Parking behaviour under the influence of alcohol.

Real-life driving studies evaluating the impact of alcohol influence on the ability to park a car are rare but necessary to assess a possible impairment to drive a car in the event of prosecution. In this study, 29 test persons (13 m, 16 f) completed three test drives with real cars, each made up of three different parking situations. While four test persons remained sober, the majority drank a previously calculated amount of alcohol before the second drive; the aim was to reach a blood alcohol concentration (BAC) of 1.1 g/kg. The third drive took place about 2 h later without any further ingestion of alcohol. The impact of BAC on the number of accidents, time needed to finish the drive, the amount of correction moves and quality of the final parking position (in the centre of the parking space) were analysed. Furthermore, pressure measuring films were applied to the test cars, measuring the average pressure and load in the areas of the accident impact. A significant increase of accidents could be noted with rising BAC. While a single accident happened to both sober and drivers under the influence of alcohol, more than one accident was only seen in drivers after the ingestion of alcohol (> 0.63 g/kg). The BAC had no impact on the other considered aspects. Concludingly, more than one impact site or accident while parking a car can serve as an indication for alcohol impairment of the driver at the time of the accident.

Skeletal injuries after cesarean section - a rare differential diagnosis of child abuse.

Birth-related fractures are an important differential diagnosis of child abuse in early infancy. While fractures associated to vaginal deliveries are well known, cesarean section is not necessarily known to cause such injuries. Nevertheless neonatal fractures have been described after cesarean sections. To give an overview over the frequency and typical locations of such fractures, the appearance of symptoms and the timespan until diagnosis, a literature research was conducted via Google scholar and Pubmed, using the key words "cesarean section" and "fractures". Birth-related fractures after cesarean sections are rare but can occur, with the long bones being particularly affected. Therefore, birth injuries should always be considered in the forensic medical assessment of fractures in early infancy, even after cesarean section. To enable a differentiation between birth trauma and physical abuse, birth and operation records should be checked for surgical manoeuvres, possible difficulties during the procedure or other risk factors. Birth-related fractures are usually detected early; in rare cases, the diagnosis is made only weeks after birth.

Altered DNA methylation at age-associated CpG sites in children with growth disorders: impact on age estimation?

Age estimation based on DNA methylation (DNAm) can be applied to children, adolescents and adults, but many CG dinucleotides (CpGs) exhibit different kinetics of age-associated DNAm across these age ranges. Furthermore, it is still unclear how growth disorders impact epigenetic age predictions, and this may be particularly relevant for a forensic application. In this study, we analyzed buccal mucosa samples from 95 healthy children and 104 children with different growth disorders. DNAm was analysed by pyrosequencing for 22 CpGs in the genes PDE4C, ELOVL2, RPA2, EDARADD and DDO. The relationship between DNAm and age in healthy children was tested by Spearman's rank correlation. Differences in DNAm between the groups "healthy children" and the (sub-)groups of children with growth disorders were tested by ANCOVA. Models for age estimation were trained (1) based on the data from 11 CpGs with a close correlation between DNAm and age (R ≥ 0.75) and (2) on five CpGs that also did not present significant differences in DNAm between healthy and diseased children. Statistical analysis revealed significant differences between the healthy group and the group with growth disorders (11 CpGs), the subgroup with a short stature (12 CpGs) and the non-short stature subgroup (three CpGs). The results are in line with the assumption of an epigenetic regulation of height-influencing genes. Age predictors trained on 11 CpGs with high correlations between DNAm and age revealed higher mean absolute errors (MAEs) in the group of growth disorders (mean MAE 2.21 years versus MAE 1.79 in the healthy group) as well as in the short stature (sub-)groups; furthermore, there was a clear tendency for overestimation of ages in all growth disorder groups (mean age deviations: total growth disorder group 1.85 years, short stature group 1.99 years). Age estimates on samples from children with growth disorders were more precise when using a model containing only the five CpGs that did not present significant differences in DNAm between healthy and diseased children (mean age deviations: total growth disorder group 1.45 years, short stature group 1.66 years). The results suggest that CpGs in genes involved in processes relevant for growth and development should be avoided in age prediction models for children since they may be sensitive for alterations in the DNAm pattern in cases of growth disorders.

Evidence for differences in DNA methylation between Germans and Japanese.

As a contribution to the discussion about the possible effects of ethnicity/ancestry on age estimation based on DNA methylation (DNAm) patterns, we directly compared age-associated DNAm in German and Japanese donors in one laboratory under identical conditions. DNAm was analyzed by pyrosequencing for 22 CpG sites (CpGs) in the genes PDE4C, RPA2, ELOVL2, DDO, and EDARADD in buccal mucosa samples from German and Japanese donors (N = 368 and N = 89, respectively).Twenty of these CpGs revealed a very high correlation with age and were subsequently tested for differences between German and Japanese donors aged between 10 and 65 years (N = 287 and N = 83, respectively). ANCOVA was performed by testing the Japanese samples against age- and sex-matched German subsamples (N = 83 each; extracted 500 times from the German total sample). The median p values suggest a strong evidence for significant differences (p < 0.05) at least for two CpGs (EDARADD, CpG 2, and PDE4C, CpG 2) and no differences for 11 CpGs (p > 0.3).Age prediction models based on DNAm data from all 20 CpGs from German training data did not reveal relevant differences between the Japanese test samples and German subsamples. Obviously, the high number of included "robust CpGs" prevented relevant effects of differences in DNAm at two CpGs.Nevertheless, the presented data demonstrates the need for further research regarding the impact of confounding factors on DNAm in the context of ethnicity/ancestry to ensure a high quality of age estimation. One approach may be the search for "robust" CpG markers-which requires the targeted investigation of different populations, at best by collaborative research with coordinated research strategies.

Age estimation based on different molecular clocks in several tissues and a multivariate approach: an explorative study.

Several molecular modifications accumulate in the human organism with increasing age. Some of these "molecular clocks" in DNA and in proteins open up promising approaches for the development of methods for forensic age estimation. A natural limitation of these methods arises from the fact that the chronological age is determined only indirectly by analyzing defined molecular changes that occur during aging. These changes are not linked exclusively to the expired life span but may be influenced significantly by intrinsic and extrinsic factors in the complex process of individual aging. We tested the hypothesis that a combined use of different molecular clocks in different tissues results in more precise age estimates because this approach addresses the complex aging processes in a more comprehensive way. Two molecular clocks (accumulation of D-aspartic acid (D-Asp), accumulation of pentosidine (PEN)) in two different tissues (annulus fibrosus of intervertebral discs and elastic cartilage of the epiglottis) were analyzed in 95 cases, and uni- and multivariate models for age estimation were generated. The more parameters were included in the models for age estimation, the smaller the mean absolute errors (MAE) became. While the MAEs were 7.5-11.0 years in univariate models, a multivariate model based on the two protein clocks in the two tissues resulted in a MAE of 4.0 years. These results support our hypothesis. The tested approach of a combined analysis of different molecular clocks analyzed in different tissues opens up new possibilities in postmortem age estimation. In a next step, we will add the epigenetic clock (DNA methylation) to our protein clocks (PEN, D-Asp) and expand our set of tissues.

Sudden unexpected cardiac death and postmortem identification of a novel RYR2 gene mutation.

A 13-year-old female was found lifeless at home. The autopsy and consecutive histological and toxicological examinations showed blood-rich and edematous lungs and foamy bloody content in the airways. No morphologic pathological findings were seen, especially no bleeding sources. Toxicological findings were unremarkable. The specific cause of death remained unclear. Due to reported losses of consciousness, a moleculargenetic postmortem testing was performed. A so far undescribed mutation in the cardiac ryanodine receptor gene RyR2 was detected. This mutation is suitable to explain the case history as well as the morphological findings. The cardiac ryanodine receptor gene RyR2 encodes the ryanodine receptor type 2, an ion channel in the cardiomyocytes. The ion channel regulates the influx of calcium ions and thus influences myocardial activity. Mutations in this channel may result in the catecholaminergic polymorphic ventricular tachycardia (CPVT), a cardiac arrhythmia that can lead to syncope and sudden cardiac death. This case demonstrates the usefulness and need of molecular autopsy, in particular to identify and treat possibly affected family members.

Dityrosine as a marker of acute myocardial infarction? Experiments with the isolated Langendorff heart.

The isolated Langendorff heart was used to evaluate dityrosine as a marker of acute myocardial infarctions. The animal model allowed the generation of local infarctions with defined survival times, as well as infarctions with and without reperfusion. The results showed that dityrosine, at least under the conditions of the animal model, occurs very shortly after early ischemia and infarctions, since positive staining results were already obtained after a survival time of only 5 min. Furthermore, it could be proved that the occurrence of dityrosine does not depend on a reperfusion of the ischemic muscle area and that there are no differences in the staining patterns of infarctions with and without reperfusion. Positive staining results for dityrosine in control hearts without infarctions had to be considered when evaluating the tissue samples of the study hearts. In part, the positive staining results of the control hearts seemed to be an artefact of the Langendorff system, easily identifiable by a distinctive staining pattern. Positive staining results in tissue samples of hearts that suffered from arrhythmia on the other hand implied that the occurrence of dityrosine is not specific for myocardial infarctions. Taking into account the results of previous works on human tissue samples, however, these findings did not question the use of dityrosine as a diagnostic tool; they simply showed that myocardial damage due to oxidative stress might occur under various pathologic conditions.

The identification of living persons on images: A literature review.

Personal identification in the forensic context commonly concerns unknown decedents. However, recently there has been an increase in cases which require identification of living persons, especially from surveillance systems. These cases bring about a relatively new challenge for forensic anthropologists and pathologists concerning the selection of the most suitable methodological approaches with regard to the limitations of the photographic representation of a given person for individualization and identity. Facial features are instinctively the primary focus for identification approaches. However, other body parts (e.g. hands), and body height and gait (on videos) have been considered in cases of personal identification. This review aims at summarizing the state-of-the-art concerning the identification of the living on images and videos, including a critical evaluation of the advantages and limitations of different methods. Recommendations are given in order to aid forensic practitioners who face cases of identification of living persons.

Quantitative proteomic analysis of formalin-fixed, paraffin-embedded clear cell renal cell carcinoma tissue using stable isotopic dimethylation of primary amines.

BACKGROUND: Formalin-fixed, paraffin-embedded (FFPE) tissues represent the most abundant resource of archived human specimens in pathology. Such tissue specimens are emerging as a highly valuable resource for translational proteomic studies. In quantitative proteomic analysis, reductive di-methylation of primary amines using stable isotopic formaldehyde variants is increasingly used due to its robustness and cost-effectiveness. RESULTS: In the present study we show for the first time that isotopic amine dimethylation can be used in a straightforward manner for the quantitative proteomic analysis of FFPE specimens without interference from formalin employed in the FFPE process. Isotopic amine dimethylation of FFPE specimens showed equal labeling efficiency as for cryopreserved specimens. For both FFPE and cryopreserved specimens, differential labeling of identical samples yielded highly similar ratio distributions within the expected range for dimethyl labeling. In an initial application, we profiled proteome changes in clear cell renal cell carcinoma (ccRCC) FFPE tissue specimens compared to adjacent non-malignant renal tissue. Our findings highlight increased levels of glyocolytic enzymes, annexins as well as ribosomal and proteasomal proteins. CONCLUSION: Our study establishes isotopic amine dimethylation as a versatile tool for quantitative proteomic analysis of FFPE specimens and underlines proteome alterations in ccRCC.

Cancer cell invasion and EMT marker expression: a three-dimensional study of the human cancer-host interface.

Cancer cell invasion takes place at the cancer-host interface and is a prerequisite for distant metastasis. The relationships between current biological and clinical concepts such as cell migration modes, tumour budding and epithelial-mesenchymal transition (EMT) remains unclear in several aspects, especially for the 'real' situation in human cancer. We developed a novel method that provides exact three-dimensional (3D) information on both microscopic morphology and gene expression, over a virtually unlimited spatial range, by reconstruction from serial immunostained tissue slices. Quantitative 3D assessment of tumour budding at the cancer-host interface in human pancreatic, colorectal, lung and breast adenocarcinoma suggests collective cell migration as the mechanism of cancer cell invasion, while single cancer cell migration seems to be virtually absent. Budding tumour cells display a shift towards spindle-like as well as a rounded morphology. This is associated with decreased E-cadherin staining intensity and a shift from membranous to cytoplasmic staining, as well as increased nuclear ZEB1 expression.

Towards a method for determining age ranges from faces of juveniles on photographs.

The steady increase in the distribution of juvenile pornographic material in recent years strongly required valid methods for estimating the age of the victims. At the present in fact forensic experts still commonly use the assessment of sexual characteristics by Tanner staging, although they have proven to be too subjective and deceiving for age estimation. The objective of this study, inspired by a previous EU project involving Italy, Germany and Lithuania, is to verify the applicability of certain anthropometric indices of faces in order to determine age and to create a database of facial measurements on a population of children in order to improve face ageing techniques. In this study, 1924 standardized facial images in frontal view and 1921 in lateral view of individuals from 7 age groups (3-5 years, 6-8 years, 9-11 years, 12-14 years, 15-17 years, 18-20 years, 21-24 years) underwent metric analysis. Individuals were all of Caucasoid ancestry and Italian nationality. Eighteen anthropometric indices in the frontal view and five in the lateral view were then calculated from the obtained measurements. Indices showing a correlation with age were ch-ch/ex-ex, ch-ch/pu-pu, en-en/ch-ch and se-sto/ex-ex in the frontal view, se-prn/se-sn, se-prn/se-sto and se-sn/se-sto in the lateral view. All the indices increased with age except for en-en/ch-ch, without relevant differences between males and females. These results provide an interesting starting point not only for placing a photographed face in an age range but also for refining the techniques of face ageing and personal identification.

STAT3 expression, activity and functional consequences of STAT3 inhibition in esophageal squamous cell carcinomas and Barrett's adenocarcinomas.

Signal transducer and activator of transcription 3 (STAT3) is altered in several epithelial cancers and represents a potential therapeutic target. Here, STAT3 expression, activity and cellular functions were examined in two main histotypes of esophageal carcinomas. In situ, immunohistochemistry for STAT3 and STAT3-Tyr705 phosphorylation (P-STAT3) in esophageal squamous cell carcinomas (ESCC, n=49) and Barrett's adenocarcinomas (BAC, n=61) revealed similar STAT3 expression in ESCCs and BACs (P=0.109), but preferentially activated P-STAT3 in ESCCs (P=0.013). In vitro, strong STAT3 activation was seen by epidermal growth factor (EGF) stimulation in OE21 (ESCC) cells, whereas OE33 (BAC) cells showed constitutive weak STAT3 activation. STAT3 knockdown significantly reduced cell proliferation of OE21 (P=0.0148) and OE33 (P=0.0243) cells. Importantly, STAT3 knockdown reduced cell migration of OE33 cells by 2.5-fold in two types of migration assays (P=0.073, P=0.015), but not in OE21 cells (P=0.1079, P=0.386). Investigation of transcriptome analysis of STAT3 knockdown revealed a reduced STAT3 level associated with significant downregulation of cell cycle genes in both OE21 (P<0.0001) and OE33 (P=0.01) cells. In contrast, genes promoting cell migration (CTHRC1) were markedly upregulated in OE21 cells, whereas a gene linked to tight-junction stabilization and restricted cell motility (SHROOM2) was downregulated in OE21 but upregulated in OE33 cells. This study shows frequent, but distinct, patterns of STAT3 expression and activation in ESCCs and BACs. STAT3 knockdown reduces cell proliferation in ESCC and BAC cells, inhibits migration of BAC cells and may support cell migration of ESCC cells. Thereby, novel STAT3-regulated genes involved in ESCC and BAC cell proliferation and cell migration were identified. Thus, STAT3 may be further exploited as a potential novel therapeutic target, however, by careful distinction between the two histotypes of esophageal cancers.

The juvenile face as a suitable age indicator in child pornography cases: a pilot study on the reliability of automated and visual estimation approaches.

OBJECTIVE: In cases of suspected child pornography, the age of the victim represents a crucial factor for legal prosecution. The conventional methods for age estimation provide unreliable age estimates, particularly if teenage victims are concerned. In this pilot study, the potential of age estimation for screening purposes is explored for juvenile faces. In addition to a visual approach, an automated procedure is introduced, which has the ability to rapidly scan through large numbers of suspicious image data in order to trace juvenile faces. METHODS: Age estimations were performed by experts, non-experts and the Demonstrator of a developed software on frontal facial images of 50 females aged 10-19 years from Germany, Italy, and Lithuania. To test the accuracy, the mean absolute error (MAE) between the estimates and the real ages was calculated for each examiner and the Demonstrator. RESULTS: The Demonstrator achieved the lowest MAE (1.47 years) for the 50 test images. Decreased image quality had no significant impact on the performance and classification results. The experts delivered slightly less accurate MAE (1.63 years). Throughout the tested age range, both the manual and the automated approach led to reliable age estimates within the limits of natural biological variability. CONCLUSIONS: The visual analysis of the face produces reasonably accurate age estimates up to the age of 18 years, which is the legally relevant age threshold for victims in cases of pedo-pornography. This approach can be applied in conjunction with the conventional methods for a preliminary age estimation of juveniles depicted on images.

Pitfalls at the root of facial assessment on photographs: a quantitative study of accuracy in positioning facial landmarks.

In the last years, facial analysis has gained great interest also for forensic anthropology. The application of facial landmarks may bring about relevant advantages for the analysis of 2D images by measuring distances and extracting quantitative indices. However, this is a complex task which depends upon the variability in positioning facial landmarks. In addition, literature provides only general indications concerning the reliability in positioning facial landmarks on photographic material, and no study is available concerning the specific errors which may be encountered in such an operation. The aim of this study is to analyze the inter- and intra-observer error in defining facial landmarks on photographs by using a software specifically developed for this purpose. Twenty-four operators were requested to define 22 facial landmarks on frontal view photographs and 11 on lateral view images; in addition, three operators repeated the procedure on the same photographs 20 times (at distance of 24 h). In the frontal view, the landmarks with less dispersion were the pupil, cheilion, endocanthion, and stomion (sto), and the landmarks with the highest dispersion were gonion, zygion, frontotemporale, tragion, and selion (se). In the lateral view, the landmarks with the least dispersion were se, pronasale, subnasale, and sto, whereas landmarks with the highest dispersion were gnathion, pogonion, and tragion. Results confirm that few anatomical points can be defined with the highest accuracy and show the importance of the preliminary investigation of reliability in positioning facial landmarks.

Quality improvement of fingerprints of decayed corpses by local thanatopractical processing (Thanatoprint).

Thanatopractical processing allows morphological reconstruction of even advanced decayed bodies. By extracting fluids from the body's tissue antemortem tenseness and volume can be restored. If bodies are partly subject to thanatopractical processing in the hand region ("Thanatoprint"), fingerprints of high quality can be gathered even in cases of advanced decay. Without this treatment fingerprinting can be extremely difficult, if not impossible. Thanatopractical processing could be applied successfully in cases of partial to subtotal detachment of the epidermis as well. In an interdisciplinary study 400 fingerprints of bodies in various states of decay were examined after application of Thanatoprint. In 76.75% fingerprints were applicable for data entry into AFIS (Automated Fingerprint Identification System); another 11.00% of the fingerprints could be used for the process of non-elimination. Further advantages of the method are low invasivity while maintaining the integrity of the corpse, less time- and material requirement as well as its long-lasting effect.

Age changes of facial measurements in European young adult males: implications for the identification of the living.

Metric and morphological analyses of facial features are currently applied in cases of personal identification of the living on images acquired from video surveillance systems. However, facial assessment in the forensic context needs to be based on reliable comparative data for facial measurements. Facial changes in the age range of early adulthood (20-30 years) have been rarely described so far, although such knowledge would be beneficial for comparative personal identification on images. This study investigates changes in facial measurements in European males aged between 20 and 30 years in order to identify metric characters that can be used for personal identification in young adults. A sample of 404 males of European ancestry, aged between 20 and 30 years from Germany, Italy and Lithuania were recruited for this project. Fourteen facial measurements were taken and correlation coefficients were calculated for each cranial measurement with age. Only two measurements - labial width and physiognomic ear length - seem to change between 20 and 30 years with a positive statistically significant correlation (p<0.05). These results suggest caution for what may concern personal identification by assessment of ear and mouth morphology.

DREEM on, dentists! Students' perceptions of the educational environment in a German dental school as measured by the Dundee Ready Education Environment Measure.

OBJECTIVES: The educational climate in which future doctors are trained is an important aspect of medical education. In contrast to human medicine, it has been rather neglected in dental educational research. The aim of the study was to supplement this lack by applying and validating the Dundee Ready Education Environment Measure (DREEM) for the first time in a German-speaking sample of dental students. METHODS: All dental students at the Medical Faculty of Heinrich-Heine-University Düsseldorf were asked to complete a German adaptation of the DREEM and the Düsseldorf Mission Statement Questionnaire (DMSQ) in a paper-pencil survey. Data from 205 participants were analysed. Psychometric validation included analysis of item homogeneity and discrimination, test reliability, criterion and construct validity (convergent, factorial). RESULTS: DREEM item parameters were satisfactory, reliability (α = 0.87) and convergent validity (r = 0.66 with DMSQ) were also high. Factor analyses, however, yielded dimensions which did not fully correspond to the original DREEM subscales. Overall perception of the educational environment was positive (DREEM total score = 122.95 ± 15.52). Students in the clinical part of course rated the atmosphere more negatively, but their academic self-perception more positively than preclinical students. CONCLUSIONS: Showing satisfactory psychometric properties, DREEM proved suitable for assessing educational environments among dental students. Given the right circumstances, e.g., small and early clinically oriented classes, traditional curricula can generate positive environments.