Abstract
Dental X-rays are widely used in clinical practice, since the technique is an important approach for diagnosing diseases in dental and periodontal tissues. However, it is widely known that radiation is capable of causing damage to cellular systems, such as genotoxicity or cytotoxicity. Thus, the aim of this review was to present a critical analysis regarding the studies published on genotoxicity and cytotoxicity induced by dental X-rays in oral mucosa cells. Such studies have revealed that some oral cell types are more sensitive than others following exposure to dental X-rays. Certainly, this review will contribute to a better understanding of this matter as well as to highlighting perspectives for further studies. Ultimately, such data will promote better safety for both patients and dental professionals.
- Dental X ray
- genotoxicity
- cytotoxicity
- review
Genotoxicity is the ability of physical, chemical or biological agents to induce damage to genetic material (1). This means that such an agent needs to interact directly with the genome in order to promote genotoxicity. Cytotoxicity is defined as the capacity for inducing cell death (2). Several biological mechanisms of cell death have been described in the literature such as necrosis, apoptosis and autophagy, among others.
Dental X-rays are widely used in clinical practice because the technique is an important approach for diagnosing diseases of oral tissues (3). Among its advantages, it is fast, has low cost and great ease of obtaining images. In recent decades, some advances in obtaining images from soft and mineralized tissues have been made in dental practice, with the introduction of computed tomography and cone beam computed tomographic techniques. In fact, these methods present a significant improvement in the presentation of images of dental structures, as they use a three-dimensional coordinate system, despite having a high cost when compared to conventional radiographs (4).
Although dental X-rays have great benefits in clinical practice, it has been widely documented that radiation is able to induce injury to eukaryotic cells (5). This is due to the fact that exposure to radiation induces the formation of reactive oxygen species in eukaryotic cells, which in turn lead to damage to genetic material, lipids and proteins (6). This inevitably leads to genotoxicity and cytotoxicity in a dose-dependent manner.
Nowadays, there are several recognized methodologies that are able to detect and, therefore, measure DNA strand breaks under different protocols and paradigms (7). Additionally, several signaling pathways have been investigated in order to better understand the processes involved in cell death (2). These methodologies are of fundamental importance in verifying the ability of any agent to interact negatively with the genome or activate signaling pathways closely related to cell death. This information predicts the risk for human populations which are exposed to toxic compounds (8). It is important to stress that biological processes participate in the development of oral cancer, since they are involved to the initiation and promotion phases of carcinogenesis (9).
The aim of this review was to present a critical analysis regarding published studies on genotoxicity and cytotoxicity induced in oral mucosa cells by dental X-rays. Certainly, this work will contribute to a better understanding of this matter as well as to highlighting new perspectives for further studies. Ultimately, such data will help increase safety for both patients and dental professionals.
For this purpose, a comprehensive literature search for studies on DNA damage, genetic damage, genotoxicity, cytotoxicity, cellular death and dental X-rays was performed between 2000 and 2017. In brief, a search of PubMed, MEDLINE, Embase and Google Scholar for a variety of articles (all publications until June 2017) was carried out using the key words mentioned above. Case reports and articles not written in English were excluded from the review. All articles were identified and included in this review.
Genotoxicity, Cytotoxicity and Dental X-Ray Exposure
After reviewing the scientific literature, we found few published articles regarding genotoxicity, cytotoxicity and dental X-ray exposure. Such findings are summarized in Table I and Figure 1.
The first study investigating genotoxicity and cytotoxicity in oral mucosal cells was published in 2004 by Cerqueira et al. (10). In this study, the authors evaluated genetic and cytotoxic injury in buccal mucosa cells before and after panoramic dental X ray exposure. The evaluation was conducted by micronucleus test in adults. The findings showed that frequencies of micronuclei, karyolysis and pycnosis were similar following dental X-ray exposure, whereas chromatin condensation and karyorrhexis increased significantly after exposure (10). Others who evaluated the genotoxic and cytotoxic potential of panoramic radiographs using the micronucleus test also demonstrated the same results found above (11-13).
Recently, new technologies for evaluating the micronucleus assay were applied, such as the cythome assay in individuals submitted to intraoral periapical radiography, conventional and digital panoramic X-ray exposure. The results also indicated significant differences in the micronucleated cell frequency when comparing before and after dental X-ray exposure (14). Moreover, X-ray exposure increased other nuclear alterations indicative of cytotoxicity, such as karyorrhexis, pyknosis and karyolysis (14). It is important to stress that the new techniques allow for a more accurate evaluation of nuclear changes when compared to the conventional micronucleus test using conventional light microscopy. Conversely, one study demonstrated genotoxic effect following panoramic radiography exposure since a high increase of the micronuclei frequency was detected in buccal epithelial cells (15). In this study, a non-specific technique for staining nucleic acids was used. This procedure interferes with the recognition of micronucleated cells and leads to the occurrence of false-positive results.
In order to clarify if some dental X-ray techniques are more sensitive than others, the micronucleus test was applied to healthy patients submitted to lateral and frontal cephalometric X-ray referred for orthodontic therapy. The results revealed no statistically significant differences in the frequency of micronucleated oral mucosa cells after X-ray exposure (16). However, exposure to dental X-rays was able to increase other nuclear alterations closely related to cytotoxicity, such as karyorrhexis, pyknosis and karyolysis (16). These data indicate that dental radiography does not induce genotoxicity as a result of chromosomal breakage or loss, but it is able to promote cytotoxicity as depicted by the increase of some nuclear changes indicative of cellular death. Cone beam computed tomography was found to induce the same findings as conventional dental X-ray. Carlin et al. showed that cone beam computed tomography did not induce micronuclei formation in buccal mucosa cells. However, increased cytotoxicity was noted by the authors (17).
To further elucidate the biological mechanisms involved in cellular death, some authors collected exfoliated oral epithelial cell samples from healthy individuals undergoing routine dental radiographic examination (2.34 cGy) after radiograph exposure. The results showed that the phosphorylation of histone and cyclin-dependent kinase 2 were induced in oral cells after exposure as assessed by immunohistochemical assay (18). Apart from these results, biomarkers related to cell-cycle regulation may also be useful in assessing the harmful effects of dental X-rays at the molecular level. Therefore, such results provide other information on the mechanisms of cytotoxicity induced by dental X-ray as detected by the micronucleus test.
Smoking has been recognized as one of the leading causes of oral cancer (19). This is because cigarette smoke contains several components capable of promoting genotoxicity and cytotoxicity, contributing to the process of oral carcinogenesis (19). Herein, it would be interesting to evaluate whether and to what extent smokers are more susceptible than non-smokers with respect to the effects of dental X-rays. This approach is coherent and relevant, particularly because a concomitant exposure to agents from physical and chemical sources is able to potentiate the harmful outcomes in the oral mucosa. Following this rationale, Angelieri et al. performed a study to evaluate DNA damage (micronucleus) and cellular death (pyknosis, karyolysis, and karyorrhexis) in exfoliated oral mucosa cells from smokers and non-smokers submitted to panoramic X-ray for orthodontic purposes using two anatomic sites: the buccal mucosa and lateral border of the tongue (20). The results showed no significant statistically differences in micronucleated oral mucosa cells before versus after X-ray exposure for both oral sites evaluated in smokers and non-smokers (20). Nevertheless, panoramic X-ray exposure was able to increase other nuclear alterations closely related to cytotoxicity such as karrhyorexis, pyknosis, and karyolysis (20).This was confirmed by others investigating the same issue (12). Curiously, the most pronounced effects for cytotoxicity were found in the lateral border of the tongue of smokers (20). Similar effects were observed in non-smokers exposed to panoramic dental X-ray in a dose-response fashion (21). In summary, these data indicate that panoramic X-ray is able to induce cellular death of oral mucosa cells. It seems that the lateral border of the tongue is more sensitive than other sites to cytotoxic insult induced by ionizing radiation combined with continuous exposure to cigarette smoke (20).
When gingival cells were evaluated, discordant findings were obtained when comparing to buccal mucosa cells. For example, panoramic dental radiography was able to induce genotoxic effects on epithelial gingival cells as a result of increased frequency of chromosomal damage and nuclear alterations indicative of apoptosis, such as karyorrhexis and chromatin condensation (22). Recently, Sheikh et al. demonstrated that micronucleus frequency in gingival epithelial cells was also higher after exposure to panoramic dental radiography (23). Using an in vitro test system, human gingival cells showed DNA strand breaks after exposure to high single-dose irradiation used for tumor therapy (24). In buccal mucosa cells, the frequency was not statistically different (23). A significant correlation was observed between the age of individuals and micronucleus frequency (11, 15, 23). This finding has been reported in the scientific literature since there is an increase in the frequency of micronuclei with advancing age (25).
Taking into consideration that children are developing organisms (26), these individuals are more susceptible to the insult triggered by genotoxic and cytotoxic agents. For example, it has been established that xenobiotic metabolizing enzyme systems and DNA repair systems are not fully developed in children (27). Orthodontic therapy is frequently performed during childhood; radiographic documentation is mandatory for clinical diagnosing and subsequent planning of orthodontic therapy. In this context, some researchers have struggled to elucidate if children are more susceptible to the effects of dental X-rays. No increase of micronucleated oral mucosa cells were noticed after exposure to radiation induced by panoramic dental X-rays (28-30). On the other hand, radiation did cause other nuclear alterations closely related to cytotoxicity, such as karyorrhexis, pyknosis or karyolysis (28). In an early study conducted by Lorenzoni et al., healthy children undergoing orthodontic therapy were submitted to a complete set of orthodontic radiographs (lateral cephalographic, posteroanterior cephalographic, panoramic, full periapical exam, and bitewing). There were no statistically significant differences in micronucleated buccal mucosa cells after exposure to radiation. However, radiation was able to promote cytotoxicity in buccal mucosa cells (31).
When a comparative analysis was performed between adults and children, the results showed similar findings between groups (33). No statistically significant differences in micronucleated oral mucosa cells before and after dental X-ray exposure were noted. On the other hand, X-rays increased other nuclear alterations closely related to cytotoxicity such as karyorrhexis, pyknosis and karyolysis in both groups (33). Cone beam computed tumography led to similar findings as those found for conventional dental X-rays. No statistically significant differences were found in micronucleated buccal mucosa cells after exposure to cone beam radiation (32). However, the radiation technique induced other nuclear alterations closely related to cytotoxicity, including karyorrhexis, pyknosis, and karyolysis (32). Further studies are welcomed to elucidate this issue.
Taken together, it is assumed that children do not present a more susceptible group when evaluating genotoxicity and cytotoxicity induced in oral mucosa cells by dental X-ray.
Concluding Remarks and Future Perspectives
In this review, we conclude that dental X-rays are not able to induce genetic damage, but can promote cellular death. It is important to stress that gingival and tongue cells are more sensitive to insults induced by dental X-rays when compared to buccal mucosa cells.
After searching the literature, it was found that all published in vivo studies were conducted using the micronucleus test. Taking into consideration that the micronucleus assay detects chromosomal damage only, it is very important to employ other methodologies for evaluating genotoxicity induced by dental X-rays, especially those able to detect DNA single- and double-strand breaks, DNA adducts, point mutations and others. Such an approach is particularly relevant because cytotoxicity may cause an underestimation of the micronucleus frequency due to cell death. Moreover, further studies evaluating cell-cycle control are also important to clarify the effect of dental X-rays on oral mucosa cells at the cellular and molecular levels. For this purpose, in vitro studies using different dental X-ray techniques are interesting to better understand the role of cell signaling pathways in oral mucosa cells.
In another context, few studies have been published evaluating the genotoxic and cytotoxic potential of dental X-rays in smokers. Such information is also very important, since cigarette smoke is the main chemical inducer of oral cancer. Finally, children do not represent a high risk group for exposure to dental X-rays.
In summary, this is an area that warrants additional investigation not only to understand the real risks presented by dental X-rays to oral mucosa cells, but also to ensure better safety of patients and dental professionals.
Acknowledgements
DAR is a recipient of the National Council for Scientific and Technological Development (CNPq) fellowship.
Footnotes
Conflicts of Interest
None declared.
- Received July 7, 2017.
- Revision received July 28, 2017.
- Accepted August 1, 2017.
- Copyright© 2017, International Institute of Anticancer Research (Dr. George J. Delinasios), All rights reserved