唾液斑口腔上皮细胞端粒长度变化在法医学实践中的应用研究

Application of telomere length of oral epithelial cells of saliva spot in forensic practice

目的:探讨人类唾液斑中口腔上皮细胞端粒长度变化与年龄的相关性及其稳定性,探讨2种方式提取的检材及其不同放置时间后口腔上皮细胞端粒长度是否有差异,拟为法医学通过微量易得检材推断年龄提供依据。方法:选取40例不同年龄段健康人唾液,分为童年组(0~6岁)、少年组(7~17岁)、青年组(18~40岁)、中年组(41~65岁)、老年组(>66岁)5个年龄组,每组8人,分别制成新鲜口腔拭子和不同时间段(1、7、14、28 d)的唾液斑,用磁珠法提取样本中的DNA,通过荧光PCR(聚合酶链反应)相对定量法检测端粒DNA限制性酶切段长度变化。结果:不同组间端粒长度有明显差异(F=7.138,P=0.018),年龄回归方程:Y=-75.006X+101.193,端粒TRF平均长度随年龄增加而逐渐缩短,并呈不均衡趋势。检材提取方式及检材新旧程度对检测到的TRF结果没有显著影响(P=0.740,P=0.740,P=0.958,P=0.324)。结论:唾液斑中口腔上皮细胞端粒长度随年龄增加而缩短,并且呈规律性改变;2种方式采集的样本及其不同放置时间后的样本端粒长度无明显差异,说明唾液斑中口腔上皮细胞端粒长度具有一定的稳定性,故可通过唾液斑中口腔上皮细胞端粒长度推断个体年龄。

Objective：To explore the relationship and stability between age and telomere length of oral epithelial cells of saliva spot,and to discuss whether freshness of sample or sample collected by two methods affects the length change,in order to provide forensic science with evidence to deduce age by means of micro-sample. Methods：Forty fresh saliva samples were collected from healthy individuals,which was divided into five groups,aged from 0 to 100 years old. Those saliva samples were made into fresh oral swabs and saliva spots at different periods（1,7,14,28 d）. DNA was extracted by paramagnetic particle method and the terminal restriction frag-ment length was measured by quantitative polymerase chain reaction with the analysis of linear regression. Results：The average TRF length showed a shortening tendency with aging,while samples collected by two methods and freshness of sample did not affect this parameter. The regression equation for age estimation was Y=-75.006 X＋101.193. Conclusion：The terminal restriction fragment length and age of donors show an inverse correction and there is no difference in samples collected by two methods and freshness of sample possible to infer individuals age by terminal restriction fragment length assay.