大鼠尸体组织电阻抗的死后变化及与死亡时间的关系

目的探讨躯体组织的电阻抗的死后变化规律，寻找推断死后经过时间的新方法。方法用阻抗测量仪测量大鼠胃壁、腹壁、躯干、胸部、腹部、腹部皮肤、头皮等处不同死后经过时间的电阻抗。结果大鼠躯体组织的电阻抗随着死后经过时间的延长存在规律性变化。这些结果告诉我们，有望通过测量死后人体躯体组织的电阻抗，利用其死后变化规律，推断人体死后经过时间。     

Estimation of postmortem interval. Temperature-correction of extracellular abdominal impedance during the first 21 days of death

Extracellular impedance of the intact abdomen of rats increased from 79.0+/-7.4Omega, 1h postmortem (i.e. 0.04 day), to 130.5+/-14.4Omega at postmortem interval (PMI)=1 day. Impedance then decreased with time, reaching 64.2+/-9.9Omega at PMI=21 days. The time taken for mean abdominal impedance to decrease below the value at PMI=0.04 day averaged 16 days. It is therefore impossible, using extracellular abdominal impedance alone, to distinguish (in terms of interpolating PMI) between numerically equal impedances on the rising and falling phases of curves depicting impedance as a function of PMI.Correction of measured impedances to their theoretically-predicted values at an arbitrarily chosen temperature of 40 degrees C appreciably diminished the magnitude of the increase in impedance following death. Thus, temperature-corrected abdominal impedance increased from 56.2+/-4.8Omega at PMI=0.04 day to 59.5+/-6.2Omega at PMI=1 day. Impedance then decreased, reaching 29.2+/-4.1Omega at PMI=21 days. The time taken for mean, temperature-corrected abdominal impedance to decrease below the value at PMI=0.04 day averaged 3 days (as opposed to 16 days (see above) in the absence of temperature-correction).These findings are believed to improve the usefulness of extracellular abdominal impedance as a potential tool for estimation of postmortem interval.     

利用尸食性蚤蝇推断死亡时间的法医学研究进展

死亡时间的准确推断一直是法医学的难题。昆虫学方法已被认为是死亡时间推断的有效方法。尸食性蝇类发育生物学在死亡时间推断中有着重要地位。蚤蝇体形微小,在相对密闭的空间,可成为尸体上主要的甚至是唯一的昆虫证据。本文从尸食性蚤蝇的作用、种属鉴定以及日龄推断方面,综述了利用尸食性蚤蝇推断死亡时间的法医学研究进展。     

The use of Megaselia abdita (Diptera: Phoridae) in forensic entomology

This case study demonstrates the importance of the Phorid, Megaselia abdita (Schmitz), as an indicator for post-mortem interval estimation in criminal investigations involving forensic entomology where it is usually the more frequently occurring Calliphorids that are most useful. A case example is discussed where the temperatures were low for the period of time the deceased was missing.     

损伤皮肤中组胺荧光显微定位定量与损伤时间推断的实验研究

本文应用显微荧光分光光度计法研究大白鼠皮肤创缘组织中组胺的分布和含量,并用甲苯胺蓝法观察创缘肥大细胞形态和数量的变化。结果发现,创缘真皮乳头层出现扩散的细胞外黄色荧光和肥大细胞脱颗粒现象为生前伤的重要特点;组胺荧光分布范围及增多的程度与损伤时间密切相关。从而为损伤时间推断提供了一种新的方法。     

The application of amino acid racemization in the acid soluble fraction of enamel to the estimation of the age of human teeth

Estimation of age-at-death for skeletonised forensic remains is one of the most significant problems in forensic anthropology. The majority of existing morphological and histological techniques are highly inaccurate, and show a bias towards underestimating the age of older individuals. One technique which has been successful in forensic age estimation is amino acid racemization in dentine. However, this method cannot be used on remains where the post-mortem interval is greater than 20 years. An alternative approach is to measure amino acid racemization in dental enamel, which is believed to be more resistant to change post-mortem. The extent of amino acid racemization in the acid soluble fraction of the enamel proteins was determined for modern known age teeth. A strong correlation was observed between the age of the tooth and the extent of racemization. No systematic bias in the direction of age estimation errors was detected. For the majority of teeth analyzed, the presence of dental caries did not affect the results obtained. In a minority of cases, carious teeth showed a higher level of racemization than would be expected given the age of the individual. These results indicate that amino acid racemization in enamel has the potential to be used in age estimation of skeletal remains.     

Kinetics of ethanol degradation in forensic blood samples

To determine ethanol in human post-mortem blood samples is problematic, largely due to the inappropriate and variable methods of preserving and storing, which can cause decomposition and loss of alcohol concentration. In this study, four crucial parameters of sample conservation were studied: temperature (T), percentage of air chamber in container (%CA), ethanol concentration in blood and post-mortem time. Blood samples from post-mortem cases were stored under different conditions (ethanol levels were known in all cases); factorial design variables: (%CA) 0, 5, 20, 35, 65%; storage temperature: 25, 4 and -10 degrees C; in a total of 15 experiments. No preserving agent was used in samples. Quantification of ethanol in blood was carried out by gas chromatography with head-space FID detector. Initial ethanol concentration ranged from 0.50 to 4.30 g/L. The kinetics of degradation observed was pseudo-first-order. The parameter that characterised the kinetics of ethanol degradation (k(0)) ranged from (4 x 10(-4) and 5.0 x 10(-1) day(-1)), depending on storage conditions. A strong dependence between ethanol degradation and the content of the air chamber was observed and this dependence was found to be stronger than that between degradation and temperature; there was an experimental relation between (k(0)) and (%CA). Activation energy for different conditions, i.e. 0, 5, 20, 35 and 65 (%CA), were calculated and contour plots were made. A mathematical equation relating air chamber, temperature and ethanol concentration at a certain time was determined. This equation allowed estimation of initial concentrations of ethanol with minimal error. A good correlation between experimental data and data calculated with the equation was obtained (r(2) = 0.9998). The best storage conditions were: 0% CA and storage at -10 degrees C, obtaining an ethanol degradation of 0.01% after 15 days. However, 33% of ethanol degradation was obtained with 35% CA at 25 degrees C after 15 days. This equation is useful in forensic cases in which original concentration of ethanol has to be estimated under different sample storage conditions.     

A comparison of endogenous and exogenous RNA reference marker as relevant for accurate Post-Mortem Interval estimation

The Post Mortem-Interval (PMI), also known as time since death, is of relevance for forensic routine casework and serves primarily as the basis for the determination of the time that has passed since the time of death. The existing, “classical” approaches are characterised by certain limitations because they are predominantly based on morphological examinations.The present study deals with the assessment of two molecular messengerRNA-markers in different kinds post-mortem human specimens concerning their ability to use them for the estimation of the time of death. Moreover, a comparison between carefully selected endogenous and one exogenous reference marker for the normalisation of the gene expression data as relevant for accurate PMI estimation is drawn.The evaluation showed that the used approach is methodologically useful but on consideration of the achieved results it was only possible to determine slightly meaningful tendencies. The tested exogenous reference marker was not suitable for sufficient data normalisation whereas the endogenous reference markers showed a stable expression. However, it was not possible to recognise a specific expression level of the genes within a PMI group when endogenous reference markers were used.     

Evaluation of hypostasis using a colorimeter measuring system and its application to assessment of the post-mortem interval (time of death)

Hypostasis was measured in 93 cadavers using a tristimulus colorimeter in order to investigate its relationship with the time of death. The intensity (lightness) of the hypostasis in each case was measured over a period of 4 h and the rate of change in lightness derived. When examined against the time of death, it was found that there was a good correlation between the two. Namely, that the rate of change of lightness (and it can be inferred that this represents displaceability) decreases as the post-mortem period increases. The shift in hypostasis was particularly marked in the first 12 h and decreased thereafter. However, hypostasis could be useful for time of death estimation for up to 48 h. After this time, the degree of change was small or non-existent and by 72 h hypostasis had become fixed in the majority of cases, within our measuring period of 4 h.     

Supplementary pathway for vitality of wounds and wound age estimation in bruises using the electric impedance spectroscopy technique

Determination of wound vitality and estimation of wound age are central issues in daily forensic practice. The objective of this study was to develop a new and rapid method for determining wound vitality and estimating wound age in bruises using electric impedance spectroscopy. Forty Sprague-Dawley rats (140-170 g) were divided into five groups: group 1 (n=8): controls, group 2 (n=8): postmortem bruises, group 3 (n=8): bruises 1 h before death, group 4 (n=8): bruises 3 h before death, group 5 (n=8): bruises 6 h before death. Measurements of the right gluteus maximus muscle were taken at 6, 24, and 48 h after the rats were sacrificed by cervical dislocation. The results from this study indicate that electric impedance spectroscopy is clearly sensitive enough to differentiate between vital and postmortem wound infliction and to determine the survival time after the infliction of an injury.     

Post-mortem analysis of apoptotic changes associated with human skin bruises

The estimation of the age of skin bruises is of importance in forensic medicine, especially in child abuse cases. Time-dependent changes in bruise colour and/or associated histological features have been used with a limited degree of success. An increased rate of apoptosis in the injured tissue has been considered as a novel time-dependent marker of cell death, by injury inflicted in a rat model. The object of the present study was to apply the TUNEL method of DNA end labelling to identify and enumerate apoptotic cells in bruised and normal skin in order to study the relationship of apoptotic cell density with the age of the bruise. A commercially available DNA end labelling kit, TUNEL method, was standardised, validated and used for this purpose. Twenty unselected post-mortem cases with bruises due to a variety of causes were studied. The apoptotic cells stained with TUNEL reaction were counted in 10 high power fields in the epidermis, as well as in the dermis of formaldehyde-fixed paraffin-embedded skin specimens. The mean positive cell densities (+/- 1 S.E.) were compared with respect to the age of the bruise. In the epidermis, the mean apoptotic cell count was statistically significantly greater in the bruised skin compared to normal skin in 2- to 6-day-old bruises; whilst in the dermis the same was true in 3- to 8-day-old bruises. The overall findings suggest that there is a quiescent period prior to the increase in the apoptotic cell activity that is seen following skin bruising. This is so provided the post-mortem skin samples were collected within a lapse of 6 days or less between the time of death and formalin fixation and paraffin embedding to avoid the bias made by the difference of length of post-mortem interval.     

The possible influence of micro-organisms and putrefaction in the production of GHB in post-mortem biological fluid

In recent years, the post-mortem production of the drug of abuse gamma-hydroxybutyric acid (GHB) in biological fluids (e.g. blood and urine) has caused various interpretative problems for toxicologists. Previously, other researchers have shown certain microbial species (Pseudomonas spp. and Clostridium aminobutyricum) possess the necessary enzymes to convert GABA to GHB. A preliminary investigation involving putrefied post-mortem blood indicated there was no observed relationship between "endogenous" GHB concentrations and concentrations of common putrefactive markers (tryptamine and phenyl-2-ethylamine). Microbiological analysis identified the presence of various micro-organisms: Clostridia spp., Escherichia coli, Proteus vulgaris, Enterococcus faecalis and Aeromonoas spp. Equine plasma, human blood and urine samples were inoculated with these and an additional micro-organism (Pseudomonas aeruginosa) and incubated at 22 degrees C for 1 month. Following comparison with control samples and pre-inoculation concentrations, the data indicated an apparent production of GHB in unpreserved P. aeruginosa inoculated blood (2.3 mg/l). All other fluoride-preserved and unpreserved samples (including controls) had GHB concentrations <1mg/l. Although this concentration is lower than is typically associated with "endogenous" post-mortem GHB concentrations, this paper proposes a potential microbial production of GHB with time.     

大鼠挫伤骨骼肌中M1/M2型巨噬细胞时间相关性研究

目的 研究大鼠骨骼肌挫伤后M1/M2型巨噬细胞浸润的动态变化规律结合组织学改变建立损伤时间推断方法.方法 建立大鼠骨骼肌挫伤模型,挫伤后6h、12h、24h、36h、48h、60h、72h、96h、144h、216h、336h为实验组,对照组不进行任何处理.采用HE染色法和免疫荧光多重染色方法观察骨骼肌挫伤后修复规律,...     

Time of death of victims found in cold water environment

Limited data is available on the application of post-mortem temperature methods to non-standard conditions, especially in problematic real life cases in which the body of the victim is found in cold water environment. Here we present our experience on two cases with known post-mortem times. A 14-year-old girl (rectal temperature 15.5 degrees C) was found assaulted and drowned after a rainy cold night (+5 degrees C) in wet clothing (four layers) at the bottom of a shallow ditch, lying in non-flowing water. The post-mortem time turned out to be 15-16 h. Four days later, at the same time in the morning, after a cold (+/- 0 degrees C) night, a young man (rectal temperature 10.8 degrees C) was found drowned in a shallow cold drain (+4 degrees C) wearing similar clothing (four layers) and being exposed to almost similar environmental and weather conditions, except of flow (7.7 l/s or 0.3 m/s) in the drain. The post-mortem time was deduced to be 10-12 h. We tested the applicability of five practical methods to estimate time of death. Henssge's temperature-time of death nomogram method with correction factors was the most versatile and gave also most accurate results, although there is limited data on choosing of correction factors. In the first case, the right correction factor was close to 1.0 (recommended 1.1-1.2), suggesting that wet clothing acted like dry clothing in slowing down body cooling. In the second case, the right correction factor was between 0.3 and 0.5, similar to the recommended 0.35 for naked bodies in flowing water.     

Entomological identification of the post-mortem colonization of wolf cadavers in different decomposition stages

Human-driven biodiversity loss is progressively becoming a problem with dramatic consequences for the conservation of vital ecosystems. The increasing number of illegal killings of the grey wolf (Canis lupus, Linnaeus, 1758), a threatened species, displays the need for investigation and prosecution of such offences. Forensic entomology makes use of the knowledge about necrophagous insects to estimate a minimum time-since-death interval of the deceased person or animal, which can give important information on a possible perpetrator. The cadaver fauna along five decomposition stages of wolves in Germany was investigated in the period 2014–2021. The insects from 70 wolf cadavers, originating from all over Germany, were provided by the Leibniz Institute for Zoo and Wildlife Research Berlin. The accumulated degree day (ADD) model was applied for the post-mortem interval estimation on wolf cadavers for the first time. A total of 20 coleopteran species and 14 different dipteran species were discovered and identified. Almost 99 % of all insect specimens were from the order of Diptera, and beetles (Coleoptera) accounted for only 1 % of the cadaver fauna. The blowflies (Calliphoridae) are of particular importance for forensic issues, accounting for about 66 % of all families. Carrion beetles (Silphidae) were found as the second most abundant family (about 21 %). In addition, combining all cases, a steadily increasing insect species richness S was detected from early decay to advanced decay (fresh S = 8; bloated S = 12; active decay S = 21; advanced decay S = 34). In the following remains stage, the species number decreased again (S = 24). However, no significant difference in the number of species was found between the stages of decay when the cases were considered individually. The temporal pattern of insect appearance was found to be congruent with those of previous studies. Furthermore, a time of death was determined for each case and compared to the pathologist's estimates. This study provides insights into the arthropod fauna of wolf remains for the first time, applies the ADD-Model for post-mortem interval estimation, and discusses the suitability of forensic entomology for wildlife death investigations.     

Fibronectin EIIIA splicing variant: a useful contribution to forensic wounding interval estimation

A fibronectin splicing variant, Fn containing an extra type III domain (EIIIA in rat, ED1 or EDA in human) has recently attracted more attention because of its sensitive response in injury of adult tissue. The characteristic that this form is absent in adult tissue while it is commonly expressed in fetal tissues and injured adult tissue is useful in estimation of injury interval in forensic science. The regulation of fibronectin splicing was studied by immunohistochemistry by using monoclonal antibody to EIIIA on sections from paraffin embedded rat skin sample after contusion. The results indicated that epidermic cells and hair follicle epithelium of rat skin were found positive staining at 6h after injury, and the color became darker with prolonged wounding time. Based on the above result, we invented a type of test paper assayed for EIIIA-Fn splicing variant by using immune colloidal gold technique. After dipping one end of test paper in the supernatant fluid of tissue homogenates of injury skin for few minutes, detecting line could be found. It showed that all experimental injured samples revealed positive staining; the darkness of positive staining was dependent on the injury time, while control normal skin cannot found positive staining. It is concluded that there is a close relationship between the expression of EIIIA-Fn splicing variant and wounding interval, and that the gold-labeled test paper can be useful in distinguishing ante- and post-mortem injury, and in estimation of wounding interval in forensic science.     

人脾细胞核几何参数变化与早期死亡时间的相关性研究

目的运用图像分析技术探讨人脾脏细胞核几何参数变化与死亡时间的关系。方法选31例已知死亡时间的人离体脾组织,在死后36h内每h细胞学涂片、Feulgen-Vans染色,对人脾细胞核的异形指数等5个几何参数进行图像分析。结果脾脏细胞核异形指数在死后5～36h呈上升趋势,r=0.983,线性回归方程y=11.334x-33.339,面积、等效直径、平均直径、周长均与死亡时间无明显的相关性。结论异形指数能较客观地反映人脾细胞核DNA降解后的离散分布状况,与死亡时间有明显相关性,可能是推断早期准确死亡时间的较好形态学指标。     

Estimating the pre-appearance interval from temperature in Necrodes littoralis L. (Coleoptera: Silphidae)

While estimating post-mortem interval from entomological evidence it is useful to divide it into two separate intervals: the pre-appearance interval (PAI) and the development interval in the case of immature insects and PAI and the presence interval in the case of adult insects. An accurate estimate of PAI is critically important particularly in the case of those insects which appear on remains late in decomposition. In this paper the possibilities of PAI estimation from temperature are evaluated in late-arriving, forensically important carrion beetle Necrodes littoralis. The relationship between PAI of adult and larval N. littoralis and temperature was modelled by using results of succession experiments in forests of Central Europe. In both stages PAI was found to be strongly, inversely dependent on mean daily ground level temperatures averaged for the duration of PAI. Several methods of PAI estimation were proposed and their accuracy was initially evaluated with external successional data.     

Glycated hemoglobin: a useful post-mortem reference marker in determining diabetes

Glycated hemoglobin (HbA(1c)) has been demonstrated to be a useful marker for long-term glucose control in diabetes. This parameter characterizes each non-enzymatic fixation of glucose on hemoglobin. It is a useful test in addition to periodic glycemia controls since it reflects the mean glycemia of the past 60 days. We studied the conservation of HbA(1c) at 4 degrees C as a function of time with different anti-coagulants and preservatives (3, 6 months, 1 year). A total of 106 tests were performed using the high performance liquid chromatography (HPLC) method dedicated to the semi-automatic analysis of HbA(1c) (Bio-Rad) and we applied the method in forensic cases. Conservation at 4 degrees C was good for as long as 3 months in blood samples collected with fluoride and 6 months in samples collected in a dry or in a heparinized tube. In non-diabetic subjects, HbA(1c) reference values obtained from forensic samples were identical to those of living controls (3.5-6.25% of total hemoglobin). All positive HbA(1c) results were confirmed by a medical evaluation. This method was successfully applied to five forensic cases. In cases of increased acetonemia, acetone or isopropanol are easily measured. However, in some unexplained post-mortem circumstances, increased HbA(1c) permits to differentiate alcoholic or starvation ketoacidosis from the diabetic cases. Glycated hemoglobin should, therefore, be considered the forensic marker of choice in the post-mortem diagnosis of a diabetic disorder and demonstrates its usefulness in post-mortem validation.     

Rapid determination of carboxyhemoglobin in blood by Oximeter

Different methods to determine carboxyhemoglobin (COHb) in blood are described in the literature. In our laboratory three methods to analyze COHb in post-mortem blood samples were compared: the spectrophotometric method of Maehly, a gas chromatographic method with a thermal conductivity detector (GC-TCD) and the Oximeter. Several COHb containing blood samples of deceased persons were analyzed. Results of all three methods were comparable for low concentrations (ca. 10% COHb) as well as for high concentrations (ca. 80% COHb) regardless of the viscosity of the blood samples. The advantages of the Oximeter when compared to Maehly's method and GC-TCD are extreme short time of analysis (<1min), very small blood volume required (<0.1ml) and easy handling. In our opinion application of the Oximeter is not limited to analyses of blood samples from living persons (e.g. in clinical toxicology); it can as well be used for the determination of COHb in post-mortem blood samples. Hence it is a useful and time saving tool in forensic toxicology.