Comparative analysis of cytotoxic, genotoxic and antioxidant effects of 2,2,4,7-tetramethyl-1,2,3,4-tetrahydroquinoline and ethoxyquin on human lymphocytes

2,2,4,7-Tetramethyl-1,2,3,4-tetrahydroquinoline (THQ) is a new synthetic compound with potential antioxidant activity. In this study, cytotoxic, genotoxic and antioxidant activities of THQ were studied on human lymphocytes with the use of the trypan blue exclusion assay, the TUNEL method, the comet assay and the micronucleus test. The activities of THQ were compared with those of a structurally similar compound-ethoxyquin (1,2-dihydro-6-ethoxy-2,2,4-trimethylquinoline, EQ), which is used in animal feeds as a preservative. Cytotoxic effects of THQ were observed after 1-h treatment at the concentration of 500 microM and after 24-h treatments at the concentrations of 250-500 microM. Although the micronucleus test did not reveal a genotoxic effect of THQ, in the comet assay the statistically significant increase in DNA damage was observed as compared with the control. On the other hand, the protection of human lymphocytes against DNA damage induced by hydrogen peroxide suggests an antioxidant activity of THQ. The comparative analysis of THQ and EQ activities performed in these studies revealed that THQ was less cytotoxic and less genotoxic than EQ. Slightly lower antioxidant activity of THQ was also shown in the comet assay when it was used at the lower studied doses (1-5 microM), but for the highest one (10 microM) its efficiency was similar to that of EQ. In the micronucleus assay THQ was more effective than EQ in protecting the cultured lymphocytes from clastogenicity of H2O2. We believe that THQ is worthy of further detailed studies on its antioxidant properties to confirm its usefulness as a preservative.     

Evaluation of genotoxicity, cytotoxicity and cytostasis in human lymphocytes exposed to patulin by using the cytokinesis-block micronucleus cytome (CBMN cyt) assay

Patulin (PAT) is a fungal secondary metabolite commonly present in apples and apple products. In the present study, PAT was evaluated for its genotoxic, cytotoxic and cytostatic effects to human peripheral blood lymphocytes by using the cytokinesis-block micronucleus cytome (CBMN Cyt) assay. Lymphocyte cultures were treated with PAT at the following concentrations, 0.1, 0.3, 0.5, 1.0, 2.5, 5.0, and 7.5 μM, as well as 0.5 μM mitomycin c (MMC) as a positive control and dimethyl sulfoxide (DMSO) as a vehicle control. PAT was found to induce nucleoplasmic bridges (NPBs) at 5.0 and 7.5 μM concentrations (P?<?0.05), apoptotic cells at 0.1, 1.0, 5.0 μM (P?<?0.05), 7.5 μM concentrations (P?<?0.01) and necrotic cells at 0.3 and 2.5 μM (P?<?0.05), 5.0 and 7.5 μM (P?<?0.01) concentrations in human lymphocytes. The 2.5, 5.0, and 7.5 μM PAT concentrations also led to a clear decrease in the nuclear division index (NDI) (P?<?0.05). PAT caused a significant dose-dependent increase in the number cells of NPBs, in the frequency of apoptotic and necrotic cells, and a significant dose-dependent decrease in the NDI values in lymphocytes. These results indicate that PAT at high concentrations is genotoxic, cytotoxic and cytostatic in cultured human lymphocytes.     

Mutagenic and genotoxic effects of Anilofos with micronucleus,chromosome aberrations,sister chromatid exchanges and Ames test

We aimed to evaluate the mutagenic effect of Anilofos, organophosphate pesticide, by using Ames/Salmonella/microsome test. Its cytotoxic and genotoxic effects were also determined by chromosome aberration (CA), sister chromatid exchange (SCE) and micronucleus (MN) test in human peripheral blood lymphocytes. In the Ames test, five different concentrations of Anilofos were examined on TA97, TA98, TA100 and TA102 strains in the absence and presence of S9 fraction. According to the results all concentrations of this pesticide have not shown any mutagenic activity on TA97, TA100 and TA102 strains in the absence and presence of S9 fraction. But, 10, 100 and 1000 µg/plate concentrations of Anilofos were determined to be mutagenic on TA98 strain without S9 fraction. Lymphocytes were treated with various concentrations (25, 50, 100 and 200 µg/ml) of Anilofos for 24 and 48 h. The results of the assays showed that Anilofos did not induce SCE frequency, replication index and MN formation at all concentrations for both treatment periods. Anilofos significantly increased CA frequency at 100 and 200 µg/ml concentrations at 24 h treatment periods and at 50, 100 and 200 µg/ml concentrations in 48 h treatment periods. Additionally, it was determined that this pesticide decreased mitotic index and nuclear division index significantly. It was concluded that Anilofos has genotoxic and cytotoxic effects in human peripheral lymphocytes.     

Mutagenicity and genotoxicity of dicapthon insecticide

Mutagenic and genotoxic effects of dicapthon were investigated by using the bacterial reverse mutation assay in Salmonella typhimurium TA97, TA98, TA100 and TA102 strains with or without metabolic activation system (S9 mix), and chromosome aberrations (CAs), sister chromatid exchanges (SCEs), and micronucleus (MN) tests in human peripheral blood lymphocytes in vitro. Dicapthon was dissolved in dimethyl sulfoxide for all test systems. 0.1, 1, 10 and 100 μg/plate doses of dicapthon were found to be weakly mutagenic on S. typhimurium TA 98 without S9 mix. The human peripheral lymphocytes were treated with four experimental concentrations of dicapthon (25, 50, 100, and 200 μg/mL) for 24 and 48 h. Dicapthon increased the frequency of SCE only at the 100 μg/mL concentration for the 24 and 48 h applications. Dicapthon also induced abnormal cell frequency, CA/cell ratio and frequency of MN dose dependently for 24 and 48 h. Dicapthon showed a statistically significant cytotoxic effect by decreasing the mitotic index in all concentrations and a cytostatic effect by decreasing nuclear division index in 100 and 200 μg/mL concentrations for both treatment periods when compared with both untreated and solvent controls. These values decreased also in a dose dependent manner.     

The genotoxic and anti-genotoxic effects of <Emphasis Type="Italic">Stachys petrokosmos</Emphasis> leaf extract in human lymphocytes using microsomal fractions

The genotoxic and anti-genotoxic effects of Stachys petrokosmos leaf extracts (Sp) were investigated in human lymphocytes. The cells were treated with 1.5, 3.0 and 6.0 μL/mL concentrations of Sp leaf extracts for 24 and 48 h treatment periods in the absence and presence of metabolic activator (S9mix). In the absence of S9mix, Sp alone did not induce chromosome aberrations and formation of micronucleus while inducing the mean sister chromatid exchange at the highest concentration. In addition, Sp decreased the mutagenic effect of mitomycin-c. Sp alone showed a cytotoxic effect determined by a decrease in the proliferation index, mitotic index and nuclear division index. On the other hand a mixture of Sp and mitomycin-c resulted in a higher cytotoxic effect especially for 48 h treatment period. In the presence of S9mix, Sp was not genotoxic and cytotoxic however, it showed an anti-genotoxic effect by decreasing the effects of cyclophosphamide.     

The effects of 4-thujanol on chromosome aberrations,sister chromatid exchanges and micronucleus in human peripheral blood lymphocytes

4-Thujanol, a bicyclic monoterpene alcohol, is present in the essential oils of many medicinal and aromatic plants. It is commonly used as a fragrance and flavouring ingredient in a lot of different products. The potential genotoxic effects of 4-thujanol on human peripheral blood lymphocytes (PBLs) were investigated in vitro by the chromosome aberrations (CAs), sister chromatid exchanges (SCEs), and micronucleus (MN) tests. The cells were treated with 13, 26 and 52 μg/mL 4-thujanol in the presence and absence of a metabolic activator (S9 mix). 4-Thujanol induced CA (P < 0.001) and MN formation (P < 0.05) at all concentrations (13, 26 and 52 μg/mL) in the presence and absence of the S9 mix without a concentration-dependent manner. However, the treatment of peripheral lymphocytes with 4-thujanol did not produce a statistical difference in the frequency of SCEs when compared with control group. Furthermore, this monoterpene did not significantly decrease the mitotic index (MI), proliferation index (PI), and nuclear division index (NDI). In conclusion, 4-thujanol had a significant clastogenic effect at the tested concentrations (13, 26 and 52 μg/mL) for human PBLs. In addition, no cytotoxic and/or cytostatic effects were observed regardless of the concentrations used. This work presents the first report on genotoxic properties of 4-thujanol.     

Mutagenic activity of estradiol evaluated by an in vitro micronucleus assay. Short communication

The objective of the present study was to evaluate possible genetic changes in cultured human lymphocytes treated with estradiol, using the cytokinesis block micronucleus assay. Eight experimental concentrations of estradiol were used (range from 10(-10) M to 0.7 x 10(-4) M). The obtained results indicate that estradiol exhibits aneugenic and/or clastogenic effects, expressed as increased frequency of micronucleated lymphocytes at two highest experimental concentrations used in this investigation. In addition to genotoxic effects, these concentrations decreased the cytokinesis block proliferation index (CBPI) and percentage of binucleated cells, indicating the cell cycle delay and possible cytotoxic effects. In conclusion, estradiol treatment might represent a human health risk, especially if overdosed or used for a prolonged period of time.     

Genotoxic effects of styrene-7,8-oxide in human white blood cells: comet assay in relation to the induction of sister-chromatid exchanges and micronuclei

Styrene is used in the production of plastics, resins and rubber. The highest human exposures to styrene take place by inhalation during the production of fiberglass reinforced plastics. Styrene is metabolized mainly in the liver to styrene-7,8-oxide (SO), its principal in vivo mutagenic metabolite. In this study, human peripheral white blood cells were exposed to several SO concentrations (10-200 microM) in order to evaluate its genotoxic properties by means of comet assay, sister-chromatid exchanges (SCE) and cytokinesis-blocked micronucleus (MN) test, in addition to determine its clastogenic or aneugenic properties by combining MN with fluorescence in situ hybridization (FISH) procedures. Our results show that SO induces DNA damage, SCE and MN in human leukocytes in vitro at concentrations above 50 microM, and that there is a strong relationship between DNA damage, as measured by the comet assay, and cytogenetic damage induced by SO at the doses employed. SO shows preferentially a clastogenic activity and produces a cytostatic effect at high doses, reflected by the significant decrease of the calculated proliferation indices. A good dose-effect relationship is obtained in the three tests performed at the concentration range assayed.     

Assessment of in vitro cytotoxic and genotoxic activities of some trimethoprim conjugates

Trimethoprim, a commonly used antibacterial agent, is widely applied in the treatment of variety of infections in human. A few studies have demonstrated an extensive exposure of man to antibiotics, but there is still a lack of data for cytotoxic effects including nephrotoxicity, gastrointestinal toxicity, hematotoxicity, neurotoxicity and ototoxicity. The main purpose behind this study was to determine cytotoxic and genotoxic activities of trimethoprim (1), trimethoprim with maleic acid (2) and trimethoprim in conjugation with oxalic acid dihydrate (3). The cytotoxic effects of these three conjugates were elucidated by employing 3-(4,5-dimethylthiazol-2-yl)-2,5-diphenyltetrazoium bromide (MTT) assay using embryonic rat fibroblast-like cell line (F2408) and H-ras oncogene activated embryonic rat fibroblast-like cancer cell line (5RP7). Additionally, determination of genotoxic activity of these three compounds were studied by using cytokinesis blocked micronucleus assay (CBMN) in human lymphocytes. The results demonstrated that trimethoprim alone and its combination with other compounds are able to induce both cytotoxic and genotoxic damage on cultured cells (F2408, 5RP7, human lymphocytes).     

In vitro genotoxicity of rocuronium bromide in human peripheral lymphocytes

Rocuronium bromide (RB), an aminosteroid type neuromuscular blocking agent, acts by reducing or inhibiting the depolarising effect of acetylcholine on the terminal disc of the muscle cell. To our knowledge, there is no adequate information on the genotoxic effects of RB, up to now. In the present study, possible genotoxic effects of RB have been determined by means of sister chromatid exchange (SCE), chromosome aberration (CA) and micronucleus (MN) analyses in human peripheral blood lymphocytes. The human peripheral blood lymphocytes were exposed to three different concentrations of RB (60, 80 and 100 μg/mL) for 24- and 48-h. In this study, RB increased the frequency of CAs, however, did not increase the frequency of SCEs. RB did not decrease the proliferation index (PI) and mitotic index (MI). Accordingly, RB increased the frequency of micronucleus (MN) but did not decrease the nuclear division index (NDI). Findings from this study suggest that rocuronium bromide is clastogenic but not cytotoxic to cultured human peripheral blood lymphocytes.     

Effects of resveratrol on the G(0)-G(1) transition and cell cycle progression of mitogenically stimulated human lymphocytes

Resveratrol (RSV) has been suggested to have cancer preventive properties, on the basis that it suppresses proliferation and induces apoptosis in various tumor cells. Here we test its cytostatic effects on peripheral blood human lymphocytes. RSV (up to 50 microM) had no detectable effects on resting lymphocytes. With the mitogen phytohemagglutin (PHA), however, RSV elicited concentration- and time-dependent responses in lymphocytes. RSV (>/=50 microM) prevented cell entry into the cell cycle, resulting in 99% suppression at 100 microM. The arrested lymphocytes following 24h treatment with 50 microM RSV had minimal RNA content, the feature characteristic of G(0) cells, and were blocked at the stage past the induction of cyclins D2 and D3 and prior to induction of cyclin E. Prolonged treatment (72h) of PHA-stimulated lymphocytes with 100 microM RSV showed a pronounced decrease in the expression of pRb, cyclins E and B, and reduction in p34cdc2 and PCNA. The activation-induced apoptosis was also reduced in the presence of >/=50 microM RSV. These data suggest that studies designed to test RSV efficacy as a chemopreventive agent should include evaluation of its immunomodulatory effect revealed by suppression of lymphocyte stimulation as well as its effect on apoptosis of stimulated lymphocytes.     

Use of HepG2 cell line for direct or indirect mutagens screening: comparative investigation between comet and micronucleus assays

In the present study, DNA-damage and clastogenic or aneugenic effects of genotoxic compounds were examined in a metabolically competent human cell line (HepG2 cells) using the micronucleus and the comet assays. Compounds with various action mechanisms were tested: direct mutagens such as 4-nitroquinoline-N-oxide (4-NQO) and methyl methanesulfonate (MMS) and indirect mutagens requiring biotransformation to be active such as N-nitrosodimethylamine (NDMA), benzo[a]pyrene (B[a]P) and 2-acetylaminofluorene (2-AAF). The compounds were first tested for cytotoxicity by measuring their effects on RNA synthesis inhibition in HepG2 cells. 4-NQO, B[a]P and 2-AAF were the most potent compounds; their IC(50) values were, respectively, 1.9 micro M (4h contact), 3.4 and 112 micro M after 20 h. MMS was mildly cytotoxic (IC(50)=0.9 mM) and NDMA had a weak effect (IC(50)=110 mM) after 4h contact. In the micronucleus and comet assays, concentrations required to obtain a significant genotoxic effect in HepG2 cells varied over a broad range, NDMA being active only at very high concentrations. To compare the sensitivity of the two assays, we measured the so-called FIC(2)-the concentration necessary to induce a 2-fold increase of the measured genotoxicity parameter. The data show that genotoxic effects were consistently observed at lower concentrations in the micronucleus test, except in the case of MMS. The measured FIC(2) values were 0.12 micro M (4-NQO), 0.17 micro M (2-AAF), 0.26 micro M (B[a]P) and 6.4mM (NDMA). MMS had such a weak effect in the HepG2 cells that we could not calculate its FIC(2) value. In the comet assay, FIC(2) values were observed, respectively, at 1.48 micro M (4-NQO), 3.67 micro M (B[a]P), 13.42 micro M (MMS) and 27 mM (NDMA). 2-AAF failed to induce DNA-damage in this assay. The present study shows that HepG2 cells could be a suitable tool for assessing the genotoxicity of direct and indirect mutagens and for establishing the lowest genotoxic concentration.     

The genotoxic effect of the new acaricide etoxazole

Etoxazole is a member of the diphenyl oxazoline class of insecticide was newly developed for use on pome fruits, cotton and strawberries as a acaricide. In the present study, genotoxic effects of acaricide etoxazole (ETX) (miticide/ovicide) were investigated using chromosome aberration (CA) test, sister chromatid exchange (SCE) test and micronucleus test in human lymphocytes. ETX induced the CAs at all concentrations (5, 10 and 20 microg/ml) for 24 h and also induced the CA at the highest concentration (20 microg/ml) for 48 h only. The inducing the CAs for 48 hours treatment period was dose-dependent. Besides, it induced the SCE at all concentrations and treatment periods in a dose-dependent manner as well. Although, ETX decreased the mitotic index (MI) at all concentration and treatment periods dose-dependently, while it did not decrease the replication index (RI) when compared to the negative and solvent controls. In addition, ETX induced the micronucleus at all concentrations except 5 microg/ml for 48 h. This inducing was in a dose-dependent manner as well. In conclusion, it can be concluded that ETX has a potential genotoxic effects in cultured human peripheral lymphocytes.     

Tests for genotoxicity and mutagenicity of furan and its metabolite cis-2-butene-1,4-dial in L5178Y tk+/- mouse lymphoma cells

Furan is found in various food items and is cytotoxic and carcinogenic in the liver of rats and mice. Metabolism of furan includes the formation of an unsaturated dialdehyde, cis-2-butene-1,4-dial (BDA). In view of the multifunctional electrophilic reactivity of BDA, adduct formation with protein and DNA may explain some of the toxic effects. Short-term tests for genotoxicity of furan in mammalian cells are inconclusive, little is known for BDA. We investigated BDA generated by hydrolysis of 2,5-diacetoxy-2,5-dihydrofuran for genotoxicity in L5178Y tk+/- mouse lymphoma cells using standard procedures for the comet assay, the micronucleus test, and the mouse lymphoma thymidine kinase gene mutation assay, using 4-h incubation periods. Cytotoxicity was remarkable: cell viability at concentrations>or=50 microM was reduced to <50%. In the dose range up to 25 microM, viability was >90%. Measures of comet-tail length and thymidine-kinase mutant frequency were increased 1.6- and 2.4-fold above control, respectively. Analysis of three fully independent replicates with a linear mixed-effects model showed a highly significant increase with concentration for both endpoints. Compared to methyl methanesulfonate used as a positive control, BDA was of similar potency with respect to genotoxicity, but it was much more cytotoxic. Furan added to cell cultures at doses that resulted in time-averaged effective concentrations of up to 3100 microM was neither cytotoxic nor genotoxic. A potential cross-linking activity of BDA was investigated by checking whether gamma radiation-induced DNA migration in the comet assay could be reduced by pre-treatment with BDA. In contrast to the effect of the positive control glutaraldehyde, BDA treatment did not reduce the comet tail length. On the contrary, an increase was observed at >or=100 microM BDA, which was attributable to early apoptotic cells. Although BDA was found to be a relatively potent genotoxic agent in terms of the concentration necessary to double the background measures, cytotoxicity strongly limited the concentration range that produced interpretable results. This may explain some of the inconclusive results and indicates that non-genotoxic effects must be taken into account in the discussion of the modes of toxic and carcinogenic action of furan.     

Apoptotic effect of gossypol on human lymphocytes

The overall objective of this study was to determine the effect of gossypol on human lymphocytes. Blood samples were taken from healthy donors and lymphocytes were cultured with various concentrations of gossypol (25-150 microM). The percentage of apoptotic cells was determined by assessing DNA fragmentation by agarose gel electrophoresis; morphological features of apoptosis were assessed by light microscopy. The concentrations of gossypol required to induce apoptosis in human lymphocytes without causing necrosis through cytotoxic effects were between 25-50 microM.     

Synthesis,anti-inflammatory activity and modeling studies of cycloartane-type terpenes derivatives isolated from Parthenium argentatum

The 12-O-tetradecanoylphorbol-13-acetate (TPA)-induced edema model in mice determined the anti-inflammatory activities in vivo of argentatins A, B and D, the main cycloartenol-type triterpenes present in Parthenium argentatum. Our results showed that argentatin B (ED₅₀ = 1.5 × 10⁻⁴ mmol/ear) and argentatin A (ED₅₀ = 2.8 × 10⁻⁴ mmol/ear) were more potent anti-inflammatory agents than indomethacin (ED₅₀ = 4.5 × 10⁻⁴ mmol/ear), the reference drug. Based on these findings, we decided to evaluate 13 derivatives of argentatins A and B. All the derivatives showed anti-inflammatory activity in the TPA-induced edema model in mice. The most active compound was 25-nor-cycloart-3, 16-dione-17-en-24-oic acid, obtained from argentatin A (ED₅₀ = 1.4 × 10⁻⁴ mmol/ear). Argentatin B was assayed as inhibitor of COX-2 activity one of the key enzymes involved in the TPA assay. The results showed that argentatin B at 15 μM doses inhibited 77% COX-2 activity. Docking studies suggest that argentatin B interacts with Arg 120, a key residue for COX-2 activity.     

Hydroquinone-induced genotoxicity and oxidative DNA damage in HepG2 cells

Hydroquinone (HQ) is used as an antioxidant in rubber industry and as a developing agent in photography. HQ is also an intermediate in the manufacture of rubber, food antioxidant and monomer inhibitor. However, the mechanisms of the effects, in particular those related to its genotoxicity in humans, are not well understood. The aim of this study was to assess the genotoxic effects of HQ and to identify and clarify the mechanisms, using human hepatoma HepG2 cells. DNA strand breaks and DNA-protein crosslinks (DPC) were measured by the proteinase K-modified alkaline single cell gel electrophoresis (SCGE) assays. Using the SCGE assay, a significant dose-dependent increment in DNA migration was detected at concentrations of HQ (6.25-25 microM); but at the higher tested concentrations (50 microM), a reduction in the migration compared to the maximum migration at 25 microM was observed. Post-incubation with proteinase K significantly increased DNA migration in cells exposed to higher concentrations of HQ (50 microM). A significant increase of the frequency of micronuclei was found in the range from 12.5 to 50 microM in the micronucleus test (MNT). The data suggested that HQ caused DNA strand breaks, DPC and chromosome breaks. To elucidate the oxidative DNA damage mechanism, the 2,7-dichlorofluorescein diacetate (DCFH-DA) and o-phthalaldehyde (OPT) were chosen to monitor the levels of reactive oxygen species (ROS) and glutathione (GSH), respectively. The present study showed that HQ induced the increased levels of ROS and depletion of GSH in HepG2 cells, the doses being 25-50 and 6.25-50 microM, respectively. Moreover, HQ significantly caused 8-hydroxydeoxyguanosine (8-OHdG) formation in HepG2 cells at concentrations from 12.5 to 50 microM. All these results demonstrate that HQ exerts genotoxic effects in HepG2 cells, probably through DNA damage by oxidative stress. GSH, as a main intracellular antioxidant, is responsible for cellular defense against HQ-induced DNA damage.     

Effect of the phytoestrogen, genistein-8-C-glucoside, on Chinese hamster ovary cells in vitro

Genistein-8-C-glucoside (G8CG) belongs to isoflavones, which are a subclass of flavonoids, a large group of polyphenolic compounds widely distributed in plants. A number of studies on flavonoids show their cardioprotective and antiosteoporosis properties in in vitro and in vivo models. As a phytoestrogen, genistein has recently generated interest as a potential anticancer and antiatherogenic agent. Several flavonoids are known as antioxidants and scavengers of free oxygen radicals. In the current investigation we used glycosylated genistein (genistein-8-C-glucoside) from flowers of lupine (Lupinus luteus L.). Many authors have found that the action of genistein is not so simple, although many reports conducted in vitro have demonstrated that it is cytotoxic and genotoxic. Therefore, the cytotoxic and genotoxic effects of this compound in Chinese hamster ovary cells (line CHO) were studied. A colorimetric MTT assay to assess cytotoxicity and a Comet assay for the detection of DNA damage were used. Apoptosis was determined by the Hoechst 33258/propidium iodide staining technique. We have also demonstrated antioxidant properties of G8CG. The level of reactive oxygen species generated by G8CG alone and/or H2O2 was evaluated with fluorescence probes: dichlorofluorescein-diacetate (DCFDA) by flow cytometry. The cells were exposed to various concentrations of genistein-8-C-glucoside (1-290 microM) and hydrogen peroxide (10-130 microM) and the effect of G8CG alone or in combination with H2O2 was determined. The results reveal that G8CG at concentrations higher than 10 microM significantly reduced cell viability, induced apoptosis and DNA damage. However at lower concentrations (5 and 7.5 microM), G8CG showed antioxidant properties, but had no cytotoxic or genotoxic activity.     

A study on the genotoxic effects of 8-Cl-cAMP on human lymphocytes in vitro

8-chloro-cyclic adenosine 3',5'-monophosphate (8-Cl-cAMP) is the most potent cAMP analogue that selectively inhibits a variety of cancer cell lines in vitro and tumors in vivo. Its action toward a variety of tumors, especially when coupled with other antitumor agents, have lead to phase I clinical investigations and recently phase II clinical investigations. Until today very little was done to evaluate its genotoxic potential. In order to evaluate its genotoxic potential we used the cytogenetic and cytokinesis block micronucleus assay in vitro on peripheral blood lymphocytes of healthy individuals. Using three concentrations (1 microM, 5 microM and 15 microM), 8-Cl-cAMP in normal human peripheral blood lymphocytes did not induce any cytogenetic aberrations of the structural type [chromatid breakage, isochromatid breakage and gaps], but did induce premature centromere separation (PCS) in all respective doses and increased the frequency of micronuclei (p <0.05) only in the highest dose (15 microM). Antiproliferative action of 8-Cl-cAMP was estimated by using the cytokinesis block nuclear division index (NDI). The results showed a decrease in the NDI of cells exposed to all doses of 8-Cl-cAMP when compared to control. Therefore, the overall results show a genotoxic potential of 8-Cl-cAMP in peripheral blood lymphocytes in vitro.     

Micronucleus assay in human lymphocytes after exposure to alloxydim sodium herbicide in vitro

This study evaluates the cytotoxic and genotoxic potential of alloxydim sodium using micronucleus (MN) assay, in human peripheral lymphocytes. MN assay was used to investigate the genotoxic effects of alloxydim sodium in human peripheral lymphocytes treated with 250, 500, 750, 1,000 µg/ml concentrations of alloxydim sodium for 24 and 48 h. Solvent, negative and positive controls were also used in the experiments in parallel. The obtained results were evaluated in statistical analyses by using Dunnett-t test (two sided) and p < 0.05 was accepted as significant. Alloxydim sodium significantly increased the MN formation compared with the negative control, at both 750 and 1,000 µg/ml concentrations and treatment periods. We also evaluated the nuclear division index (NDI) for cytotoxicity of this pesticide in the experiment, and finally observed a significant decrease of the NDI values at all concentrations of alloxydim sodium and at both treatment periods.