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Age estimation in adult human sound and periodontally affected teeth using tooth cementum annulations
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Available online at www.sciencedirect.com
ScienceDirect Tanta Dental Journal xx (2015) 1e9 www.elsevier.com/locate/tdj
Age estimation in adult human sound and periodontally affected teeth using tooth cementum annulations W.Y. Alghonamy*, O.M. Gaballah, D.A. Labah Faculty of Dentistry, Tanta University, Oral Biology Department, Egypt Received 16 June 2015; revised 3 July 2015; accepted 26 July 2015
Abstract Background: There are many tools for identification of human remains, via radio graphical imaging, histological processing & microscopical examination of dental remains. Purpose: to estimate the age of both female and male adults using tooth cementum annulations (TCA) method, evaluate the accuracy of TCA of both sound and periodontally affected teeth, and compare the estimated age (Est.age) with the chronological age (Chr.age) of an individual by the aid of; light microscope (LM), polarized microscope (PM) and phase contrast microscope (PHM). Material and methods: 60 teeth divided into four equal experimental groups; group I contained sound teeth extracted from female patients, group II contained sound teeth extracted from male patients, group III contained periodontally affected teeth extracted from female patients and group IV contained periodontally affected teeth extracted from male patients. Longitudinal ground section for all groups were prepared & observed under LM, PM and PHM. Afterward, images were taken for each section then the average of TCA for each section was counted. The age was estimated using the formula {Estimated age ¼ Numbers of incremental lines þ Eruption age of tooth}. Results: There was no significant difference between Est.age and Chr.age of both male and female sound teeth (group I and group II) while in periodontally affected teeth of both male and female (group III and group IV) there was significant difference. Also, there was a strong correlation between the Est. & Chr. age when PHM was used followed by LM then PM. Finally, there was no sex difference. Conclusion: TCA is reliable for sound but not for periodontally affected teeth using either LM, PM or PHM. Moreover, PHM is a more accurate microscope for TCA in contrast to LM & PM. Interestingly, a novel regression formula was developed to enable estimating age in periodontally affected teeth by TCA method. © 2015, Hosting by Elsevier B.V. on behalf of the Faculty of Dentistry, Tanta University.
Keywords: Age estimation; Tooth cementum annulations; Periodontitis; Light microscope; Polarized microscope; Phase contrast microscope
* Corresponding author. Alshahid fekry, Berkit el sabee, Minufia, Egypt. Tel.: þ20 0482977257, þ20 01066754527 (mobile). E-mail addresses: [email protected] (W.Y. Alghonamy), [email protected] (O.M. Gaballah), [email protected] (D.A. Labah). Peer review under the responsibility of the Faculty of Dentistry, Tanta University.
1. Introduction Forensic odontology (Forensic dentistry) is a specialized branch of forensic medicine which, in the interests of justice, deals with examination of dental
http://dx.doi.org/10.1016/j.tdj.2015.07.002 1687-8574/© 2015, Hosting by Elsevier B.V. on behalf of the Faculty of Dentistry, Tanta University. Please cite this article in press as: Alghonamy WY et al., Age estimation in adult human sound and periodontally affected teeth using tooth cementum annulations, Tanta Dental Journal (2015), http://dx.doi.org/10.1016/j.tdj.2015.07.002
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evidence from which a proper evaluation and presentation of dental findings can be made [1]. Age determination plays an important role in forensic medicine, not only in identification of bodies but also in connection with crimes. When the subjects have undergone changes so extensive that external characteristics yield no information, the teeth are often the only means of identification. Also, Human tooth can be preserved for a long time after death without gross changes so; it serves as an important tool in forensic science [2]. Various methods are utilized for determination of age from dentition. These may be described in four categories namely, clinical, radiographic, histological, physical and chemical analysis [3]. Apart from above mentioned techniques, a histological method based on counting the incremental lines of tooth cementum has shown promise for age estimation method in adult as the biological factors of cementum formation is well established so far. Tooth cementum is the calcified ectomesenchymal tissue that surrounds the dentine and forms the attachment site for the periodontal ligaments that support the tooth to the surrounding alveolar bone [4]. Microscopically, cementum appears as a series of alternating light and dark lines or bands that have the appearance of tree annual rings called TCA [5,6]. In acellular cementum, many fine incremental lines of about 0.5 micron width were observed that ran perpendicularly or obliquely to Sharpey's fibers. Also, these lines are not straight lines but had wave-like structure. Additionally, the fine incremental lines showing a close relationship to the degree of calcification, they also seemed to show a strong relationship to the quality of the organic component of acellular cementum [7]. In respect of cementum position, it has not been used to the extent of enamel and dentin to estimate the age of unidentified individuals. However, many studies recommended that counting of TCA in thin histological sections and adding them to the mean eruption age of the tooth may offer a more accurate method for age estimation in human beings and can be used as a more reliable age marker than any other morphological or histological traits in the human skeleton based on the biological factors of TCA [8]. For many years, it was reported that age determination using TCA could be an accurate means of age determination in animals that live on a relatively constant diet throughout life [9,10]. On the other hand, numerous studies were done to estimate ages by TCA method in human sound teeth using different types of microscopes and considered this method to be extremely valuable in forensic medicine, forensic dentistry, and anthropology [11,12].
Surprisingly, It has been shown that, few studies were done on age estimation using TCA method in periodontally affected teeth and these studies exhibited many contrasts. Therefore, it is of prime importance to study the efficiency of using TCA in age estimation of adult male and female using both sound and periodontally affected teeth with the aid of three microscopes; LM, PM & PHM and to compare the Est. ages with the Chronological ages of the individuals. 2. Materials and methods 2.1. The sample
Sixty extracted single rooted teeth were used in this study including both sound and early periodontally affected teeth. They were obtained from the Oral Surgery and Maxillofacial Department, Faculty of Dentistry, Tanta University. Teeth were extracted as a part of essential clinical care from patients ranging from 20 to 70 years old of both sexes with well known Chr. age and well known date and reason of extraction. Both written and oral consents were obtained from the patients. The protocol was approved from ethical committee at Faculty of Dentistry, Tanta University. Afterward, the selected teeth were divided into four equal experimental groups each one consists of fifteen teeth. Group I: sound single rooted teeth extracted from female patients, Group II: sound single rooted teeth extracted from male patients, Group III: periodontally affected single rooted teeth extracted from female patients and Group IV: periodontally affected single rooted teeth extracted from male patients. 2.2. The method 2.2.1. Histological study
The extracted teeth were preserved in 10% buffered formalin then they were washed under running water. Thin longitudinal ground histological sections were done as reported by Aggarwal et al. [13]. After excluding unbeneficial sections, the ground sections in each group were examined by light, polarized and phase contrast microscopes. Afterward, images were taken for the middle third of the root of each section using the three microscopes at magnification 400 for areas of the cementum showing uniform thickness, clear definition of layers from the cementodentinal junction (CDJ) to the outer border of the root then images magnified by computer. The average of TCA for each section was counted by image analysis system as follow: Counting the dark lines manually at the
Please cite this article in press as: Alghonamy WY et al., Age estimation in adult human sound and periodontally affected teeth using tooth cementum annulations, Tanta Dental Journal (2015), http://dx.doi.org/10.1016/j.tdj.2015.07.002
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monitor based on digitally enhanced images using the measurement overlay (Image J program).1 Counting the TCA had been done by three observers to avoid inter-observer variability. When there was discrepancy between different counts, the mean value was used for statistical analysis. Then, each observation was repeated after one week to evaluate intra-observer error. One lightedark pair was considered as one annulation which corresponds to one year age of the individual [14]. Formula for estimation of age at death or time of tooth extraction was calculated by adding average age of tooth eruption in years for each tooth as represented by Schour and Massler [15] to the counted number of incremental lines [16]. {Estimated age (E) ¼ Numbers of incremental lines (n) þ Eruption age of tooth (t)}. 2.2.2. Statistical study
All the data was recorded, tabulated and statistically analyzed using software SPSS V.20. Statistical presentation and analysis of the present study was conducted using the mean and standard deviation in the following tests: A) Analysis of variance [ANOVA] tests, B) Post Hoc test., C) Regression analysis and equation [16]. 3. Results 3.1. Histological results Histological study revealed that TCA were more clearly visible under the PHM followed by LM then PM. Interestingly, microscopic observations of the cementum in group I & group II gave similar results as they showed uniform thickness, clear definition of TCA from the CDJ to the outer borders of the root that appeared as a series of alternating light and dark lines or bands and each one lightedark pair was considered as one annulation. As well, calculation of Est. age by adding eruption age of the tooth to total number of TCA illustrated accordance between the Est. age by TCA and Chr. age for both groups but for patients older than 50 years there was some divergence (Fig. 1 &Fig. 2). On the other hand, microscopic observations of the cementum in group III& group IV exhibited variability in cementum thickness even in different areas of the same section. Moreover, some areas of cementum showed resorption (saucer shaped) that could be considered as a manifestation of periodontitis. Also, counting TCA and calculation of Est. age 1 This is a software tool allows marking each line by mouse click which is then summed.
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according to the previous equation illustrated divergence between the Est. age and Chr. age of the patients (Fig. 3 & Fig. 4). 3.2. Statistical results 3.2.1. Comparison between the Est. ages using TCA & the Chr. ages in each group by ANOVA test See Table 1. 3.2.2. Comparison between the Est. ages with different microscopes (LM, PM & PHM) & the Chr. ages in each group by post hoc test See Table 2. 3.2.3. Comparison between the Est. ages by each microscope (LM, PM or PHM) among the four different groups by post hoc test See Table 3. 3.2.4. Regression analysis and equation 1-Age estimation of group III with L.M (See Table 4). 2-Age estimation of group III with PM (See Table 5). 3-Age estimation of group III with PHM (See Table 6). 4-Age estimation of group IV with L.M (See Table 7). 5-Age estimation of group IV with PM (See Table 8). 6-Age estimation of group IV with PHM (See Table 9). 4. Discussion Teeth are particularly useful in age estimation as they display a number of observable age related variables and they tend to remain intact under circumstances which might alter or obliterate the rest of the skeleton. The present study hypothesized that the cementum annulations may serve as a powerful method for age determination in both male and female sound single rooted teeth and how this method may be valuable even in periodontally affected teeth. Briefly, this discussion covered two essential points first; the validity and accuracy of TCA methods in age estimation of both sound and periodontally affected male and female single rooted teeth. Second, the most accurate microscope that achieve our goal. Using TCA in age estimation was based on the facts reported by Lieberman [14] that during cementum formation, the hyper mineralized layers of extracellular matrix alternate with less mineralized layers causing the cementum to appear as a series of alternating light
Please cite this article in press as: Alghonamy WY et al., Age estimation in adult human sound and periodontally affected teeth using tooth cementum annulations, Tanta Dental Journal (2015), http://dx.doi.org/10.1016/j.tdj.2015.07.002
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Fig. 1. Micrographs of (group I) sound upper right central incisor of 60 years old female patient. A, LM depicts the total number of TCA is 46 (yellow dots). B, PM shows the total number of TCA is 45 (yellow dots). C, PHM shows the total number of TCA is 51 (red dots), (L.S. microscopic magnification.X 400). The Est. age by LM ¼ 46 þ 8 ¼ 54 years, by PM ¼ 45 þ 8 ¼ 53 years and by PHM ¼ 51 þ 8 ¼ 59 years.
and dark lines or bands and each one lightedark pair was considered as one annulation which corresponds to one year age of the individual. Thus, counting these lines in thin histological sections and adding them to the mean eruption age of the tooth will give the Chr. age of the individual [17]. In the current study, the sample included extracted teeth from patients ranging between 20 and 70 years old since by the age of 20 years growth and dental development were completed as confirmed by Someda et al. [18]. Notably, examination of TCA using LM allowed good observation of TCA. This is in agreement with
Stott et al. [11]. In contrary, Radovic [19] who found that incremental lines in the cementum could not be read from one third of an archeological sample when using LM. On the other hand, the present work illustrated that observation of TCA using PM gave a good observation for cementum annulations in all groups in most of the samples but in some samples PM couldn't enable clear TCA observation. The proper PM observation was advocated by Wedel [20]. Moreover, In the current study, LM revealed more obvious observations than PM. and PHM revealed excellent observation of annulations. The clarity of
Fig. 2. Micrographs of (group II) sound upper right lateral incisor of 34 years old male patient A, LM depicts the total number of TCA is 25. B PM shows the total number of TCA is 24. C, PHM shows the total number of TCA is 25. (L.S. microscopic magnification. 400). The Est. age by LM ¼ 25 þ 9 ¼ 34 years, PM ¼ 24 þ 9 ¼ 33 years and PHM ¼ 25 þ 9 ¼ 34 years. Please cite this article in press as: Alghonamy WY et al., Age estimation in adult human sound and periodontally affected teeth using tooth cementum annulations, Tanta Dental Journal (2015), http://dx.doi.org/10.1016/j.tdj.2015.07.002
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Fig. 3. Micrographs of (group III) for periodontally affected lower right central incisor for 70 years old female patient A, LM shows the total number of TCA is 42 (yellow dots). B, PM shows the total number of TCA (yellow dots) is 36 in the thick part and 21 in the thin part (the resorbed area) with the mean of TCA ¼ 28.5. C, PHM shows the total number of TCA (yellow dots) is 38 in the thick part and 31in the thin part with the mean of TCA is 34.5. (L.S. microscopic magnification. 400). The Est. age by LM ¼ 42 þ 7 ¼ 49 years, by PM ¼ 28.5 þ 7 ¼ 35.5 years and PHM ¼ 34.5 þ 7 ¼ 41.5 years.
TCA by PHM was explained by Sanderson [21] who discussed how PHM gave the proper distinguishing between two types of layers with different optical properties. He found that PHM depends on refraction of light by the specimen as it utilizes the difference between light rays propagating directly from the light source and light rays refracted by the specimen when light passes through it to add bright/dark contrast to images of transparent specimens and this done by the aid of a phase-contrast objective and a condenser fitted with PHM for observations. He depicted that PHM is
suitable for viewing colorless and transparent specimens (as cementum). In the present histological study, counting TCA using LM, PM and PHM for the same cementum area in each ground section followed by calculation of Est. age by adding eruption age of the tooth to total number of TCA illustrated accordance between the Est. age and Chr. age in sound teeth of female and male patients less than 50 years however, there was some divergence between the Est. age and Chr. age in elders. This is in agreement with Condon et al. [22] who depicted that
Fig. 4. Micrographs of (group IV) for periodontally affected lower left canine of 46 years old male patient. A, LM shows total number of TCA is 38. B, PM shows total number of TCA is 34. C, PHM shows total number of TCA is 37. (L.S. microscopic magnification. 400). The Est. age by LM ¼ 38 þ 10 ¼ 48 years, by PM ¼ 34 þ 10 ¼ 44 years and by PHM ¼ 37 þ 10 ¼ 47 years. Please cite this article in press as: Alghonamy WY et al., Age estimation in adult human sound and periodontally affected teeth using tooth cementum annulations, Tanta Dental Journal (2015), http://dx.doi.org/10.1016/j.tdj.2015.07.002
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Table 1 Comparison of mean ± SD values of Est. ages & Chr. ages in each group using ANOVA test. N Mean ± SD Group I
Chr.age Est.age with Est.age with Est.age with Total Group II Chr.age Est.age with Est.age with Est.age with Total Group III Chr.age Est.age with Est.age with Est.age with Total Group IV Chr.age Est.age with Est.age with Est.age with Total
15 60.07 ± LM 15 52.60 ± PM 15 49.40 ± PHM 15 53.27 ± 60 53.83 ± 15 47.87 ± LM 15 46.33 ± PM 15 44.07 ± PHM 15 47.27 ± 60 46.38 ± 15 59.47 ± LM 15 51.33 ± PM 15 44.60 ± PHM 15 53.07 ± 60 52.12 ± 15 53.20 ± LM 15 43.67 ± PM 15 40.73 ± PHM 15 47.07 ± 60 46.17 ±
14.723 15.231 13.979 14.280 14.717 13.799 14.356 12.533 13.946 13.403 9.273 14.226 10.176 12.792 12.661 7.476 5.912 7.166 8.860 8.616
Range p-value 74e30 77e27 68e25 71e26 77e25 60e20 64e20 59e23 65e25 65e20 70e45 76e30 59e24 84e33 84e24 68e44 53e32 53e24 71e36 71e24
NS
0.246
0.880NS
0.011*
0.000**
(*) Means there is a significant difference when P. value < 0. 05. (**) means highly significant difference when P. value < 0.001. (NS) means no significant difference when P. value > 0.05.
annual deposition of cementum rings exhibited a decrease with increasing age. On the other hand in the present histological work, periodontally affected teeth that were taken from female and male patients exhibited variability in cementum shape with some noticeable areas of Table 2 Comparison of Est. ages with LM, PM, & PHM and Chr. ages in each group using post hoc test. Chr. Age Est. age Est. age Est. age by L.M by PM by PHM Group I
Chr. Age Est. age by Est. age by Est. age by Group II Chr. Age Est. age by Est. age by Est. age by Group III Chr. Age Est. age by Est. age by Est. age by Group IV Chr. Age Est. age by Est. age by Est. age by
e L.M 0.166NS PM 0.050NS PHM 0.206NS e L.M 0.760NS PM 0.450NS PHM 0.905NS e L.M 0.064NS PM 0.001* PHM 0.143NS e L.M 0.001* PM 0.000** PHM 0.028*
Table 3 Comparison of Est. age by LM, PM or PHM in all groups using post hoc test.
e 0.550NS e 0.901NS 0.470NS e e 0.652NS e 0.852NS 0.524NS e e 0.123NS e 0.689 NS 0.054NS e e 0.284NS e 0.215NS 0.023*
e
(*) Means there is a significant difference when P. value < 0. 05. (**) means highly significant difference when P. value < 0.001. (NS) means no significant difference when P. value > 0.05.
Group I Group II Group III Group IV Est. age by LM
Group Group Group Group Est. age Group by PM Group Group Group Est. age Group by PHM Group Group Group
I II III IV I II III IV I II III IV
e 0.253NS 0.036* 0.019* e 0.362NS 0.019* 0.020* e 0.629NS 0.027* 0.031*
e 0.017* 0.033*
e 0.360NS
e
e 0.043* 0.010*
e 0.241NS
e
e 0.011* 0.026*
e 0.094NS
e
(*) Means there is a significant difference when P. value < 0. 05. (**) means highly significant difference when P. value < 0.001. (NS) means no significant difference when P. value > 0.05.
cementum resorption that was described as a manifestation of periodontal diseases [23]. Moreover, counting TCA using LM, PM and PHM and calculation of Est. age using periodontally affected teeth illustrated variety in divergence between the Est. age and Chr. age of the patients since some exhibited some Table 4 Regression analysis for age estimation of group III with L.M. Model
Unstandardized coefficients B
Std. Error
(Constant) Est. age with LM
27.386 0.582
7.883 0.140
Equation for age estimation with L.M (Est. age LM ¼ 27.386 þ 0.582 £ Est. age with LM in group III.).
Sig.
0.004 0.001 with
Table 5 Regression analysis for age estimation of group III with PM. Model
(Constant) Est. age with PM
Unstandardized coefficients B
Std. Error
23.144 0.688
14.136 0.265
Equation for age estimation with PM (Est. age PM ¼ 23.144 þ 0.688 x ¼ Est. age with P in group III.).
Sig.
0.126 0.022 with
Table 6 Regression analysis for age estimation of group III with PHM. Model
Unstandardized coefficients B
Std. Error
(Constant) Est. age with PHM
25.945 0.636
13.197 0.248
Sig.
0.071 0.023
Equation for age estimation with PHM (Est. age with PHM ¼ 25.945 þ 0.636 £ Est. age with PHM in group III).
Please cite this article in press as: Alghonamy WY et al., Age estimation in adult human sound and periodontally affected teeth using tooth cementum annulations, Tanta Dental Journal (2015), http://dx.doi.org/10.1016/j.tdj.2015.07.002
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W.Y. Alghonamy et al. / Tanta Dental Journal xx (2015) 1e9 Table 7 Regression analysis for age estimation of group IV with L.M. Model
(Constant) Est. age with LM
Unstandardized coefficients B
Std. Error
14.845 1.446
11.863 0.251
Sig.
0.233 0.000
Equation for age estimation with L.M (Est. age with LM ¼ ¡14.845 þ 1.446 £ Est. age with LM in group IV). Table 8 Regression analysis for age estimation of group IV with PM. Model
Unstandardized coefficients B
Std. Error
(Constant) Est. age with PM
13.726 0.846
12.716 0.271
Equation for age estimation with PM (Est. age PM ¼ 13.726 þ 0.846 £ Est .age with PM in group IV).
Sig.
0.300 0.008 with
Table 9 Regression analysis for age estimation of group IV with PHM. Model
(Constant) Est. age with PHM
Unstandardized coefficients B
Std. Error
26.911 0.559
8.391 0.175
Sig.
0.007 0.007
Equation for age estimation with PHM (Est. age with PHM ¼ 26.911 þ 0.559 £ Est. age with PHM in group IV).
deviation between Est. age and Chr. age while others showed great divergence. These results could be explained by those reported by Kagrer and Grupe [24] who depicted that periodontally affected teeth with sufficient nutritional support of their roots showed minimal deviation of histological age from the known. Interestingly in this study, statistical analysis was performed to compare the mean ± SD values of Est. ages & Chr. ages in each group using ANOVA test and it revealed no significant difference between Chr. ages and Est. ages in female and male sound single rooted teeth. However, a significant difference was depicted in female and male periodontally affected single rooted teeth. This indicated that age estimation using TCA method was accurate in female & male sound single rooted teeth but inaccurate for periodontally affected single rooted teeth of either female or male. Considering sound teeth, the present result was in accordance with others who depicted that counting of TCA strongly correlated with the Chr. age and hence TCA is a reliable age marker that could be used in age determination [6,7]. But, Kasetty et al. [25] concluded that TCA was not sufficiently accurate for prediction of age and they suggested that the procedural difficulties
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like indistinct annulations and sometimes absence of annulations complicate the calculation based on this method. Nonetheless considering periodontally affected teeth, the present result was showed that TCA is applicable to periodontally sound teeth but not for periodontally diseased teeth. The inability to use TCA as age estimation method in periodontally affected teeth might be related to that in periodontitis there was destruction in collagen fibers causing the cementum to be thinner than normal and hence less incremental lines of cementum was detected that consequently could affects the TCA result [26,27]. Interestingly, Swaminathan and Thomas [28] found that in early and moderate periodontitis the coronal half of the root, containing acellular cementum, was usually affected whereas in advanced cases, damage extends to cellular cementum. In accordance with the depicted observation of TCA under the LM, PM or PHM that illustrated in this study, the statistical results of comparing the Est. ages with LM, PM, & PHM and Chr. ages in each group using post hoc test revealed that age estimation using TCA method by either LM, PM and PHM was accurate in female & male sound single rooted teeth with the best results obtained by PHM. These findings matched with Pundir et al. [29] who found strong positive correlation between Est. and Chr. age when using PHM and less correlation when using PM and LM. On the other hand, age estimation using TCA method by either LM, PM and PHM in female periodontally affected single rooted teeth showed that both LM and PHM were accurate, with the best results obtained by PHM, but PM was inaccurate. However, in male periodontally affected single rooted teeth all of the three microscopes were inaccurate which confirmed the inability to use TCA as age estimation method in periodontally affected teeth. Upon comparison between the Est. ages by LM, PM or PHM among the four different groups using post hoc test revealed no difference between females and males in either sound or periodontally affected teeth (i.e. no sex difference). The more interesting point in the present work is the regression analysis and equations of female and male periodontally affected teeth. Since a significant difference was depicted between Chr. ages and Est. ages using TCA of female and male periodontally affected single rooted teeth, absolute multiple regression equations were done to equalize their results with the non significant ones and enabled age estimation by TCA method using L.M, PM& PHM with any tooth
Please cite this article in press as: Alghonamy WY et al., Age estimation in adult human sound and periodontally affected teeth using tooth cementum annulations, Tanta Dental Journal (2015), http://dx.doi.org/10.1016/j.tdj.2015.07.002
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included in these groups. Similar study was performed by Kvaal and Solheim [4] who found that teeth extracted because of periodontal disease showed a weaker correlation with age than did the sound teeth. Hence, they did a regression analysis and equation to enable age estimation using TCA by fluorescence microscope. Also, Kasetty et al. [25] performed an absolute regression equation for age estimation using TCA by PM. In contrast with Condon et al. [22] who used regression analysis in their study but unfortunately they couldn't give absolute equation of age estimation using TCA. However, they calculated the error of age estimates using TCA and they found that the standard error of the estimates ranged from 4.7 to 9.7 years depending on sex and health status of the tooth. Regardless, periodontally affected teeth weren't included in their study sample. 5. Conclusion and recommendation i The results of the present study could support the reliability of TCA as an age estimation method for both female and male individuals who have sound teeth. ii TCA not applicable for periodontally affected teeth neither female nor male. iii The data showed that TCA can be used with female or male periodontally affected teeth using specific novel regression formula for each microscope. From the present study we could recommend the followings: i When using TCA as age estimation method it is preferred to use LM, PM or PHM in observing cementum with the best one of them is PHM. ii In age estimation protocol it is preferred to use more than one method for good results. iii Further studies are needed to determine the availability of TCA for individuals older than 50 years.
Acknowledgment It is pleasure to express my deepest gratitude to Prof. Dr. Khalid Abo-Eisha Prof of Microbiology and Immunology Department, Faculty of Pharmacy, German University in Cairo (GUC) for his scientific and personal contribution when using his laboratory for photographing our slides under phase contrast microscope.
References [1] Singh A, Gorea RK, Singla U. age estimation from physiological changes of teeth. J Indian Assoc Forensic Med 2004;26:971e4. [2] Stein TJ, Corcoran JF. Pararadicular cementum deposition as a criterion for age estimation in human beings. Oral Surg Oral Med Oral Pathol 1994;77:266e70. [3] Willems G, Romsee MC, Solheim T. Non destructive dental age calculation methods in adults: intra-and inter-observer effects. Forensic Sci Int 2002;126:221e6. [4] Kvaal SI, Solheim T. Incremental lines in human dental cementum in relation to age. Eur J Oral Sci 1995;103:225e30. [5] Lieberman D, Meadow R. The biology of cementum increments (with an archaeological application). Mammal Rev 1992;22:57e77. [6] Renz H, Schaefer V, Duschner H, Radlanski RJ. incremental lines in root cementum of human teeth: an approach to their ultrastructural nature by microscopy. Adv Dent Res 1997;11:472e7. [7] Li YJ. Observations of the structure of incremental lines in acellular cementum. Kokubyo Gakkai Zasshi 1996;63:489e515. [8] Tencate A. The estimation of age of skeletal remains from color of roots of teeth. J Can Dent Assoc 1977;2:83e6. [9] Stoneberg RP, Jonkel CJ. Age determination of black bears by cementum layers. J Wldlfmngmt 1966;30:411e4. [10] Stott GG, Sis RF, Levy BM. Cemental annulation as an age criterion in the common Marmoset (Callithrix jacchus). J Med Primatol 1980;9:274e85. [11] Stott GG, Sis RF, Levy BM. Cemental annulation as an age criterion in forensic dentistry. J Dent Res 1982;61:814e7. [12] Avadhani A, Tupkari JV, Khambaty A. Cementum annulations and age determination. J Forensic Dent Sci 2009;1:73e6. [13] Aggarwal P, Saxena S, Bansal P. Incremental lines in root cementum of human teeth: an approach to their role in age estimation using polarizing microscopy. Indian J Dent Res 2008;19:326e30. [14] Lieberman DE. The biological basis for seasonal increments in dental cementum and their application to archaeological research. Archaeol Sci 1994;21:525e39. [15] Schour I, Massler M. Development of human dentition. J Am Dent Assoc 1941;20:379e427. [16] Scott AJ. Illusions in regression analysis. Int J Forecast Forthcom 2012;28:689e95. [17] Dias P, Beaini T, Melani RF. Age estimation from dental cementum incremental lines and periodontal disease. J Forensic Odontostomatol 2010;28:13e21. [18] Someda H, Saka A, Matsunaga S, Nakahara K, Hirata S, Hashimoto M. Age estimation based on three-dimensional measurement of mandibular central incisors in Japanese. Forensic Sci Int 2009;185:110e4. [19] Radovic MB. Ageing in the Danube gorges population (9500e5500 BC) e tooth cementum annulation method. CATAPHAP 2012;12:9e18. [20] Wedel VL. Determination of season at death using dental cementum increment analysis. J Forensic Sci 2007;52:1e4. [21] Sanderson J. The theory of contrast control in microscope. Quekett J Microsc 2000;38:617e27. [22] Condon K, Charles DK, Cheverud JM, James MC, Jane EB. Cementum annulation and age determination in Homo sapiens II estimates and accuracy. Am J Phys Anthropol 1986;71:321e30. [23] Nanci A, Bosshardt DD. Structure of periodontal tissues in health and disease. Periodontol 2000;2006(40):11e28.
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W.Y. Alghonamy et al. / Tanta Dental Journal xx (2015) 1e9 [24] Kagrer P, Grupe G. Age at death diagnosis and determination of life history parameters by incremental lines in human dental cementum as an identification aid. Forensic Sci Int 2001;118:75e82. [25] Kasetty S, Rammanohar M, Ragavendra RT. Dental cementum in age estimation: a polarized light and stereomicroscopic study. J Forensic Sci 2010;55:779e80. [26] Sottosanti JR, Garret JR. A mannual for root preparation - a scanning electron microscopic study of deseased cementum. J Periodont Res 1975;46:628e9.
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[27] Barton NS, Van Swol RL. Periodontally diseased versus normal roots as evaluated by scanning electron microscopy and electron probe analysis. J Periodont Res 1987;58:634e8. [28] Swaminathan Y, Thomas JT. What makes cementum a unique structure? IJSER 2013;4. 2229e5518. [29] Pundir S, Saxena S, Aggrawal P. Estimation of age based on tooth cementum annulations using three different microscopic methods. J Forensic Dent Sci 2009;1:82e7.
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Available online at www.sciencedirect.com
ScienceDirect Tanta Dental Journal xx (2015) 1e9 www.elsevier.com/locate/tdj
Age estimation in adult human sound and periodontally affected teeth using tooth cementum annulations W.Y. Alghonamy*, O.M. Gaballah, D.A. Labah Faculty of Dentistry, Tanta University, Oral Biology Department, Egypt Received 16 June 2015; revised 3 July 2015; accepted 26 July 2015
Abstract Background: There are many tools for identification of human remains, via radio graphical imaging, histological processing & microscopical examination of dental remains. Purpose: to estimate the age of both female and male adults using tooth cementum annulations (TCA) method, evaluate the accuracy of TCA of both sound and periodontally affected teeth, and compare the estimated age (Est.age) with the chronological age (Chr.age) of an individual by the aid of; light microscope (LM), polarized microscope (PM) and phase contrast microscope (PHM). Material and methods: 60 teeth divided into four equal experimental groups; group I contained sound teeth extracted from female patients, group II contained sound teeth extracted from male patients, group III contained periodontally affected teeth extracted from female patients and group IV contained periodontally affected teeth extracted from male patients. Longitudinal ground section for all groups were prepared & observed under LM, PM and PHM. Afterward, images were taken for each section then the average of TCA for each section was counted. The age was estimated using the formula {Estimated age ¼ Numbers of incremental lines þ Eruption age of tooth}. Results: There was no significant difference between Est.age and Chr.age of both male and female sound teeth (group I and group II) while in periodontally affected teeth of both male and female (group III and group IV) there was significant difference. Also, there was a strong correlation between the Est. & Chr. age when PHM was used followed by LM then PM. Finally, there was no sex difference. Conclusion: TCA is reliable for sound but not for periodontally affected teeth using either LM, PM or PHM. Moreover, PHM is a more accurate microscope for TCA in contrast to LM & PM. Interestingly, a novel regression formula was developed to enable estimating age in periodontally affected teeth by TCA method. © 2015, Hosting by Elsevier B.V. on behalf of the Faculty of Dentistry, Tanta University.
Keywords: Age estimation; Tooth cementum annulations; Periodontitis; Light microscope; Polarized microscope; Phase contrast microscope
* Corresponding author. Alshahid fekry, Berkit el sabee, Minufia, Egypt. Tel.: þ20 0482977257, þ20 01066754527 (mobile). E-mail addresses: [email protected] (W.Y. Alghonamy), [email protected] (O.M. Gaballah), [email protected] (D.A. Labah). Peer review under the responsibility of the Faculty of Dentistry, Tanta University.
1. Introduction Forensic odontology (Forensic dentistry) is a specialized branch of forensic medicine which, in the interests of justice, deals with examination of dental
http://dx.doi.org/10.1016/j.tdj.2015.07.002 1687-8574/© 2015, Hosting by Elsevier B.V. on behalf of the Faculty of Dentistry, Tanta University. Please cite this article in press as: Alghonamy WY et al., Age estimation in adult human sound and periodontally affected teeth using tooth cementum annulations, Tanta Dental Journal (2015), http://dx.doi.org/10.1016/j.tdj.2015.07.002
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evidence from which a proper evaluation and presentation of dental findings can be made [1]. Age determination plays an important role in forensic medicine, not only in identification of bodies but also in connection with crimes. When the subjects have undergone changes so extensive that external characteristics yield no information, the teeth are often the only means of identification. Also, Human tooth can be preserved for a long time after death without gross changes so; it serves as an important tool in forensic science [2]. Various methods are utilized for determination of age from dentition. These may be described in four categories namely, clinical, radiographic, histological, physical and chemical analysis [3]. Apart from above mentioned techniques, a histological method based on counting the incremental lines of tooth cementum has shown promise for age estimation method in adult as the biological factors of cementum formation is well established so far. Tooth cementum is the calcified ectomesenchymal tissue that surrounds the dentine and forms the attachment site for the periodontal ligaments that support the tooth to the surrounding alveolar bone [4]. Microscopically, cementum appears as a series of alternating light and dark lines or bands that have the appearance of tree annual rings called TCA [5,6]. In acellular cementum, many fine incremental lines of about 0.5 micron width were observed that ran perpendicularly or obliquely to Sharpey's fibers. Also, these lines are not straight lines but had wave-like structure. Additionally, the fine incremental lines showing a close relationship to the degree of calcification, they also seemed to show a strong relationship to the quality of the organic component of acellular cementum [7]. In respect of cementum position, it has not been used to the extent of enamel and dentin to estimate the age of unidentified individuals. However, many studies recommended that counting of TCA in thin histological sections and adding them to the mean eruption age of the tooth may offer a more accurate method for age estimation in human beings and can be used as a more reliable age marker than any other morphological or histological traits in the human skeleton based on the biological factors of TCA [8]. For many years, it was reported that age determination using TCA could be an accurate means of age determination in animals that live on a relatively constant diet throughout life [9,10]. On the other hand, numerous studies were done to estimate ages by TCA method in human sound teeth using different types of microscopes and considered this method to be extremely valuable in forensic medicine, forensic dentistry, and anthropology [11,12].
Surprisingly, It has been shown that, few studies were done on age estimation using TCA method in periodontally affected teeth and these studies exhibited many contrasts. Therefore, it is of prime importance to study the efficiency of using TCA in age estimation of adult male and female using both sound and periodontally affected teeth with the aid of three microscopes; LM, PM & PHM and to compare the Est. ages with the Chronological ages of the individuals. 2. Materials and methods 2.1. The sample
Sixty extracted single rooted teeth were used in this study including both sound and early periodontally affected teeth. They were obtained from the Oral Surgery and Maxillofacial Department, Faculty of Dentistry, Tanta University. Teeth were extracted as a part of essential clinical care from patients ranging from 20 to 70 years old of both sexes with well known Chr. age and well known date and reason of extraction. Both written and oral consents were obtained from the patients. The protocol was approved from ethical committee at Faculty of Dentistry, Tanta University. Afterward, the selected teeth were divided into four equal experimental groups each one consists of fifteen teeth. Group I: sound single rooted teeth extracted from female patients, Group II: sound single rooted teeth extracted from male patients, Group III: periodontally affected single rooted teeth extracted from female patients and Group IV: periodontally affected single rooted teeth extracted from male patients. 2.2. The method 2.2.1. Histological study
The extracted teeth were preserved in 10% buffered formalin then they were washed under running water. Thin longitudinal ground histological sections were done as reported by Aggarwal et al. [13]. After excluding unbeneficial sections, the ground sections in each group were examined by light, polarized and phase contrast microscopes. Afterward, images were taken for the middle third of the root of each section using the three microscopes at magnification 400 for areas of the cementum showing uniform thickness, clear definition of layers from the cementodentinal junction (CDJ) to the outer border of the root then images magnified by computer. The average of TCA for each section was counted by image analysis system as follow: Counting the dark lines manually at the
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monitor based on digitally enhanced images using the measurement overlay (Image J program).1 Counting the TCA had been done by three observers to avoid inter-observer variability. When there was discrepancy between different counts, the mean value was used for statistical analysis. Then, each observation was repeated after one week to evaluate intra-observer error. One lightedark pair was considered as one annulation which corresponds to one year age of the individual [14]. Formula for estimation of age at death or time of tooth extraction was calculated by adding average age of tooth eruption in years for each tooth as represented by Schour and Massler [15] to the counted number of incremental lines [16]. {Estimated age (E) ¼ Numbers of incremental lines (n) þ Eruption age of tooth (t)}. 2.2.2. Statistical study
All the data was recorded, tabulated and statistically analyzed using software SPSS V.20. Statistical presentation and analysis of the present study was conducted using the mean and standard deviation in the following tests: A) Analysis of variance [ANOVA] tests, B) Post Hoc test., C) Regression analysis and equation [16]. 3. Results 3.1. Histological results Histological study revealed that TCA were more clearly visible under the PHM followed by LM then PM. Interestingly, microscopic observations of the cementum in group I & group II gave similar results as they showed uniform thickness, clear definition of TCA from the CDJ to the outer borders of the root that appeared as a series of alternating light and dark lines or bands and each one lightedark pair was considered as one annulation. As well, calculation of Est. age by adding eruption age of the tooth to total number of TCA illustrated accordance between the Est. age by TCA and Chr. age for both groups but for patients older than 50 years there was some divergence (Fig. 1 &Fig. 2). On the other hand, microscopic observations of the cementum in group III& group IV exhibited variability in cementum thickness even in different areas of the same section. Moreover, some areas of cementum showed resorption (saucer shaped) that could be considered as a manifestation of periodontitis. Also, counting TCA and calculation of Est. age 1 This is a software tool allows marking each line by mouse click which is then summed.
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according to the previous equation illustrated divergence between the Est. age and Chr. age of the patients (Fig. 3 & Fig. 4). 3.2. Statistical results 3.2.1. Comparison between the Est. ages using TCA & the Chr. ages in each group by ANOVA test See Table 1. 3.2.2. Comparison between the Est. ages with different microscopes (LM, PM & PHM) & the Chr. ages in each group by post hoc test See Table 2. 3.2.3. Comparison between the Est. ages by each microscope (LM, PM or PHM) among the four different groups by post hoc test See Table 3. 3.2.4. Regression analysis and equation 1-Age estimation of group III with L.M (See Table 4). 2-Age estimation of group III with PM (See Table 5). 3-Age estimation of group III with PHM (See Table 6). 4-Age estimation of group IV with L.M (See Table 7). 5-Age estimation of group IV with PM (See Table 8). 6-Age estimation of group IV with PHM (See Table 9). 4. Discussion Teeth are particularly useful in age estimation as they display a number of observable age related variables and they tend to remain intact under circumstances which might alter or obliterate the rest of the skeleton. The present study hypothesized that the cementum annulations may serve as a powerful method for age determination in both male and female sound single rooted teeth and how this method may be valuable even in periodontally affected teeth. Briefly, this discussion covered two essential points first; the validity and accuracy of TCA methods in age estimation of both sound and periodontally affected male and female single rooted teeth. Second, the most accurate microscope that achieve our goal. Using TCA in age estimation was based on the facts reported by Lieberman [14] that during cementum formation, the hyper mineralized layers of extracellular matrix alternate with less mineralized layers causing the cementum to appear as a series of alternating light
Please cite this article in press as: Alghonamy WY et al., Age estimation in adult human sound and periodontally affected teeth using tooth cementum annulations, Tanta Dental Journal (2015), http://dx.doi.org/10.1016/j.tdj.2015.07.002
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Fig. 1. Micrographs of (group I) sound upper right central incisor of 60 years old female patient. A, LM depicts the total number of TCA is 46 (yellow dots). B, PM shows the total number of TCA is 45 (yellow dots). C, PHM shows the total number of TCA is 51 (red dots), (L.S. microscopic magnification.X 400). The Est. age by LM ¼ 46 þ 8 ¼ 54 years, by PM ¼ 45 þ 8 ¼ 53 years and by PHM ¼ 51 þ 8 ¼ 59 years.
and dark lines or bands and each one lightedark pair was considered as one annulation which corresponds to one year age of the individual. Thus, counting these lines in thin histological sections and adding them to the mean eruption age of the tooth will give the Chr. age of the individual [17]. In the current study, the sample included extracted teeth from patients ranging between 20 and 70 years old since by the age of 20 years growth and dental development were completed as confirmed by Someda et al. [18]. Notably, examination of TCA using LM allowed good observation of TCA. This is in agreement with
Stott et al. [11]. In contrary, Radovic [19] who found that incremental lines in the cementum could not be read from one third of an archeological sample when using LM. On the other hand, the present work illustrated that observation of TCA using PM gave a good observation for cementum annulations in all groups in most of the samples but in some samples PM couldn't enable clear TCA observation. The proper PM observation was advocated by Wedel [20]. Moreover, In the current study, LM revealed more obvious observations than PM. and PHM revealed excellent observation of annulations. The clarity of
Fig. 2. Micrographs of (group II) sound upper right lateral incisor of 34 years old male patient A, LM depicts the total number of TCA is 25. B PM shows the total number of TCA is 24. C, PHM shows the total number of TCA is 25. (L.S. microscopic magnification. 400). The Est. age by LM ¼ 25 þ 9 ¼ 34 years, PM ¼ 24 þ 9 ¼ 33 years and PHM ¼ 25 þ 9 ¼ 34 years. Please cite this article in press as: Alghonamy WY et al., Age estimation in adult human sound and periodontally affected teeth using tooth cementum annulations, Tanta Dental Journal (2015), http://dx.doi.org/10.1016/j.tdj.2015.07.002
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Fig. 3. Micrographs of (group III) for periodontally affected lower right central incisor for 70 years old female patient A, LM shows the total number of TCA is 42 (yellow dots). B, PM shows the total number of TCA (yellow dots) is 36 in the thick part and 21 in the thin part (the resorbed area) with the mean of TCA ¼ 28.5. C, PHM shows the total number of TCA (yellow dots) is 38 in the thick part and 31in the thin part with the mean of TCA is 34.5. (L.S. microscopic magnification. 400). The Est. age by LM ¼ 42 þ 7 ¼ 49 years, by PM ¼ 28.5 þ 7 ¼ 35.5 years and PHM ¼ 34.5 þ 7 ¼ 41.5 years.
TCA by PHM was explained by Sanderson [21] who discussed how PHM gave the proper distinguishing between two types of layers with different optical properties. He found that PHM depends on refraction of light by the specimen as it utilizes the difference between light rays propagating directly from the light source and light rays refracted by the specimen when light passes through it to add bright/dark contrast to images of transparent specimens and this done by the aid of a phase-contrast objective and a condenser fitted with PHM for observations. He depicted that PHM is
suitable for viewing colorless and transparent specimens (as cementum). In the present histological study, counting TCA using LM, PM and PHM for the same cementum area in each ground section followed by calculation of Est. age by adding eruption age of the tooth to total number of TCA illustrated accordance between the Est. age and Chr. age in sound teeth of female and male patients less than 50 years however, there was some divergence between the Est. age and Chr. age in elders. This is in agreement with Condon et al. [22] who depicted that
Fig. 4. Micrographs of (group IV) for periodontally affected lower left canine of 46 years old male patient. A, LM shows total number of TCA is 38. B, PM shows total number of TCA is 34. C, PHM shows total number of TCA is 37. (L.S. microscopic magnification. 400). The Est. age by LM ¼ 38 þ 10 ¼ 48 years, by PM ¼ 34 þ 10 ¼ 44 years and by PHM ¼ 37 þ 10 ¼ 47 years. Please cite this article in press as: Alghonamy WY et al., Age estimation in adult human sound and periodontally affected teeth using tooth cementum annulations, Tanta Dental Journal (2015), http://dx.doi.org/10.1016/j.tdj.2015.07.002
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Table 1 Comparison of mean ± SD values of Est. ages & Chr. ages in each group using ANOVA test. N Mean ± SD Group I
Chr.age Est.age with Est.age with Est.age with Total Group II Chr.age Est.age with Est.age with Est.age with Total Group III Chr.age Est.age with Est.age with Est.age with Total Group IV Chr.age Est.age with Est.age with Est.age with Total
15 60.07 ± LM 15 52.60 ± PM 15 49.40 ± PHM 15 53.27 ± 60 53.83 ± 15 47.87 ± LM 15 46.33 ± PM 15 44.07 ± PHM 15 47.27 ± 60 46.38 ± 15 59.47 ± LM 15 51.33 ± PM 15 44.60 ± PHM 15 53.07 ± 60 52.12 ± 15 53.20 ± LM 15 43.67 ± PM 15 40.73 ± PHM 15 47.07 ± 60 46.17 ±
14.723 15.231 13.979 14.280 14.717 13.799 14.356 12.533 13.946 13.403 9.273 14.226 10.176 12.792 12.661 7.476 5.912 7.166 8.860 8.616
Range p-value 74e30 77e27 68e25 71e26 77e25 60e20 64e20 59e23 65e25 65e20 70e45 76e30 59e24 84e33 84e24 68e44 53e32 53e24 71e36 71e24
NS
0.246
0.880NS
0.011*
0.000**
(*) Means there is a significant difference when P. value < 0. 05. (**) means highly significant difference when P. value < 0.001. (NS) means no significant difference when P. value > 0.05.
annual deposition of cementum rings exhibited a decrease with increasing age. On the other hand in the present histological work, periodontally affected teeth that were taken from female and male patients exhibited variability in cementum shape with some noticeable areas of Table 2 Comparison of Est. ages with LM, PM, & PHM and Chr. ages in each group using post hoc test. Chr. Age Est. age Est. age Est. age by L.M by PM by PHM Group I
Chr. Age Est. age by Est. age by Est. age by Group II Chr. Age Est. age by Est. age by Est. age by Group III Chr. Age Est. age by Est. age by Est. age by Group IV Chr. Age Est. age by Est. age by Est. age by
e L.M 0.166NS PM 0.050NS PHM 0.206NS e L.M 0.760NS PM 0.450NS PHM 0.905NS e L.M 0.064NS PM 0.001* PHM 0.143NS e L.M 0.001* PM 0.000** PHM 0.028*
Table 3 Comparison of Est. age by LM, PM or PHM in all groups using post hoc test.
e 0.550NS e 0.901NS 0.470NS e e 0.652NS e 0.852NS 0.524NS e e 0.123NS e 0.689 NS 0.054NS e e 0.284NS e 0.215NS 0.023*
e
(*) Means there is a significant difference when P. value < 0. 05. (**) means highly significant difference when P. value < 0.001. (NS) means no significant difference when P. value > 0.05.
Group I Group II Group III Group IV Est. age by LM
Group Group Group Group Est. age Group by PM Group Group Group Est. age Group by PHM Group Group Group
I II III IV I II III IV I II III IV
e 0.253NS 0.036* 0.019* e 0.362NS 0.019* 0.020* e 0.629NS 0.027* 0.031*
e 0.017* 0.033*
e 0.360NS
e
e 0.043* 0.010*
e 0.241NS
e
e 0.011* 0.026*
e 0.094NS
e
(*) Means there is a significant difference when P. value < 0. 05. (**) means highly significant difference when P. value < 0.001. (NS) means no significant difference when P. value > 0.05.
cementum resorption that was described as a manifestation of periodontal diseases [23]. Moreover, counting TCA using LM, PM and PHM and calculation of Est. age using periodontally affected teeth illustrated variety in divergence between the Est. age and Chr. age of the patients since some exhibited some Table 4 Regression analysis for age estimation of group III with L.M. Model
Unstandardized coefficients B
Std. Error
(Constant) Est. age with LM
27.386 0.582
7.883 0.140
Equation for age estimation with L.M (Est. age LM ¼ 27.386 þ 0.582 £ Est. age with LM in group III.).
Sig.
0.004 0.001 with
Table 5 Regression analysis for age estimation of group III with PM. Model
(Constant) Est. age with PM
Unstandardized coefficients B
Std. Error
23.144 0.688
14.136 0.265
Equation for age estimation with PM (Est. age PM ¼ 23.144 þ 0.688 x ¼ Est. age with P in group III.).
Sig.
0.126 0.022 with
Table 6 Regression analysis for age estimation of group III with PHM. Model
Unstandardized coefficients B
Std. Error
(Constant) Est. age with PHM
25.945 0.636
13.197 0.248
Sig.
0.071 0.023
Equation for age estimation with PHM (Est. age with PHM ¼ 25.945 þ 0.636 £ Est. age with PHM in group III).
Please cite this article in press as: Alghonamy WY et al., Age estimation in adult human sound and periodontally affected teeth using tooth cementum annulations, Tanta Dental Journal (2015), http://dx.doi.org/10.1016/j.tdj.2015.07.002
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W.Y. Alghonamy et al. / Tanta Dental Journal xx (2015) 1e9 Table 7 Regression analysis for age estimation of group IV with L.M. Model
(Constant) Est. age with LM
Unstandardized coefficients B
Std. Error
14.845 1.446
11.863 0.251
Sig.
0.233 0.000
Equation for age estimation with L.M (Est. age with LM ¼ ¡14.845 þ 1.446 £ Est. age with LM in group IV). Table 8 Regression analysis for age estimation of group IV with PM. Model
Unstandardized coefficients B
Std. Error
(Constant) Est. age with PM
13.726 0.846
12.716 0.271
Equation for age estimation with PM (Est. age PM ¼ 13.726 þ 0.846 £ Est .age with PM in group IV).
Sig.
0.300 0.008 with
Table 9 Regression analysis for age estimation of group IV with PHM. Model
(Constant) Est. age with PHM
Unstandardized coefficients B
Std. Error
26.911 0.559
8.391 0.175
Sig.
0.007 0.007
Equation for age estimation with PHM (Est. age with PHM ¼ 26.911 þ 0.559 £ Est. age with PHM in group IV).
deviation between Est. age and Chr. age while others showed great divergence. These results could be explained by those reported by Kagrer and Grupe [24] who depicted that periodontally affected teeth with sufficient nutritional support of their roots showed minimal deviation of histological age from the known. Interestingly in this study, statistical analysis was performed to compare the mean ± SD values of Est. ages & Chr. ages in each group using ANOVA test and it revealed no significant difference between Chr. ages and Est. ages in female and male sound single rooted teeth. However, a significant difference was depicted in female and male periodontally affected single rooted teeth. This indicated that age estimation using TCA method was accurate in female & male sound single rooted teeth but inaccurate for periodontally affected single rooted teeth of either female or male. Considering sound teeth, the present result was in accordance with others who depicted that counting of TCA strongly correlated with the Chr. age and hence TCA is a reliable age marker that could be used in age determination [6,7]. But, Kasetty et al. [25] concluded that TCA was not sufficiently accurate for prediction of age and they suggested that the procedural difficulties
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like indistinct annulations and sometimes absence of annulations complicate the calculation based on this method. Nonetheless considering periodontally affected teeth, the present result was showed that TCA is applicable to periodontally sound teeth but not for periodontally diseased teeth. The inability to use TCA as age estimation method in periodontally affected teeth might be related to that in periodontitis there was destruction in collagen fibers causing the cementum to be thinner than normal and hence less incremental lines of cementum was detected that consequently could affects the TCA result [26,27]. Interestingly, Swaminathan and Thomas [28] found that in early and moderate periodontitis the coronal half of the root, containing acellular cementum, was usually affected whereas in advanced cases, damage extends to cellular cementum. In accordance with the depicted observation of TCA under the LM, PM or PHM that illustrated in this study, the statistical results of comparing the Est. ages with LM, PM, & PHM and Chr. ages in each group using post hoc test revealed that age estimation using TCA method by either LM, PM and PHM was accurate in female & male sound single rooted teeth with the best results obtained by PHM. These findings matched with Pundir et al. [29] who found strong positive correlation between Est. and Chr. age when using PHM and less correlation when using PM and LM. On the other hand, age estimation using TCA method by either LM, PM and PHM in female periodontally affected single rooted teeth showed that both LM and PHM were accurate, with the best results obtained by PHM, but PM was inaccurate. However, in male periodontally affected single rooted teeth all of the three microscopes were inaccurate which confirmed the inability to use TCA as age estimation method in periodontally affected teeth. Upon comparison between the Est. ages by LM, PM or PHM among the four different groups using post hoc test revealed no difference between females and males in either sound or periodontally affected teeth (i.e. no sex difference). The more interesting point in the present work is the regression analysis and equations of female and male periodontally affected teeth. Since a significant difference was depicted between Chr. ages and Est. ages using TCA of female and male periodontally affected single rooted teeth, absolute multiple regression equations were done to equalize their results with the non significant ones and enabled age estimation by TCA method using L.M, PM& PHM with any tooth
Please cite this article in press as: Alghonamy WY et al., Age estimation in adult human sound and periodontally affected teeth using tooth cementum annulations, Tanta Dental Journal (2015), http://dx.doi.org/10.1016/j.tdj.2015.07.002
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included in these groups. Similar study was performed by Kvaal and Solheim [4] who found that teeth extracted because of periodontal disease showed a weaker correlation with age than did the sound teeth. Hence, they did a regression analysis and equation to enable age estimation using TCA by fluorescence microscope. Also, Kasetty et al. [25] performed an absolute regression equation for age estimation using TCA by PM. In contrast with Condon et al. [22] who used regression analysis in their study but unfortunately they couldn't give absolute equation of age estimation using TCA. However, they calculated the error of age estimates using TCA and they found that the standard error of the estimates ranged from 4.7 to 9.7 years depending on sex and health status of the tooth. Regardless, periodontally affected teeth weren't included in their study sample. 5. Conclusion and recommendation i The results of the present study could support the reliability of TCA as an age estimation method for both female and male individuals who have sound teeth. ii TCA not applicable for periodontally affected teeth neither female nor male. iii The data showed that TCA can be used with female or male periodontally affected teeth using specific novel regression formula for each microscope. From the present study we could recommend the followings: i When using TCA as age estimation method it is preferred to use LM, PM or PHM in observing cementum with the best one of them is PHM. ii In age estimation protocol it is preferred to use more than one method for good results. iii Further studies are needed to determine the availability of TCA for individuals older than 50 years.
Acknowledgment It is pleasure to express my deepest gratitude to Prof. Dr. Khalid Abo-Eisha Prof of Microbiology and Immunology Department, Faculty of Pharmacy, German University in Cairo (GUC) for his scientific and personal contribution when using his laboratory for photographing our slides under phase contrast microscope.
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Please cite this article in press as: Alghonamy WY et al., Age estimation in adult human sound and periodontally affected teeth using tooth cementum annulations, Tanta Dental Journal (2015), http://dx.doi.org/10.1016/j.tdj.2015.07.002