A study on Amygdalin's genotoxicological safety and modulatory activity in human peripheral lymphocytes in vitro.

Amygdalin (AMY), a plant secondary metabolite containing nitrile, is a major component of the seeds of Rosaceae family plants. It is known that this compound has many pharmacological activities such as cancer prevention, antipyretic, and cough suppressant. In this study, the genotoxic and modulatory effects of amygdalin were assessed by chromosomal aberration (CA), sister chromatid exchange (SCE), and cytokinesis-block micronucleus assay (CBMN) assays using human peripheral lymphocytes (HPLs) in the absence and presence of metabolic activator (S9 mix). Lymphocytes were exposed to various concentrations of amygdalin (0.86, 1.72, 3.43, 6.86, and 13.75 µg/mL) alone and in combination with mitomycin-C (MMC, 0.20 µg/mL) or cyclophosphamide (CP, 12 µg/mL). The mitotic index (MI), replication index (RI), cytokinesis-block proliferation index (CBPI), and cytostasis were also evaluated to determine cytotoxicity. Amygdalin alone did not exhibit genotoxic and cytotoxic effects at all the tested concentrations both in the absence and presence of the S9 mix. In contrast, amygdalin significantly reduced the frequencies of CA (especially at 48 h treatments), SCE, and MN (except 0.86 µg/mL in pre- and simultaneous treatment) induced by MMC in all the tested concentrations and treatment protocols. It has also considerably decreased CP-induced CA and SCE frequencies at all the concentrations (except 0.86 µg/mL) in simultaneous treatment. This study demonstrated that amygdalin alone was not genotoxic, on the contrary, it has revealed modulatory effects against chemotherapy agents that induced genomic damage in human lymphocytes, suggesting its chemopreventive potential.

Antigenotoxic effect of hyperoside against Mitomycin C and hydrogen peroxide-induced genotoxic damage on human lymphocytes.

Hyperoside is a flavonol glycoside isolated from various plant genera such as Hypericum and Crataegus. It has an important place in the human diet and is used medically to relieve pain and ameliorate cardiovascular functions. However, a comprehensive profile of the genotoxic and antigenotoxic effects of hyperoside is not known. The current study aimed to investigate the genotoxic and antigenotoxic effects of hyperoside against genetic damages induced by two genotoxins (MMC and H2O2) using chromosomal aberrations (CAs), sister chromatid exchanges (SCEs), and micronucleus (MN) assays in human peripheral blood lymphocytes in vitro. Blood lymphocytes were incubated with 7.8-62.5 µg/mL concentrations of hyperoside alone and simultaneously with 0.20 µg/mL Mitomycin C (MMC) or 100 µM Hydrogen peroxide (H2O2). Hyperoside did not exhibit genotoxic potential in the CA, SCE, and MN assays. Moreover, it did not cause a decrease in mitotic index (MI) which is an indicator of cytotoxicity. On the other hand, hyperoside significantly decreased CA, SCE, and MN (except for MMC treatment) frequencies induced by MMC and H2O2. Hyperoside, increased mitotic index against both mutagenic agents at 24-h treatment when compared to positive control. Our results demonstrate that hyperoside exhibited antigenotoxic effects rather than genotoxic in vitro human lymphocytes. Therefore, hyperoside may be a potential preventive agent in inhibiting chromosomal and oxidative damage induced by genotoxic chemicals.

Determination of genotoxic damages of picloram and dicamba with comet assay in Allium cepa rooted in tissue culture and distilled water.

BACKGROUND: Many genotoxicity tests allow us to understand the mechanism of damages on genetic material occurring in living organisms against various physical and chemical agents. One of them is the Comet test. The current study aimed to evaluate genotoxic caused by picloram and dicamba to root meristems of Allium cepa utilizing comet assay. METHODS: Two different protocols were used for rooting and auxin/pesticide application. (i) A. cepa bulbs were rooted in MS medium and then treated with Murashige and Skoog (MS) medium (control) and 0.67, 1.34, 2.01, 2.68, 3.35, 4.02, and 8.04 mg/L of picloram and dicamba using aseptic tissue culture techniques. (ii) A. cepa bulbs were then rooted in bidistilled water and treated with 0 (control), 0.67, 1.34, 2.01, 2.68, 3.35, 4.02, and 8.04 mg/L of picloram and dicamba in distilled water. The A. cepa root tip cells in both treatment groups were examined using comet test to find the possible DNA damaging effects of picloram and dicamba. RESULTS: The results obtained at all the concentrations were statistically compared with their control groups. Almost at all the concentrations of Picloram and dicamba increased comet tail intensity (%) and tail moment in roots treated in MS medium. Two highest concentrations revealed toxic effect. On the other hand, DNA damaging effect of both auxins was only noted on the highest (> 4.02 mg/L) in roots treated in distilled water. CONCLUSIONS: This study approve and confirm genotoxic effects of how growth regulators on plants. These findings give an evidence of DNA damage in A. cepa. Therefore, both picloram and dicamba should only be used in appropriate and recommended concentrations in agriculture to conserve ecosystem and to pose minimum threat to life.

Evaluation of the genotoxic and antigenotoxic effects of exopolysaccharide pullulan in human lymphocytes in vitro.

Pullulan is a biocompatible and water-soluble exo-polysaccharide produced by primary strains of the fungus Aureobasidium pullulans. It is frequently used in the pharmaceutical and food industries. In this study, possible cytotoxic effect of pullulan was assessed using the MTT assay in the human breast cancer (MCF-7) cell line. Micronucleus (MN), micronucleus-FISH (MN-FISH), random amplified polymorphic DNA (RAPD-PCR), and comet assays were used to investigate genotoxic and antigenotoxic effects of pullulan against mitomycin C (MMC) (at MN assay) and hydrogen peroxide (at comet assay) in human lymphocytes. Antigenotoxicity was determined using two different applications: 1 h pretreatment and simultaneous treatment. In the MTT assay, pullulan significantly reduced the cell viability at 15.6-2000 µg/mL compared to the control. No significant alterations in MN rates were found in human lymphocytes treated with different concentrations of pullulan compared to the control. In contrast, co-treatment of pullulan and MMC decreased the frequency of MN in almost all the treatment concentrations and durations compared to the MMC. No significant change was observed in the frequency of the centromere-positive C + or negative C- MNi compared to the positive control. In comet assay, pullulan did not affect comet tail intensity compared to the negative control. On the contrary, pullulan in combination with H2O2 significantly decreased tail intensity at almost all the concentrations compared to the positive control. The changes occurring in RAPD-PCR profiles following pullulan treatments included an increase or decrease in band intensity and gain or loss of bands. These results indicate that exopolysaccharide Pullulan is not genotoxic; moreover, it possesses a protective effect against MMC and H2O2 induced genotoxicity. In breast cancer cells, pullulan induced cytotoxic/anti-proliferative effect.

Genotoxic and antigenotoxic potential of amygdalin on isolated human lymphocytes by the comet assay.

Amygdalin is a cyanogenic glycoside, mainly present in the seeds of the Rosaceae family such as apricots, peaches, and bitter almond. In this study, in vitro genotoxic and antigenotoxic effects of amygdalin have been investigated on human peripheral blood lymphocytes using the comet assay. The antigenotoxic effect of amygdalin was performed against hydrogen peroxide (H2 O2 ) using three different treatment types (pre-, simultaneous, and post-treatment). The isolated lymphocytes were incubated with different concentrations of amygdalin (0.86-13.75 µg/ml) alone and in combination with H2 O2 (100 µM). The results indicated that amygdalin exhibited an antigenotoxic effect against H2 O2 , but it did not induce the genotoxic effect alone in tested concentrations in vitro on human lymphocytes. PRACTICAL APPLICATIONS: Amygdalin is a natural compound used in alternative medicine as an anti-cancer, antipyretic, and cough suppressant. The comet assay which is relatively simple, rapid, sensitive, and economically efficient, measures the changes in genomic stability. Assessment of amygdalin alone has no genotoxic effect on human lymphocytes. Moreover, antigenotoxicity applications (pre-, simultaneous, and post-treatments) of amygdalin significantly reduced the DNA damage induced by H2 O2 on isolated human lymphocytes. In conclusion, amygdalin is not genotoxic, also, it exhibited antigenotoxic activity against oxidatively damaged DNA due to its antioxidant properties on human lymphocytes.

Evaluation of the cytotoxic and genotoxic effects of mycotoxin fusaric acid.

Fusaric acid (FA) is produced by several Fusarium species and is commonly found in grains. This investigation was performed to evaluate the cytotoxic and genotoxic effects of FA either in human cervix carcinoma (HeLa) cell line using 3-(4,5-dimethylthiazolyl-2)-2,5 diphenyltetrazolium bromide (MTT) assay and in human lymphocytes using chromosome aberrations (CAs), sister chromatid exchanges (SCEs), micronuclei (MN) as well as comet assay in vitro. The cells were treated with 0.78, 1.56, 3.125, 6.25, 12.50, 25, 50, 100, 200, and 400 µg/mL concentrations of FA. It has potent cytotoxic effect on HeLa cell line measured by MTT assay especially at higher concentrations (200, 400 µg/mL). The half of inhibitory concentration (IC50) evidenced by FA in the HeLa cells was 200 µg/mL at 24 h and between 200 and 400 µg/mL at 48 h. It was also observed that FA produced a significant decrease in mitotic index (MI) at 12.50 µg/mL compared to solvent control. Furthermore, it indicated a cytotoxic effect at the concentrations ranging from 25 to 400 µg/mL in human lymphocytes. The results of this research point out that being exposed to FA at high concentrations show cytotoxicity. Besides FA induced comet tail intensity at 3.125, 6.25, and 12.50 µg/mL concentrations in isolated human lymphocytes. On the other hand, no genotoxic effects were seen in human lymphocytes in vitro using CA, SCE and MN assays.

In vitro genotoxic and antigenotoxic effects of cynarin.

ETHNOPHARMACOLOGICAL RELEVANCE: Cynarin is an artichoke phytochemical that possesses a variety of pharmacological features including free-radical scavenging and antioxidant activity. The origin of artichoke species appears to be Mediterranean region. Two of these species, globe artichoke (Cynara cardunculus var. scolymus L.) and cardoon (Cynara cardunculus var. altilis DC), are widely cultivated and consumed. This vegetable, as the basis of the mediterranean diet, has been used as herbal medicine for its therapeutic effects since ancient times. Therefore, this study was performed to determine genotoxic and antigenotoxic effects of cynarin against MMC (mitomycin C) and H2O2 (hydrogen peroxide) induced genomic instability using chromosome aberrations (CAs), sister chromatid exchanges (SCEs), micronucleus (MN), and comet assays in human lymphocytes. MATERIALS AND METHODS: Lymphocytes obtained from two healthy volunteers (1 male and 1 female) were exposed to different concentrations of cynarin (12-194 µM) alone and the combination of cynarin and MMC (0.60 µM) or cynarin and H2O2 (100 µM, only for comet assay). RESULTS: Cynarin alone did not induce significant genotoxic effect in the CA, SCE (except 194 µM), MN, and comet assays. The combination of some concentrations of cynarin and MMC decreased the frequency of CAs, SCEs and MN induced by MMC. Furthermore, the combination of cynarin and H2O2 reduced all comet parameters at all the concentrations compared to H2O2 alone. While the highest concentrations of cynarin significantly decreased mitotic index (MI), the combination of cynarin and MMC increased the reduction of MI induced by MMC alone. CONCLUSION: All the results obtained in this study demonstrated that cynarin exhibited antigenotoxic effects rather than genotoxic effects. It is believed that cynarin can act as a potential chemo-preventive against genotoxic agents.

Assessment of cytotoxic and genotoxic effects of enniatin-A in vitro.

Enniatin A (EN-A) is a Fusarium mycotoxin which is a common contaminant in grains and especially in maize and it causes serious loss of product. The aim of this study was to investigate the cytotoxic effects using 3-(4,5-dimethylthiazolyl-2)-2,5 diphenyltetrazolium bromide (MTT) assay in human cervix carcinoma (HeLa) cell line, and genotoxic effects of EN-A using chromosome aberrations (CAs), sister chromatid exchanges (SCEs), micronuclei (MN) and comet assays in human lymphocytes. The cells were treated with 0.07, 0.14, 0.29, 0.57, 1.15, 2.29, 4.59 and 9.17 µM concentrations of EN-A. It exhibited cytotoxic effects in HeLa cell lines especially when the concentrations were increased. The half-inhibitory value (IC50) was determined as 1.15 µM concentration for 24 h and 0.57 µM concentration for 48 h. However, EN-A failed to affect the frequency of CAs, SCEs and MN in human lymphocytes. Only a slight increase was observed in the frequency of SCEs at 0.57 µM concentration over 48 h. The replication (RI) and nuclear division (NDI) indices were not affected. On the contrary, EN-A decreased the mitotic index (MI) significantly at all concentrations compared to the negative control and solvent control (except at 0.29 µM for 24 h, and except at 0.14, 0.29 and 0.57 µM for 48 h). Treatments over 2.29 µM showed toxic effects in human lymphocytes. EN-A significantly increased comet tail intensity (except at 0.07 and 0.57 µM) in isolated human lymphocytes. The results of this study demonstrate that EN-A has an obvious cytotoxic effect especially when the EN-A concentration was increased. In addition, EN-A could exhibit a mild genotoxic effect.