bluuebunny/crossref_metadata_2025 huggingface dataset:
Partial Hepatectomy and Bile Duct Ligation in Rainbow Trout (Oncorhynchus mykiss): Histologic, Immunohistochemical and Enzyme Histochemical Characterization of Hepatic Regeneration and Biliary Hyperplasia
Authors: Okihiro Mark S. , Hinton David E.
Published: February 2008

Hepatic regeneration following partial hepatectomy (PH) and biliary hyperplasia subsequent to bile duct ligation (BDL) were characterized in rainbow trout (Oncorhynchus mykiss) by light microscopy using routine and special (immunohistochemical and enzyme histochemical) stains. Both PH and BDL involved initial hypertrophy and hyperplasia of bile preductular epithelial cells (BPDECs). BPDECs are small oval cells that form junctional complexes with hepatocytes and bile ductular cells and are commonly found in hepatic tubules of teleost liver. Proliferating BPDECs transitioned through intermediate cell types before final differentiation into large basophilic hepatocytes (following PH) or biliary epithelial cells (after BDL). Normal BPDECs and hepatocytes were both negative for cytokeratin intermediate filaments in control fish when screened with the monoclonal antibody AE1/AE3. In contrast, hyperplastic BPDECs and their progeny (intermediate cells, immature hepatocytes, ductal epithelial cells) were all strongly cytokeratin positive. Cytokeratin expression was transient in newly differentiated hepatocytes (expression decreased as hepatocytes acquired characteristics consistent with full differentiation) but was permanent in biliary epithelial cells (expression was very strong in large mature ducts). BPDECs, intermediate cells, and immature ductal cells were also strongly positive for alkaline phosphatase following BDL. Chronology of histologic events and cytokeratin and enzyme expression all support the hypothesis that BPDECs possess the capacity to differentiate into either hepatocytes or biliary epithelial cells. Thus, BPDECs may be the teleost equivalent of a bipolar hepatic stem cell in mammals.
Thresholds for Genotoxic Carcinogens
Authors: O'Connor Peter J. , Manning Francis C.R. , Gordon Anthony T. , Billett Michael A. , Cooper Donald P. , Elder Rhoderick H. , Margison Geoffrey P.
Published: February 2008

DNA damage is a critical factor in the initiation of chemically induced toxicities (including cancer), and the repair of this damage represents the cell's first line of defense against the deleterious effects of these agents. The various mechanisms of DNA repair are reviewed briefly and the actions of the DNA repair protein 06-alkylguanine-DNA alkyltransferase (ATase) are used to illustrate how DNA repair can protect cells against alkylating agent-induced toxicities, mutagenesis, clastogenesis, and carcinogenesis. The effectiveness of this repair protein can be measured based on its ability to deplete levels of its promutagenic substrate O6-methylguanine ( O6-meG) in the DNA of cells. These studies reveal that the repair of O6-meG from DNA occurs heterogeneously, both intra- and intercellularly. Even in cells that repair O6-meG hyperefficiently, certain regions of chromatin DNA are repaired with difficulty, and in other regions they are not repaired at all; most likely this lack of repair is a result of the location of the lesion in the DNA sequence. When individual cells are compared within a tissue, some cells are clearly repair deficient, because the O6-meG can persist in DNA for many weeks, whereas in other cells, it is removed within a matter of hours. The role of these repair-deficient cells as targets for alkylating agent-induced carcinogenesis is considered. The mechanisms of the homeostatic control of DNA repair function in mammalian cells are not yet well understood. Because there are now indications of the mechanisms by which the level of DNA damage may be sensed (and so influence the activity of the ATase repair protein), this is an important area for future study.
Industry Viewpoint on Thresholds for Genotoxic Carcinogens
Authors: Morelli Michael A.
Published: February 2008

Modem chemical control of pests has contributed to a dramatic improvement in public welfare since its introduction 50 years ago. Millions of lives have been saved through the control of disease vectors, and millions more have been improved by the use of chemicals to produce an inexpensive and abundant food supply. Hundreds of pesticidally active ingredients are in commercial use today, and among these are found genotoxic and nongenotoxic carcinogens. In the United States, the Environmental Protection Agency regulates carcinogens using threshold and nonthreshold approaches depending upon the outcome of a weight-of-evidence determination. More than one-half of all pesticides with some evidence of carcinogenic potential are regulated by the nonthreshold approach. The limitations on product use imposed by this approach have restricted the number of products available to growers and to the public. This restriction has had a direct impact on industry with respect to commercial success and financial returns on investment as well as an indirect impact on the industry's ability to fund the discovery and development of new compounds. This paper explores the question of how well regulation by the nonthreshold approach has achieved the goal of protecting public health, whether it does this better than the alternative use of the threshold approach, and whether the incremental protection it affords is a meaningful public benefit that justifies the aforementioned impacts on industry.
DNA Adducts: Endogenous and Induced
Authors: Povey Andrew C.
Published: February 2008

Human exposure to DNA damaging agents can arise from exogenous sources or endogenous processes that occur normally or in pathological states. DNA isolated from human tissues, obtained from the very young to the old, contains detectable amounts of a number of different types of DNA adducts that reflect exposure to both known carcinogens and as yet unidentified genotoxic agents. The levels of DNA damage observed in human studies as a result of exogenous exposures (noniatrogenic) is of the order of 1 adduct per 107-109 normal DNA bases, whereas that arising from endogenous exposures may potentially be several orders of magnitude higher. Large interindividual variations in DNA adduct levels have been reported, and these are probably the result of host and environmental factors, although variation in analytical and sampling procedures may also play a role. It is important to recognize that the presence of DNA adducts in a tissue does not necessarily indicate a specific tumorigenic risk for that tissue, as other factors downstream of DNA adduct formation (including DNA repair and cell proliferation) play an important role in determining overall risk.
Validation of In Vitro and In Vivo Methods for Assessing Endocrine Disrupting Chemicals
Authors: Ashby John
Published: February 2008

The concepts that require validation in terms of the subject of endocrine disruption are listed and discussed. The main mechanisms by which endocrine disruption can occur are identified, and the assays required for the detection of adverse endocrine disruption toxicities associated with these mechanisms are discussed. The process of assay validation is considered. The validation of structure-activity relationships, the need for reference chemicals, and the problems recently encountered when attempting to reproduce endocrine disruption data are also explored. The most important conclusions derived from this analysis are that given the immature state of research into endocrine disruption toxicity, testing strategies and the types of assay employed should be kept under constant review; inevitably researchers need to accept the fact that future revision of each assay will be required. Second, given the current absence of any chemical that is universally accepted to be devoid of endocrine toxicity, assay specificity will be difficult to assess, and that imposes the need for alternative objective criteria for assessing the value of individual assays.
Cell Death and Cell Proliferation in the Control of Normal and Neoplastic Tissue Growth
Authors: Foster John R.
Published: February 2008

The development of reliable methodology for the assessment of rates of cell replication and cell death has enabled the study of how these 2 fundamentally opposed processes work to form and maintain tissue and to remodel tissue following diseases resulting in cell loss. The balance between these 2 processes and the consequences of an imbalance are fundamental to a clearer understanding of how hyperplasia and neoplasia develop in tissues under the influence of chemicals and drugs. An understanding of the changes that occur in target organs and tissues following chemical or drug exposure has enabled a better understanding of the mechanism by which these chemicals are able to induce cancer after prolonged exposure. Studies of the control of cell replication and the changes that occur following drug exposure have defined 2 types of response, 1 in which the cell replicative response is sustained and the other in which the cell replicative response is transient and occurs during the first few days of exposure. Although regulatory and scientific opinion appears ready to accept sustained cell replicative processes as an increased risk factor in the development of cancer, the role played by transient increases in cell replication remains unclear. Concurrent events in target organs following treatment with chemicals that induce transient increases in cell replication have revealed that the rates of apoptosis are suppressed at the same time as the cell replication levels are induced. Additional evidence suggests that growth and antigrowth factors are central in controlling these responses. Escape from the regulatory action of these factors is postulated to be one of the ways in which nongenotoxic carcinogenic chemicals, such as the peroxisome proliferators and sodium phenobarbitone, may induce cancer, with apoptosis playing a key role in the process.
Histopathologic Approaches to Detect Changes Indicative of Immunotoxicity
Authors: Kuper C. Frieke , Harleman Johannes H. , Richter-Reichelm Hans B. , Vos Joseph G.
Published: February 2008

Toxicologic pathology is crucial in the identification and characterization of health effects following exposure to xenobiotics, mainly in toxicity experiments in rodents. Regarding regulatory toxicology, histopathology of lymphoid organs and tissues is a cornerstone in the identification of immunotoxic compounds. A 2-tier testing system is usually employed in which the first tier is a general screen for (immuno)toxicity and the second tier consists of specific immune function studies, including host resistance tests or mechanistic studies. The role attributed to histopathology of lymphoid organs in the updated Organisation for Economic Cooperation and Development and Food and Drug Administration guidelines requires improvement and standardization of the histopathology procedures. Optimalization and standardization was started in an international collaborative immunotoxicity study (ICICIS). However, several problems were left unaddressed, mostly because of the few compounds tested in this study. Based on the results of the ICICIS study and the morphologic changes induced by immunotoxic/immunomodulatory compounds observed in other investigations, suggestions are given to further improve the identification and (semi)quantification of histopathologic changes in lymphoid organs and tissues.
Responses of the Immune System to Injury
Authors: Descotes Jacques , Choquet-Kastylevsky Genevieve , Van Ganse Eric , Vial Thierry
Published: February 2008

Three categories of immunotoxic effects are identified: direct immunotoxicity, hypersensitivity, and autoimmunity. Direct immunotoxicity consists of immunosuppression and immunostimulation. Total abrogation of the immune response (immunosuppression) results in more frequent, severe, and often atypical and relapsing infections and lymphomas. Immunostimulation is associated with febrile reactions, the induction/facilitation of autoimmune diseases and allergic reactions to unrelated allergens, and impaired hepatic drug biotransformation. Hypersensitivity is manifested by a variety of symptoms involving either antigen-specific or non-antigen-specific humoral and cellular adverse responses. Autoimmune reactions are divided into organ-specific and systemic reactions. Because of the involvement of many redundant mechanisms, it is difficult to predict responses of the immune system to a given immunotoxic injury. In laboratory animals, histologic but also functional changes are necessary to show evidence of and to predict such adverse responses.
Review Article: Use of Transgenic Animals for Carcinogenicity Testing: Considerations and Implications for Risk Assessment
Authors: Gulezian Donna , Jacobson-Kram David , Mccullough C. Bruce , Olson Harry , Recio Leslie , Robinson Denise , Storer Richard , Tennant Raymond , Ward Jerrold M. , Neumann David A.
Published: February 2008

Advances in genetic engineering have created opportunities for improved understanding of the molecular basis of carcinogenesis. Through selective introduction, activation, and inactivation of specific genes, investigators can produce mice of unique genotypes and phenotypes that afford insights into the events and mechanisms responsible for tumor formation. It has been suggested that such animals might be used for routine testing of chemicals to determine their carcinogenic potential because the animals may be mechanistically relevant for understanding and predicting the human response to exposure to the chemical being tested. Before transgenic and knockout mice can be used as an adjunct or alternative to the conventional 2-year rodent bioassay, information related to the animal line to be used, study design, and data analysis and interpretation must be carefully considered. Here, we identify and review such information relative to Tg.AC and rasH2 transgenic mice and p53+/- and XPA -/- knockout mice, all of which have been proposed for use in chemical carcinogenicity testing. In addition, the implications of findings of tumors in transgenic and knockout animals when exposed to chemicals is discussed in the context of human health risk assessment.
The Nongenotoxic Hepatocarcinogens Diethylhexylphthalate and Methylclofenapate Induce DNA Synthesis Preferentially in Octoploid Rat Hepatocytes
Authors: Hasmall Susan C. , Roberts Ruth A.
Published: February 2008

Diethylhexylphthalate (DEHP), a rodent carcinogen, and 1,4-dichlorobenzene (DCB), a noncarcinogen in rat liver, are potent hepatomitogens. We have reported previously that 7-day dosing with DEHP induced a higher bromodeoxyuridine labeling index (LI) in binuclear octoploid (2×4N) rat hepatocytes than did DCB, suggesting that induction of DNA synthesis in 2×4N hepatocytes might represent a more substantial carcinogenic risk. We compared 2 additional rodent hepatocarcinogens, methylclofenapate (MCP) and phenobarbitone, with ethylene thiourea (ETU), a noncarcinogenic hepatomitogen in rat. All 3 chemicals increased hepatic LI; the 8N population had the highest LI, but only the carcinogens increased LI in the 2×4N and 4N populations. To identify the target population for induction of DNA synthesis, we used a 1-hour pulse label at the peak of induction. The results were consistent with the 7-day data, and again the highest LI was in the 8N population. The nongenotoxic rodent carcinogens MCP and DEHP induced a significant increase in the LI in the 2×4N population, whereas ETU and DCB did not. These data support the hypothesis that increased DNA synthesis within the minority 2×4N population may be more significant for subsequent hepatocarcinogenesis.
Morphological Changes in the Kidney of Dogs Chronically Exposed to Exogenous Growth Hormone
Authors: Molon-Noblot Sylvain , Laroque Philippe , Prahalada Srinivasa , Stabinski Lea G. , Peter Chennekatu P. , Duprat Pierre , Van Zwieten Matthew J.
Published: February 2008

Porcine growth hormone was administered subcutaneously to beagle dogs at doses of 0.025,0.1, and 1 IU/kg/d for 14 weeks, markedly elevating serum growth hormone (GH) and insulin-like growth factor-1 (IGF-1) levels. This was accompanied by a significant increase in body weight gain and kidney weights in both male and female dogs. The increase in kidney weight (6 to 54%) was slightly greater than the increase in body weight (6 to 40%). By light microscopy, glomerular deposits, mesangial thickening, and very slight cellular infiltration in glomeruli were seen in mid- and high-dose groups. Based on morphometric evaluation, there was an increase in the renal glomerular area, which was statistically significant ( p ≤ 0.05) in the mid- and high-dose males and in the high-dose females. This was associated with a statistically significant ( p ≤ 0.05) increase in the number of total glomerular cells in the mid- and high-dose males. By transmission electron microscopy, thickening of the glomerular basal lamina and diffuse increase of the mesangial matrix were observed in both male and female dogs in the mid- and high-dose groups. Immunohistochemical reactions were negative for IgG, IgM, and C3. The morphological changes in the kidney of dogs resemble the diffuse glomerulosclerosis described in human diabetic nephropathy.
Overexpression of Grb2 in Inflammatory Lesions and Preneoplastic Foci and Tumors Induced by N-Nitrosodimethylamine in Helicobacter hepaticus—Infected and -Noninfected A/J Mice
Authors: Diwan Bhalchandra A. , Ramakrishna Gayatri , Anderson Lucy M. , Ramljak Danica
Published: February 2008

Growth factors bind to membrane receptor tyrosine kinases, resulting in autophosphorylation and subsequent binding to proteins with SH2 domains, including growth factor receptor-bound protein 2 (Grb2). Grb2 bridges receptors to tyrosine kinase substrates such as SHC and SOS, which in turn facilitate the activation of downstream signaling pathways, including Ras and mitogen-activated protein kinase (MAPK). Overexpression of Grb2 has been demonstrated in several types of neoplasia but has not been investigated in liver tumorigenesis. Here we investigated Grb2 expression in liver lesions in N-nitrosodimethylamine (NDMA)-treated Helicobacter hepaticus-infected and -noninfected A/J mice at 1 year of age. Previously, we reported (6) that infection promotes the development of these NDMA-initiated tumors. In controls, Grb2 immunostaining was absent from normal hepatic tissues, whereas the inflammatory lesions in infected livers were positive for cytoplasmic Grb2 in both hepatocytes and infiltrating leukocytes. All preneoplastic foci (7 of 7), 15 of 27 adenomas, and 3 of 7 carcinomas were positive for Grb2 by immunostaining in both infected and noninfected NDMA-initiated livers. Involvement of Grb2 was confirmed by immunoblotting of similarly infected mice at 9 to 18 months of age, showing a 2.5- to 3.3-fold increase in Grb2 protein in infected livers (p < 0.05 compared with uninfected controls) as well as in preneoplastic foci, adenomas, and carcinomas. These livers also showed a 2.5- to 2.8-fold increase in total Ras protein. The results suggest that upregulation of Grb2 is an early event in liver carcinogenesis, whether caused by the bacterial infection or by NDMA. Concomitant upregulation of Ras p21 would ensure transmission of amplified signal from growth factors via Grb2.
Morphogenesis of Craniopharyngeal Derivatives in the Neurohypophysis of Fisher 344 Rats: Abnormally Developed Epithelial Tissues Including Parotid Glands Derived from the Stomatodeum
Authors: Iwata Hijiri , Hosoi Masayo , Miyajima Rumiko , Yamamoto Shinji , Mikami Shinichi , Yamakawa Seiki , Enomoto Makoto , Imazawa Takayoshi , Mitsumori Kunitoshi
Published: February 2008

Morphogenesis of craniopharyngeal derivatives of the neurohypophysis found in 14 Fischer 344 (F344) rats was studied. The incidence of the craniopharyngeal derivatives was 0.17% in male (7 out of 4,200) and 0.16% in female (7 out of 4,450) F344 rats. Neither a sex-related difference in their incidence nor a strain-related difference in their morphological features was observed. Craniopharyngeal derivatives were composed of aberrant epithelial structures consisting of serous acinar and tubular and fusiform cell structures, and most of these derivatives were associated with Rathke's cleft cysts, which are suggestive of a congenital background. The acinar structures were positive for periodic acid-Schiff reaction and negative for Alcian blue stain. Immunohistochemically, cells forming these structures were positive for cytokeratin, and basal cells of the acinar or tubular structures and some of the fusiform cells showed positive staining for alpha-smooth muscle actin. Electron microscopically, these spindle-shaped basal cells had intracytoplasmic myofilaments with focal density in their cytoplasm, and they were regarded to be myoepithelial cells. These findings strongly indicate that the craniopharyngeal derivatives are not a neoplastic lesion but rather are a developmental aberration derived from the stomatodeum, which is known to be the origin of both nasal and oral epithelial tissues, including the parotid glands, other than Rathke's pouch.
Possible Involvement of bcl-2 Suppression in Wild-Type p53 Gene-Dependent Cell Growth Repression in Rat Osteosarcoma Cells
Authors: Honoki Kanya , Tsujiuchi Toshifumi , Tsutsumi Masahiro , Kido Akira , Morishita Toru , Yoshimoto Masatoshi , Miyauchi Yoshizumi , Mii Yoshio , Tamai Susumu , Konishi Yoichi
Published: February 2008

We recently obtained 3 cloned cell lines demonstrating the p53 mutation from a lung metastatic nodule of a rat transplantable osteosarcoma. In this study, we applied wild-type p53 gene transfer to the rat osteosarcoma cells by lipofection to investigate the effects on cell growth, expression of genes such as waf1/p21, bcl-2, and bax, and nucleosomal DNA fragmentation due to apoptosis. Reconstitution of the p53 gene inhibits cellular growth, and this growth-suppressive effect is partly due to apoptosis involving bcl-2 gene suppression in this tumor type. This rat osteosarcoma model is similar in biologic behavior to human cases and thus is very suitable for further investigation of tumorigenesis and gene therapy for osteosarcoma.
The Prophylactic Effects of Natural Water-Soluble Antioxidant from Spinach and Apocynin in a Rabbit Model of Lipopolysaccharide-Induced Endotoxemia
Authors: Lomnitski Liat , Carbonatto Michela , Ben-Shaul Varda , Peano Sergio , Conz Angelo , Corradin Lorena , Maronpot Robert R. , Grossman Shlomo , Nyska Abraham
Published: February 2008

Radical-scavenging antioxidants, as part of the cellular defense system, function to inhibit the formation and propagation of free radicals and active oxygen species formation. In previous studies we demonstrated that endotoxin lipopolysaccharide (LPS) promotes oxidative stress and associated pathological changes in a rat model and that use of selected antioxidants was effective in reducing LPS-related lipid peroxidation product formation in the liver, as well as LPS-related pathological changes in different organs. In this study, several toxicological parameters (ie, clinical signs, blood chemistry, and histopathological changes) were compared among groups of male New Zealand rabbits injected with LPS following prophylactic pretreatment with either of 2 antioxidants, a group injected with LPS without pretreatment with antioxidants, groups injected with either of the 2 antioxidants only, and an untreated control group. The antioxidants used were a water-soluble natural antioxidant (NAO) from spinach and the NADPH oxidase inhibitor, apocynin. Exposure to LPS alone was associated clinically with depression, tachypnea, outer ear vasodilation, and iris congestion; biochemically with a significant increase in blood total bilirubin, transaminase activity, and glucose, total cholesterol, and triglyceride levels; macroscopically with multiple whitish areas in the liver; and histologically with hepatocellular focal necrosis and acute inflammation, thymic and splenic lymphoid necrosis and depletion, acute uveitis and hemorrhages in the ciliary processes, and decreased adrenal cortical cytoplasmic vacuolation considered consistent with depletion of steroidal hormone contents. The NAO had more effective prophylactic capacities than the apocynin. The protective effects were obvious in all investigated parameters. The results indicate the possible therapeutic efficacy of NAO in the treatment of clinical endotoxemia associated with gram-negative bacterial sepsis that is known to be associated with oxidative stress.
Carcinogenicity of Chloral Hydrate Administered in Drinking Water to the Male F344/N Rat and Male B6C3F1 Mouse
Authors: George Michael H. , Moore Tanya , Kilburn Steve , Olson Greg R. , Deangelo Anthony B.
Published: February 2008

Male B6C3F1 mice and male F344/N rats were exposed to chloral hydrate (chloral) in the drinking water for 2 years. Rats: Measured chloral hydrate drinking water concentrations for the study were 0.12 g/L, 0.58 g/L, and 2.51 g/L chloral hydrate that yielded time-weighted mean daily doses (MDDs) of 7.4, 37.4, and 162.6 mg/kg per day. Water consumptions, survival, body weights, and organ weights were not altered in any of the chloral hydrate treatments. Life-time exposures to chloral hydrate failed to increase the prevalence (percentage of animals with a tumor) or the multiplicity (tumors/animal) of hepatocellular neoplasia. Chloral hydrate did not increase the prevalence of neoplasia at any other organ site. Mice: Measured chloral hydrate drinking water concentrations for the study were 0.12 g/L, 0.58 g/L, and 1.28 g/L that gave MDDs of 13.5, 65.0, and 146.6 mg/kg per day. Water consumptions, survival, body and organ weights, were not altered from the control values by any of the chloral hydrate treatments. Enhanced neoplasia was observed only in the liver. Prevalence and multiplicity of hepatocellular carcinoma (HC) were increased only for the high-dose group (84.4%; 0.72 HC/animal; p ≤ 0.05). Values of 54.3%; 0.72 HC/animal and 59%; 1.03 HC/animal were observed for the 13.5- and 65.0-mg/ kg per day treatment groups. Prevalence and multiplicity for the control group were 54.8%; 0.74 HC/animal. Hepatoadenoma (HA) prevalence and multiplicity were significantly increased ( p ≤ 0.05) at all chloral hydrate concentrations: 43.5%; 0.65 HA/animal, 51.3%; 0.95 HA/animal and 50%; 0.72 HA/animal at 13.5, 65.0, and 146.6 mg/kg per day chloral hydrate compared to 21.4%; 0.21 HA/animal in the untreated group. Altered foci of cells were evident in all doses tested in the mouse, but no significant differences were observed over the control values. Hepatocellular necrosis was minimal and did not exceed that seen in untreated rats and mice. Chloral hydrate exposure did not alter serum chemistry and hepatocyte proliferation in rats and mice or increase hepatic palmitoyl CoA oxidase in mice at any of the time periods monitored. It was concluded that chloral hydrate was carcinogenic (hepatocellular neoplasia) in the male mouse, but not in the rat, following a lifetime exposure in the drinking water. Based upon the increased HA and combined tumors at all chloral hydrate doses tested, a no observed adverse effect level was not determined.
Accumulation of Iron in the Rat Lung after Tracheal Instillation of Diesel Particles
Authors: Ghio Andrew J. , Richards Judy H. , Carter Jacqueline D. , Madden Michael C.
Published: February 2008

Oxidant generation catalyzed by metals has been postulated to account for a lung injury following exposure to air pollution particles. In particles that are predominantly carbonaceous, it is difficult to implicate such an oxidative stress as the responsible mechanism, since concentrations of metals can be extremely low. Comparable to these air pollution particles, mineral oxide particles can include only minute amounts of metal, but lung injury following their exposure can be associated with an accumulation of endogenous iron from the host and an oxidative stress. We tested the hypothesis that diesel exhaust particulate (DEP) effects an accumulation of biologically active iron in the rat lung, with both oxidative stress and a lung injury resulting. Characterization of the DEP confirmed a high concentration of carbon, whereas metals were low in quantity. The concentration of total lavage iron in animals instilled with saline was low, but this concentration increased with exposure to DEP. Non-heme iron in lung tissue was similarly elevated after instillation of the diesel product. Particle instillation was associated with a decrease in lavage ascorbate concentration supporting an oxidative stress. Relative to saline exposure, DEP resulted in elevated lavage concentrations of the inflammatory mediators macrophage inflammatory protein-2 and tumor necrosis factor. Finally, an injury after particle instillation was evident with increased neutrophils and an elevation of lavage protein and lactic dehydrogenase. We conclude that DEP exposure effected an accumulation of iron in the rat lung. This accrual of iron was associated with an oxidative stress, release of oxidant-sensitive mediators, and a neutrophilic lung injury.
Cecal Torsion in Rodents Associated with Chronic Administration of Clinafloxacin
Authors: Courtney Cynthia L.
Published: February 2008

The chronic toxicity of clinafloxacin, a fluoroquinolone antibacterial agent, was evaluated in multiple strains of mice and rats. In 5 separate studies, mice and rats that were orally administered up to 1,000 mg/kg of clinafloxacin from 9 to 104 weeks developed dose-related cecal dilatation and deaths that were attributable to cecal torsion. Cecal rupture was observed in association with torsion. Although cecal dilatation is commonly observed in rodents given antibacterials such as fluoroquinolones, cecal torsion has not been a reported sequelae to dilatation.
Short-Term Pulmonary Response to Inhaled JP-8 Jet Fuel Aerosol in Mice
Authors: Robledo Raymond F. , Young R. Scott , Lantz R. Clark , Witten Mark L.
Published: February 2008

B6.A.D. ( Ahrd/Nat s) mice were utilized to investigate the short-term pulmonary response to JP-8 jet fuel (JP-8) aerosol inhalation. Mice were nose-only exposed to atmospheres of 0 to 118 mg/m3 for 1 h/d over a period of 7 days to further test the hypothesis that JP-8 concentrations below the permissible exposure level (PEL) of 350 mg/m3 will induce lung injury. At 24 to 30 hours after the final exposure, pulmonary function and respiratory permeability were measured on anesthetized mice and then randomly assigned for bronchoalveolar lavage or histopathology. Bronchoalveolar lavage fluid (BALF) was analyzed for total protein, lactic dehydrogenase (LDH), N-acetyl-β-D-glucosaminidase (NAG), and cytology. Respiratory permeability increases were observed following doses of 48 and 118 mg/m3 and were supported by concomitant BALF increases in total protein and LDH. Conversely, NAG and alveolar macrophage levels decreased following the same exposure concentrations. Morphological lung injury was characterized by the targeting of bronchiolar epithelium and consisted of perivascular edema, Clara cell vacuolization, and necrosis. Alveolar injury included sporadic pulmonary edema, intra-alveolar hemorrhage, and alterations in type II epithelial cells. These results indicate that repeated inhalation of aerosolized JP-8 induces physiological, biochemical, cellular, and morphological lung injury. This study also provides evidence for the reevaluation of the 350 mg/m3 PEL for more volatile petroleum distillates with regard to respirable aerosols.
Immunohistochemical Localization of Transforming Growth Factor Alpha in Chemically Induced Rat Hepatocellular Carcinomas with Reference to Differentiation and Proliferation
Authors: Kobayashi Kiyoshi , Kusakabe Manami , Okada Miyoko , Sakairi Tetsuya , Goto Kazuhiro , Tsuchiya Takayuki , Sugimoto Jiro , Sano Fumiko , Mutai Mamoru
Published: February 2008

Hepatocellular carcinomas (HCCs) were induced in male Fischer 344 rats with dietary 3'-methyl-4-(dimethylamino)-azobenzene treatment and were classified into solid, glandular (well- or poorly differentiated), and trabecular types. Investigation of cell proliferation kinetics and immunohistochemical localization of transforming growth factor alpha (TGF-alpha) demonstrated all solid (n = 24) and poorly differentiated glandular type (n = 6) HCCs to have TGF-alpha-positive nuclei. Nuclear staining of TGF-alpha was also observed in 13 of 28 (46%) trabecular-type HCCs, whereas 12 (43%) exhibited cytoplasmic staining, and 3 (11%) were negative. As for well-differentiated glandular HCCs, 7 of 20 (35%) were positively stained in their nucleus, another 7 (35%) demonstrated antibody binding in the cytoplasm, and 6 (30%) were negative. The order for growth rate evaluated by bromodeoxyuridine (BrdU) labeling was solid (38.22%), poorly differentiated glandular (26.82%), trabecular (7.98%), and well-differentiated glandular (2.57%) types. For trabecular HCCs with nuclear, cytoplasmic, or negative TGF reactions, values were 13.39% (n = 13), 3.61% (n = 12), and 2.01% (n = 3), respectively. Likewise, BrdU-labeling indices for the counterpart groups of well-differentiated glandular type HCCs were 4.53, 1.91, and 1.29%, respectively. The results indicate that TGF-alpha expression might be linked to histopathological differentiation and cell proliferation in rat HCCs.
Quantitative Characterization of Acid- and Alkali-Induced Corneal Injury in the Low-Volume Eye Test
Authors: Jester James V. , Molai AL , Petroll W. Matthew , Parker Ron D. , Carr Gregory J. , Cavanagh H. Dwight , Maurer James K.
Published: February 2008

Defining the extent of initial injury has proven to be a useful basis for differentiating the ocular irritation potential of surfactants; however, the applicability of this method to other types of irritants has not been demonstrated. In the following studies we characterized the extent of corneal injury following exposure to different concentrations of acetic acid and sodium hydroxide (NaOH) in the rabbit low-volume eye test. Groups of rabbits received 3% acetic acid, 10% acetic acid, 2% NaOH, or 8% NaOH and were evaluated in vivo by macroscopic and in vivo confocal microscopic examination and postmortem using a live/dead staining kit and scanning laser confocal microscopic examination. Quantitative assessment of macroscopic scores, corneal surface epithelial cell size, corneal epithelial thickness, corneal thickness, depth of stromal injury, corneal light scattering (confocal microscopy through focusing, CMTF), and number of dead cells was conducted at various times, including the following: at 3 hours and at 1, 3, 7, 14, and 35 days. Based on macroscopic scores, the order of ocular irritancy potential was 3% acetic acid < 2% NaOH < 10% acetic acid < 8% NaOH. Evaluation of the quantitative in vivo and postmortem microscopic live/dead data revealed a slight decrease in epithelial thickness and an increase in dead epithelial cell numbers with 3% acetic acid. With 2% NaOH, significant focal changes in epithelial cell size, epithelial thickness, corneal thickness, and number of dead surface epithelial cells occurred at 3 hours and at 1 day, with injury to only a very small number of corneal stromal keratocytes, despite the presence of epithelial denudation. Changes with 10% acetic acid were similar to those noted with 2% NaOH at 3 hours and 1 day, but these changes were more diffuse and included stromal injury to a depth of 7.2 ± 9.3% of the corneal thickness, with significant numbers of dead keratocytes. Eight percent NaOH, on the other hand, caused focally extensive injury that averaged 26.3 ± 18.4% of the corneal thickness at 1 day, with significant light scattering from the cornea, which did not return to normal by 35 days postinjury. Overall, these data indicate that ocular irritation as a result of acetic acid and NaOH was associated with changes similar to those observed with surfactants (ie, slight irritants damage the corneal epithelium, mild and moderate irritants damage the corneal epithelium and anterior stromal cells, and severe irritants damage the corneal epithelium and deep stroma). To our knowledge, this is the first time that the ocular irritation potential for different types of materials (acid/alkali, surfactants) has been shown to be primarily dependent on the initial area and depth of injury.
Microscopic Changes with Acetic Acid and Sodium Hydroxide in the Rabbit Low-Volume Eye Test
Authors: Maurer James K. , Parker Ronald D.
Published: February 2008

Differences in ocular irritancy have been hypothesized to reflect differences in the extent of initial injury. Although differences in the processes leading to tissue damage may exist, extent of injury is believed to be the principal factor determining final outcome of ocular irritation. Previous studies characterizing the pathology of surfactant-induced ocular irritation support this premise. The purpose of this study was to begin to determine the applicability of this premise in terms of nonsurfactants; we planned to accomplish this by assessing the ocular irritancy of different concentrations of an acid and an alkali. Ten microliters of 3 or 10% acetic acid (C2H4O2) or 2 or 8% sodium hydroxide (NaOH) were directly applied to the cornea of the right eye of each test rabbit. Untreated left eyes served as the controls. Eyes and eyelids were macroscopically examined for signs of irritation beginning 3 hours after dosing and periodically until recovery or day 35. Eyes and eyelids from animals in each group were collected for microscopic examination after 3 hours and on days 1, 3, and 35. The macroscopic and microscopic changes were consistent with slight (3% C2H4O2), mild (2% NaOH, 10% C2H4O2), and severe (8% NaOH) irritancy. The spectra of changes were similar to those previously reported for surfactants of differing types and irritancies. As with surfactants, as the extent of initial injury increased, the intensity and duration of the subsequent responses increased. These results indicate that our hypothesis also applies to nonsurfactants. The results also support our belief that the initial extent of injury associated with ocular irritation may be used to predict the subsequent responses and final outcome. Finally, our results further indicate that such an approach may be applicable to the development of alternative assays that are based on either injury to ex vivo eyes or injury to an in vitro corneal equivalent system.
Dandy-Walker Like Malformation in a Fischer-344 Rat
Authors: Kuwamura Mitsuru , Morikawa Takeshi , Yasui Hisae , Yamate Jyoji , Kotani Takao
Published: February 2008

A spontaneous cerebellar malformation was found in a 32-day-old male Fischer 344 rat. The cerebellar malformation was composed of a vermis defect and markedly dilated fourth ventricle. The cerebellar hemispheres were separated, with the left hemisphere being smaller than the right one. Degenerative/inflammatory lesions consisting of macrophage/lymphocyte infiltration, spheroidal calcium depositions, and astrocytic gliosis were seen adjacent to the cerebral aqueduct. Abnormally arranged hyperplastic ependymal cells were observed beneath the fourth ventricle in the medulla oblongata. The gross findings of the present case resemble those of the human Dandy-Walker malformation. While a precise mechanism remains to be elucidated, degenerative/inflammatory lesions in the present rat may be involved in the pathogenesis of this malformation.
Neoplasia in Zebrafish (Danio rerio) Treated with 7,12-Diniethylbenz[a]anthracene by Two Exposure Routes at Different Developmental Stages
Authors: Spitsbergen Jan M. , Tsai Hsi-Wen , Reddy Ashok , Miller Tom , Arbogast Dan , Hendricks Jerry D. , Bailey G.S.
Published: February 2008

Using zebrafish, Danio rerio, initial pioneering work in the 1960s revealed carcinogen responsiveness of fish, yet very few subsequent tumorigenesis investigations have utilized this species. We exposed embryos (60 hours postfertilization) and fry (3 week posthatch) to 7,12-dimethylbenz[a]anthracene (DMBA) by immersion in aqueous solutions for 24 hours, at concentrations of 0-1 or 0-5 ppm (mg/L), respectively. Juvenile zebrafish 2 months posthatch were fed a diet containing 0-1,000 ppm DMBA for 4 months. Fish were sampled for histologic evaluation at 7-12 months after the onset of carcinogen treatment. Fry were most responsive to DMBA and showed the widest diversity of target tissues and histologic types of neoplasia, having several types of epithelial, mesenchymal, and neural neoplasia. The principal target tissues for carcinogenic response were liver following embryo or fry exposure, with gill and blood vessel the second and third most responsive tissues in fry. Intestine was the primary target and gill a secondary target in fish that received dietary DMBA as juveniles. These studies indicate that young zebrafish are most responsive to DMBA, showing a greater diversity of neoplasm types than rainbow trout. Thus, zebrafish are a valuable model system in which to study mechanistic aspects of the carcinogenesis process.
Fulminant Hepatocyte Apoptosis In Vivo Following Microcystin-LR Administration to Rats
Authors: Hooser Stephen B.
Published: February 2008

Microcystin-LR (MCLR) is a cyanobacterial toxin responsible for human and livestock deaths worldwide. MCLR has also been implicated as a contributing factor in hepatocellular carcinoma. Following absorption, MCLR is taken up via a hepatocyte-specific bile acid carrier. Inside hepatocytes, MCLR selectively binds to protein phosphatases 1 and 2A, resulting in rapid, massive liver damage. However, the apoptotic nature of this toxicosis in rats has not been fully characterized as such at appropriate time points utilizing light and electron microscopy, terminal deoxynucleotidyl transferase-mediated dUTP-biotin nick end labeling (TUNEL), and electrophoresis of hepatic DNA. Rats were administered intraperitoneal saline or MCLR at 500 μg/kg (0.5 μmol/kg) and necropsied at 3 or 9 hours. Light microscopy at 3 hours revealed massive, widespread apoptotic necrosis of the majority of hepatocytes. Hepatocytes were rounded and disassociated, with cell shrinkage, increased eosinophilia, and margination of nuclear chromatin or pyknosis. The apoptotic index increased from 0.03% ± 0.02% in controls to 205% ± 12% in MCLR-treated animals (p ≤ 0.0001). At 3 hours, transmission electron microscopy revealed hepatocellular changes typical of apoptotic necrosis: rounding and disassociation of hepatocytes, loss of microvilli, and margination and condensation of nuclear chromatin. Laddering of hepatic DNA by electrophoresis and widespread TUNEL staining of hepatocytes were consistent with apoptosis. These results demonstrate that in rats, hepatic damage caused by MCLR is due to extremely rapid induction and progression of apoptosis in virtually every hepatocyte in the liver. This model of fulminant hepatic necrosis should be useful for increased characterization and understanding of the relationship between protein phosphatase inhibition and apoptosis.
Induction of Lung Lesions in Female Rats Following Chronic Exposure to 2,3,7,8-Tetrachlorodibenzo-p-dioxin
Authors: Tritscher Angelika M. , Mahler Joel , Portier Christopher J. , Lucier George W. , Walker Nigel J.
Published: February 2008

2,3,7,8,-Tetrachlorodibenzo- p-dioxin (TCDD) has been classified as a known human carcinogen, and epidemiologic studies identify the lung as one of the target organs. Few experimental studies have attempted to characterize pulmonary effects of TCDD exposure. In this study, we characterize the induction of lesions in the lung by chronic oral TCDD exposure in diethylnitrosamine (DEN)-initiated or noninitiated female Sprague-Dawley rats. Two or 18 weeks after initiation, rats were treated with TCDD continuously for 14, 30, or 60 weeks by biweekly oral gavage (1,750 ng TCDD/kg) at a dose equivalent to 125 ng/kg body weight per day (controls received corn oil). To assess the time dependence and reversibility of potential changes, some groups included withdrawal periods of 16 or 30 weeks after 30 weeks of TCDD treatment. TCDD treatment alone for 60 weeks caused significant increases in alveolar-bronchiolar (AB) metaplasia. TCDD treatment of DEN-initiated animals for 60 weeks resulted in a significant increase in bronchiolar epithelial hyperplasia. These increases were not observed in animals treated with TCDD for 30 weeks followed by corn oil for 30 weeks, indicating that the development of these lesions required continuous exposure to TCDD. AB hyperplasia increased in an age-dependent manner after DEN initiation but was unaffected by TCDD treatment. Expression of the aromatic hydrocarbon receptor (AHR) and induction of CYP1A1 was observed only in bronchiolar Clara and ciliated cells, indicating that the mechanism of induction of AB metaplasia may be mediated by the AHR. TCDD elimination half-life was monophasic in the lung, and serum and was estimated to be 39.7 days and 44.6 days, respectively, independent of age, tissue TCDD concentration, or body weight. This is the first report to identify the AB region as a target for TCDD-induced metaplastic and proliferative changes after chronic oral exposure.
Evolution of Alachlor-Induced Nasal Neoplasms in the Long-Evans Rat
Authors: Genter Mary Beth , Burman Dawn M. , Dingeldein Michael W. , Clough Ian , Bolon Brad
Published: February 2008

The chloracetanilide herbicide alachlor (2-chloro-2',6-diethyl- N-(methoxymethyl)-acetanilide) induces nasal neoplasms in rats following chronic dietary exposure. The present study sought to identify the cellular origin and mechanisms of tumor induction and progression. Male Long-Evans rats were fed alachlor (0 or 126 mg/kg/day) beginning at 6 weeks of age. Following 1 month of alachlor ingestion, neither histological abnormalities nor enhanced cell division (assessed by BrdU incorporation) occurred in any region of the nasal cavity. Six months of alachlor exposure resulted in proliferation of basal and nonbasal cells in the olfactory mucosa while inducing nasal masses in 7 of 15 animals. Tumors ranged from dysplastic plaques to polypoid adenomas and originated in the olfactory regions of the nasal cavity. Neoplasms were associated with regions of respiratory metaplasia and were often covered with a low cuboidal, poorly ciliated epithelium. Tumor cells did not express characteristics of the olfactory mucosa, including olfactory marker protein (OMP, for neurons) and NMa (antibody recognizing cytochrome P450 [CYP] 2A3, found in Bowman's glands). Sites of plaque and tumor development coincided with regions of NMa immunoreactivity. These data suggest that local metabolism is important in alachlor-induced olfactory tumors and support the concept that metaplastic respiratory epithelial cells give rise to the observed neoplasms.
Spontaneous Testicular Lesions in Purpose-Bred Beagle Dogs
Authors: Rehm S.
Published: February 2008

Spontaneous testicular lesions were assessed in 50 control purpose-bred male beagle dogs. They were selected from 13 toxicology studies conducted over the period 1988-1999. Age of the dogs at study termination varied from 8-20 months with an average age of 13 months. Regardless of age, the most significant finding was bilateral segmental hypospermatogenesis in 15/50 (30%) of the dogs. Cross sections of tubules with hypospermatogenesis were distributed randomly throughout the testes and were characterized by reduced proportions of germ cells, tubular shrinkage, and Sertoli cell prominence. These changes were occasionally associated with giant cells, with cellular debris, and in 6/15 (40%) with atrophic tubules devoid of germ cells, indicating a degenerative process. Focal subcapsular tubular atrophy or hypoplasia (tubules lined by Sertoli cells only) was also found in 9/35 (26%) of dogs without hypospermatogenesis. Inhibited spermiation with retention of mature sperm in tubules was seen in 6/50 dogs, 3 of which also showed hypospermatogenesis. Other findings of high incidence but low prevalence included tubules with multinucleated giant cells, swollen spermatocytes, or apoptotic germ cells. These latter changes are probably a constituent of normal spermatogenesis. In conclusion, about 30% of control beagle dogs show segmental hypospermatogenesis, which may be associated with degenerative changes, and an additional 18% of the dogs exhibit focal tubular atrophy/hypoplasia in otherwise normal testes. These changes have to be distinguished from compound-related toxic effects.