoxide ring and in position 1 of heptachlor epoxide, respectively.     

UDP-glucuronosyltransferase, epoxide hydrolase and glutathione S-transferase activities in the liver of diabetic mice

The activities of UDPglucuronosyltransferase, microsomal epoxide hydrolase and cytosolic glutathione S-transferase were measured in the liver of spontaneously (db/db and ob/ob) or streptozotocin-induced diabetic mice. An important (2-3-fold) increase of most phase II activities was observed in streptozotocin-treated animals, whereas slighter changes were detected in spontaneously diabetic animals. The latter also exhibited physico-chemical modifications of the liver microsomal membranes, as shown by the temperature-induced variations of epoxide hydrolase activity.     

Expression of human liver epoxide hydrolase in Saccharomyces pombe.

Human liver microsomal epoxide hydrolase cDNA was inserted into the yeast expression vector pEVP11. The resulting recombinant plasmid was introduced into Saccharomyces pombe. The epoxide hydrolase protein and enzymic activity was subsequently expressed and identified in the 105,000 g pellet after centrifugal fractionation of homogenized yeast cells. This method will provide a useful source of human liver epoxide hydrolase, avoiding the problems of obtaining human tissue.     

Cytosolic epoxide hydrolase from liver of control and clofibrate-treated mice. Structural comparison by HPLC peptide mapping

Cytosolic epoxide hydrolases purified from livers of control and clofibrate-induced male C57B1/6 mice were compared. The proteins were reduced, alkylated and cleaved with trypsin and chymotrypsin. The digests were analyzed by HPLC and no qualitative differences were observed in the peptide mapping profiles of the two types of epoxide hydrolase preparation. The amino acid compositions and N-terminal residues of selected tryptic peptides also gave identical results for the control and clofibrate-induced mice. Both intact proteins have e-amino-blocked N-termini. The two enzyme forms are concluded to have highly similar, if not identical, primary structures.Abbreviations HPLC high-performance liquid chromatography - DABITC dimethylaminoazobenzene isothiocyanate     

Histopathology of the pituitary gland in neonatal little (lit) mutant mice.

The pituitary gland in the little (lit) mutant mouse was analyzed with respect to the cytoarchitecture of the pars distalis and the volumetric density of immunoreactive growth hormone (GH) cell granules in neonatal lit/lit and normal C57BL mice. At 8 days postnatally the volume of GH granules/total tissue was significantly less in the lit/lit pars distalis, and the cells were loosely arranged, as compared with the normal pars distalis. In newborn mice a statistically significant difference could not be detected between normal and lit/lit mice with respect to the volumetric density of GH granules; however, differences occurred in the cytoarchitectural organization of the pars distalis. These differences included prominent vascular channels and well-defined cords and clusters of cells in the normal newborn mice, in contrast to indistinct vascular elements and a more diffuse arrangement of cells in lit/lit.     

Immunohistochemical localization of epoxide hydratase in rat liver

An antibody produced against epoxide hydratase (EC 4.2.1.63) which had been purified to apparent homogeneity from hepatic microsomes of phenobarbital pretreated rats was employed in an unlabeled antibody peroxidase-antiperoxidase technique to localize the enzyme at the light microscopic level in the livers of untreated rats. Immunohistochemical staining for epoxide hydratase was detected in parenchymal cells throughout the liver lobule. Cells within the centrilobular regions, however, were observed to be stained more intensely than were those within the midzonal and periportal regions of the lobule. The results of this immunohistochemical study thus demonstrate that epoxide hydratase does not exhibit a uniform pattern of distribution within the liver lobule in untreated rats.     

The regulation of cytosolic epoxide hydrolase in mice

The concentration of cytosolic epoxide hydrolase in untreated and clofibrate-treated mouse liver extracts was estimated by immunoblotting. Clofibrate treatment of mice was found to increase liver cytosolic epoxide hydrolase concentration by two fold, showing that the increase in cytosolic epoxide hydrolase in mouse liver after clofibrate treatment is primarily due to induction. The induced and uninduced cytosolic epoxide hydrolase, and epoxide hydrolase in the cytosolic and mitochondrial fractions were compared and found to be identical or very similar. Cytosolic epoxide hydrolases in kidney and liver were similar in molecular weight and antigenic properties.     

Properties of cytosolic epoxide hydrolase purified from the liver of untreated and clofibrate-treated mice. Purification procedure and physiochemical characterization of the pure enzymes

Cytosolic epoxide hydrolase was purified from the liver of untreated and clofibrate-treated male C57Bl/6 mice. The purification procedure involves chromatography on DEAE-cellulose, phenyl-Sepharose and hydroxyapatite, takes two days to perform and results in a 120-fold purification and approximately 35% yield of the enzyme from untreated mice. The purified enzyme is a dimer with a molecular mass of 120 kDa, a Stokes' radius of 4.2 nm, a frictional ratio of 1.0 and an isoelectric point of 5.5. The subunits behave identically upon isoelectric focusing in 8 M urea and only one band with a molecular mass of 60 kDa is seen after sodium dodecyl sulfate/polyacrylamide gel electrophoresis. The form purified from clofibrate-treated mice had very similar properties and was apparently identical to the control form as judged by amino acid analysis and peptide mapping as well. These analyses also demonstrated that the cytosolic enzyme is clearly different from microsomal epoxide hydrolase isolated from rat liver. Furthermore, Ouchterlony immunodiffusion using antibodies raised in rabbits towards the control form of cytosolic epoxide hydrolase revealed identity between the two forms of cytosolic epoxide hydrolase, but no reaction with the microsomal epoxide hydrolase was observed. These findings indicate large structural differences between the cytosolic and microsomal forms of epoxide hydrolase in the liver.