Role of cytoprotectants and nitric oxide inhibition in nonsteroidal anti-inflammatory drug-induced gastroduodenal injury in the rat.

BACKGROUND AND PURPOSE: Nonsteroidal anti-inflammatory drugs (NSAIDs) induce gastroduodenal injury and ulceration. The pathogenesis is uncertain, although reductions in cytoprotective prostaglandins and nitric oxide (NO) have been proposed. The effects of several cytoprotective agents on inhibition of gastroduodenal ulcerogenesis induced by CI-987, a novel NSAID, were evaluated in Wistar rats. METHODS: Male Wistar rats were given CI-987 orally (p.o.) at a dosage of 300 or 450 mg/kg of body weight or subcutaneously (s.c.) (3 x 50 mg/kg), alone or with misoprostol pretreatment (2 x 1 mg/kg, p.o.). In a second experiment, rats were pre-treated with 2 ml of gelusil p.o., 500 mg of sucralfate/kg, p.o., 100 mg of ranitidine/kg s.c., or 200 mg of N omega-nitro-L-arginine methyl ester (L-NAME)/kg, s.c.. Duodenal injury was induced by administration of 450 mg of CI-987/kg, p.o., 3 x 50 mg of CI-987/kg, s.c., or 300 mg of cysteamine/kg, s.c. Animals were euthanized within 24 to 48 hours, and the gastrointestinal tract was examined for evidence of gross or microscopic change. RESULTS: The L-NAME significantly reduced the incidence and severity of gastroduodenal injury induced by CI-987 and cysteamine. Prostaglandin ameliorated duodenal lesions induced by CI-987 given s.c., and Gelusil, ranitidine, and sucralfate were without effect on duodenal lesions induced by NSAID. CONCLUSIONS: Preemptive blockade of NO synthase is important in preventing NSAID-induced duodenal injury in rats. Inhibition of cytoprotective prostaglandins and enhanced acid-induced damage are unlikely to be primary mechanisms underlying NSAID-induced duodenal injury in rats.

Reference values for young normal Sprague-Dawley rats: weight gain, hematology and clinical chemistry.

1. Arterial blood from 63 male (315-500 g) and 60 female (210-290 g) healthy Sprague-Dawley rats was analyzed for 16 hematological and 22 clinical chemistry parameters. 2. Nine of these parameters were associated with growth and developmental changes in males between 38-78 days and females 49 - 89 days of age. 3. Weight gain in both sexes followed patterns consistent with other studies using this strain, but interexperimental variation was as much as +/-22%. 4. Rectal temperatures of females averaged 37.28 degrees C, being statistically greater (P < 0.05) than the average male at 36.99 degrees C. 5. The data provide reference values for use in toxicological and other investigations.

Biochemical effects of Pembina Cardium crude oil exposure in cattle.

Crude oil pollution at drilling sites located within or in close proximity to agricultural pasture lands poses serious health risks to cattle raised on these lands. To investigate the clinical and systemic biochemical effects, cattle (8/group) were administered single oral doses of Pembina Cardium crude oil (PCCO) at 16.7, 33.4, and 67.4 g/kg, or water (control group) at 80 g/kg. Cattle exposed to PCCO showed dose-dependent clinical effects. At the lowest dosage, PCCO caused transient and minimal clinical effects; however, high dosages caused varied clinical signs which included tremors, nystagmus, vomiting, and pulmonary distress. On posttreatment day 7 or 30, four cattle from each treatment group were sacrificed and biochemical parameters were assayed in liver, lungs, and kidney cortex. In cattle monitored on posttreatment day 7, the PCCO-treated groups showed marked alterations from the control group in hepatic cytochrome P-450 (P-450), and in aryl hydrocarbon hydroxylase (AHH) and 7-ethoxycoumarin-O-deethylase (ECOD) activities of these tissues. Administration of PCCO caused significant increases (> 100%) in hepatic P-450, but produced variable effects on AHH and ECOD activities in each tissue. The activity of AHH was increased in all tissues; however, the effect was highest in kidney cortex (> 5000%), followed by liver (> 500%) and lungs (> 250%). The activity of ECOD was altered in a differential manner. It was either increased markedly (>1300%) in kidney cortex or increased slightly (20-30%) in liver, but decreased (> 80%) in lungs. The activities of respiratory chain enzymes (succinate-cytochrome c reductase, NADH-cytochrome c reductase and cytochrome oxidase), or NADPH-cytochrome c reductase and glutathione transferase were not changed significantly in any tissues. The alterations in P-450, AHH, and ECOD observed on day 7 were markedly reversed in cattle examined on day 30 posttreatment, indicating a recovery from induced changes. Studies in vitro with hepatic microsomal preparations from day 7 posttreatment groups showed that increases in AHH and ECOD activity in PCCO-treated cattle were due to induction of new isoforms of P-450, as evidenced by (1) the appearance of a 448-nm spectral peak, and (2) differential inhibitory effects of metyrapone and 7,8-benzoflavone on AHH and ECOD activities.

Cytotoxic effects of hydrogen sulfide on pulmonary alveolar macrophages in rats.

Respiratory rates (basal and zymosan-stimulated) and cell viability were monitored in pulmonary alveolar macrophages (PAM) from rats exposed to 0, 70, 280, and 560 mg/m3 (0, 50, 200, and 400 ppm) hydrogen sulfide (H2S) gas for 4 h. Zymosan-stimulated respiratory rates were markedly reduced in PAM collected from rats exposed to 280 and 560 mg/m3 H2S; however, their basal respiratory rates were not affected. Significant decrease in cell viability was also observed in samples from 560 mg/m3 H2S-treated rats, but it remained high and unchanged in other treatments. In vitro incubation of PAM from control rats with sulfide (a precursor of H2S) and its two oxidation products, sulfite and sulfate, showed that sulfide was markedly more inhibitory to both respiratory rates than sulfite or sulfate. These treatments did not affect cell viability.

Effects of hydrogen sulfide exposure on lung mitochondrial respiratory chain enzymes in rats.

Fischer-344 rats were exposed for 4 hr to various concentrations of hydrogen sulfide (H2S) gas and killed either immediately or at 1, 24, or 48 hr after exposure. Mitochondrial fractions from lung tissues were assayed for the activities of respiratory chain enzymes. Exposure of rats to a low concentration (10 ppm) of H2S caused no significant changes in the activities of lung mitochondrial enzymes. However, exposure to sublethal concentrations of H2S (50-400 ppm) produced marked and highly significant depressions in the activities of cytochrome c oxidase and succinate oxidase complexes of the respiratory chain. The inhibition of cytochrome c oxidase activity in lungs was most severe (greater than 90%) in rats that died from acute exposure to greater than 500 ppm H2S. In rats exposed to 200 and 400 ppm H2S, a marked recovery in cytochrome c oxidase activity of lungs was observed at 24 and 48 hr postexposure. Studies in vitro with rat lung mitochondria showed that low concentrations of sulfide also caused a similar and selective inhibition of cytochrome c oxidase activity. This effect was reversed upon removal of sulfide either by washing or by oxidation with methemoglobin. The nature of sulfide inhibition of cytochrome c oxidase was noncompetitive with respect to ferrocytochrome c. Because the activities of NADH-cytochrome c reductase and succinate-cytochrome c reductase were not significantly altered by H2S exposure and in vitro treatments with low concentrations of sulfide, it is concluded that under physiological conditions H2S would block the respiratory chain primarily by inhibiting cytochrome c oxidase. Such a biochemical impairment would lead to functional (histotoxic) hypoxia in the lung tissues.

Effects of dichlorvos on blood cholinesterase activities of cattle.

Inhibitory effects of dichlorvos (2,2-dichlorovinyl dimethyl phosphate, DDVP) [corrected] on erythrocyte acetylcholinesterase (AChE) and plasma cholinesterase (ChE) activities of steers were characterized after treatments in vitro and in vivo (cutaneous application). The rates of in vitro inhibition were markedly influenced by DDVP concentration and incubation time. The activities of inhibited enzymes failed to reactivate spontaneously and had little response to treatment with 2-pyridine aldoxime methiodide (2-PAM). After gel-filtration chromatography, however, the inhibited enzymes had remarkable spontaneous reactivation and reactivation by 2-PAM treatment, indicating interference of excess unreacted DDVP in the reactivation process. Repeated cutaneous applications of a DDVP-containing livestock spray caused marked and characteristic decreases of AChE and ChE activities in blood of treated steers; however, the effects were transient because activities of both enzymes regenerated gradually. The nature of these in vivo trends suggests that spontaneous and de novo synthetic mechanisms could be responsible for complete recovery of both enzyme activities.

Induction of hepatic cytochrome P-450 and xenobiotic metabolizing enzymes in rats gavaged with an Alberta crude oil.

Changes in body weight gain and in biochemical parameters of blood and liver were assessed in Sprague-Dawley rats after multiple oral administration of three test doses of an Alberta crude oil (ACO). Rats treated with ACO (1.25-5 ml/kg) did not show statistically significant (p greater than .05) differences from control, corn-oil treated (5 ml/kg) rats, in body weight gains, liver weight, and blood biochemical indicators of liver (alanine aminotransferase, gamma glutamyltransferase), kidney (blood urea nitrogen, creatinine), and erythrocyte (adenosine 5'-triphosphate, 2,3-diphosphoglyceric acid, reduced glutathione) cytotoxicity. Treatment with ACO, however, caused statistically significant (p less than .05) and dose-related increases from control in (1) microsomal protein and cytochrome P-450 content, and NADPH-cytochrome c reductase, aryl hydrocarbon hydroxylase (AHH), and 7-ethoxycoumarin-O-deethylase (7-ECOD) activities, and (2) cytosolic glutathione transferase activity of liver. The induction of hepatic cytochrome P-450 and xenobiotic-metabolizing enzymes in microsomes of ACO-treated rats was probably associated with dose-related changes in isozymic forms of cytochrome P-450, as evidenced by (1) appearance of a 448-nm spectral peak in microsomes of ACO-treated rats and (2) differences in the inhibition pattern of AHH and 7-ECOD activities in microsomes of control and ACO-treated rats upon treatment with metyrapone and 7,8-benzoflavone.

The toxicology of detergents, bleaches, antiseptics and disinfectants in small animals.

Small animals are exposed to a variety of household products. The most common route of exposure is oral by direct ingestion or from grooming soiled fur and feet. The majority of household products are complex chemical mixtures. The clinician must determine the body burden of toxic or potentially toxic substances and the potential for a combination of chemicals to produce intoxication in a particular species and then plan a course of action. Toxicoses of selected household and commercial products are discussed in detail. Animals are exposed to a variety of antiseptics by numerous routes. Potential for iatrogenic intoxication from antiseptics is also discussed. An abridged version of this article will be published in the 10th edition of CURRENT VETERINARY THERAPY: SMALL ANIMAL PRACTICE, RW Kirk, editor, WB Saunders Co, Philadelphia, PA.

Histologic and ultrastructural alterations in lungs of rats exposed to sub-lethal concentrations of hydrogen sulfide.

Fischer-344 rats were killed 1, 18, and 42 hr after a single 4-hr exposure to an atmosphere of 0, 116, or 615 mg m-3 of hydrogen sulfide (H2S). Lungs, fixed by the intratracheal route, were examined by light and electron microscopy. Histologic changes were transient and mainly present in rats exposed to 615 mg m-3 H2S. Lesions included severe but transitory pulmonary edema and fibrinocellular alveolitis which was restricted to the proximal alveolar region of the lung. Electron microscopically, ciliated bronchiolar cells were the only cells that developed necrosis; they were rapidly replaced by mitosis. Alveolar endothelium had cytoplasmic blebs, but alveolar epithelium had minor changes. No mast cell degranulation was detected in lungs with edema. A 4-hr exposure to 615 mg m-3 is markedly edematogenic for the lung but only moderately cytotoxic for pulmonary cells.

In vitro and in vivo effects of dichlorvos on blood cholinesterase activities of cattle.

Studies were conducted to ascertain in vitro effects and effects of percutaneous application (in vivo) of dichlorvos (2,2-dichlorovinyl dimethyl phosphate; DDVP) on cholinesterase activities in bovine erythrocytes and plasma. Treatment in vitro of erythrocytes and plasma with DDVP resulted in concentration- and time-dependent inhibition of erythrocyte acetylcholinesterase (AChE) and plasma cholinesterase (ChE) activities. Mean (+/- SD) DDVP concentrations required to cause 50% enzyme inhibition were 15.7 +/- 3.3 microM and 43.1 +/- 5.7 microM for AChE and ChE, respectively; however, these values required to achieve this inhibition were markedly decreased with increasing incubation time. The inhibited AChE activity failed to be reactivated after incubation of erythrocytes with 2-pyridine aldoxime methiodide (2-PAM); however, limited reactivation of inhibited AChE and ChE activities was observed with excess concentration of 2-PAM. Percutaneous application of a DDVP-containing livestock spray on cattle also caused a marked decrease in the in vivo activities of AChE and ChE; however, the inhibited enzyme activities were reactivated rapidly after incubation with 2-PAM.

Biochemical and cytologic alterations in the respiratory tract of rats exposed for 4 hours to hydrogen sulfide.

Fischer-344 rats were killed by exsanguination 1, 20, and 44 hr after a single 4-hr exposure to an atmosphere of 0, 10, 200, and 400 ppm of hydrogen sulfide (H2S). Alterations in the activities of lactate dehydrogenase and alkaline phosphatase, and cytomorphology of epithelial cells in fluids obtained by nasal and bronchoalveolar lavage were used as indicators of cell injury. Changes in the number of leukocytes were used as indicators of inflammatory response, and changes in the concentration of protein were used as indicators of altered vascular permeability. Inhalation of H2S resulted in 139, 483, and 817% increased cellularity in the nasal cavity of rats exposed to 10, 200, and 400 ppm, respectively. This was due to marked exfoliation of degenerated epithelial cells and exudation of neutrophils. The high dose of H2S resulted in a moderate increase in lactate dehydrogenase and protein in nasal passages; values returned to baseline levels 20 hr later. Bronchoalveolar cell counts were decreased in rats exposed to 400 ppm and unchanged in those exposed to 10 and 200 ppm. Enzymatic activities in lung lavage fluid were moderately elevated (up to 90%), yet protein concentrations were increased by more than 3000% and remained significantly elevated up to 44 hr after exposure to 400 ppm. It was concluded that inhalation of H2S has a severe cytotoxic effect on the nasal epithelium and a severe edematogenic effect on lung parenchyma. These results are in agreement with autopsy findings of individuals killed by accidental exposure to H2S-containing sour gas.

Profiles of type-II pneumocytes in rats inoculated intratracheally with bacterial lipopolysaccharide.

Ultrastructural and morphometric profiles of type-II pneumocytes (P-II) were investigated in rats killed 18 or 24 hours after a single intratracheal inoculation of bacterial (Escherichia coli) lipopolysaccharide (LPS). Inoculation with LPS induced pulmonary injury and inflammation, as measured by increased lactate dehydrogenase and alkaline phosphatase activities and increased numbers of polymorphonuclear neutrophils in fluid collected by bronchoalveolar lavage. Marked ultrastructural changes and desquamation of a few P-II developed at the time of high activity of lactate dehydrogenase and alkaline phosphatase in bronchoalveolar lavage fluid. Ultrastructural changes included swollen mitochondria and localized cisternal dilatation of the endoplasmic reticulum in which was contained membrane-bound homogenous material of medium electron density. Twenty-four hours after LPS inoculation, point-count stereologic analysis and digitizing morphometry revealed greater than 50% increase in P-II size. Changes in cell size corresponded with ultrastructural finding of swollen cells. Results obtained by point-count stereologic analysis and digitizing morphometry were highly correlated (r = 0.95). Lamellar bodies (LB) comprised 12 to 15% of P-II volume. Volume density and number of LB remained unaltered in LPS-injured P-II, and evidence of accelerated release of LB was not detected after LPS inoculation. Exudated polymorphonuclear neutrophils and pulmonary alveolar macrophages were involved actively in the phagocytosis of LB originating from necrotic and desquamated P-II. On the basis of measurement of enzyme activity (enzymes released into the bronchoalveolar space), considerable ultrastructural alterations developed in P-II when maximal LPS-induced pulmonary cell injury took place.

Effects of high dietary sulphur on enzyme activities, selenium concentrations and body weights of cattle.

This study was designed to assess the effects of a moderate increase in dietary sulphur (S) in cattle. Twelve animals were initially fed a basal concentrate (S = 0.2%) and then divided into two groups; one fed basal and the other high S (S = 0.75%) concentrates. Health, body weight gains, and activities of erythrocyte enzymes-glutathione peroxidase (GSH-Px), superoxide dismutase (SOD), glucose-6-phosphate dehydrogenase (G6PD), acetylcholinesterase (AChE), plasma- asparate aminotransferase (AST), and whole blood concentrations of selenium (Se) were monitored at various stages of the study. Marked increases in the activities of GSH-Px, SOD and G6PD from the pretrial values were observed upon initial feeding of basal concentrate diet. Sex related differences were not evident in enzyme activities and Se concentrations of the blood. A high linear correlation (r = 0.92) between averages of GSH-Px activity and Se concentration of blood was observed in both sexes. Increasing the amount of S in the concentrate diet (from 0.2 to 0.75%) did not produce any statistically significant change in enzyme activities and Se concentrations, body weight gains, and health of the cattle during the 85 days feeding period. The results indicate that a moderate increase in the dietary S would not impair Se and copper status or cause related disorders in cattle.

Effect of sex and age on the activities of lactate dehydrogenase and alkaline phosphatase in the lungs of rats.

Since toxicity studies among different laboratories generally involve rats of different sex and age, this study was conducted to investigate the effect of sex, age and animal to animal variation in the activities of lactate dehydrogenase and alkaline phosphatase from bronchoalveolar lavage fluid, bronchoalveolar cell lysate and lung homogenate. Correlation between numbers of bronchoalveolar cells recovered from lungs and enzyme activity in bronchoalveolar cell lysate or lung homogenate supernatant were also investigated. Male rats showed significantly (p less than 0.05) higher activities of alkaline phosphatase in the bronchoalveolar lavage fluid and lung homogenate. Animal to animal variation for lactate dehydrogenase and alkaline phosphatase was higher in lungs than in serum. The number of bronchoalveolar cells recovered from lungs revealed a significant (p less than 0.01) positive correlation with the activities of both enzymes in the supernatant of cell lysates but not in the bronchoalveolar fluid. These results indicated that in an inhalation study interindividual variation in the levels of pulmonary enzymes should be considered in order to minimize the numerous possible sources of experimental error.

Adiaspiromycosis in striped skunks in Alberta, Canada.

Pulmonary adiaspiromycosis was diagnosed in seven of 25 striped skunks (Mephitis mephitis) in east-central Alberta. The infection varied from mild, where only microscopic lesions were seen, to severe, where gross lesions of grayish-white nodules were observed in the lung parenchyma. Mild lesions were restricted to the lung, while severe lesions extended to the tracheobronchial and mediastinal lymph nodes. Histologically, the lesions were characterized by a centrally located fungal spherule, surrounded by granulomatous inflammation. The morphology of the fungal spherules was consistent with that of Emmonsia crescens. By electron microscopy, the fungal cells had an outer thick fibrillar wall and an inner cytoplasm filled with large lipid vacuoles with relatively few mitochondria, ribosomes or glycogen inclusions. The absence of endosporulation and budding suggested that each fungal cell in the lung represented a separate inhaled spore. Infection was by inhalation, nevertheless adiaspores may disseminate to the regional lymph nodes.

A mouse model for estimation of Pasteurella haemolytica deposition in calf lungs following aerosol exposure.

A method used to calculate the number of Pasteurella haemolytica reaching the lungs of calves during an aerosol exposure is described. This method is based on a linear relationship of bacterial deposition in lungs of mice and calves when exposed to the same bacterial aerosol.