To clarify whether apoptosis is involved in doxorubicin (DXR)-induced testicular toxicity and to identify the target germ cell type, adult Sprague-Dawley rats were treated with a single intravenous dose of DXR (8 or 12 mg/kg) and euthanized at 3, 6, 12, 24, and 48 h subsequently. Histologically, germ cell degeneration was first found 6 h after dosing in meiotically dividing spermatocytes and early round spermatids of seminiferous tubules at stage 1, and subsequently observed in spermatogonia at stages I-VI showing ultrastructural characteristics of apoptosis. Coincident with the appearance of morphological changes, degenerating germ cells were shown to be undergoing apoptosis as revealed by in situ terminal deoxynucleotidyl transferase-mediated dUTP nick end labeling (TUNEL). The frequency of TUNEL-labeled germ cells increased in a stage- and cell type-specific manner, the peak of frequency gradually progressing from stage I of seminiferous tubules to later stages with time after dosing, suggesting that the damaged germ cells, especially spermatogonia, gradually underwent the processes leading to apoptosis. DNA laddering on gel electrophoresis was apparent 24 and 48 h after dosing. The results demonstrate that apoptosis plays an important role in the induction of testicular toxicity caused by DXR with meiotically dividing spermatocytes and type A and intermediate spermatogonia as highly vulnerable target cells.

Animals↗

PubMedDate not supplied

Simultaneous determination of a new anticancer drug galocitabine and its metabolites in blood by high-performance liquid chromatography.

A relatively simple and sensitive high-performance liquid chromatographic (HPLC) method is described for measuring galocitabine (Ro 09-1390) and its meatbolites, i.e. 5'-deoxy-fluorocytidine (5'-DFCR), 5'-deoxy-fluorouridine (5'-DFUR) and 5-fluorouracil (5-FU), in blood for the purpose of studying pharmacokinetics and toxicokinetics in small animals. The procedure for blood includes deproteinization with acetonitrile. Blood components were separated on a reversed-phase C18 column with a linear gradient of acetonitrile and water and detected at a wavelength of 270 nm. The between-day relative standard deviation (RSD) was less than 10% for all compounds at concentrations of 10-100 micrograms ml-1. The calibration curves obtained from the analysis of blood samples were linear and the correlation coefficients ranged from 0.997 to 0.999. The calculated determination limits were 6.9 micrograms ml-1 for galocitabine, 3.0 micrograms ml-1 for 5'-DFCR, 4.0 micrograms ml-1 for 5'-DFUR and 3.7 micrograms ml-1 for 5-FU.

Animals↗

PubMedDate not supplied

Immunohistochemical and ultrastructural studies of 2,6-dimethylaniline-induced nasal proliferative lesions in a rat two-stage nasal carcinogenesis model initiated with N-bis(2-hydroxypropyl)nitrosamine.

Proliferative lesions induced by 2,6-dimethylaniline (DMA) in a two-stage rat nasal carcinogenesis model were immunohistochemically and ultrastructurally investigated. Male F344 rats received diet containing 3,000 ppm DMA for 52 weeks after initiation with a single subcutaneous injection of 2400 mg/kg of N-bis(2-hydroxypropyl)nitrosamine (DHPN). Histopathologically, proliferation of Bowman's glands, glandular hyperplasias, dysplastic foci, adenomas, and carcinomas were observed in treated rats. These nasal lesions mostly arose in the olfactory mucosa of the nasal cavity. Immunohistochemically, they were positive for cytokeratin and/or collagen type IV antibodies. Ultrastructurally, intracytoplasmic dense secretory granules (200-850 nm in diameter), identical to those in normal Bowman's glands, were observed in all the lesions, providing further support from an origin from these glands. Based on their cellular characterization, growth pattern and/or proliferative activity, two morphological continua were evident, one from dysplastic foci to carcinomas and the other from proliferation of Bowman's glands to glandular hyperplasias and adenomas. These results suggest that dysplastic foci arise from Bowman's glands and progress to carcinomas, while proliferation of Bowman's glands result in glandular hyperplasias and adenomas.

Adenoma↗

PubMedDate not supplied

Novel control system for blood glucose using a model predictive method.

We developed a novel blood glucose control system, using a model predictive method, to achieve optimal control of the blood glucose level in severely diabetic or pancreatectomized patients. This system is designed to predict glucose level changes in advance, considering delayed response time and the administered doses of insulin. This method is also designed to calculate the most appropriate insulin infusion rate by considering differences in individual response to insulin. In this study, we compared our system with a conventional proportional and differential controller (PD controller) to determine whether the new system could regulate the glucose level efficiently in pancreatectomized dogs. The model predictive control method resulted in a significant reduction of mean insulin infusion rate compared with the conventional PD controller (0.71 mU/kg per min vs. 1.81 mU/kg per min, p = 0.0005), when the glucose level in both methods reached the planned target level (100 mg/dl). The new system also tended to have a reduced mean glucose infusion rate for compensating for overshooting of the glucose level compared with the PD controller (0.7 mg/kg per min vs. 1.1 mg/kg per min, p = 0.16). These results indicate that the new system should be a useful tool for regulating the glucose level in severely diabetic patients.

Animals↗

PubMedDate not supplied

Expression of insulin-like growth factor-2 can predict the prognosis of human colorectal cancer patients: correlation with tumor progression, proliferative activity and survival.

Expression of insulin-like growth factor-2 (IGF-2) has been reported in tissue specimens and cell lines of human colorectal cancers. However, the effects of IGF-2 in colorectal cancer patients are not well known. In this study, IGF-2 staining was performed on tissue samples from 92 patients with colorectal cancer, and the relationship of IGF-2 staining to clinicopathological variables, proliferating cell nuclear antigen (PCNA) staining and patient survival was analyzed. IGF-2 staining was correlated with tumor progression, PCNA staining and patient survival. Our results suggest that IGF-2 plays an important role in tumor progression and that IGF-2 staining is useful as a prognostic factor in colorectal cancer patients.

Adult↗

PubMedDate not supplied

Dual effects of prolonged ACTH stimulation on 4-hydroxyaminoquinoline 1-oxide-induced adrenocortical lesions in rats.

The effects of a long-acting synthetic ACTH on 4-hydroxyaminoquinoline 1-oxide (4HAQO)-induced adrenocortical lesions were investigated in female rats. A total of 140 6-week-old rats were divided into 4 equal groups, given a single s.c. injection of 7 mg/kg 4HAQO or vehicle, followed by repeated sc administration of the synthetic ACTH or no further treatment. Subgroups of 10 rats in each group were sequentially sacrificed at weeks 20, 30, and 40. Adenomas and adenomatous nodules developed in the adrenal cortex of animals receiving 4HAQO and the chronic ACTH stimulation. Both lesions were located in the deeper zones of the adrenal cortex adjacent to the medulla and were composed of large-sized, clear-type cells. From week 20, middle zone, cortical cystic degeneration, which mimics the age-associated degenerative change named adrenal peliosis, was frequently observed in the adrenal glands of animals treated with 4HAQO alone. Its development was inhibited by ACTH. In the control animals, peliotic changes occurred at low incidence and only at the termination of experiment. These results indicate that long-term stimulation of ACTH promotes the development of adrenocortical tumors but suppresses the occurrence of adrenal peliosis in rats treated with 4HAQO.

4-Hydroxyaminoquinoline-1-oxide↗

PubMedDate not supplied

Liver tumor-promoting effect of beta-naphthoflavone, a strong CYP 1A1/2 inducer, and the relationship between CYP 1A1/2 induction and Cx32 decrease in its hepatocarcinogenesis in the rat.

Interrelationships among induction of cytochrome P-450 (CYP) 1A1/2, decrease in connexin 32 (Cx32), and liver tumor-promoting activity by beta-naphthoflavone (BNF) in the promotion stage were examined in a 2-stage liver carcinogenesis model. A total of 20 male Fischer 344 rats were initiated with a single intraperitoneal injection of 150 mg/kg of diethylnitrosamine (DEN) or were given the saline vehicle alone. Starting 2 weeks later, they were fed a diet containing 2%, 1%, or 0% BNF for 6 weeks. All animals were subjected to a two-thirds partial hepatectomy at week 3 and were sacrificed at week 8. Absolute and relative liver weights were significantly increased in the DEN+BNF groups as compared to the DEN-alone group. Diffuse hepatocellular hypertrophy with cytoplasmic eosinophilia, sometimes accompanied by development of adenoma-like hepatic foci, was observed in the BNF-treated rats. Remarkable induction of cytochrome CYP 1A1/2 and significant increase in CYP 2E1 were noted in the DEN+BNF groups, and positive immunohistochemical staining for both was observed diffusely. The areas of Cx32-positive spots per hepatocyte in the centrilobular areas of livers of the BNF-treated rats were significantly decreased, but no changes were observed in periportal areas. The numbers and areas of foci positive for glutathione S-transferase placental form were increased in the BNF-treated groups. These results suggest that BNF is a liver tumor promoter that, unlike phenobarbital, does not induce CYP 2B1/2 isozymes, and there seems to be no direct relationship between CYP 1A1/2 induction and Cx32 reduction in BNF hepatocarcinogenesis.

Animals↗

PubMedDate not supplied

A mechanistic study of ovarian carcinogenesis induced by nitrofurazone using rasH2 mice.

In order to clarify whether the ovarian tumors induced in a long-term carcinogenicity study of nitrofurazone (NF) in mice can be also produced in a short-term model using transgenic (Tg) mice carrying the human c-Ha-ras gene (rasH2 mice), the following 3 experiments were performed. In experiment 1, both rasH2 mice and their wild CB6F1 littermates carrying no c-Ha-ras gene (non-Tg mice) that were fed a diet containing 500 to 1,000 ppm NF for 7 weeks demonstrated ovarian atrophy characterized by decreased labeling indices (LIs) for proliferating cell nuclear antigen (PCNA) in granulosa cells. In experiment 2, increased numbers of atretic follicles and decreased PCNA LIs in granulosa cells were recognized in rasH2 mice given diets containing 250 or 500 ppm NF for 26 weeks, but no tumor induction was grossly observed. In experiment 3, similar ovarian atrophy was observed in association with increased serum luteinizing hormone (LH) levels in both rasH2 and non-Tg mice given diet containing 1,000 ppm NF for 11 days. These results indicate that long-term NF treatment induces ovarian tumors in mice, possibly by continuous stimulation with gonadotropins such as LH via a negative-feedback phenomenon secondary to ovarian atrophy (as the tumor-induction mechanism), although we could not completely rule out a genotoxic mechanism.

Animals↗

PubMedDate not supplied

Sequential observation of spontaneous endometrial adenocarcinoma development in Donryu rats.

Sequential observation of spontaneous endometrial adenocarcinoma development revealed a clear, hormone-dependent, histogenetic pathway in Donryu rats. The first histological changes of the uterine endometrium appeared in both the lining epithelium and uterine gland of the endometrium at 6 mo of age, along with the beginning of persistent estrus. These changes included areas of tall columnar epithelium and gland formation in the lining epithelium as well as metaplastic change in the uterine gland. At 8 mo of age, endometrial hyperplasias were found, with subsequent increase in both incidence and degree. At 8-10 mo of age, hyperplasias were all within the limit of grade ++. After 12 mo of age, however, severe hyperplasias (grade ) began to increase markedly, and adenocarcinomas developed at 15 mo of age. The findings thus suggest that uterine endometrial adenocarcinomas arise from hyperplastic lesions, which should therefore be regarded as preneoplastic, as in the human case. Sequential analysis of plasma gonad steroids also ascertained a link between the appearance of these lesions and an increased estrogen:progesterone ratio, suggesting that estrogen may play an important role in development of both hyperplastic and neoplastic lesions. In Fischer-344 rats used for comparative assessment of strain differences, neither advanced histological changes nor hormonal changes were evident.

Adenocarcinoma↗

PubMedDate not supplied

Relationship between the development of pulmonary fibrosis and lung tumors in Syrian golden hamsters induced by N-methyl-N-nitrosourethane.

To cast light on the relationship between the development of pulmonary fibrosis and lung cancer, female Syrian golden hamsters were given 5 sc injections of 0.6 mg/animal of N-methyl-N-nitrosourethane (MNUR) at 2-wk intervals and then maintained without any treatment for 26 wk. Bronchiolo-alveolar cell tumors and hyperplasias, which were recognized from week 4 after termination of treatment, were each subdivided morphologically into 3 types. Bronchiolo-alveolar cell tumors included papillary tumors consisting of basophilic cells, papillary tumors consisting of clear cells, and papillary/solid tumors consisting of pleomorphic cells, with papillary tumors consisting of basophilic cells predominating. Bronchiolo-alveolar cell hyperplasias encountered were glandular metaplasia, simple hyperplasia, and papillary hyperplasia. Glandular metaplasia was the most common of the hyperplastic lesions. To some extent, all the proliferative lesions were associated with inflammatory cell infiltration, but this varied greatly. The rate for papillary tumors consisting of basophilic cells with connective tissue proliferation was 35%. Among the hyperplastic lesions, the cell proliferative activity of papillary hyperplasia (12.5%) was significantly higher than in other hyperplastic lesions, suggesting that this lesion might be a preneoplastic change. None of the lung proliferative lesions showed any unequivocal immunoreactivity for the p53 protein. The present study suggests that most lung tumors may develop from pulmonary inflammatory lesions induced by MNUR, but the possibility that DNA injuries to the respiratory epithelial cells by MNUR may cause lung tumors cannot be precluded.

Animals↗

PubMedDate not supplied

Comparison of ultrastructural changes in thyrotrophs of the rat pituitary between intermittent and continuous treatments with sulfadimethoxine.

To clarify relationships between serum thyroid-stimulating hormone (TSH) levels and ultrastructural changes in thyrotrophs caused by intermittent or continuous treatments with antithyroid compound, male Fischer-344 rats initiated with N-bis(2-hydroxypropyl)nitrosamine (DHPN) were given water containing 0.1% sulfadimethoxine (SM) for 8 wk and then sacrificed (Group 1). Additional groups were examined 2 wk after withdrawal (Group 2), after 4 wk retreatment following a 2-wk withdrawal period (Group 3), and after 14 wk continuous exposure (Group 4). Control rats (Group 5) remained untreated for 8 wk after the DHPN initiation. Microscopic examination revealed hypertrophy of thyrotrophs and depletion of TSH-positive material in Groups 1, 3, and 4 but a return to normal in Group 2. Electron microscopic examination of thyrotrophs in the anterior pituitary in Groups 1, 3, and 4 revealed dilated rough endoplasmic reticulum (ER) cisternae with intracisternal dense granules as well as diminished numbers of intracytoplasmic secretory granules, these changes being most marked in Group 1 and least pronounced in Group 4. The number of intracytoplasmic secretory granules in Group 3 was much lower than in Group 4, as demonstrated by morphometric analysis. In Group 2, thyrotrophs showed dilated rough ER cisternae without intracisternal dense granules and essentially the same component of intracytoplasmic secretory granules as Group 5. The present study suggest that while prolonged continuous treatment with SM results in gradual acclimation to an increased demand for TSH, intermittent treatment elicits a persistent state of reduced TSH storage in thyrotrophs due to a continued strong feedback through the hypothalamus-pituitary-thyroid axis.

Animals↗

PubMedDate not supplied

Mice with focal pulmonary fibrosis caused by monocrotaline are insensitive to urethane induction of lung tumorigenesis.

To establish the characteristics of an optimized pulmonary fibrosis model, male ICR mice were given 4 weekly sc injections of 150 or 0 mg/kg monocrotaline (MCT) and maintained without further treatment for 33 wk (Experiment 1). The final mortality in the MCT group was 64%. Epithelial cells with large bizarre nuclei and an increased incidence of alveolar/bronchiolar hyperplasias were typically observed. In areas of pulmonary fibrosis, the PCNA labeling index (LI) in the alveolar/airway epithelium was significantly elevated. DNA content analysis demonstrated a larger range (4-8C) for the ploidy pattern of alveolar epithelium with large bizarre nuclei than in the normal epithelium (2C). In Experiment 2, the relationship between pulmonary fibrosis development and lung tumorigenesis was investigated. Mice were given 4 weekly sc injections of 150 and 0 mg/kg MCT, followed by a single i.p. injection of 1,000 or 500 mg/kg urethane (UR) on week 7, then maintained without further treatment for an additional 15 wk. UR following MCT-induced inflammatory changes, fibrosis, and epithelia with large bizarre nuclei but no tumorous lesions, in spite of the fact that treatment with UR alone caused a high incidence of pulmonary tumors. Hyperplasias were seen in all groups, but the multiplicity in the combined groups tended to be decreased by the MCT pretreatment. The present study demonstrated that this new protocol is more suitable than previous one for the experimental production of pulmonary fibrosis. Furthermore, the induction of lung tumors by UR was completely depressed in mice with MCT-induced pulmonary fibrosis, suggesting that alveolar epithelial cells are resistant to this lung carcinogen under these conditions.

Animals↗

PubMedDate not supplied

Liver tumor promoting effects of fenbendazole in rats.

In order to examine whether fenbendazole has tumor-promoting activity, a total of 70 male Fischer 344 rats were initiated with a single intraperitoneal injection of 100 mg/kg of diethylnitrosamine (DEN) or were given the saline vehicle alone; beginning 1 wk later, rats were given a diet containing 3,600; 1,800; 600; 200; 70; or 0 ppm of fenbendazole for 8 wk. Subgroups of 5 rats each from the DEN+ 1,800; DEN+0; 1,800; and 0 ppm groups were euthanatized after 1 wk of fenbendazole treatment, and the remaining animals were euthanatized at 8 wk. After 1 wk, relative liver weights (ratios to body weights) were significantly increased in the DEN+ 1,800 and 1,800 ppm groups, and based on light microscopy, periportal hepatocellular hypertrophy was evident in these groups. After 8 wk, relative liver weights were significantly increased in the groups given > or =600 ppm with or without DEN initiation. Periportal hepatocellular hypertrophy, characterized by a marked increase in smooth endoplasmic reticulum, was observed in the groups given > or =600 ppm with or without DEN initiation. Induction of cytochrome P-450 (CYP) 1A2, 2B1, or 4A1 was noted in the fenbendazole-treated groups with or without DEN initiation; that associated with CYP 1A2 was most marked. Positive immunostaining for anti-CYP 1A1/2 or CYP 2B1/2 was observed diffusely in the livers of animals in the DEN+1,800 and DEN+3,600 ppm groups. The numbers and areas of connexin 32 (Cx32)-positive spots per square centimeter in centrilobular hepatocytes were significantly decreased in an almost dose-dependent manner with fenbendazole treatment after DEN initiation. In situ hybridization for Cx32 mRNA revealed a remarkable decrease in its expression in the centrilobular hepatocytes in the DEN+70 ppm group. The numbers of glutathione S-transferase placental-form positive single cells (plus mini foci) were significantly increased in the DEN+ 1,800 and DEN+3,600 ppm groups. Since those agents that induce CYP 2B1/2 isozymes and reduce Cx32 in centrilobular hepatocytes have been suggested to be liver tumor promoters, the present results indicate that fenbendazole may be a liver tumor promoter.

Alkylating Agents↗

PubMedDate not supplied

Experimental induction of uterine adenocarcinoma in rats by estrogen and N-methyl-N-nitrosourea.

To induce uterine endometrial adenocarcinoma, Donryu rats were given subcutaneous injections of 100 micrograms/kg estradiol dipropionate (ED) four times once a week from 3 months old, followed by weekly intraperitoneal injections of 20 mg/kg N-methyl-N-nitrosourea (MNU) four times. In the ED+MNU group, the incidences of marked hyperplasias and adenocarcinomas were higher than those in the control and ED or MNU alone groups, when the incidences were compared in survival cases only up to the age of about 15 months, while the survival rate showed a specially low value of 35% in this group. These results may suggest the possibility of early and high occurrence of uterine adenocarcinoma in this rat strain by estrogen and MNU.

Adenocarcinoma↗