Proliferative lesions of ovarian granulosa cells and reversible hormonal changes induced in rats by a selective estrogen receptor modulator.

This study assessed the effects of raloxifene, a selective estrogen receptor modulator (SERM), on ovarian morphology and circulating hormone levels in rats. Female Fischer-344 rats (65/group) were given dietary raloxifene for 6 months at average daily doses of 0, 15, 75, and 365 mg/kg. Morphologic evaluation of ovaries was conducted on 25 rats/group at the end of the treatment period and from 20 rats per group after 1 and 3 months withdrawal from treatment. Plasma hormone analyses were conducted on 10 rats pergroup at the end of the treatment period and aftereach withdrawal period. Treatment with raloxifene for 6 months resulted in disruption of the hypothalamic-pituitary-ovarian axis, manifested by increased plasma concentrations of luteinizing hormone (LH) and estradiol-17beta (E2), and failure of ovulation, manifested by ovarian follicular prominence (retained anovulatory follicles), lack of corpora lutea (CL), and depressed plasma progesterone (P4). Many (56% to 80%) rats in all raloxifene treated groups had focal, minimal to slight hyperplasia of granulosa cells within individual retained follicles. A few treated rats in the mid- and high-dose groups (2 of 25 and 3 of 25, respectively) had more extensive focal proliferation of granulosa cells. These foci were approximately 3 to 6 mm in overall size and were characterized by moderate papillary proliferation of large granulosa cells associated with cystic spaces, often with hemorrhage. In 4 of the 5 rats with this focal cystic granulosa cell hyperplasia, the remainder of the involved ovary and the contralateral ovary were atrophic. After 1 or 3 months of drug withdrawal, most previously treated rats examined had morphologic evidence of ovarian cyclic changes. including developing follicles, various stages of CL, and normal plasma levels of LH, E2, and P4. Continued lack of cyclic changes was limited to 4 of 20 rats from the low-dose group after 1 month of recovery and to 1 low dose rat after 3 months. Intrafollicular granulosa cell hyperplasia was not seen in rats in the reversibility phase. Areas of prior focal cystic granulosa cell hyperplasia were represented by focal sclerosis that included hemorrhage and/or hemosiderin. The foci of sclerosis were associated with cystic spaces after 1 month and were solid after 3 months. A granulosa cell tumor, approximately 12-13 mm diameter, was present in a high-dose rat in the 3-month reversibility group. This tumor effaced 1 ovary and was characterized by proliferative granulosa cells, usually in papillary formations and cords within cystic spaces. This rat had atrophy of the uninvolved ovary, excessive plasma levels of E2 and prolactin, and high P4 levels considering the absence of CL. The results of this study indicate that ovarian granulosa cells in rats are susceptible to proliferative changes when stimulated chronically with excessive trophic hormones. Most of these proliferative changes were reversible upon cessation of the hormonal stimulation. However, the proliferative lesion in one treated rat progressed to apparent autonomous (neoplastic) growth.

Proliferative lesions of ovarian granulosa cells and reversible hormonal changes induced in rats by a selective estrogen receptor modulator.

This study assessed the effects of raloxifene. a selective estrogen receptor modulator (SERM), on ovarian morphology and circulating hormone levels in rats. Female Fischer-344 rats (65/group) were given dietary raloxifene for 6 months at average daily doses of 0, 15, 75, and 365 mg/kg. Morphologic evaluation of ovaries was conducted on 25 rats/group at the end of the treatment period and from 20 rats per group after 1 and 3 months withdrawal from treatment. Plasma hormone analyses were conducted on 10 rats per group at the end of the treatment period and after each withdrawal period. Treatment with raloxifene for 6 months resulted in disruption of the hypothalamic-pituitary-ovarian axis, manifested by increased plasma concentrations of luteinizing hormone (LH) and estradiol-17beta (E2), and failure of ovulation, manifested by ovarian follicular prominence (retained anovulatory follicles), lack of corpora lutea (CL), and depressed plasma progesterone (P4). Many (56% to 80%) rats in all raloxifene treated groups had focal, minimal to slight hyperplasia of granulosa cells within individual retained follicles. A few treated rats in the mid- and high-dose groups (2 of 25 and 3 of 25, respectively) had more extensive focal proliferation of granulosa cells. These foci were approximately 3 to 6 mm in overall size and were characterized by moderate papillary proliferation of large granulosa cells associated with cystic spaces, often with hemorrhage. In 4 of the 5 rats with this focal cystic granulosa cell hyperplasia, the remainder of the involved ovary and the contralateral ovary were atrophic. After 1 or 3 months of drug withdrawal, most previously treated rats examined had morphologic evidence of ovarian cyclic changes, including developing follicles, various stages of CL, and normal plasma levels of LH, E2, and P4. Continued lack of cyclic changes was limited to 4 of 20 rats from the low-dose group after 1 month of recovery and to 1 low dose rat after 3 months. Intrafollicular granulosa cell hyperplasia was not seen in rats in the reversibility phase. Areas of prior focal cystic granulosa cell hyperplasia were represented by focal sclerosis that included hemorrhage and/or hemosiderin. The foci of sclerosis were associated with cystic spaces after 1 month and were solid after 3 months. A granulosa cell tumor, approximately 12-13 mm diameter, was present in a high-dose rat in the 3-month reversibility group. This tumor effaced 1 ovary and was characterized by proliferative granulosa cells, usually in papillary formations and cords within cystic spaces. This rat had atrophy of the uninvolved ovary, excessive plasma levels of E2 and prolactin, and high P4 levels considering the absence of CL. The results of this study indicate that ovarian granulosa cells in rats are susceptible to proliferative changes when stimulated chronically with excessive trophic hormones. Most of these proliferative changes were reversible upon cessation of the hormonal stimulation. However, the proliferative lesion in one treated rat progressed to apparent autonomous (neoplastic) growth.

Developmental toxicology studies of vancomycin hydrochloride administered intravenously to rats and rabbits.

Pregnant CD rats were given vancomycin intravenously in doses of 0, 40, 120, or 200 mg/kg on Gestation Days (GD) 6-15; pregnant New Zealand white rabbits were given 0, 40, 80, or 120 mg/kg intravenously on GD 6-18. Cesarean sections were performed on rats and rabbits on GD 20 and 28, respectively. In rats, maternal toxicity was indicated in the 120- and 200-mg/kg treatment groups by cortical tubular nephrosis. Maternal body weight gain and food consumption and fetal viability, weight, and morphology were not adversely affected by vancomycin. Maternal and developmental no observed adverse effect levels (NOAELs) in the rat were 40 and 200 mg/kg, respectively. In rabbits, maternal toxicity was indicated by cortical tubular nephrosis in the 80- and 120-mg/kg treatment groups; a single death and depression of body weight gain and food consumption occurred in the 120-mg/kg treatment group. Developmental toxicity was indicated by depression of fetal weight in the 120-mg/kg treatment group; fetal viability and morphology were not adversely affected by vancomycin. Maternal and developmental NOAELs in the rabbit were 40 and 80 mg/kg, respectively. Based on these data, vancomycin did not exhibit selective toxicity toward the developing rat or rabbit conceptus.

Use of vancomycin hydrochloride for treatment of Clostridium difficile enteritis in Syrian hamsters.

As part of an 18-month carcinogenicity study, 680 Syrian hamsters (Mesocricetus auratus) received daily gavage doses of fenazaquin, an experimental miticide. Mortality associated with severe enteritis was noticed beginning when the hamsters were 4 months old and ranged from one to five deaths per month until the hamsters were about 10 months old, when 41 deaths occurred in a 1-month period. Ante- and postmortem findings were consistent with those reported for antibiotic-induced enteritis in hamsters. Clostridium difficile was isolated from 12 of the 13 samples of cecal contents analyzed. Toxin assays of C. difficile isolates collected from 11 affected animals were positive for both cyto- and enterotoxins. Daily oral administration of vancomycin hydrochloride at a dose of 20 mg/kg was initiated when the hamsters were about 10 months old. Deaths due to C. difficile enteritis were significantly decreased within 2 weeks, and treatment was continued for 3 months. A trial withdrawal period for a subset of 64 hamsters (approximately 16% of the total population) was initiated to evaluate survival after discontinuation of the antibiotic treatment. Clostridium difficile enteritis recurred within 2 weeks and caused 19 deaths during the next month; therefore, these hamsters were returned to daily vancomycin treatment for the remainder of the study. With the exception of severe gaseous distention of the ceca, which caused death in 17 (< 4% of the total population) of the affected hamsters, vancomycin treatment did not cause any major adverse effects.(ABSTRACT TRUNCATED AT 250 WORDS)

Incidence of spontaneous neoplastic and nonneoplastic lesions in Charles River CD-1 mice varies with breeding origin.

Three separate control lifetime studies were conducted with untreated Crl:CD-1 (ICR)BR mice using a total of 400 mice/sex maintained to 21 mo of age. Similar husbandry practices and environmental conditions were used for all 3 studies. It was noted after study initiation that the Charles River breeding facility of origin was different for each study. The aggregate range of survival and incidence of neoplasms for the combined studies was similar to that previously reported. However, these 3 groups of mice had prominent variation in survival and in the incidence of pulmonary adenomas and systemic amyloidosis in males and females, and in the incidence of hepatocellular neoplasms in males. The present studies indicate that consistent procurement of test animals is an additional variable to be considered in the establishment of a valid database within a test facility when using an outbred mouse.

Carcinogenicity studies of fluoxetine hydrochloride in rats and mice.

The antidepressant drug fluoxetine HCl was tested for carcinogenicity in three well designed and controlled studies in Fischer rats and C57BL/6 x C3H F1 mice. The compound was administered to the animals for 24 months at dietary doses of approximately 0, 0.5, 2.0, or 10.0 mg/kg body weight in rats and 1.0, 5.0, or 10.0 mg/kg in mice. The highest dose tested was a maximum tolerated dose for both species as evidenced by clinical signs (rats and mice) and some mortality (mice) referable to central nervous system pharmacological effects, decreased weight gain (rats), and histopathological changes of phospholipidosis (rats) and hepatic fatty change (mice). There was no evidence of an increased incidence of any type of unusual or commonly occurring spontaneous neoplasm in either rats or mice. There were statistically significant decreases in a few commonly occurring neoplasms. The data reported herein provide convincing evidence that fluoxetine is neither a complete carcinogen nor a tumor promoter.

Comparative toxicity of oral cephalosporin antibiotics in the rabbit and in a rabbit kidney cell line (LLC-RK(1)).

The nephrotoxic potential of four oral cephalosporin antibiotics, cephalexin, cefaclor, LY195885 and LY171217, was determined in rabbits given single oral doses of 250-500 mg/kg body weight. Histopathological changes, blood chemistry, and ex vivo renal slice function were evaluated 48 hr after dosing. Additionally, the viability of rabbit renal cells in culture (LLC-RK(1)) was evaluated by nigrosin dye exclusion after 48 hr exposure to each antibiotic at concentrations of 0.5-2.0 mg/ml. Only LY171217 was significantly nephrotoxic in vivo. Prominent lesions were observed at 500 mg/kg body weight and were accompanied by marked increases in blood urea nitrogen and serum creatinine, and decreases in ex vivo renal slice gluconeogenesis and p-aminohippurate and tetraethylammonium uptake. In vitro toxicity to renal cells correlated well with the in vivo results yielding TC(50) values (TC(50) = concentration producing 50% lethality) > 1.0 mg/ml for cephalexin, LY195885 and cefaclor. LY171217, on the other hand, was significantly toxic in vitro (TC(50) = < 0.5). These results suggest that renal cells in culture may provide a useful method for examining the nephrotoxic potential of oral cephalosporins before in vivo studies.

A species comparison of the toxicity of nabilone, a new synthetic cannabinoid.

Acute, subchronic, and chronic studies were conducted in various species to evaluate and compare the toxicity of nabilone, a new synthetic 9-ketocannabinoid that is orally effective for the treatment of nausea and vomiting induced by cancer chemotherapy agents. The oral LD50 in mice and rats for nabilone formulated as a polyvinylpyrrolidone (PVP) codispersion was in excess of 1000 mg/kg. Among nonrodents, rhesus monkeys had a higher tolerance to the CNS depression induced by single oral doses of nabilone-PVP than did dogs. Rats fed dietary mixtures of nabilone-PVP which provided approximate daily nabilone doses of 1 to 93 mg/kg tolerated treatment for 3 months with no deaths. Treatment-related changes (at doses greater than or equal to 5 mg/kg) were limited to reduced body temperature, slight-to-moderate decreases in weight gain, and behavioral changes (e.g., hyperactivity, hyperirritability to touch, and hypoactivity). All dogs treated for 3 months with daily oral doses of up to 1.0 mg/kg survived; treatment-related effects were limited to transient episodes of ataxia and anorexia. Nabilone treatment of rats and dogs for 3 months produced no evidence of systemic toxicity in the clinical chemistry, hematology, or pathology parameters examined. Chronic treatment of dogs with daily oral doses of nabilone-PVP equal to 0.5, 1.0, or 2.0 mg of nabilone/kg produced cumulative toxicity; by the end of 7 months, 2, 6, and 7 dogs in the respective dose groups had died. In a number of instances, death was preceded by one or more convulsive episodes. In contrast to the dog, the toxic potential of nabilone was minimal in rhesus monkeys treated with nabilone-PVP for 1 year at daily oral nabilone doses of up to 2.0 mg/kg. The enzymatic reduction of the 9-keto group of nabilone to form carbinol metabolites was a major metabolic pathway for nabilone in dogs but not in rhesus monkeys. The carbinols were long-lived metabolites in the plasma of dogs and accumulated in the plasma compartment with time. Furthermore, the carbinol metabolites were found to concentrate in the brain tissues of treated dogs. Although the precise mechanism for this marked species difference in chronic toxicity is not known, the metabolic differences responsible for the presence of the carbinol metabolites at high concentrations in the plasma and brain over time may play a role in the toxicity observed in the dog.

Nasal delivery of polypeptides I: nasal absorption of enkephalins in rats.

The serum levels of two enkephalins after various routes of administration were compared in rats. The results indicated that serum levels of metkephamid after nasal administration were not significantly different than levels after intravenous injection. The oral administration of metkephamid resulted in undetectable serum levels. The effects of a promoter and variations in the peptide dose on nasal absorption were studied. Depending on the stability of the polypeptide and its susceptibility to enzymatic degradation, nasal absorption of peptides can be influenced by the presence of a promoting agent in the formulation. A linear relationship between the dose and the AUC was observed in the range of concentrations studied. The absorption mechanism appears to be passive diffusion. Microscopic examinations of nasal mucosa in rats revealed degrees of irritation which, considering the experimental exposure, were slight and probably repairable. The data indicate that enkephalins can be absorbed through the nasal mucosa into the systemic circulation, and the onset of absorption was rapid. Nasal administration may offer an attractive alternative for the delivery of proteins and/or polypeptides which are, in general, absorbed poorly when given orally.

Nasal drug delivery system of a quaternary ammonium compound: clofilium tosylate.

The blood levels of the [14C]clofilium ion in rats after various routes of administration of clofilium tosylate were compared. The results indicate that the blood levels after nasal administration were not statistically different from levels after intravenous administration (p greater than 0.05). Administration by the oral route resulted in considerably lower blood levels. Nasal administration of clofilium tosylate appeared to be superior to oral administration. Histological examinations of nasal mucosa were conducted. At the lower concentration, mild necrosis was observed, and large areas of mucosa were unaffected. However, necrosis of large areas of mucosa occurred after exposure to the higher concentration. Levels of radioactivity in heart, liver, lung, and kidney tissue, as a function of time, were also studied. Unlike the blood levels after nasal administration, the levels of radioactivity were persistent in heart tissue. The data suggest that the [14C]clofilium ion and/or metabolite concentrate in the heart and that blood levels of radioactivity may not be an accurate index of cardiac levels or biological response.

Exploration of the negative correlation between proliferative hepatocellular lesions and lymphoma in rats and mice--establishment and implications.

Data from 1858 mice at Lilly and eight studies from the literature were used to establish a negative correlation between proliferative hepatic lesions and malignant lymphoma in rodents. This negative correlation implies that the rodent model is flawed in the sense that interpretation of hepatic lesions without consideration of malignant lymphoma can lead to the incorrect conclusion that there is something unique about the compound. In fact it is the rodent model that has a unique feature--the negative correlation.

Positive correlation of body weight with pituitary tumor incidence in rats.

Data based on 4700 Wistar rats in 13 two-year studies were used to calculate the correlation between body weight at 12 months and pituitary tumor incidence. The positive correlations of 0.754 for male rats and 0.828 for female rats were both highly significant (p less than 0.0001). The results indicate that any treatment which causes increased body weight will probably be associated with an increased incidence of pituitary tumors.