Enhancement of ribonucleic acid synthesis by chromium(III) in mouse liver

The effect of Cr(III) administration on hepatic RNA synthesis in mice was studied. It was found that Cr accumulated in mouse liver. Forty-eight hours after intraperitoneal injection of CrCl3 (0.005-5 mg Cr/kg body weight) approximately 10% of the administered dose per g of tissue remained. The accumulated Cr was still retained 64 days after administration (5 mg Cr/kg) with only a slight decrease. Approximately 20% of the hepatic Cr was detected in the nuclei. By administering CrCl3. RNA synthesis in mouse liver was markedly enhanced without altering the pool size of nucleotides. This enhancement was dose-dependent and statistically significant at doses of 0.05 (p less than 0.05), 0.5 (p less than 0.01), and 5 mg Cr/kg (p less than 0.01), and remained so for at least 16 days after administration of 5 mg Cr/kg. The synthesis of DNA and protein in mouse liver were not significantly changed by CrCl3 administration. On the other hand, Cr(VI) administration did not enhance but rather inhibited RNA synthesis in mouse liver. These results suggest that Cr(III) specifically enhances RNA synthesis in mouse liver.     

Pharmacodynamics of mefloquine and praziquantel combination therapy in mice harbouring juvenile and adult Schistosoma mansoni

Praziquantel (PZQ) is currently the only drug widely used for the treatment of schistosomiasis, but the antimalarial drug mefloquine (Mef) possesses interesting antischistosomal properties. Combination therapy with these two drugs has been suggested as a strategy for transmission control, as PZQ is active against adult worms and Mef is active against schistosomula. To examine the efficacy of combination therapy, Schistosoma mansoni-reinfected mice were separated into seven groups: untreated (I), treated with PZQ in doses of 200 mg/kg (II) or 1,000 mg/kg (III), treated with Mef in doses of 200 mg/kg (IV) or 400 mg/kg (V); each dose was divided equally and given on two consecutive days. Group VI was treated with doses of PZQ + Mef as in groups II and IV, respectively, while group VII was treated with PZQ + Mef as in groups III and V, respectively. PZQ + Mef at the reduced doses of 200 mg/kg each enhanced the therapeutic efficacy over the reduced PZQ dose alone as shown by a very high reduction in the total numbers of mature worms (95% vs. 49%), immature worms (96% vs. 29%) and the complete eradication of immature females, mature females and immature eggs. The reduction in worm burden was associated with the healing of hepatic granulomatous lesions and the normalisation of all liver enzymes. Therefore, the use of Mef with PZQ is more effective than PZQ alone and should be considered for clinical trials in humans as a potential treatment regimen to prevent treatment failures in areas with high rates of schistosomiasis.     

Prolongation of skin xenograft survival with modified cultured fibroblasts

Cyclosporine A, one of the most potent immunosuppressive drugs, mediates some of its immunosuppressive and nephrotoxic effects by enhancing transforming growth factor-beta secretion and receptor expression. In this experimental study, the effect of cyclosporine pretreatment of cultured dermal fibroblasts on xenogeneic tissue rejection after microimplantation beneath skin grafts was investigated. The effects of the site-specific immunosuppressive strategy on skin xenograft survival were tested. Because the skin is an immunological indicator of the rejection of composite tissue allografts, it was considered that this strategy could be used as a supportive therapy for composite tissue allotransplantation in the future. In the first stage of the study, fibroblast cultures obtained from skin biopsy samples from five rats were treated with different single doses of cyclosporine (100 to 3000 ng/ml), and transforming growth factor-beta levels were measured in culture supernatants after 72 hours. In the second stage, 60 Sprague-Dawley rats were divided into six groups, as follows. For group I (sham), only the standard grafting procedure was performed. For group II, after the standard grafting procedure, rats were treated with intraperitoneal injections of 30 mg/kg (n = 5) or 10 mg/kg (n = 5) cyclosporine for 10 days. For group III, cultured fibroblasts obtained from skin biopsy samples from rats were treated with 100 or 500 ng/ml cyclosporine, and the cells were collected by light trypsinization and centrifugation after 72 hours. After the standard skin grafting procedure, modified fibroblasts were implanted between the graft and the recipient bed with a Pasteur pipet. For group IV, the same procedures as for group III were performed and then rats were treated with 10 mg/kg cyclosporine, administered intraperitoneally, for 10 days. For group V, in addition to standard grafting, unmodified fibroblasts (not treated with cyclosporine) were implanted between the graft and the recipient bed. For group VI, the same procedures as for group V were performed and then rats were treated with 10 mg/kg cyclosporine, administered intraperitoneally, for 10 days. The rejection process was observed macroscopically, and statistical significance was determined with the Mann-Whitney test (p < 0.01). Graft survival times were significantly prolonged in groups III and IV, compared with groups I, II, V, and VI (p < 0.001). No difference between groups III and IV was observed. The novel finding of this investigation is that xenogeneic skin graft survival times could be prolonged with microimplantation of cyclosporine-treated cultured fibro-blasts.     

Severe embryolethality of artesunate related to pharmacokinetics following intravenous and intramuscular doses in pregnant rats

Artesunate (AS), a rapid, effective, and safe antimalarial drug, has been used for the treatment of malaria for decades. However, severe embryolethality was found for injectable AS in pregnant animals. In the present study, pregnant rats were selected and dosed with AS (GMP product) intravenously (IV) and intramuscularly (IM) at varied doses daily for 13 days from gestation day (GD) 6 to 18. In addition, a toxic dose of 1.2 mg/kg/day was subsequently tested in the GD 6–10, GD 11–15, and GD 16–20 periods of rat pregnancy. A pharmacokinetic study was also conducted to evaluate the bioavailability of AS following the IM administrations. Results showed that no significant adverse effects were found in maternal rats. All of the fetuses were either damaged or reabsorbed by placentas in treated pregnant rats, but doses did not show an adverse effect at 0.4 and 0.5 mg/kg after IV and IM administrations, respectively. The survival rate of fetuses is dose-dependent and the 50% fetus re-absorption doses (FRD₅₀) were 0.61 and 0.60 mg/kg following the IV and IM, respectively. The most drug-sensitive period, showing severe embryotoxicity, was between GD 11 and 15 for injectable AS. When calculated with total concentrations of AS and dihydroartemisinin, an active metabolite of AS, the bioavailability of 97.8% after intramuscular injection was fulfilled to a bioequivalence of that in intravenous treatment. The fact that injectable AS exhibited severe embryolethality after both IV and IM injections seems related to their comparable pharmacokinetic profiles that indicate high peak concentrations in pregnant animals. Birth Defects Res (Part B) 86:385–393, 2009. Published 2009 Wiley-Liss, Inc.     

Dose response kinetics of serum vitellogenin, liver DNA, RNA, protein and lipid after induction by estradiol-17 beta in male flounders (Platichthys flesus L.).

1. Male flounders receiving 100 micrograms estradiol each second day were fully induced to vitellogenin synthesis within 11 days, while fishes given 5 micrograms doses continued to accumulate vitellogenin in the serum at a progressive rate through 17 days. 2. Liver DNA per unit fish remained constant, while RNA per unit fish in flounders given 100 and 5 micrograms doses attained values 80 and 25% respectively, above the values found in control animals. 3. Liver RNA per unit DNA increased at maximal rate within 6 days in fishes receiving 100 micrograms doses. RNA synthesis continued at a progressive rate through 17 days in fishes given 5 micrograms doses of estradiol. 4. Liver protein per unit DNA elevated at a plateau 60% above control within 6 days with 100 micrograms doses. Doses of 5 micrograms had only little effect on liver protein. 5. Estradiol had a lipogenic effect on the liver. Cellular lipid rose 120 and 60% above control after treatment with 100 and 5 micrograms respectively. 6. Liver dry weight per unit DNA increased 60 and 55% above control with 100 and 5 micrograms doses respectively. Cellular hypertrophy in fishes receiving the smaller dose was primarily associated with an increase in lipid concentration, while protein and lipid contributed almost equally to cellular growth in fishes receiving the high dose.     

Assays to predict the genotoxicity of the chromosomal mutagen etoposide -- focussing on the best assay

The topoisomerase II inhibitor etoposide is used routinely to treat a variety of cancers in patients of all ages. As a result of its extensive use in the clinic and its association with secondary malignancies it has become a compound of great interest with regard to its genotoxic activity in vivo. This paper describes a series of assays that were employed to determine the in vivo genotoxicity of etoposide in a murine model system. The alkaline comet assay detected DNA damage in the bone marrow mononuclear compartment over the dose range of 10--100mg/kg and was associated with a large and dose dependent rise in the proportion of cells with severely damaged DNA. In contrast, the bone marrow micronucleus assay was found to be sensitive to genotoxic damage between the doses of 0.1--1mg/kg without any corresponding increases in cytotoxicity. An increase in the mutant frequency was undetectable at the Hprt locus at administered doses of 1 and 10mg/kg of etoposide, however, an increase in the mutant frequency was seen at the Aprt locus at these doses. We conclude that the BMMN assay is a good short-term predictor of the clastogenicity of etoposide at doses that do not result in cytotoxic activity, giving an indication of potential mutagenic effects. Moreover, the detection of mutants at the Aprt locus gives an indication of the potential of etoposide to cause chromosomal mutations that may lead to secondary malignancy.     

Differential recovery of intestine, bone marrow, and thymus of rats with solid tumors following 5-fluorouracil administration.

Time relationships for recovery of several host organs from toxic effects of 5-fluorouracil were determined in ACI rats bearing Morris hepatoma 3924A. A single injection of 150 mg/kg body weight 5-fluorouracil (the LD10) resulted in loss of 90% of the tibial bone marrow, 60% of the intestinal mucosa, and 90% of the thymus as measured by total DNA content of the organs. Organ DNA contents following 150 mg/kg of the drug were minimal on day 3 for intestine and on day 5 for marrow and thymus. A return to pretreatment or higher levels of DNA was observed by day 4 for intestine, day 11 for tibial marrow, and day 19 for thymus. Incorporation of 3H-deoxyuridine into host organ DNA after 150 mg/kg 5-fluorouracil was inhibited 36 hrs for intestine, 3 days for thymus, and 5 days for tibial bone marrow. Inhibition of 3H-deoxyuridine incorporation into DNA was similar for 50, 100, and 150 mg/kg doses both in tumor and in host organs, but recovery of 3H-deoxyuridine incorporation and DNA content of host organs began later with the higher doses of 5-fluorouracil. Maximal incorporation of 3H-deoxyuridine into DNA was observed on day 4 for intestine, day 8 for marrow, and day 9 for thymus after treatment with 150 mg/kg 5-fluorouracil. Animal lethality following the second of two 150 mg/kg injections of 5-fluorouracil was related to the extent of recovery of intestinal mucosa and bone marrow at the time of the second injection. Survival decreased to 0% for normal rats when the interval between injections was 3-4 days, improved at 5 days and was 100% when the interval was 10-11 days.     

Antitumor effects of the molecule-downsized immunoconjugate composed of lidamycin and Fab′fragment of monoclonal antibody directed against type IV collagenase

Type IV collagenase plays an important role in tumor invasion and metastasis through cleaving type IV collagen in the basement membrane and extracellular matrix. In this study a molecule-downsized immunoconjugate (Fab'-LDM) was constructed by linking lidamycin (LDM), a highly potent antitumor antibiotic, to the Fab' fragment of a monoclonal antibody directed against type IV collagenase and its antitumor effect was investigated. As assayed in 10% SDS-PAGE gel, the molecular weight of Fab'-LDM conjugate was 65 kD with a 1 : 1 molecular ratio of Fab' and LDM. The Fab'-LDM conjugate maintained most part of the immunoreactivity of Fab' fragment to both type IV collagense and mouse hepatoma 22 cells by ELISA. By MTT assay, Fab'-LDM conjugate showed more potent cytotoxicity to hepatoma 22 cells than that of LDM. Administered intravenously, Fab'-LDM conjugate proved to be more effective against the growth of subcutaneously transplanted hepatoma 22 in mice than free LDM in two experiment settings. In Experiment     

Effect of chronic D-fenfluramine administration on rat hypothalamic serotonin levels and release

D-fenfluramine, an anorectic agent in rats and man, was administered daily at doses 1.25, 2.5, 5 or 10 mg/kg/day for 10 days, and sacrificed 6 days later. Hypothalamic serotonin (5-HT) levels were unchanged in rats receiving 1.25-5 mg/kg/day of d-fenfluramine but reduced by 22% (p less than 0.01) at the highest drug dose (10 mg/kg/day); hypothalamic 5-hydroxyindole acetic acid (5-HIAA) levels were reduced by 15% (p less than 0.05) or 28% (p less than 0.01) in rats receiving 5 or 10 mg/kg/day of the drug, respectively. Hypothalamic slices prepared from rats which were previously treated with any of the drug doses spontaneously released endogenous 5-HT at rates that did not differ from those of vehicle-treated rats. 5-HT released with electrical field-stimulation was unaffected by prior d-fenfluramine treatment at doses of 1.25-5 mg/kg/day, and was reduced by 20% (p less than 0.05) from slices prepared from rats which received 10 mg/kg/day. 5-HIAA efflux was also attenuated by the highest drug dose. These data indicate that chronic administration to rats of customary anorectic doses of d-fenfluramine (i.e. 0.06-1.25 mg/kg) fail to cause long-lasting reductions in brain 5-HT release.     

Low dose intravenous minocycline is neuroprotective after middle cerebral artery occlusion-reperfusion in rats

Background

Minocycline, a semi-synthetic tetracycline antibiotic, is an effective neuroprotective agent in animal models of cerebral ischemia when given in high doses intraperitoneally. The aim of this study was to determine if minocycline was effective at reducing infarct size in a Temporary Middle Cerebral Artery Occlusion model (TMCAO) when given at lower intravenous (IV) doses that correspond to human clinical exposure regimens.

Methods

Rats underwent 90 minutes of TMCAO. Minocycline or saline placebo was administered IV starting at 4, 5, or 6 hours post TMCAO. Infarct volume and neurofunctional tests were carried out at 24 hr after TMCAO using 2,3,5-triphenyltetrazolium chloride (TTC) brain staining and Neurological Score evaluation. Pharmacokinetic studies and hemodynamic monitoring were performed on minocycline-treated rats.

Results

Minocycline at doses of 3 mg/kg and 10 mg/kg IV was effective at reducing infarct size when administered at 4 hours post TMCAO. At doses of 3 mg/kg, minocycline reduced infarct size by 42% while 10 mg/kg reduced infarct size by 56%. Minocycline at a dose of 10 mg/kg significantly reduced infarct size at 5 hours by 40% and the 3 mg/kg dose significantly reduced infarct size by 34%. With a 6 hour time window there was a non-significant trend in infarct reduction. There was a significant difference in neurological scores favoring minocycline in both the 3 mg/kg and 10 mg/kg doses at 4 hours and at the 10 mg/kg dose at 5 hours. Minocycline did not significantly affect hemodynamic and physiological variables. A 3 mg/kg IV dose of minocycline resulted in serum levels similar to that achieved in humans after a standard 200 mg dose.

Conclusions

The neuroprotective action of minocycline at clinically suitable dosing regimens and at a therapeutic time window of at least 4–5 hours merits consideration of phase I trials in humans in view of developing this drug for treatment of stroke.

     

Stage-specific inhibition of deoxyribonucleic acid synthesis and induction of apoptosis by antracyclines in cultured rat spermatogenic cells

A rapid in vitro method has been developed to detect early effects of cytostatic drugs on rat spermatogenesis. The induction of programmed cell death (apoptosis) and changes in DNA synthesis induced by doxorubicin and idarubicin were measured in specific stages of the cycle of seminiferous epithelium including mitotic (stage V) and meiotic (stage VIII-IX) S-phase cells. The model was used to investigate the protective effect of an organic thiophosphate, amifostine, against the toxicity of antracyclines. Premitotic DNA synthesis was found to be more sensitive than premeiotic DNA synthesis to antracyclines. Idarubicin was more toxic than doxorubicin to germ cells in inducing apoptosis and suppressing DNA synthesis. Amifostine had no protective effect against doxorubicin- or idarubicin-induced inhibition of DNA synthesis. In contrast, a significant stimulation of DNA synthesis in premitotic cells by amifostine was found, suggesting that this compound may have a stimulative effect on spermatogenic stem cells. These data show that stage-specific dissection of the seminiferous tubules and their in vitro exposure to predetermined doses of drugs may give us a unique possibility to detect drug action and protection against the cytotoxicity of antineoplastic agents at the cellular level of the spermatogenic cycle.     

Female to female stimulation of ovarian activity in the ewe

The ovulatory and oestrus responses of seasonally anovulatory ewes to the presence of ewes with synchronised oestrus was evaluated. The experiment was carried between 4 June and 1 July when the ewes were in seasonal anoestrus. Two hundred adult Suffolk and Dorset ewes were used. The animals were randomly divided into five groups balanced according to breed: Group I (treated) consisted of 25 ewes induced to cycle by the treatment for 10 days with vaginal sponges containing 40 mg of fluorogestone acetate and an injection of 200 IU of pregnant mares' serum gonadotropin (PMSG) at the time of sponge removal. Group II (mixed) consisted of 25 untreated ewes housed in the same pen as the treated ewes throughout the experiment. Groups III, IV and V each consisted of 50 untreated ewes located in adjacent pens progressively more distant from the pen which contained the treated animals. The ewes in Group III had contact with the treated animals through the fence, while those in Groups IV and V were separated from the treated ewes by one and two pens respectively. Day 0 of the experiment was defined as the day in which the sponges were removed from the treated ewes. Blood samples for progesterone determination were obtained from 25 animals from each group on days 6, 10 and 13. Oestrus was detected twice a day using vasectomised rams introduced to each pen for 15 min in the morning and 15 min in the evening. As expected, the proportion of ewes with luteal activity was higher (P < 0.01) in the treated group than in the other four groups on days 6, 10 and 13. By day 13 progesterone levels were elevated in 87.5%, 52%, 37.5%, 32% and 13% of the ewes sampled in Groups I, II, III, IV and V respectively. There was a direct relationship between the proportion of non-treated ewes with ovarian activity and the intensity of contact with the treated ewes, being maximal in the ewes that remained mixed with the synchronised animals, and lowest in the ewes located in the most distant pen. The proportion of ewes that showed oestrus during the first 14 days after sponge removal was significantly higher in the treated (92%) and mixed (40%) groups than in Groups III (10%), IV (8%) and V (4%). It is concluded that the presence of a large number of ewes in oestrus can stimulate ovarian activity in seasonally anoestrous ewes. This female to female stimulation could be mediated by olfactory, visual and/or auditory stimuli.     

In vivo genotoxicity of nitrates and nitrites in germ cells of male mice. II. Unscheduled DNA synthesis and sperm abnormality after treatment of spermatids

Effects of both sodium nitrate (doses of 600 and 1200 mg/kg/day for 3 days) and sodium nitrite (doses of 60 and 120 mg/kg/day for 3 days) on spermatids of mice were investigated by measuring unscheduled DNA synthesis (UDS) 17 days after the end of treatment, and sperm-head abnormality 11 and 17 days after the end of treatment. Neither chemical induced the UDS response in early to mid spermatids (17 days). The only positive result in the sperm-head abnormality test was obtained for the dose of 120 mg/kg/day of sodium nitrite both at 11 and 17 days after treatment. The results presented are in accordance with those of our earlier experiments with the same chemicals, suggesting their nonmutagenic action on the tested germ-cell stages of male mice.     

Procollagen production by rat hepatocytes in primary culture

Hepatocytes were obtained from rat liver and maintained in primary culture for periods up to 14 days. Collagen synthesis was maximal after 3–5 days and declined thereafter. The rate of collagen production was appox. one-tenth that observed by the rat skin fibroblasts of the same animals after 3–5 passages. Type I procollagen, the major macromolecular collagenous species, was identified as a 450 000 dalton molecule which was converted to 120 000 dalton, denatured, reduced procollagen chains. Prior pepsin digestion of the native procollagen released 95 000 dalton collagen chains identified as α1(I) and α2(I) by co-migration with carrier rat skin type I collagen chains. The production of type III procollagen was also tentatively identified by DEAE-cellulose chromatography. This material was isolated and identified with type-specific antibodies developed against the amino-terminal extension peptide of bovine skin type III procollagen. The relative distribution of type I:type III procollagen was estimated at 7:3 similar to the ratio previously found in whole rat liver. No evidence of type IV or type V procollagen biosynthesis was observed. These results suggest that rat hepatocytes in primary culture are capable of interstitial type I and type III collagen biosynthesis in a ratio similar to that found in their parent hepatic tissue in situ. They also suggest that the less abundant type IV (basement membrane-associated) or type V are nor major collagenous products of these cells.     

Pharmacodynamics and pharmacokinetics of the insulin-mimetic agent vanadyl acetylacetonate in non-diabetic and diabetic rats

The objectives of this study were to evaluate the pharmacodynamics and pharmacokinetics of vanadyl acetylacetonate (VAC) in rats. Pharmacodynamic study was carried out using non-diabetic and diabetic rats by subcutaneous (s.c.) and intragastric (i.g.) administrations at single dose or multiple doses. Pharmacokinetic study was performed using non-diabetic rats. Results showed that VAC resulted in a significant decrease of plasma glucose levels in diabetic rats in all dosing levels, and nearly restored hyperglycemic values to normal values after s.c. injection at a single dose of 2, 4, and 8 mg vanadium (V)/kg, or after i.g. administration at multiple doses of 3 and 6 mg V/kg once daily for seven consecutive days, respectively. The VAC could be rapidly absorbed and T(max) values ranged from 0.9 +/- 0.3 h for s.c. injection to 3.0 +/- 0.9 h for i.g. administration. The average absolute bioavailabilities for i.g. administrations at a single dose of 3, 6, and 10 mg V/kg were 34.7%, 28.1%, and 22.8%, respectively. After i.g. administration at a single dose of 10 mg V/kg, the average elimination half-lives obtained from non-diabetic rats were very long ranging from 144.7 +/- 8.7 h in plasma to 657.3 +/- 34.8 h in femur tissue. In conclusion, VAC widely distributed in various tissues and accumulated more in the femur tissue. The time to reach maximal vanadium level after s.c. injection or i.g. administration was not coincident with the time to reach maximal hypoglycemic effect. The accumulated vanadium in bone, kidney or other tissues may gradually release and exert a longer action. In present dosing levels and administration routes, VAC was effective for lowering plasma glucose levels in diabetic rats and could reverse the higher triglyceride and cholesterol levels to the normal ranges. VAC did not influence the insulin levels in plasma and not cause obvious toxic signs like diarrhea.     

Different diets and amino acid supplementation do not affect the voluntary consumption of ethanol by rats

The effects of four different diets (control diet: 19.5% protein, 60.5% carbohydrate, 10% fat; diet I: 65% protein, 10% carbohydrate, 10% fat; diet II: 5% protein, 76% carbohydrate, 10% fat; diet III: 20% protein, 69% carbohydrate, 1% fat; diet IV: 69% protein, 15% carbohydrate, 1% fat) and supplementation with 3 amino acids (tryptophan: 150 mg/kg/d; arginine: 400 mg/kg/d; taurine: 380 mg/kg/d) on the voluntary consumption of ethanol were investigated in rats using the 2 bottle method. First, rats received the control diet and diets I, II, III and IV for 20 days with a choice of ethanol for the last 6 days only. Ethanol consumption was similar in all dietary groups. Second, rats received the control diet for 8 days followed by diets I, II and IV for another 8 days. Ethanol was offered throughout both periods. The switch to the special diets did not affect ethanol consumption. Third, rats received a control diet with arginine, tryptophan or taurine added to the drinking fluids for 16 days with a choice of ethanol for the last 5 days; thereafter supplementation stopped but the ethanol choice remained. No difference in the voluntary intake of ethanol was noted but ethanol consumption fell after cessation of arginine supplementation. In conclusion, diets differing greatly in their composition or supplementation with these 3 amino acids did not affect the voluntary choice of ethanol by rats in a significant manner.     

Dose-dependent increase or decrease of somatic intrachromosomal recombination produced by etoposide

Chromosomal inversions and deletions can occur via somatic intrachromosomal recombination (SICR), a mechanism known to be important in mutagenesis and carcinogenesis. Here, we demonstrate a dose-dependent increase or decrease in SICR inversion frequency both in vivo and in vitro after treatment with etoposide, using the pKZ1 mouse mutagenesis model. pKZ1 mice received a single intraperitoneal injection of etoposide dose ranging from 0.0005 to 50mg/kg. Animals were sacrificed 3 days after treatment and the spleen was analysed for SICR. A significant 1.4-3.1-fold induction of SICR inversion events was detected in pKZ1 mice after treatment with etoposide doses ranging from 0.05 to 50 mg/kg etoposide. However, inversion frequencies after treatment with 0.0005 and 0.005 mg/kg etoposide decreased significantly to 0.67 and 0.43 of the levels observed in control animals, respectively. A pKZ1 mouse hybridoma cell line was exposed to etoposide (1-1000 nM) and a similar pattern of SICR response to that detected in vivo was observed. A significant 2.3-4.6-fold induction of SICR inversions was observed in pKZ1 cells treated with 100 and 1000 nM etoposide. Inversion frequencies after treatment with 1 and 10nM etoposide decreased significantly to 0.31 and 0.5 of the level observed in control cell lines. Our in vitro studies complement our in vivo studies and exclude a kinetic phenomenon as the responsible mechanism of reduction in SICR in response to low dose etoposide. Determination of the exact mechanism and significance of recombination suppression at low doses of etoposide treatment requires further investigation.     

Antiestrogenic and antitumor properties of the new triphenylethylene derivative toremifene in the rat

The effects of toremifene, a new triphenylethylene derivative, on the uterus and DMBA-induced mammary tumors in rats were compared to tamoxifen. The ability of toremifene to compete with [3H]estradiol for cytoplasmic estrogen receptor from rat uterus was similar to tamoxifen, the IC50 being 26 and 23 microM respectively. In immature intact rats the two compounds, administered orally for three consecutive days, had similar intrinsic partial estrogenic efficacy, at 50 mg/kg, about 40% of that of estradiol benzoate (EB). However, at doses less than or equal to 10 mg/kg, the estrogenic effect of toremifene was seen at doses about 40 times higher than that of tamoxifen. The two compounds, administered together with a standard dose of EB, expressed the same maximal antiestrogenic efficacy (about 65% inhibition) at 50 mg/kg. However, the minimal effective antiestrogenic dose of toremifene was about 10 times that of tamoxifen and the ratio between antiestrogenic/estrogenic properties was favourable to toremifene. The duration of the antiestrogenic (antiuterotrophic) effect of a single oral dose (10 mg/kg) of the two compounds proved similar: at least 4 days in intact rats and 3 days in ovariectomized rats. In DMBA-induced tumor bearing rats toremifene was administered p.o., 6 times/week for 4 weeks at 0.08, 0.4, 2, 10 and 50 mg/kg. It was effective at the doses of 2, 10 and 50 mg/kg, inducing 39, 35 and 46% tumor regressions. The activity of toremifene at the minimal effective dose of 2 mg/kg was then compared with that of tamoxifen given at the same dose level. The compounds had comparable activity (47 vs 44% tumor regressions).     

Use of an antiarrhythmic drug against acute selenium toxicity

ObjectiveSelenium is an essential trace element. But, selenium may have toxic effects in high doses. There are no proven antidotes or curative treatments for acut selenium toxicity. Treatment involves stopping the exposure and providing supportive care for symptoms. Therefore, it is necessary to find more effective substances in the treatment of selenium toxicity. The aim of this study was to increase the survival rate of animals by supporting the heart with amiodarone and to determine the effect of amiodarone on the pathological, hematological and biochemical parameters in acute selenium intoxication.Methods64 Wistar-Albino rats were divided into four groups. Group I was given only distilled water, Group II was given 18 mg/kg dose of amiodarone, Group III was given 18 mg/kg amiodarone and 10 mg/kg sodium selenite and Group IV was given sodium selenite 10 mg/kg (LD₅₀ dose)orally.Results11 of the 16 animals in Group IV died within the first 48 h of drug administration. However, no deaths were observed in the rats in Group III. No hematological changes were observed. Biochemically, CK, CK-MB and LDH levels of Group IV were higher than the other groups on both the 2nd and 10th days. In Groups II and III, this serum level decreased, and vitamin B12 levels increased. In macroscopic inspections of the organs of Groups III and IV, slight paleness was detected. Histopathologically, degenerative changes in tissue were observed, especially in Group IV.ConclusionThis study shows that amiodarone application has a reducing effect on selenium toxicity. This was because amiodarone protected the heart by reducing CK and CK-MB levels and increased vitamin B12 levels, which play a role in the synthesis of S-adenosyl methionine that converts selenium into a nontoxic form.     

Relative sensitivity of 32P-postlabelling of DNA and the autoradiographic UDS assay in the liver of rats exposed to 2-acetylaminofluorene (2AAF)

Groups of male Alderley Park rats were dosed concomitantly with 2-acetylaminofluorene (2AAF) by gavage at doses between 0.01 mg/kg and 40 mg/kg, and livers sampled 2-72 h later. The liver of one group of animals was perfused to yield hepatocytes which were assayed in vitro for unscheduled DNA synthesis (UDS) via incorporation of tritiated thymidine and autoradiography. DNA was extracted from the livers of the other group and DNA adduct levels determined using the 32P-postlabelling technique. The major C-8 2-aminofluorene/guanosine adduct and 3 minor adducts were quantitated, enabling the relative sensitivity of the 2 techniques to be compared. A dose- and time-related UDS response was observed, which, at the most sensitive time-point (12 h) enabled DNA repair to be discerned at a dose level of 0.1-1 mg/kg of 2AAF, a response classified as formally positive at 5 mg/kg 2AAF. Only the C-8 adduct, as determined by 32P-postlabelling, was discernible at 0.01 mg/kg of 2AAF, although other adducts were visible on autoradiograms at higher dose levels. It is concluded that as part of a well-defined dose response, UDS can be discerned with confidence for doses of 2AAF between approximately 0.1 and 5 mg/kg, and DNA adducts for doses of 2AAF between approximately 0.01 and 1 mg/kg. Discernible UDS for 2AAF in the rat liver is apparent at approximately 13 DNA (total) adducts/10(8) nucleotides, or approximately 8 DNA (C-8) adducts/10(8) nucleotides. The presumed C-8 2-acetylaminofluorene/guanosine adduct, prepared by reaction of 2-acetoxy-2-acetylaminofluorene (2AAAF) with DNA, was a significant but unreliable marker of 2AAF/DNA adducts in the rat liver in vivo. DNA repair did not appear to remove DNA adducts selectively, and adducts remained in DNA when discernible DNA repair had ceased.