Effects of multiple dosing of phenacetin in the micronucleus test

As a part of the international cooperative study to identify the most sensitive regimen in the micronucleus test, phenacetin was given i.p. to male CD-1 mice at doses of 37.5, 75, 150, 300, 400, and 600 mg/kg once, twice, thrice or four times and the bone marrow cells were harvested 24 h after the final dosing. Positive responses were seen at 600 mg/kg after single and triple dosing and at 400 and 600 mg/kg after double dosing. No dose level gave a positive response after quadruple dosing. A repeated-dosing effect was detected at double and triple dosing. Although triple dosing gave the highest magnitude of micronuclei at 600 mg/kg, double dosing showed a sufficient sensitivity and was more convenient from the viewpoint of selecting a suitable test dose and carrying out the micronucleus test.     

The evaluation of a multiple dosing protocol for the mouse bone-marrow micronucleus assay using benzidine and 2,6-xylidine

Male ICR mice were treated with 1, 2 or 3 daily doses of either benzidine or 2,6-xylidine. Groups of 5 animals were sacrificed 24 h after the last dose and the bone marrow examined for micronuclei. Benzidine was given at dose levels of 40 and 200 mg/kg and 2,6-xylidine was given at dose levels of 75 and 375 mg/kg. These doses represent 10 and 50% of the respective median lethal doses. Benzidine produced a significant (p less than 0.001) dose related increase in the incidence of micronucleated polychromatic erythrocytes (MPE), while 2,6-xylidine had no effect on the frequency of micronucleated cells. Statistical analyses of the data indicated that the incidence of MPE was independent of the number of doses administered prior to bone marrow harvest.     

Effect of multiple dosing on the analgesic action of diflunisal in rats

This investigation was designed to compare the analgesic effect of the initial dose of a repetitively dosed non-narcotic analgesic with the analgesic effect of a subsequent dose given 3 days later. To exclude gradual drug accumulation as a variable, the first ("loading") dose was larger than the maintenance doses. Male Sprague-Dawley rats received 100 mg/kg diflunisal i.v. as the first dose and 70 or 75 mg/kg every 12 hours thereafter. The analgesic effect of the first and seventh doses was determined as the pain threshold (voltage) upon electrical stimulation of the tail every 15 to 30 minutes from the third to the ninth hour after dosing. Blood samples for drug assay were obtained at 3 and 9 hours. A control group received injections of solvent for 6 doses and 100 mg/kg diflunisal as the seventh dose. There were no statistically significant differences between the area under the total or free (unbound) drug concentration versus time curves of the first and seventh dose but the average analgesic effect (area under the voltage increase versus time) of the seventh dose was only 28 percent that of the first dose. The areas under the drug concentration and analgesic effect versus time curves of the diflunisal dose given as the seventh injection to the control rats were similar to those produced by the first dose given to the multiple dosed rats. The results of this investigation show that the analgesic effect of a non-narcotic drug decreases substantially during repeated dosing in an animal model of experimental pain. This change in pharmacologic response has the characteristics of functional rather than pharmacokinetic tolerance in that there was no change in the drug concentration profile with time and no effect of the manipulations as such (i.e., repeated pain threshold determinations and blood withdrawals) on diflunisal-induced analgesia. These observations may have important implications for the evaluation and use of non-narcotic analgesics in the management of clinical pain.     

Evaluation of micronucleus induction in a Chilean population environmentally exposed to arsenic

In the present study we have evaluated whether or not environmental exposure to arsenic in ground drinking-water results in a significant increase in the frequency of micronuclei (MN) in peripheral blood lymphocytes. Thus, 106 individuals from the Antofagasta region (North Chile), together with 111 individuals from the area of Concepción, were used in this investigation. In the Antofagasta area, arsenic levels in drinking-water as high as 0.750 mg/L were measured. In Concepción, located about 2500 km towards the south and used as reference area, arsenic levels in tap water were as low as 0.002 mg/L. The total content of arsenic in fingernails was determined as a biomarker of individual exposure. The cytogenetic results obtained in this study indicate that in the exposed group the overall frequency of binucleated micronucleated cells (BNMN) is higher than in the reference group, the difference being statistically significant. In addition, no differences were found between the exposed and the reference groups, regarding the cytokinesis-block proliferation index (CBPI). No association was observed between BNMN and arsenic content in water or arsenic in fingernails. On the other hand, when the exposed group was divided according to their Atacameno or Caucasian ethnicity, no significant differences were observed between them. In addition, as usually found in other human biomonitoring studies, sex and age are factors that modulate the frequency of MN in both exposed and reference populations.     

Effects of chemically- and environmentally-induced hypothermia on micronucleus induction in rats

We investigated micronucleus induction in rats treated with chlorpromazine and reserpine, drugs that induce hypothermia. We administered chlorpromazine (31.3--250mg/kg) or reserpine (500--2000 mg/kg) intraperitoneally and measured temperature rectally. Chlorpromazine at 62.5-250mg/kg and reserpine at all doses significantly decreased rectal temperature, although the hypothermic response was weaker than previously reported in mice. Only chlorpromazine at 250mg/kg decreased rectal temperature transiently to <33 degrees C for 20h and induced a statistically significant increase in micronucleated polychromatic erythrocyte frequency. When rats treated with reserpine at 500mg/kg were exposed to an environmental temperature of 16 degrees C for 6, 12, or 24h to keep their body temperature under 33 degrees C, only the 24h treatment group significantly induced micronuclei. In addition, relatively large micronuclei (diameter of micronucleus> or = 1/4 diameter of cytoplasm) accounted for 33.0% of the induced micronuclei, suggesting that hypothermia affected the mitotic apparatus. The hypothermic response to chlorpromazine and reserpine was weaker in rats than in mice, and it was correspondingly more difficult to induce micronuclei in rats with those drugs.     

Effect of a pre-exercise energy supplement on the acute hormonal response to resistance exercise

The effect of a pre-exercise energy sport drink on the acute hormonal response to resistance exercise was examined in eight experienced resistance trained men. Subjects were randomly provided either a placebo (P: maltodextrin) or the supplement (S: combination of branched chain amino acids, creatine, taurine, caffeine, and glucuronolactone). Subjects performed 6 sets of no more than 10 repetitions of the squat exercise at 75% of their 1 repetition maximum (1RM) with 2 minutes of rest between sets. Blood draws occurred at baseline pre-exercise, immediately post- (IP), 15 minutes post- (15P), and 30-minutes post (30P) exercise for measurement of serum growth hormone, total and free testosterone, cortisol, and insulin concentrations. Although significant differences were seen only at set 5, the total number of repetitions and training volume tended (p = 0.08) to be higher with S compared to P. Serum growth hormone and insulin concentrations were significantly higher at 15P and IP, respectively, in S compared to P. Results suggest that a pre-exercise energy S consumed 10 minutes before resistance exercise can enhance acute exercise performance by increasing the number of repetitions performed and the total volume of exercise. The enhanced exercise performance resulted in a significantly greater increase in both growth hormone and insulin concentrations, indicating an augmented anabolic hormone response to this pre-exercise S.     

Effects of heavy metal contamination of soils on micronucleus induction in Tradescantia and on microbial enzyme activities: a comparative investigation

The aim of this study was to investigate correlation between genotoxic effects and changes of microbial parameters caused by metal contamination in soils. In total, 20 soils from nine locations were examined; metal contents and physicochemical soil parameters were measured with standard methods. In general, a pronounced induction of the frequency of micronuclei (MN) in the Tradescantia micronucleus (Trad-MN) assay was seen with increasing metal concentration in soils from identical locations. However, no correlations were found between metal contents and genotoxicity of soils from different locations. These discrepancies are probably due to differences of the physicochemical characteristics of the samples. Also, the microbial parameters depended on the metal content in soils from identical sampling locations. Inconsistent responses of the individual enzymes were seen in soils from different locations, indicating that it is not possible to define a specific marker enzyme for metal contamination. The most sensitive microbial parameters were dehydrogenase and arylsulfatase activity, biomass C, and biomass N. Statistical analyses showed an overall correlation between genotoxicity in Tradescantia on the one hand and dehydrogenase activity, biomass C, and the metabolic quotient on the other hand. In conclusion, the results of the present study show that the Trad-MN assay is suitable for the detection of genotoxic effects of metal contamination in soils and furthermore, that the DNA-damaging potential of soils from different origin cannot be predicted on the basis of chemical analyses of their metal concentrations.     

Japanese encephalitis virus differentially modulates the induction of multiple pro-inflammatory mediators in human astrocytoma and astroglioma cell-lines

Astrocytes become activated in response to many CNS pathologies. The process of astrocyte activation remains rather enigmatic and results in so-called reactive gliosis, a reaction with specific structural and functional characteristics. Astrocytes play a vital role in regulating aspects of inflammation and in the homeostatic maintenance of the CNS. However, the responses of different human astroglial cell-lines in viral encephalitis mediated inflammation are not well documented. We have shown that Japanese encephalitis virus (JEV) infection causes morphological and functional changes in astrocytic cell-lines. We have demonstrated that besides reactive oxygen species (ROS) JEV infection differentially regulated the induction pattern of IL-6, IL-1 beta and IL-8. IP-10, MCP-1, MIG and RANTES secretions in different astroglial cell-lines. The expression of different proteins such as astrocyte-specific glial fibrillary acidic protein (GFAP), the glutamate aspartate transporter/essential amino acid transporter-1 (GLAST/EAAT-1), glutamate transporter-1/essential amino acid transporter-2 (GLT-1/EAAT-2), Ceruloplasmin and Thioredoxin (TRX) expression level also differ in different human astrocyte cell-lines following infection.     

Effect of Cimetidine and other antihistaminics on the elimination of aminopyrine,phenacetin and caffein

Cimetidine is widely prescribed for the treatment of peptic ulcer disease and has recently been shown to inhibit the metabolism of warfarin, antipyrine and diazepam. To further examine this phenomenon we investigated the effect of various doses of cimetidine and other related drugs on ¹⁴C-aminopyrine, ¹⁴C-phenacetin and ¹⁴C-caffeine breath tests. Cimetidine caused a dose-related inhibition of the metabolism of aminopyrine and caffeine but had no effect on the phenacetin breath test. Metiamide, H₁-antihistamines, phenothiazines and local anesthetics also inhibited the aminopyrine breath test. Cyproheptadine had no effect on either phenacetin or caffeine elimination. This study demonstrates a complex drug-drug interaction which may have widespread clinical implications.     

Effect of a cellulose-containing weight-loss supplement on gastric emptying and sensory functions

CM3, a highly cross-linked cellulose in capsule form, expands in the stomach to a size several fold of its original volume. It is purported to induce a prolonged feeling of satiation and a delay in gastric emptying, thus promoting weight loss. We examined whether CM3 delays gastric emptying (using the stable isotope (13)C-octanoic breath test) and whether it influences subjective feelings of appetite sensations (using visual analog scales, VASs). We performed a double-blind randomized placebo-controlled crossover trial in 19 moderately obese but otherwise healthy subjects (mean age 55 +/- 9 years, BMI 31.1 +/- 4.6 kg/m(2)). The subjects were treated with six capsules of CM3 or matching placebo 30 min before a standardized solid meal. Breath collection and VASs were performed over 4 h every 15 min and 30 min, respectively. Half-excretion time of (13)CO(2) in breath, indicating gastric emptying half time, was the primary outcome parameter. The study was powered to detect a change in gastric emptying of 20-30 min. Mean (13)CO(2) half-excretion time changed from 2.3 +/- 0.4 to 2.4 +/- 0.33 h (mean difference +6 min, 95% confidence interval (CI) -3 to +15 min; P = 0.17). Appetite sensations (hunger, satiation, fullness, prospective food consumption, desire to eat something sweet, salty, savory, or fatty) changed over time during the course of the postprandial phase but were not influenced by CM3 (repeated measures ANOVA). In obese subjects, acute administration of the weight-loss supplement CM3 does not delay gastric emptying and does not influence subjective appetite sensations.     

In vivo photochemical micronucleus induction due to certain quinolone antimicrobial agents in the skin of hairless mice

The skin micronucleus test combined with irradiation due to a sunlight simulator having a spectrum almost identical to solar irradiation was used as a novel in vivo testing method for detecting or comparing the photochemical chromosome damage of quinolone antibacterial agents (quinolones). Eight-week-old male SKH1 hairless mice were orally administered once lomefloxacin (LFLX), a strong in vitro photochemical clastogen, at 25 or 50 mg/kg, followed by light irradiation at 7.9–9.4 J/cm² of ultraviolet A (UVA). Animals were killed on Days 2, 3, 4, 5 or 8 (the dosing day was designated as Day 1), and the incidence of micronucleus in the epidermis was determined. As results, LFLX at either dose caused significant increases in the micronucleus frequency, which peaked on Day 4. These changes tended to return to the control level on Day 8. Then, the micronucleus induction potential of the quinolone derivatives levofloxacin (LVFX) and clinafloxacin (CLFX) at 10, 20 or 40 mg/kg was assessed on Day 4 under the same experimental conditions as for LFLX. Although LVFX was negative even at 40 mg/kg, CFLX dose-dependently induced significant increases in micronucleus frequency at all doses. The correlation of magnitude among the three quinolones in the skin micronucleus test with light irradiation was similar to that in our previous in vitro photochemical clastogenicity study. No significant increase in micronucleus frequency was observed in any of three quinolones employed without light irradiation. In conclusion, the experimental method presented here would be a useful tool for detecting in vivo photochemical chromosome damage and for research on photochemical carcinogenesis of chemicals.     

In vivo photochemical micronucleus induction due to certain quinolone antimicrobial agents in the skin of hairless mice

The skin micronucleus test combined with irradiation due to a sunlight simulator having a spectrum almost identical to solar irradiation was used as a novel in vivo testing method for detecting or comparing the photochemical chromosome damage of quinolone antibacterial agents (quinolones). Eight-week-old male SKH1 hairless mice were orally administered once lomefloxacin (LFLX), a strong in vitro photochemical clastogen, at 25 or 50 mg/kg, followed by light irradiation at 7.9-9.4J/cm2 of ultraviolet A (UVA). Animals were killed on Days 2, 3, 4, 5 or 8 (the dosing day was designated as Day 1), and the incidence of micronucleus in the epidermis was determined. As results, LFLX at either dose caused significant increases in the micronucleus frequency, which peaked on Day 4. These changes tended to return to the control level on Day 8. Then, the micronucleus induction potential of the quinolone derivatives levofloxacin (LVFX) and clinafloxacin (CLFX) at 10, 20 or 40 mg/kg was assessed on Day 4 under the same experimental conditions as for LFLX. Although LVFX was negative even at 40 mg/kg, CFLX dose-dependently induced significant increases in micronucleus frequency at all doses. The correlation of magnitude among the three quinolones in the skin micronucleus test with light irradiation was similar to that in our previous in vitro photochemical clastogenicity study. No significant increase in micronucleus frequency was observed in any of three quinolones employed without light irradiation. In conclusion, the experimental method presented here would be a useful tool for detecting in vivo photochemical chromosome damage and for research on photochemical carcinogenesis of chemicals.