Highlights
- •The age-associated epi-markers were identified and validated in Chinese children for the first time.
- •Four CpG sites for boys and five CpG sites for girls were identified to be significantly associated with chronologic age.
- •DNA methylation markers in combination with skeletal age and dental age greatly improved the accuracy of age estimation in Chinese children.
Abstract
Age estimation is critical in forensic science, in competitive sports and games and
in other age-related fields, but the current methods are suboptimal. The combination
of age-associated DNA methylation markers with skeletal age (SA) and dental age (DA)
may improve the accuracy and precision of age estimation, but no study has examined
this topic. In the current study, we measured SA (GP, TW3-RUS, and TW3-Carpal methods)
and DA (Demirjian and Willems methods) by X-ray examination in 124 Chinese children
(78 boys and 46 girls) aged 6–15 years. To identify age-associated CpG sites, we analyzed
methylome-wide DNA methylation profiling by using the Illumina HumanMethylation450
BeadChip system in 48 randomly selected children. Five CpG sites were identified as
associated with chronologic age (CA), with an absolute value of Pearson’s correlation
coefficient (r) > 0.5 (p< 0.01) and a false discovery rate < 0.01. The validation of age-associated CpG sites was performed using droplet digital
PCR techniques in all 124 children. After validation, four CpG sites for boys and
five CpG sites for girls were further adopted to build the age estimation model with
SA and DA using multivariate linear stepwise regressions. These CpG sites were located
at 4 known genes: DDO, PRPH2, DHX8, and ITGA2B and at one unknown gene with the Illumina ID number of 22398226. The accuracy of
age estimation methods was compared according to the mean absolute error (MAE) and
root mean square error (RMSE). The best single measure for SA was the TW3-RUS method
(MAE = 0.69 years, RMSE = 0.95 years) in boys, and the GP method (MAE = 0.74 years, RMSE = 0.94 years) in girls. For DA, the Willems method was the best single measure for both boys
(MAE = 0.63 years, RMSE = 0.78 years) and girls (MAE = 0.54 years, RMSE = 0.68 years). The models that incorporated SA and DA with the methylation levels of age-associated
CpG sites provided the highest accuracy of age estimation in both boys (MAE = 0.47 years, R2 = 0.886) and girls (MAE = 0.33 years, R2 = 0.941). Cross validation of the results confirmed the reliability and validity of
the models. In conclusion, age-associated DNA methylation markers in combination with
SA and DA greatly improve the accuracy of age estimation in Chinese children. This
method may be applied in forensic science, in competitive sports and games and in
other age-related fields.
Abbreviations:
SA (skeletal age), DA (dental age), MAE (mean absolute error), RMSE (root mean standard error), CA (chronological age), GP (Greulich and Pyle), TW3 (Tanner and Whitehouse version 3), ddPCR (digital droplet PCR), FDR (false discovery rate), EA (estimated age)Keywords
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Article info
Publication history
Published online: November 16, 2017
Accepted:
November 11,
2017
Received in revised form:
October 15,
2017
Received:
August 24,
2017
Identification
Copyright
© 2017 Published by Elsevier Ireland Ltd.
