Human exposure to polycyclic aromatic hydrocarbons (PAH) occurs through complex mixtures such as coal tar. The effect of complex PAH mixtures on the activation of carcinogenic PAH to DNA-binding derivatives and carcinogenesis were investigated in mice treated topically with NIST (National Institute of Standards and Technology) Standard Reference Material 1597 (SRM), a complex mixture of PAH extracted from coal tar, and either additional benzo[a]pyrene (B[a]P) or dibenzo[a,l]pyrene (DB[a,l]P). In an initiation-promotion study using 12-O-tetradecanoylphorbol-13-acetate as the promoter for 25 weeks, the SRM and B[a]P co-treated mice had a similar incidence of papillomas per mouse compared with the group exposed to B[a]P alone as the initiator. PAH-DNA adduct analysis of epidermal DNA by 33P-post-labeling and reversed-phase high-performance liquid chromatography found the SRM co-treatment led to a significant decrease in the total level of DNA adducts and B[a]P-DNA adducts to less than that observed in mice treated with B[a]P alone at 6, 12 and 72 h exposure. After 24 and 48 h exposure, there was no significant difference in the levels of adducts between these groups. In the DB[a,l]P initiation-promotion study, the co-treated group had significantly fewer papillomas per mouse than mice treated with DB[a,l]P alone as initiator. Averaging over the times of exposure gave strong evidence that mice co-treated with SRM and DB[a,l]P had a significantly lower level of PAH-DNA adducts than mice treated with DB[a,l]P alone. Western immunoblots showed that both cytochrome P450 (CYP) 1A1 and 1B1 were induced by the SRM. These results are consistent with the hypothesis that two major factors determining the carcinogenic activity of PAH within a complex mixture are (i) the persistence of certain PAH-DNA adducts as well as total adduct levels, and (ii) the ability of the components present in the mixture to inhibit the activation of carcinogenic PAH by the induced CYP enzymes.

Campylobacter jejuni was inoculated intravenously into pregnant ewes on gestation days 1 14 and 123 to reproduce ovine abortion. All ewes aborted 7-1 2 days post-inoculation. High numbers of C. jejuni were isolated from ewe tissues (caruncle, bile, cecal feces), fetal tissues, and placenta. C. jejuni colonies were identified in caruncles and placenta by light microscopy and immunoperoxidase techniques. Histologically, inoculated ewes had a severe purulent endometritis with vasculitis. Placentas from inoculated ewes and field cases showed necrosis and purulent inflammation; however, placentas from inoculated ewes had large numbers of bacterial colonies compared to few bacteria found in field cases. Histologically, only one fetus from the inoculated ewes showed lesions (purulent bronchopneumonia), whereas all fetuses from field cases had a distinct bronchopneumonia, and one fetus showed multifocal hepatic necrosis. These results suggest that C. jejuni (serotypes Penner 1 and Lior 2) is an important abortifacient organism for sheep. Campylobacteriosis is a highly contagious and economically significant disease in sheep and is most often caused by Campylobacter fetus subspecies fetus (previously named C. fetus var. intestinalis).15.32 It is characterized by abortion (third trimester), stillbirths, premature births, weak lambs, and occasional ewe deaths due to m e t r i t i~. '~,~~ Grossly, aborted or stillborn fetuses may have no lesions or they may show subcutaneous serosanguineous edema, serosanguineous body fluids, discrete necrotic liver lesions, or bronchopneumonia. Placental cotyledons are enlarged, yellowish, and covered with blood-tinged exudate, while the intercotyledonary stroma is edematous. Microscopically, placental lesions, which are predominant in hilar zones, consist of septa1 necrosis, arteriolitis, leukocyte infiltration, and accumulation of high numbers of bacteria within lacunae, chorionic trophoblasts, and endothelial cells. In humans, Campylobacter jejuni is recognized to be a common cause of acute dia~-rhea,~J~ and is associated with abortion and neonatal sepsis.33 Campylobacter organisms are small, curved, highly motile, noncapsulated, microaerophilic, gram-nega

The metazoan parasitic blood f lukes, Schistosoma spp., infect over 200 million people worldwide and cause extensive human morbidity and mortality. Research strategies for development of anti-schistosomal agents are impeded by the organism's complex molluscan-mammalian life cycle, which limits experimental approaches and availability of material. We derived long-term continuously proliferative cultures of Schistosoma mansoni sporocysts capable of generating cercariae in vitro. Cultured organisms retained the ability to parasitize the host, and they exhibited developmental regulation of candidate stage-specific genes in the host-free culture system. Evidence for expression of a reverse transcriptase also was found in the cultured organisms, pointing to this activity as a possible mechanistic contributor to the dynamic relationship between the parasite and its hosts. Continuous in vitro propagation of the asexual sporocyst stage allows isolation of clonally derived parasite populations and provides a means to study schistosomal molecular genetics, metabolism, and evasion of host defenses.Schistosoma mansoni, Schistosoma haematobium, and Schistosoma japonicum create widespread disease in tropical developing countries (1). These parasites thwart attempts to produce practical and effective vaccines, and pharmacological approaches are problematic (2). Schistosome eggs are laid by females paired with males and residing in the vasculature of parasitized vertebrate hosts. To be successfully infective, eggs must penetrate the intestinal wall and pass in feces to fresh water. From these released eggs hatch free-swimming miracidia, the first larval form. Successful miracidia encounter and penetrate the appropriate species of snail and develop into sporocysts, the second larval from. In the following weeks, these primary sporocysts asexually generate first-generation and second-generation daughter sporocysts in the snail. From the later-generation sporocysts develop the third larval form, free-swimming cercaria. To be successfully infective in the human or other vertebrate host, cercaria must encounter and penetrate the skin of the host and undergo further changes to form schistosomula. These move through several further developmental stages, ultimately leading to sexual pairing in the veins of the parasitized host and egg production.Approaches to elucidate the mechanisms schistosomes employ to escape host immunity, as well as molecular genetic and pharmacological investigations, have been limited by the lack of an in vitro system for the continuous proliferative culture of the parasite in the absence of the host. Recently, Yoshino and colleagues (3, 4) have reported significant success in limited culture of the intramolluscan stages of S. mansoni and S.japonicum, but these techniques do not allow extended culture of proliferating sporocysts.Here we describe long-term cultures of S. mansoni in vitro and the development from the snail-infective miracidial stage, through generations of mother and daughter sporocysts, t...

␥-Glutamyltranspeptidase (GGT)-deficient mice (GGT ؊/؊ ) display chronic glutathione (GSH) deficiency, growth retardation, and die at a young age (<20 weeks). Using livers from these mice, we investigated the relationship between GSH content, especially mitochondrial, and mitochondrial and cellular function. We found that the GSH content of isolated liver mitochondria was diminished by >50% in GGT ؊/؊ mice when compared with wild-type mice. Respiratory control ratios (RCRs) of GGT ؊/؊ mice liver mitochondria were <60% those of wild-type mice primarily as a result of impaired state 3 respiration. Mitochondrial adenine nucleotide content was decreased by >40% in mitochondria obtained from GGT ؊/؊ mice. We observed a strong correlation between mitochondrial GSH content and RCRs. Even moderate decreases (<50%) correlated with adverse effects with respect to respiration. Electron microscopy revealed that livers from GGT ؊/؊ knockout mice were deprived of fat and glycogen, and swollen mitochondria were observed in animals that were severely deprived of GSH. Thus, GGT ؊/؊ mice exhibit a loss of GSH homeostasis and impaired oxidative phosphorylation, which may be related to the rate of adenosine triphosphate (ATP) formation and subsequently leads to progressive liver injury, which characterizes the diseased state. We also found that supplementation of GGT ؊/؊ mice with N-acetylcysteine (NAC) partially restored liver GSH, but fully restored mitochondrial GSH and respiratory function. Electron microscopy revealed that the livers of NAC-supplemented GGT ؊/؊ mice contained fat and glycogen; however, slightly enlarged mitochondria were found in some livers. NAC supplementation did not have any beneficial effect on the parameters examined in wildtype mice. (HEPATOLOGY 2000;32:740-749.)The liver is the main site of interorgan glutathione (GSH) synthesis, and also the major organ for biosynthesis of macromolecules, plasma proteins, and detoxification of toxic metabolites. GSH is known to function as an antioxidant; a physiological reservoir for cysteine, it is involved in DNA synthesis, regulation of protein synthesis, detoxification, synthesis of leukotrienes, etc. 1 The liver provides Ͼ85% of the plasma cysteine, which is essential for GSH synthesis in nonhepatic cells. This is achieved through the action of ␥-glutamyltranspeptidase (GGT) and dipeptidases in the kidney. 2 Liver GSH can be decreased as a result of inhibition of synthesis, increased demand, increased detoxification, or inhibited turnover by GGT. A decrease in liver GSH has also been found in association with a variety of diseases and age. 3 Many studies indicated that cellular GSH deficiency can lead to cellular injury and even cell death, but few attempts have been made to investigate metabolic changes preceding these 2 events. Cellular GSH deficiency affects the mitochondrial GSH pool as well as the cytosolic GSH pool. It was first shown by Meredith and Reed 4 that the onset of chemically induced cellular injury correlated with the depletion of the mitochondr...

Abstract. Serum-free mouse embryo (SFME) cells, derived in medium in which serum is replaced with growth factors and other supplements, are proastroblasts that are acutely dependent on epidermal growth factor (EGF) for survival. Ultrastructurally, an early change found in SFME cells deprived of EGF was a loss of polysomes which sedimentation analysis confirmed to be a shift from polysomes to monosomes. The ribosomal shift was not accompanied by decreased steady-state level of cytoplasmic actin mRNA examined as an indicator of cellular mRNA level. With time the cells became small and severely degenerate and exhibited nuclear morphology characteristic of apoptosis. Genomic DNA isolated from cultures undergoing EGF deprivation-dependent cell death exhibited a pattern of fragmentation resulting from endonuclease activation characteristic of cells undergoing apoptosis or programmed cell death. Flow cytometric analysis indicated that cultures in the absence of EGF contained almost exclusively Gl-phase cells. Some of the phenomena associated with EGF deprivation of SFME cells are similar to those observed upon NGF deprivation of nerve cells in culture, suggesting that these neuroectodermal-derived cell types share common mechanisms of proliferative control involving peptide growth factor-dependent survival.

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Helminth parasites are reported for the first time from northern spotted owls. Seventyone percent of a sample of Strix occidentalis caurina from western Oregon was infected. Nematodes (Porrocaecum depressum, Capillaria falconis, Microtetrameres sp. and Synhimantus hamatus) were the most prevalent parasites although cestodes (Paruterina rauschi) and acanthocephalans (Cent rorhynchus conspectus) were also represented. There was an association between components of this helminth fauna and the diet of spotted owls which is dominated by small rodents. The occurrence of P. rauschi rather than P. candelabraria in this geographic region and host-species may provide additional support for recognition of a parapatric distribution in the ranges of Paruterina spp. among strigiforms in the Nearctic.

A 31-kg mediastinal mass from a 24-year-old horse is described. Cytologic imprints contained numerous spindle cells with indistinct cytoplasm and multiple foci of palisading nuclei. Histologic sections had these same characteristics and areas of central fibrillar material between palisading nuclei (Verocay bodies). Within histologic sections, spindle cell cytoplasm had positive immunoreactivity for vimentin and S-100, whereas immunoreactivity was not detected for desmin and cytokeratin. Cytologic, histologic, and immunochemical evaluation confirmed a diagnosis of solitary schwannoma.