Vacuolation in renal tubular epithelium of Cd-1 mice: an incidental finding.

Prominent cytoplasmic vacuoles were observed in renal tubular epithelial cells of the outer medulla in several kidneys from test article-dosed mice (Crl:CD-1 (ICR)BR VAF/PLUS) during routine light microscopic (LM) examination. Because the vacuolar change was detected infrequently and was not found in any control mice from that study, it was not clear whether the vacuolation represented a drug-induced change. To address this question, kidney sections from mice from multiple unrelated studies were examined by LM for similar vacuolar changes. Vacuolation was seen by LM in 2.3% of the control and 2.8% of the test article-dosed mice. Transmission electron microscopy (TEM) was also performed on kidneys with prominent light microscopic vacuoles in 5 control mice and 2 test article-dosed mice to further characterize the vacuoles. Ultrastructurally, the vacuoles contained fibrillar and finely stipled granular material or membranous whorls. Kidneys from control mice lacking light microscopic evidence of vacuolation had smaller vacuoles containing similar material when examined by TEM. Because vacuoles were present in both control mice and test article-dosed mice, it was concluded that the vacuoles were incidental and unrelated to compound administration. These studies also demonstrated that vacuoles can be expected to be observed by LM examination in 2-3% of Crl:CD-1 (ICR)BR VAF/PLUS, mice.

Cardiovascular effects after inhalation of large doses of albuterol dry powder in rats, monkeys, and dogs: a species comparison.

Albuterol is a quickly acting beta-2 adrenergic agonist bronchodilator widely used by asthmatics. Because recent case-control studies have suggested a relationship between the increase in mortality of asthmatics over the past decade and the use of beta 2-adrenergic agonists in the control of asthma, concern has developed regarding the potential cardiotoxicity of beta 2-specific adrenergic agonists, including albuterol. The aim of this investigation was to assess the potential for cardiotoxicity of inhaled albuterol dry powder in rats, monkeys, and dogs. All species were exposed to an aerosol of albuterol 1 h per day, 7 days per week, for at least 2 weeks. Control groups were exposed to filtered conditioned air and handled in the same manner as the albuterol-exposed animals. Plasma concentrations of albuterol confirmed systemic exposure. The daily inhaled dose received by the animals was calculated based on measured respiratory minute volumes, published respiratory tract deposition data, as well as HPLC-determined particle size distribution data and aerosolized albuterol concentrations. Multiples of the maximum daily clinical dose (presentation of 15 micrograms/kg in a 70-kg human) were approximately 0.25- to 2500-fold in the rat, 9- to 100-fold in the monkey, and 0.5- to 90-fold in the dog. No findings attributed to albuterol were observed in the monkey. Tachycardia and transient hypokalemia occurred in rats at multiples of 1.5 times or greater of the maximum clinical dose. Absolute and relative heart weights increased in rats receiving multiples of 47 times or greater of the maximum human dose. In the absence of histopathologic findings, the increases in rat heart weights were considered a physiologic hypertrophic response to tachycardia. In dogs tachycardia and transient hypokalemia occurred at all doses tested. Slight to mild fibrosis in the papillary muscles of the left ventricle of the heart occurred in dogs at multiples > or = 19 times the clinical dose. The cardiovascular effects observed were consistent with the known pharmacologic action of beta 2-adrenergic agonists. Due to the lack of toxicologically relevant findings in rats and monkeys and the wide safety margin in dogs, the findings in this study do not suggest a cardiotoxicity risk in the human population after repeated exposures to clinical doses of albuterol currently used in the treatment of asthma.

A pharmacokinetic study of ethanol inhibition of micronuclei induction by urethane in mouse bone marrow erythrocytes.

Urethane (ethyl carbamate) is a genotoxic carcinogen in fermented products and alcoholic beverages. The genotoxicity of urethane requires metabolic activation. Metabolism of urethane is mediated by multiple pathways, and ethanol is known to inhibit the esterase hydrolysis pathway of urethane, which accounts for over 95% of urethane metabolism. This report shows that ethanol also inhibits the induction of micronuclei by urethane in mouse bone marrow erythrocytes, presumably by inhibiting the minor pathway that generates genotoxic metabolite(s). In this study, male CD-1 mice were administered urethane, ethanol, or urethane co-administered with increasing amounts of ethanol in single intraperitoneal injections. Bone marrow polychromatic erythrocytes (PCE) obtained 24 h after injection were scored for micronuclei. The dose of urethane was 1000 mg/kg, and the doses of ethanol were 0, 625, 1250, 2000, 2250, 2500, 3000 and 3500 mg/kg. The blood ethanol level at each dose was determined. Two pharmacokinetic parameters, Cmax and AUC, were estimated for each dose. The observed Cmax of ethanol at doses of 1250, 2000, 2250, 2500, 3000 and 3500 mg/kg were 1.39, 2.84, 3.15, 3.69, 4.13 and 4.76 mg/ml, with AUCs of 1.37, 4.84, 5.88, 7.28, 10.76 and 13.51 mg.h/ml, respectively. Urethane treatment alone markedly increased the micronucleus frequency from 0.1% in the vehicle control to 2.47%. This magnitude of increase was suppressed when urethane was co-administered with ethanol at ethanol doses of 2500 mg/kg and above. At 2500, 3000 and 3500 mg/kg, the micronucleus frequencies reduced from 2.47% to 0.9, 0.44 and 0.28%, respectively. This study shows that ethanol inhibits the induction of micronuclei by urethane.

In vitro chromosome aberration assay of formulated recombinant human granulocyte-macrophage colony-stimulating factor in human peripheral blood lymphocytes.

The clastogenic potential of a recombinant human granulocyte-macrophage colony-stimulating factor (rhGM-CSF) therapeutic protein formulation was assessed in human peripheral blood lymphocyte (HPBL) cultures. Final formulation, rather than pure protein, was evaluated to simulate exposure conditions encountered in man. Because the formulation excipients (citric acid, sodium phosphate, mannitol, human serum albumin and polyethylene glycol) were found to alter cell-cycle kinetics and interfere with S-9 metabolic activation in vitro, a novel testing sequence and protocol were used to distinguish between rhGM-CSF and excipient effects and to guard against false negative results. Initial trials were performed to establish the volume of formulation alone (with and without rhGM-CSF) that would not cause severe cell-cycle delay, interfere with S-9 metabolic activation or inhibit positive control responses. This maximum tolerated volume of formulation was then incorporated in all dosed and control groups of each respective assay phase to give the same extent of cell-cycle delay. In the main assay, 24-hr HPBL cultures were exposed to dose levels of 0, 50, 100, 200 and 350 mug rhGM-CSF/ml for 24 hr in -S-9 phases and 0, 200, 350, 500 and 650 mug/ml for 2 hr in +S-9 phases, and harvested 24 hr later. No significant increases in the percentage of cells with structural chromosomal aberrations were found in either test phase. These results show that rhGM-CSF is not clastogenic in HPBL at greater than 17,000 times human exposure levels, and demonstrate that valid cytogenetic assay data with formulations containing cytotoxic excipient can be obtained in vitro.

Aldicarb sulfoxide/aldicarb sulfone mixture in drinking water of rats: effects on growth and acetylcholinesterase activity.

A 1:1 mixture of aldicarb sulfoxide/aldicarb sulfone was administered to young adult Wistar rats via the drinking water at nominal concentrations of 19.2, 4.8, 1.2., 0.3, 0.075, or 0 ppm for 29 d. Blood was collected after 8, 15, and 29 d of treatment for plasma and erythrocyte cholinesterase determinations, and brain cholinesterase was determined at sacrifice. Body weight, food intake, and water consumption were measured weekly. Body weight gain and water consumption were reduced at 7, 14, 21, and 29 d in male and female rats at 19.2 ppm. Food consumption was reduced in males at 7, 14, 21, and 29 d but was reduced in females only on d 7. Both plasma and erythrocyte cholinesterase activity were reduced after 8, 15, and 29 d in male and female rats at 19.2 ppm. Males at 4.8 ppm showed reductions in plasma activity only after d 8 and in erythrocyte activity only after d 29. Female rats at 19.2 ppm also displayed depressions in brain cholinesterase activity not observed in similarly treated males. Since the only effects noted at 4.8 ppm were reductions in plasma and red blood cell cholinesterase activity in males only and at only one of three sampling periods, these two instances are not believed to be of any biological significance. The data suggest that 4.8 ppm in drinking water is a no observable ill-effect level for exposure of rats to aldicarb residues based on the parameters measured in this study.

Habronemic blepharoconjunctivitis in horses.

Habronemic blepharoconjunctivitis was characterized clinically by raised yellow gritty plaques in the palpebral and bulbar conjunctivae. Lid granulomas and blepharitis were observed in some cases. On histologic examination, mast cells, eosinophils, and collagenolysis was found in most sections, but if only one section was examined an erroneous diagnosis of mastocytosis could have been made. Treatment consisted of larvicidal mixtures for lid lesions and organophosphate ophthalmic drops along with corticosteroids for the conjunctivitis. If the cornea was damaged by the gritty conjunctival plaques, healing was more prolonged because corticosteroids were then contraindicated.

Acute hemolytic anemia, methemoglobinemia, and heinz body formation associated with ingestion of red maple leaves by horses.

From June 1975 through June 1979, acute hemolytic anemia developed in 11 horses from 7 New York farms. Of the 7 horses that died, 6 had methemoglobinemia. In the 4 horses that recovered, methemoglobinemia was not observed. but Heinz body formation was seen in 3 of the 4. On 2 of the premises involved, frank methemoglobinemia was observed concurrently with Heinz body formation, suggesting a relationship between the pathogenesis of methemoglobinemia and Heinz body formation in the hemolytic process. In addition to the 11 cases described, 22 clinically similar cases were reported to us during the period of this investigation by practicing veterinarians from New York, Pennsylvania, and the New England states. All 33 cases of hemolytic anemia occurred between June and October of each year, and affected horses had access to outside paddocks or fields containing a variety of native grasses, weeds, and trees. On 2 farms, hemolytic anemia developed after the horses were observed browsing fallen branches of red maple trees (Acer rubrum). Red maple leaves and bark were obtained from 1 of these farms, and approximately 1 kg of a leaf and bark mixture was fed to each of 2 ponies. Within 48 hours, both ponies became ill. The syndrome was indistinguishable from that observed in clinical patients and was characterized by methemoglobinemia and intravascular hemolysis. The ponies died 5 and 6 days after which time the packed cell volumes were 6% and 7% respectively. It was concluded that many cases of hemolytic anemia in horses in northeastern states may be related to ingestion of leaves or bark from red maple trees. The studies did not, however, define the factors that predispose to poisoning and did not exclude the possibility that other environmental toxins may have been involved.