法医学杂志

法医学杂志 ›› 2016, Vol. 32 ›› Issue (3): 165-170.DOI: 10.3969/j.issn.1004-5619.2016.03.002

• 论著 • 上一篇    下一篇

不同温度下大鼠脑组织RNA降解与早期PMI的相关性

吕叶辉1，李志宏1，托  娅1，刘  丽1，李  堃1，卞  杰1，马剑龙2，陈  龙2   

  1. （1． 上海健康医学院基础医学院，上海 201318； 2． 复旦大学基础医学院法医学系，上海 200032）
  • 发布日期:2016-06-25 出版日期:2016-06-28
  • 通讯作者: 陈龙，男，博士，主任法医师，副教授，主要从事法医病理学及法医临床学研究；E-mail：chenlong@shmu.edu.cn
  • 作者简介:吕叶辉（1989—），男，硕士，主要从事基础医学教学、科研工作；E-mail：yeyeliushu@163.com
  • 基金资助:

    上海健康医学院种子基金项目；上海高校青年教师培养资助计划项目

Correlation between RNA Degradation Patterns of Rat’s Brain and Early PMI at Different Temperatures

L?譈 YE-HUI1, LI ZHI-HONG1, TUO YA1, LIU LI1, LI KUN1, BIAN JIE1, MA JIAN-LONG2, CHEN LONG2   

  1. （1. School of Basic Medical Science, Shanghai University of Medicine & Health Science, Shanghai 201318, China; 2. Department of Forensic Medicine, School of Basic Medical Science, Fudan University, Shanghai 200032, China）
  • Online:2016-06-25 Published:2016-06-28

被引次数

5

摘要： 目的 探讨大鼠脑组织8种RNA指标，在不同温度下的表达水平与早期死亡时间（PMI）的相关性。 方法 将222只SD大鼠随机分为对照组（死后0 h）和4个实验组，实验组断颈处死后分别置于5 ℃、15 ℃、25 ℃和35 ℃的环境中，于死后1~24 h内9个时间点提取脑组织RNA，利用实时荧光定量PCR技术检测8种RNA指标（β-actin、GAPDH、RPS29、18S rRNA、5S rRNA、U6 snRNA、miRNA-9及miRNA-125b）的表达水平，geNorm软件选取合适内参，SPSS软件对内参标准化RNA指标进行回归分析，R软件构建推断PMI的数学模型，另选6只已知PMI的SD大鼠予以验证。 结果 5S rRNA、miR-9和miR-125b表达稳定，可作为内参指标。β-actin和GAPDH具有良好的时序性降解规律，在24 h内随PMI延长不断降解。R软件拟合得ΔCt值随PMI和温度变化的数学模型可用以推断PMI。运用β-actin 和GAPDH验证模型的平均误差率分别为14.1%和22.2%。 结论 β-actin和GAPDH表达水平与PMI和环境温度相关性良好。本研究建立的数学模型可为温度变化条件下的早期PMI推断提供参考。

关键词: 法医病理学, RNA, 死亡时间, 温度, 降解, 大鼠

Abstract: Objective To explore the correlation between early postmortem interval （PMI） and eight RNA markers of rat’s brain at different temperatures. Methods Total 222 SD rats were randomly divided into control group （PMI=0 h） and four experimental groups. And the rats in the experimental groups were sacrificed by cervical dislocation and respectively kept at 5 ℃, 15 ℃, 25 ℃ and 35 ℃ in a controlled environment chamber. The RNA was extracted from brain tissues, which was taken at 9 time points from 1 h to 24 h postmortem. The expression levels of eight markers, β-actin, GAPDH, RPS29, 18S rRNA, 5S rRNA, U6 snRNA, miRNA-9 and miRNA-125b, were detected using real-time fluorescent quantitative PCR, respectively. Proper internal reference was selected by geNorm software. Regression analysis of normalized RNA markers was performed by SPSS software. Mathematical model for PMI estimation was established using R software. Another 6 SD rats with known PMI were used to verify the mathematical model. Results 5S rRNA, miR-9 and miR-125b were suitable as internal reference markers for their stable expression. Both β-actin and GAPDH had well time-dependent degradation patterns and degraded continually with prolongation of PMI in 24 h postmortem. The mathematical model of the variation of ΔCt values with PMI and temperature was set up by R software and the model could be used for PMI estimation. The average error rates of model validation using β-actin and GAPDH were 14.1% and 22.2%, respectively. Conclusion The expression levels of β-actin and GAPDH are well correlated with PMI and environmental temperature. The mathematical model established in present study can provide references for estimating early PMI under various temperature conditions.

Key words: forensic pathology, RNA, postmortem intervals, temperature, degradation, rats

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