title: Evaluation of mRNA markers for estimating blood deposition time: Towards alibi testing from human forensic stains with rhythmic biomarkers.
abstract: Determining the time a biological trace was left at a scene of crime reflects a crucial aspect of forensic investigations as - if possible - it would permit testing the sample donor's alibi directly from the trace evidence, helping to link (or not) the DNA-identified sample donor with the crime event. However, reliable and robust methodology is lacking thus far. In this study, we assessed the suitability of mRNA for the purpose of estimating blood deposition time, and its added value relative to melatonin and cortisol, two circadian hormones we previously introduced for this purpose. By analysing 21 candidate mRNA markers in blood samples from 12 individuals collected around the clock at 2h intervals for 36h under real-life, controlled conditions, we identified 11 mRNAs with statistically significant expression rhythms. We then used these 11 significantly rhythmic mRNA markers, with and without melatonin and cortisol also analysed in these samples, to establish statistical models for predicting day/night time categories. We found that although in general mRNA-based estimation of time categories was less accurate than hormone-based estimation, the use of three mRNA markers HSPA1B, MKNK2 and PER3 together with melatonin and cortisol generally enhanced the time prediction accuracy relative to the use of the two hormones alone. Our data best support a model that by using these five molecular biomarkers estimates three time categories, i.e. night/early morning, morning/noon, and afternoon/evening with prediction accuracies expressed as AUC values of 0.88, 0.88, and 0.95, respectively. For the first time, we demonstrate the value of mRNA for blood deposition timing and introduce a statistical model for estimating day/night time categories based on molecular biomarkers, which shall be further validated with additional samples in the future. Moreover, our work provides new leads for molecular approaches on time of death estimation using the significantly rhythmic mRNA markers established here.
authors.affiliation: Department of Genetic Identification, Erasmus MC University Medical Centre Rotterdam, Rotterdam, The Netherlands.
authors.forename: Karolina
authors.initials: K
authors.lastname: Lech
authors.affiliation: Department of Genetic Identification, Erasmus MC University Medical Centre Rotterdam, Rotterdam, The Netherlands.
authors.forename: Fan
authors.initials: F
authors.lastname: Liu
authors.affiliation: Department of Genetic Identification, Erasmus MC University Medical Centre Rotterdam, Rotterdam, The Netherlands; EaStCHEM School of Chemistry, Biomedical Sciences Research Complex and Centre of Magnetic Resonance, University of St. Andrews, St. Andrews, United Kingdom.
authors.forename: Katrin
authors.initials: K
authors.lastname: Ackermann
authors.affiliation: Centre for Chronobiology, Faculty of Health and Medical Sciences, University of Surrey, Guildford, United Kingdom.
authors.forename: Victoria L
authors.initials: VL
authors.lastname: Revell
authors.affiliation: Department of Genetic Identification, Erasmus MC University Medical Centre Rotterdam, Rotterdam, The Netherlands; CNAG-CRG, Centro Nacional de Análisis Genómico, Parc Científic de Barcelona-Torre I, Barcelona, Spain.
authors.forename: Oscar
authors.initials: O
authors.lastname: Lao
authors.affiliation: Centre for Chronobiology, Faculty of Health and Medical Sciences, University of Surrey, Guildford, United Kingdom.
authors.forename: Debra J
authors.initials: DJ
authors.lastname: Skene
authors.affiliation: Department of Genetic Identification, Erasmus MC University Medical Centre Rotterdam, Rotterdam, The Netherlands. Electronic address: m.kayser@erasmusmc.nl.
authors.forename: Manfred
authors.initials: M
authors.lastname: Kayser
mesh_terms: D015415:Biomarkers / Q000097:blood / Q000378:metabolism; D002940:Circadian Rhythm / Q000235:genetics*; D004396:Coloring Agents / Q000737:chemistry; D053803:Forensic Genetics / Q000379:methods*; D015870:Gene Expression; D006801:Humans; D006854:Hydrocortisone / Q000378:metabolism; D008550:Melatonin / Q000097:blood / Q000378:metabolism; D011237:Predictive Value of Tests; D020871:RNA Stability; D012333:RNA, Messenger / Q000097:blood / Q000235:genetics* / Q000378:metabolism; D012463:Saliva / Q000378:metabolism
chemical_list: D015415:Biomarkers; D004396:Coloring Agents; D012333:RNA, Messenger; D008550:Melatonin; D006854:Hydrocortisone
keywords: Blood deposition time; Circadian hormones; Forensic time estimation; Rhythmic gene expression; mRNA