The epithelial cell kinetics is especially important in the interpretation of results obtained as a result of low dose exposure of x rays from orthopantamograph. Chromosomal alterations leading to nuclear anomalies occurs in dividing cells from basal layer of oral epithelium, but is only observed later in exfoliated cells after the differentiation. A case control study was conducted during July 2008-April 2009 to assess the genetic damage from exfoliated cells of oral mucosa in individuals subjected to panoramic dental radiography. The study population consisted of normal healthy subjects who were attending the extra oral radiology department in Ragas Dental College and Hospital, Chennai. The study group comprised of 35 subjects of both sexes in which, 18(51.43%) males and 17 (48.57%) females. All the subjects were adults between the age group of 16 to 38 years of age. For each subject, two sets of cytological smears were prepared immediately before and 10 days after exposure to panoramic dental radiography. The smears were stained by using Schiff’s staining and analyses were performed in a blind fashion among 2000 cells. The following nuclear alterations were considered: micronucleus, karyorrhexis, condensed chromatin, pyknosis, karyolysis, broken eggs, nuclear buds and nuclear projections. The alterations were identified under light microscope.
To summarize the results of the study:
1. In the study the occurrence of micronucleus frequencies were not altered before and after exposure with p value of 0.54 and is statistically insignificant which states that panoramic dental radiography does not produce chromosomal alterations.
2. The presence of karyorrhexis was increased after the exposure with a p value of 0.001 which is statistically significant which is indicative of apoptosis.
3. The presence of condensed chromatin was increased after the exposure with a p value of 0.001 which is statistically significant which is indicative of apoptosis.
4. The presence of pyknosis was increased after the exposure with a p value of 0.001 which is statistically significant which is indicative of apoptosis.
5. The presence of karyolysis were increased after the exposure with a p value of 0.009 which is statistically significant suggesting that the cellular response to x rays produce a cytotoxic effect which may lead to necrosis.
6. The presence of broken eggs was increased after the exposure with a p value of 0.001 which is statistically significant and should be considered as genotoxicity bio marker.
7. The presence of nuclear buds was increased after the exposure with a p value of 0.31 which is statistically not significant and is indicative of normal epithelial differentiation.
8. The presence of nuclear projections was increased after the exposure with a p value of 0.15 though not statistically significant it is indicative of mild cellular damage due to radiation.
9. On comparison between pre exposure and post exposure values among males the p value for karyorrhexis was 0.003, condensed chromatin p=0.001 pyknosis p=0.05, karyolysis p=0.02 and broken eggs p=0.002 these values are statistically significant before and after exposure. The p value for micronuclei was 0.74, nuclear buds p=0.39 and nuclear projections p=0.14, these values are statistically not significant before and after exposure.
10. On comparison between the pre exposure and post exposure values among females the p value for karyorrhexis was 0.001, condensed chromatin p=0.001 pyknosis p=0.01, and broken eggs p=0.006 these values are statistically significant before and after exposure. The p value for micronuclei was 0.74, karyolysis p= 0.16, nuclear buds p=0.39 and nuclear projections p=0.14 these values are statistically not significant before and after exposure.
11. On comparison of the pre exposure values between males and females the p value for micronuclei was 0.64, karyorrhexis p=0.51, condensed chromatin p=0.81, pyknosis p=0.16, karyolysis p= 0.16, broken eggs p=0.58, nuclear buds p=0.71 and nuclear projections p=0.81 these values are statistically not significant between males and females which indicates that sex doesn’t influence the formation of nuclear anomalies.
12. On comparison of the post exposure values between males and females the p value for micronuclei was 0.71, karyorrhexis p=0.25, condensed chromatin p=0.38, pyknosis p=0.27, karyolysis p= 0.40, broken eggs p=0.30, nuclear buds p=0.40 and nuclear projections p=0.40 these values are statistically not significant between males and females which indicates that sexually there is no difference between males and females in formation of nuclear anomalies.
The present study analyzed the epithelial cells from oral mucosa because this anatomical location is centrally located when the source of radiation moves around the head of patient in the radiographic technique we adopted. Panoramic radiographs are frequently requested by dentists and incorrect positioning of the patient may require the procedure to be repeated. In human cytogenetic studies, it is important to consider some confounding factors. Viruses, alterations in the immune system, failures in DNA repair system and individual variations have already been associated with increased frequencies of chromosome aberrations. Moreover, the influence of tobacco smoke has usually been considered as a relevant confounding factor. Thus, all adults recruited to participate in this study were non-smokers. Due to the cost factors in depth investigations like any DNA analysis, FISH analysis, nuclear alterations in lymphocytes was not assessed. According to the results from this investigation, exposure to Xrays during panoramic radiography induces genotoxic effects in oral mucosal buccal epithelial cells that increase chromosomal damage and induce apoptosis. Thus panoramic dental radiography should be requested only when necessary because it cannot be considered a risk free procedure. It is also recommended that the expanded protocol for the micronucleus test suggested by Tolbert [et al.] 1991 should be adopted, including not only micronuclei but also other types of nuclear abnormalities that are in themselves cell damage markers. The frequencies of nuclear alterations indicative of apoptosis (karyorrhexis and condensed chromatin) were significantly higher after the exposure in contrast to micronuclei results. Apoptosis is a fundamental biological process, which is genetically controlled and required for normal development and tissue homeostasis. The results from the study showed that panoramic dental radiography induced the apoptotic response, which probably interfered with the micronucleus induction.
In some cases the cells with nuclear anomalies were greater before x-rays suggesting that they may be associated with normal process of cell differentiation.
Taken as a whole, such results support the notion that X-rays are a cytotoxic agent. It is important to stress that cytotoxicity interferes with micronucleus induction since some MN are inevitably lost after cytotoxic insult, therefore confirming the lack of mutagenic effect induced by X-rays. Nevertheless, it has been postulated that repeated exposure to cytotoxicants can result in chronic cell injury, compensatory cell proliferation, hyperplasia and, ultimately, tumor development. In fact, a correlation between cell proliferation and induction of cancer is assumed. Proliferation probably increases the risk of mutations within target cells, and may also be important in selective clonal expansion of (exogenously or endogenously) initiated cells from preneoplastic foci and eventually tumors. Our results demonstrated that the micronucleus frequency did not increase following exposure to ionizing radiation.
In conclusion, the results of the present study indicate that high levels of genotoxicity and cytotoxicity in exposed tissues, expressed respectively by increased apoptotic or necrotic responses may be a factor in the low micronucleus frequencies observed after x- ray exposure suggesting that X-rays can induce cytotoxic effects in oral mucosal cells. The risks associated with dental radiographs are small but should not be overlooked.
Since cellular death is considered to be a prime mechanism in non-genotoxic mechanisms of carcinogenesis, dental X-rays should be used only when necessary. More frequent, both as substitute for and as a complement to intra oral radiographs, their indication should always follow the concept of maximum benefit with minimum risk. Panoramic radiography should be carefully performed in order to avoid to retakes and increase in radiation doses
