Lv Ye-Hui, Ma Jian-Long, Pan Hui, Zeng Yan, Tao Li, Zhang Heng, Li Wen-Can, Ma Kai-Jun, Chen Long
Department of Forensic Medicine, School of Basic Medical Sciences, Fudan University, 131 Dongan Road, Shanghai, 200032, People's Republic of China.
Shanghai University of Medicine & Health Sciences, 21 Meilong Road, Shanghai, 200030, People's Republic of China.
Forensic Sci Med Pathol. 2017 Mar;13(1):20-27. doi: 10.1007/s12024-016-9827-4. Epub 2016 Dec 28.
In our previous study, a R code-based mathematical model using RNA degradation patterns was developed for PMI determination in rat brain specimens. However, the postmortem changes of RNA are much more complicated in real cases, and there is still a huge challenge in efficiently applying information in animal data to real cases. In the present study, different RNA markers in both rat and human tissues were collected to screen valid biomarkers and the corresponding mathematical models were established and validated. With the same methodology, multi-RNA markers of myocardium and liver tissues were detected by qPCR and the Ct values of ten biomarkers generally increased with prolonged PMIs. 5S, miR-1 and miR-133a were shown to be optimum reference biomarkers that were not affected by a PMI of up to 5 or more days; however, liver-specific miR-122 began to degrade under higher temperatures and only 5S was selected as an endogenous control in the liver. Among the tested target RNAs, similar to our previous study in brain tissue, β-actin (ΔCt) was found to exhibit the best correlation coefficient with PMI and was employed to build mathematical models using R software. Following validation, the relatively low estimated error demonstrated that PMIs can be accurately predicted in human cases through comprehensive consideration of various factors and using effective biomarkers.
在我们之前的研究中,开发了一种基于R代码的数学模型,该模型利用RNA降解模式来确定大鼠脑标本中的死亡时间(PMI)。然而,在实际案例中,RNA的死后变化要复杂得多,将动物数据中的信息有效应用于实际案例仍面临巨大挑战。在本研究中,收集了大鼠和人类组织中的不同RNA标记物,以筛选有效的生物标志物,并建立和验证了相应的数学模型。采用相同的方法,通过qPCR检测心肌和肝组织的多RNA标记物,十种生物标志物的Ct值通常随PMI延长而增加。5S、miR-1和miR-133a被证明是最佳的参考生物标志物,在长达5天或更长时间的PMI下不受影响;然而,肝脏特异性miR-122在较高温度下开始降解,在肝脏中仅选择5S作为内参。在测试的目标RNA中,与我们之前在脑组织中的研究相似,发现β-肌动蛋白(ΔCt)与PMI的相关系数最佳,并使用R软件构建数学模型。经过验证,相对较低的估计误差表明,通过综合考虑各种因素并使用有效的生物标志物,可以准确预测人类案例中的PMI。