Topical pimecrolimus lacks genotoxicity and cytotoxicity by means of micronucleus erythrocyte rodent assay

Topical pimecrolimus is an alternative treatment of atopic dermatitis. However, rare cases of malignancy have been reported with their use. This study was performed to investigate the possible geno- or cytotoxic effect in mouse bone marrow caused by systemic absorption of pimecrolimus 1% cream. In order to determine this, induction of micronucleated erythrocytes (MNE) in mouse peripheral blood was determined after the cutaneous application of three different doses, daily for 5 consecutive days. No differences were found in frequencies of polychromatic erythrocytes, MNE, and micronucleated polychromatic erythrocytes in the different groups of study. In conclusion, under described conditions, no geno- or cytotoxic effects were detected after the cutaneous application of pimecrolimus.     

Topical pimecrolimus lacks genotoxicity and cytotoxicity by means of micronucleus erythrocyte rodent assay

Topical pimecrolimus is an alternative treatment of atopic dermatitis. However, rare cases of malignancy have been reported with their use. This study was performed to investigate the possible geno- or cytotoxic effect in mouse bone marrow caused by systemic absorption of pimecrolimus 1% cream. In order to determine this, induction of micronucleated erythrocytes (MNE) in mouse peripheral blood was determined after the cutaneous application of three different doses, daily for 5 consecutive days. No differences were found in frequencies of polychromatic erythrocytes, MNE, and micronucleated polychromatic erythrocytes in the different groups of study. In conclusion, under described conditions, no geno- or cytotoxic effects were detected after the cutaneous application of pimecrolimus.     

The lack of mutagenicity of aryl 4-guanidinobenzoates in the transplacental micronucleus assay in mice.

Two acrosin inhibitors, 4'-methylumbelliferyl 4-guanidinobenzoate and 2'-carbomethoxyphenyl 4-guanidinobenzoate, were tested for mutagenicity in the transplacental micronucleus assay and the mouse bone marrow micronucleus assay. The compounds were administered intraperitoneally at doses of 125 mg/kg and 250 mg/kg to pregnant mice. Fetal peripheral blood and maternal bone marrow cells were examined at 36 h for the frequency of micronucleated polychromatic erythrocytes. Neither compound induced micronuclei in maternal or fetal tissues. The ratio of polychromatic erythrocytes to normochromatic erythrocytes was not affected by the drug treatments indicating that the compounds had no effect on the cell cycle or mitosis in these tissues and that they were not cytotoxic. Both compounds, which show promise as vaginal contraceptives, were not mutagenic in this study.     

Increased micronucleus frequency in rat bone marrow after acrylamide treatment

This study was performed to investigate whether acrylamide (AA), occured during cooking carbohydrate-rich foods at high temperature, increased the frequency of micronucleated polychromatic erythrocytes (MNPCEs) in rat bone marrow. For this purpose AA, dissolved in distilled water, was administered to 8-week old male Sprague Dawley rats at single oral doses of 0, 125, 150 or 175 mg/kg b.w. After 48 h from AA treatment, the bone marrow samples were analysed for the frequency of MNPCEs. The cytotoxic effect of AA on bone marrow was also tested by assessing polychromatic erythrocyte/normochromatic erythrocyte (PCE/NCE) ratio. It was found that all three doses applied significantly increased the frequency of MNPCEs and this increase was 3.75-fold in rats given the highest administered dose of AA. In addition AA decreased the PCE/NCE ratio, which is indicative of bone marrow cytotoxicity when compared to the control group. This study displayed that AA increased the formation of micronuclei in polychromatic erythrocytes (PCEs) of rat bone marrow and this increase might have resulted from administrating the high dose level of AA to rats by gavage instead of by i.p. injection.     

Application of pharmacokinetic/pharmacodynamic and stochastic modelling to 6-mercaptopurine micronucleus induction in mouse bone marrow erythrocytes

This study investigates the kinetics of bone marrow micronucleated polychromatic erythrocytes and some mechanistic aspects of micronuclei induction using mathematical models. Female mice were administered a single intraperitoneal injection of the purine antagonist 6-mercaptopurine at 50 mg kg(-1). The time course evolution of the drug concentrations in the plasma and the micronucleated polychromatic erythrocyte kinetic rate in bone marrow were observed. Two mathematical models were developed for this study. The first model was built from a simultaneous pharmacokinetic/pharmacodynamic approach, but was invalidated after comparing its predictions to experimental data. The second model was a stochastic model based on some biological hypotheses involved in micronuclei induction. This model predicted a wavy kinetic rate of micronucleated polychromatic erythrocytes that was confirmed by a second data set obtained from a specifically built experimental design. The biological hypotheses were then discussed. It turned out from this work that mathematical modelling could be used as a tool to explore the cellular mechanisms of toxicity of the compound: for instance, the assumptions that 6-mercaptopurine induced micronuclei mainly in cells entering the S phase, and not only during the last cell cycle but during one of the earlier cycles preceding the extrusion of the main nucleus, were confirmed. Moreover, the use of the stochastic model would help to schedule more accurately the bone marrow or blood harvesting times in the in vivo rodent micronucleus test.     

Modulation of radiation-induced organs toxicity by cremophor-el in experimental animals.

Pharmacological and cytogenetic evaluations of the protective effects of polyethoxylated castor oil cremophor-EL (cremophor) against hepato, renal and bone marrow toxicity induced by gamma irradiation in normal rats were carried out. A single dose of irradiation (6 Gy) caused hepatic and renal damage manifested biochemically as an elevation in levels of serum alanine and aspartate aminotransferase as well as an increase in blood urea. Cremophor administration at a dose level of 50 microl kg-1 intravenously 1 day before exposure to irradiation (6 Gy) protected the liver and kidney as indicated by the recovery of levels of hepatic aminotransferase, urea and lipid profiles to normal values. Gamma irradiation of male rats caused a decrease in reduced glutathione and an increase in the oxidized form in rat-liver homogenate. A highly significant increase in the incidence of micronucleated normochromatic erythrocytes and micronucleated polychromatic erythrocytes was observed after irradiation exposure. The induced genotoxicity in the bone marrow cells was corrected by pretreatment with cremophor. The findings of this study suggest that cremophor pretreatment can potentially be used clinically to prevent irradiation-induced hepato, renal and bone marrow toxicity without interference with its cytotoxic activity.     

In vivo micronucleus test in the assessment of cytogenotoxicity of landfill leachates in three animal models from various ecological habitats

The in vivo micronucleus (MN) test, a standard test for the genotoxicity screening of xenobiotics, was used to evaluate the cytotoxic and genotoxic activities of landfill leachates in Clarias gariepinus, Coturnix coturnix japonica and Rattus norvegicus. These organisms were exposed to various sub-lethal concentrations (1–50 %) of Olusosun and Aba Eku landfill leachates. At post exposure, peripheral erythrocytes from catfish and quail, and bone marrow cells of quail and rat were subjected to MN analysis following standard protocols. The leachates induced significant increase in MN formation and total nuclear abnormalities (NAs) in the peripheral erythrocytes of catfish and quail. NAs occurred in the order; BN > BL > LB > NT in the catfish and BN > BudN > TLN > TN in quail. There was significant increase in MN formation in the bone marrow cells of quail, and micronucleated polychromatic erythrocytes and micronucleated normochromatic erythrocytes formation in the bone marrow of rats. The concentration dependent significant (p < 0.05) decrease in the PCE/NCE ratio in the bone marrow of the leachate treated rats suggest alterations in the bone marrow cell proliferation, leading to the suppression of immature erythrocytes (PCE). MN induction showed positive corrections with leachate concentrations in the test organisms; and it increased with exposure duration in the catfish. Indiscriminate disposal of solid waste generates leachates containing multiple xenobiotics that are capable of increasing genomic instability among vertebrates inhabiting various ecological habitats.     

Methylphenidate lacks genotoxic effects in mouse peripheral blood erythrocytes

Methylphenidate (MPH; Ritalin?; Novartis Pharmaceuticals, Inc., Basel, Switzerland) has been prescribed to treat attention deficit/hyperactivity disorder (ADHD) since its approval by the U.S. Food and Drug Administration over 50 years ago. Due to concerns that MPH might induce cytogenetic alterations in children, treatment with this drug has been a controversial issue. In the present study, we assessed the frequency of micronucleated erythrocytes (MNEs), micronucleated polychromatic erythrocytes (MNPCEs), and polychromatic erythrocytes (PCEs) in peripheral blood samples from mice treated with three different doses of MPH (30, 60, or 125 mg/kg). We found no evidence of increased MNEs or MNPCEs, nor did PCEs decline. These results add to the accumulating evidence that MPH does not induce genotoxic or cytotoxic damage.     

Methylphenidate lacks genotoxic effects in mouse peripheral blood erythrocytes

Methylphenidate (MPH; Ritalin^®; Novartis Pharmaceuticals, Inc., Basel, Switzerland) has been prescribed to treat attention deficit/hyperactivity disorder (ADHD) since its approval by the U.S. Food and Drug Administration over 50 years ago. Due to concerns that MPH might induce cytogenetic alterations in children, treatment with this drug has been a controversial issue. In the present study, we assessed the frequency of micronucleated erythrocytes (MNEs), micronucleated polychromatic erythrocytes (MNPCEs), and polychromatic erythrocytes (PCEs) in peripheral blood samples from mice treated with three different doses of MPH (30, 60, or 125?mg/kg). We found no evidence of increased MNEs or MNPCEs, nor did PCEs decline. These results add to the accumulating evidence that MPH does not induce genotoxic or cytotoxic damage.     

Effects of ethylbenzene,toluene, and xylene on the induction of micronuclei in bone marrow polychromatic erythrocytes of mice

Genotoxic effects of five widely used aromatic industrial solvents, ethylbenzene, methylbenzene (toluene), o-, m-, and p-dimethylbenzene (xylene), on bone marrow cells of male NMRI mice were studied using micronucleus test. Each compound was given to animals by IP administration of two similar doses 24 h apart. Increased formation of micronuclei within polychromatic erythrocytes of femoral bone marrow 30 h after the first injection was conducted to be due to the clastogenic effect of the test compound. Of the chemicals tested, only toluene gave a dose-dependent increase in the frequency of micronucleated polychromatic erythrocytes. This genotoxic activity of toluene was confirmed in male B6C3F1 mice.     

Flow cytometric analysis of micronuclei in peripheral blood reticulocytes III. An efficient method of monitoring chromosomal damage in the beagle dog.

Erythrocyte-based micronucleus tests have traditionally analyzed bone marrow because splenic filtration in most species removes micronucleated cells from peripheral blood. We have evaluated a flow cytometric method for monitoring micronucleated reticulocyte frequencies (%MN-RET) in the peripheral blood of beagle dogs treated with cyclophosphamide (CP) and have found that analysis of micronucleated reticulocytes (MN-RETs) in peripheral blood is a suitable surrogate for bone marrow analysis. The three-color flow cytometric method uses anti-CD71 labeling to identify reticulocytes and Plasmodium berghei-containing erythrocytes as a calibration standard. The spontaneous %MN-RET determined by flow cytometry was 0.31 +/- 0.09% (n = 22) for peripheral blood, compared with 0.38 +/- 0.13% (SD, n = 12) for bone marrow, and 0.27 +/- 0.08% (n = 12) for peripheral blood by microscopic scoring with acridine orange staining. The kinetics of appearance and disappearance of MN-RETs in blood were determined by collecting daily samples after iv treatment with CP. The maximum frequency occurred approximately 48 h after dosing. Frequencies of MN-RETs in peripheral blood at steady state following daily CP treatment were 55-68% of corresponding bone marrow values assessed by microscopy and 55-112% as assessed by flow cytometry. This difference is presumably due to splenic removal, which appears slightly less stringent than that previously reported for CP-treated Sprague-Dawley rats. Responses in bone marrow and peripheral blood were highly correlated and similar to or greater than those reported in mice and rats at equitoxic doses.     

Clastogenic effect of passive smoking on bone marrow polychromatic erythrocytes of NMRI mice

The genotoxic effect of passive inhalation of sidestream cigarette smoke on bone marrow polychromatic erythrocytes was studied using male NMRI mice. The animals were placed in individual 145.2-dm3 glass chambers resembling a room provided with normal air flow. They were exposed to the sidestream smoke of a commercial brand of cigarettes smoked by a smoking machine under standard conditions. Increased formation of micronuclei within polychromatic erythrocytes (PCEs) of femoral bone marrow 30 h after passive smoking was regarded as being due to the clastogenic effect of the smoke. Passive inhalation of the diluted sidestream smoke of a single cigarette resulted in a significant increase (P less than 0.01) in the frequency of micronucleated PCEs. This clastogenic activity was found to be dose-dependent.     

多环芳烃对金属硫蛋白缺欠小鼠微核及红细胞的影响

目的：观察两种致癌性多环芳烃化合物二甲基苯蒽（DMBA）和苯并（a）芘（BaP）对小鼠髓多染红细胞微核发生和血液循环中红细胞数量的时相影响以及金属硫蛋白的可能拮抗作用。方法：选用雄性金属硫蛋白（MT）基因敲除的转基因小鼠[MT(-/-)]和野生型小鼠[MT( / )]。在DMBA和BaP各50mg/kg1次腹腔注射染毒后24、48、72和144h，观察动物骨髓多染红细胞中的微核细胞发生率和外周血液循环中红细胞数量的变化。结果：DMBA和BaP均能引起两种小鼠骨髓多染红细胞微核发生增加和外周血液循环中红细胞数量减少。DMBA诱导微核发生的高峰在48h。在同种小鼠内DMBA诱导的微核细胞发生率大于BaP。在DMBA染毒48和72h后MT（－／－）小鼠的微核细胞率明显大于MT（＋／＋）小鼠，而在BaP染毒后两种小鼠之间差异无显著性。在DMBA染毒24h后MT（－／－）小鼠的红细胞数比染毒前明显减少。结论：在该研究 条件下，缺乏MT的小鼠的骨髓多染红细胞微核更易被DMBA诱导和发生红细胞数量减少。提示MT具有一定保护DMBA所致遗传损伤和红细胞系统损伤的功能。     

Effects of GSH and WR-2721 on induction of micronuclei by cyclophosphamide

The frequency of micronucleated polychromatic erythrocytes (MNPCEs) was assessed in the bone marrow and peripheral blood of adult male Swiss mice treated with reduced glutathione (GSH) and/or S-2-/3-aminopropylamino/ethyl phosphorothioic acid (WR-2721), at a dose of 400 mg/kg body weight, and/or with cyclophosphamide (CP), at a dose of 200 mg/kg body weight. GSH was given 60 or 15 min and/or WR-2721 was applied 30 min before CP administration. The number of MNPCEs was determined at 24 h after the drug application. After treatment of mice with CP, the frequency of MNPCEs was distinctly increased. The stronger chemoprotective effect against CP-induced cytotoxicity was obtained following GSH administration than after WR-2721 injection. WR-2721 characterized greater cytotoxicity than GSH. The combination of GSH and WR-2721 given alone, or before CP administration resulted in the most cytotoxic and chemoprotective effects, compared with the respective single-thiol treatment of mice. The most effective protection against CP-induced genotoxicity was observed in the case of treatment of mice with WR-2721and GSH, respectively, 30 and 15 min before CP administration. The most cytotoxic effect of the thiols was found when GSH given 30 min prior to WR-2721 application. The chemoprotection and cytotoxicity caused in the mouse erythroblasts by GSH and WR-2721, as indicated by the number of MNPCEs were dependent on the thiol(s) given, and the time intervals between the drug administration. The modulatory effect of the thiols GSH and WR-2721 on 'delayed apoptosis' induced in the erythropoietic system by cyclophosphamide was shown.     

Effects of pretreatment of male NMRI mice with enzyme inducers or inhibitors on clastogenicity of toluene

Five groups of young male NMRI mice were pretreated with IP injections of three known inducers of cytochrome P450, Aroclor 1254, phenobarbital and 3-methylcholanthrene, and two inhibitors, metyrapone and alpha-naphthoflavone, 5, 3, 2, 1, and 1 day(s) before receiving toluene, respectively. Toluene was given to animals by IP injections of two similar doses 24 h apart. Increased formation of micronuclei within polychromatic erythrocytes of femoral bone marrow 30 h after the first injection of toluene was recorded. None of the treatments with an inducer or inhibitor alone gave a significant increase in the frequency of micronucleated polychromatic erythrocytes. However, pretreatment of animals with each inducer or even inhibitor resulted in an enhanced clastogenic activity of toluene. Simultaneous injections of an inhibitor and toluene clearly decreased the clastogenicities observed. Enhancement of the clastogenicity of toluene was more evident among Aroclor -pretreated animals than among the other groups. Treatment of animals with a mixture of toluene and benzene did not result in an additive clastogenic activity of benzene. IP injection of a mixture of toluene and every xylene isomer resulted in an enhanced clastogenic activity of toluene, although xylene isomers are not found to be clastogenic.Abbreviation PCE polychromatic erythrocyte     

The investigation of the genotoxic effects of fenarimol and propamocarb in mouse bone marrow in vivo

In this study, we aimed to evaluate the genotoxic effects of fungicides fenarimol and propamocarb which are used to protect crops from fungi. For this reason, bone-marrow micronucleus and chromosome aberration tests were carried out in Swiss albino mice. Mice were injected with four different doses of fenarimol and propamocarb intraperitoneally; 50, 100, 200 and 400 mg/kg b.w. Fenarimol did not induce any significant increase in micronucleated erythrocytes after 24, 36, and 48 h treatment but it decreased the ratio of polychromatic/normochromatic erythrocytes at all dose groups and sampling intervals. Fenarimol did not increase the number of chromosome aberrations significantly, but it reduced the mitotic index at the higher doses (P < 0.05). Propamocarb did not increase the frequency of micronucleated erythrocytes, but decreased the polychromatic/normochromatic erythrocytes ratio at all sampling intervals. Propamocarb increased only gaps in total chromosome aberrations, but when gaps were excluded, there were no significant differences in total aberrations between the control and dose groups (P > 0.05). Propamocarb also reduced the mitotic index compared with the negative control group (P < 0.001). Contributing these results, we can suggest that fenarimol and propamocarb are non-genotoxic in mouse bone marrow in vivo but have cytotoxic effects.     

Evaluation of genotoxicity of oral exposure to tetravalent vanadium in vivo

The trace element vanadium interacts with living cells, in which it exerts a variety of biological effects depending on its chemical form and oxidation state. Tetravalent vanadium was shown to affect several genotoxicity end-points in vitro, but its genotoxic potential in vivo is not elucidated. In this study, the genotoxic effects induced in vivo by subacute oral exposure to vanadyl sulphate (VOSO4), a tetravalent vanadium salt, were investigated. To this aim male CD1 mice were administered with VOSO4 in drinking water over the dose range 2-1000 mg/l for 5 weeks. The incidence of micronucleated blood reticulocytes was measured along treatment period. At the end of treatment, micronuclei in both blood reticulocytes and bone marrow polychromatic erythrocytes were determined; in addition, DNA lesions detectable by comet assay were assessed in marrow and testicular cells. Tissue distribution of vanadium at sacrifice was determined by atomic absorption spectrometry. Comet assays and the analysis of micronuclei in polychromatic erythrocytes did not reveal treatment related effects. A slight increase in micronucleated reticulocytes, with no relationship with the administered dose, was observed in some treated groups. The determination of vanadium content in kidney, liver, spleen, bone, stomach, small intestine and testis highlighted low internal exposure, especially in soft tissues. Overall, data indicate scarce bioavailability for orally administered tetravalent vanadium, and lack of significant genotoxic potential in vivo.     

Suppressive effect of aspirin on chromosome aberration induced by mitomycin C in mice

Chromosome aberrations induced by mitomycin C (MMC) were suppressed by aspirin in a mouse micronucleus test with peripheral blood and bone marrow cells. Aspirin at doses of 0.5, 5, and 50 mg/kg was injected intraperitoneally or per administered orally 0.5, 6, or 24 h after administration of MMC and then peripheral blood and/or bone marrow cells were sampled 48 h after administration of MMC. The suppressive effect of aspirin was more pronounced in the aspirin-treated groups 24 h than 0.5 and 6 h after administration of MMC. In the aspirin-treated group at 24 h, the frequency of polychromatic erythrocytes with micronuclei was decreased by about 60-80% after intraperitoneal injection and by about 40-70% after oral administration. It is suggested that aspirin may directly act on MMC metabolites, but not on MMC itself.     

Nandrolone androgenic hormone presents genotoxic effects in different cells of mice

Nandrolone is an androgenic–anabolic steroid (AAS) with diverse medical applications but taken indiscriminately by some to rapidly increase muscle mass. The aim of this study was to evaluate the genotoxic and clastogenic potential of nandrolone (deca‐durabolin®) in vivo in different cells of mice, using the comet assay and micronucleus test, respectively. The animals received subcutaneous injection of the three doses of the steroid (1.0, 2.5 and 5.0 mg kg^?1 body weight). Cytotoxicity was assessed by scoring 200 consecutive total polychromatic (PCE) and normochromatic (NCE) erythrocytes (PCE–NCE ratio). The results showed a significant dose‐related increase in the frequency of DNA damage in leukocytes, liver, bone marrow, brain and testicle cells at the three tested doses and a significant increase of the micronucleated polychromatic erythrocytes at all tested doses. Under our experimental conditions, the nandrolone steroid hormone showed genotoxic and clastogenic effects when administered subcutaneously to mice. Copyright © 2011 John Wiley & Sons, Ltd.     

Chemoprotective effects of hesperidin against genotoxicity induced by cyclophosphamide in mice bone marrow cells

The preventive effect of hesperidin as a flavonoid was investigated in mouse bone marrow cells against genotoxicty induced by cyclophosphamide. Mice were orally (gavages) pretreated with solutions of hesperidin at four different doses (50, 100, 200, and 400 mg/kg b.w.) for five consecutive days. Mice were injected intraperitoneally on the fifth day with cyclophosphamide (50 mg/kg b.w.) and killed after 24 h for the evaluation of micronucleated polychromatic erythrocytes (MnPCEs) and the ratio of PCE/(PCE+NCE) (polychromatic erythrocyte/ polychromatic erythrocyte + normochromatic erythrocyte). Three last doses of hesperidin significantly reduced frequency of MnPCEs induced by cyclophosphamide (p<0.0001). Hesperdin at dose 200 mg/kg b.w. reduced MnPCEs 2.37 time and also completely normalized PCE/ (PCE+NCE) ratio. Histological examination of bone marrow showed that hesperidin affected on proliferation and hyper cellularity of immature myeloid elements in bone marrow that reduced by cyclophsopahmide. It is obvious that hesperidin, may with antioxidative activity, reduced the oxidative stress and genotoxicity induced by cyclophosphamide in mouse bone marrow cells.