Dietary intake of quercetin and resveratrol has been considered representative and more promising in terms of their positive effects as shown in previous studies [36, 37]. The present study was undertaken to evaluate the possible protective effects of quercetin and/ or resveratrol against acrolein-induced sister chromatid exchanges and DNA damage in male albino mice. Current results showed that the incidence of sister chromatid exchanges (SCEs) in bone marrow cells of acrolein-treated group was significantly (p < 0.05) increased when compared to the corresponding control. This confirmed the previous results of Au et al. [38] which indicated that acrolein was the most toxic metabolite of cyclophosphamide and it was responsible for the cytotoxicity of it by inducing chromosome breakage and a high SCEs rate in vitro. Also, Khoramjouy et al., Storme et al. and Uchida [39-41] observed that acrolein caused chromosomal aberrations, sister chromatid exchange, point mutations and inhibition of DNA repair in mammalian cells.
Moreover, our results showed that oral administration of acrolein 10 mg/kg for four consecutive days induced a statistically significant increase (p < 0.05) in various comet parameters including the tail length, tail DNA% and olive tail moment when compared to those of the control group which confirmed the exerted genotoxic effect of acrolein. Comet assay was included in many studies to detect genotoxic effect of many classes of chemicals in vivo due to its sensitivity for detecting low levels of DNA damage [42-44]. In addition, the previous study of Gore et al. [45] showed a significant increase of tail movement and tail DNA content in the bladder cells of mice as a consequence of cyclophosphamide treatment. Cyclophosphamide was reported as an inactive prodrug that is rapidly metabolized to active metabolites, such as acrolein and phosphoramide [46]. Additionally, acrolein was considered as the main cause of the cytotoxicity of cyclophosphamide [47]. In contrast, Kuchenmeister et al. [48] observed that treatment of rat hepatocytes with acrolein resulted in an insignificant difference in the value of tail moment when compared to that of the control group by using the alkaline comet assay and they explained that their observation was probably the consequence of the known DNA and protein cross-linking activity of acrolein which helped in reducing free DNA fragments and preventing DNA migration and comet formation.
The cytoprotective effect of quercetin was explained by the study of Bakheet [49] which observed that quercetin raised the level of glutathione (GSH) and reduced malondialdehyde (MDA) levels in mice treated with a cytotoxic drug called topotecan. In the present study, oral pretreatment of quercetin was able to protect mice bone marrow cells against the acrolein-induced clastogenicity by decreasing significantly (p < 0.05) the frequency of sister chromatid exchanges compared to those of acrolein-treated group. This result coincided with results of Farombi & Onyema and Gupta et al. [17, 19] which confirmed that quercetin was an antigenotoxic and antimutagenic agent and could reduce the clastogenic effects of antitumor agents.
Results of the present study revealed that the oral pretreatment with quercetin 50 mg/ kg b.wt. prior to acrolein treatment significantly (p < 0.05) decreased the mean of tail length, tail DNA% and olive tail moment less than those of acrolein-treated group.
Experimental studies reported that quercetin is a DNA-protective agent and it is considered as an important free radical scavenging antioxidant owing to the presence of high number of hydroxyl groups and its ability to inactivate the metal iron which responsible for the generation of reactive oxygen species [50, 51]. This finding is in accordance with Devipriya et al. [52] who studied oral administration of quercetin at 3, 6, 12, 24, and 48 µM to human peripheral blood lymphocytes and observed a dose-dependently suppression of DNA strand breaks, micronuclei frequencies and comet assay parameters (tail length, tail moment, olive tail moment and % DNA in the tail) in response to gamma radiation exposure. Also, results from the previous study of Srinivas et al. [53] suggested that oral pretreatment of quercetin 50 mg/kg reduced the cytogenotoxic effect of cyclophosphamide 40 mg/kg in bone marrow cells of rats through a mechanism related to their ability to decrease oxidative stress and inflammation. Further, Sallmyr et al. [54] observed the role of Q in preventing the consequences of oxidative damage to DNA including DNA strand breaks which followed by apoptosis or mutation. Furthermore, Noroozi et al. [55] showed that pretreatment with quercetin reduced the level of oxidative DNA damage in human lymphocytes exposed to hydrogen peroxide, indicating its high antioxidant and antigenotoxic activity. Additionally, results of the experimental study of Ramos et al. [56] showed that quercetin prevented tert-butyl hydroperoxide-induced DNA damage and increased the rate of DNA repair in cultured human hepatoma cells, suggesting a genopreventive and an anticarcinogenic potential for this compound which helped in explaining the lower cancer incidence in human population with high dietary intakes of fruits and vegetables containing such flavonoid. Muthukumaran et al. [57] observed the protective role of quercetin against nicotine-induced DNA damage in male albino rats was related to its ability to modulate the extent of lipid peroxidation and enhance antioxidant defense system which reduced glutathione (GSH) level, catalase and glutathione peroxidase activities [58, 59].
In the current study, the protective effect of oral pretreatment of resveratrol dose 12.5 mg/kg b. wt. was investigated against the genotoxicity of acrolein according to the study of Mokni et al. [60] which confirmed that the optimal protective effect of resveratrol on antioxidant enzyme activities and lipoperoxidation products was obtained at this dose. Our results observed that oral pretreatment of resveratrol significantly (p < 0.05) decreased the mean of total aberrations and the frequency of sister chromatid exchanges. This finding was confirmed by various investigators who examined the antigenotoxic effect of resveratrol. Türkez & Sisman [26] demonstrated that the high concentrations of resveratrol were not genotoxic and could minimize the frequency of chromosome aberrations and sister chromatid exchanges caused by aflatoxin in human lymphocytes. Türkez & Aydin [61] observed that the combined application of the most popular environmental pollutant, Permethrin and the plant derived antioxidant, resveratrol, significantly reduced the frequency of chromosomal aberrations and the formation of sister chromatid exchanges (SCEs) in cultured human lymphocytes in comparison with alone Permethrin treated cultures.
Also, Chen et al. [62] reported that resveratrol antagonized the genotoxicity of sodium arsenite by improving cell survival and reducing DNA damage, chromosomal damage, oxidative stress and apoptosis. Moreover, resveratrol was reported in a previous in vitro study to have antioxidant effects against acrolein-induced cytotoxicity in human retinal pigment epithelial cells [63]. According to previous studies, the protective activity of resveratrol was mainly due to its free radical scavenging activity and its property to maintain intracellular antioxidants such as glutathione reductase, glutathione peroxidase, glutathione S-transferase, dismutase and catalase [64-66]. In contrast, the previous cytogenetic study of Matsuoka et al. [67] showed that resveratrol induced SCEs induction by causing S phase arrest.
Current results showed that the oral pretreatment with resveratrol 12.5 mg/ kg b.wt. prior to acrolein treatment significantly (p < 0.05) decreased the mean of comet parameters (tail length, tail DNA% and olive tail moment) less than those of the acrolein-treated group. Mokni et al. [60] revealed that the optimal effect on antioxidant enzyme activities and lipoperoxidation was obtained with resveratrol concentration of 12.5 mg/kg b.wt.
Also, this finding agreed with a number of in vitro and in vivo studies which reported resveratrol as a polyphenolic compound, exhibited genotoxic effects in cells and animal models.
Resveratrol showed protective effects in preventing lipid peroxidation in the cell membrane and DNA damage resulted from excessive reactive oxygen species production and scavenging hydroxyl, superoxide, and metal-induced radicals [68].
Further, Koohian et al. [69] evaluated the protective effect of pretreatment with various doses of resveratrol against genotoxicity induced by γ-Irradiation in mice blood lymphocytes by using the alkaline comet assay and the results of their study showed that RES significantly inhibited radiation induced DNA damage.
Otherwise, the study of Shafie et al. [70] showed that even at high concentrations of resveratrol, no considerable protective effect was observed against toxicity of acrolein in rat liver cells. Gatz & Wiesmüller [71] suggested that the antimutagenic effect of resveratrol was probably due to one of the following mechanisms; including its effect on the metabolic activation of mutagens, de novo expression of genes that encode detoxifying proteins, inhibition of inflammation, radical scavenging properties, formation of acrolein-resveratrol adducts and blocking DNA-adducts formation.
The present findings of comet assay showed that the preventive effects of quercetin alone, resveratrol alone or a combination of quercetin/resveratrol against acrolein-induced DNA damage were similar without clear differences.
However, results of the previous study of Celik & Arinc [72] indicated that quercetin was the most powerful antioxidant in preventing DNA damage and the protective effects of several phenolic antioxidants, rutin, naringenin, resveratrol and trolox, against idarubicin (anticancer drug)-induced DNA strand breaks were significantly less than that of quercetin even at high concentrations.
But, ElAttar & Virji [73] reported that cancer chemopreventive role of resveratrol or a combination of resveratrol and quercetin were more effective than that of quercetin alone in inhibition of oral squamous carcinoma cell growth and proliferation. Moreover, results of the present study revealed that quercetin 50 mg/kg and resveratrol 12.5 mg/kg were not genotoxic, as there were no significant changes in the comet parameters over the corresponding quercetin-vehicle control group and the corresponding resveratrol-vehicle control group, respectively. This finding was in agreement with Barcelos et al. [74] who confirmed that quercetin was not genotoxic and showed considerable protection against a wide range of mutagens. Also, Juan et al. [75] demonstrated that RES was not toxic even at high concentrations as neither weight loss nor lactate dehydrogenase (LDH) release in the plasma occurred.
The current investigation revealed that neither quercetin nor resveratrol were clastogenic or cytotoxic at the doses tested. Also, Attia [18] observed that quercetin at doses equivalent to 50 or 100 mg/kg failed to induce chromosomal aberrations in bone marrow cells of mice which indicated its non-clastogenicity. Boersma et al. [76] revealed that quercetin was non-clastogenic because of degradation of its aglicone form caused by intestinal bacterial flora or O-methylation, glucuronidation and sulphatation in the gastrointestinal tract. In addition, the previous in vitro study of observed that resveratrol treatment did not cause chromosomal aberrations induction in a Chinese hamster lung (CHL) cell line [67].