Fast microglial activation after severe traumatic brain injuries.

Traumatic brain injury is among the leading causes of death in individuals under 45 years of age. However, since trauma mechanisms and survival times differ enormously, the exact mechanisms leading to the primary and secondary injury and eventually to death after traumatic brain injury (TBI) remain unclear. Several studies showed the versatile functions of microglia, the innate macrophages of the brain, following a TBI. Earlier being characterized as rather neurotoxic, neuroprotective capacities were recently demonstrated, therefore, making microglia one of the key players following TBI. Especially in cases with only short survival times, immediate microglial reactions are of great forensic interest in questions of wound age estimation. Using standardized immunohistochemical methods, we examined 8 cases which died causatively of TBI with survival times between minutes and 7 days and 5 control cases with cardiovascular failure as the cause of death to determine acute changes in microglial morphology and antigen expression after TBI. In this pilot study, we detected highly localized changes in microglial morphology already early after traumatic damage, e.g., activated microglia and phagocyted erythrocytes in the contusion areas in cases with minute survival. Furthermore, an altered antigen expression was observed with increasing trauma wound age, showing similar effects like earlier transcriptomic studies. There is minute data on the direct impact of shear forces on microglial morphology. We were able to show localization-depending effects on microglial morphology causing localized dystrophy and adjacent activation. While rodent studies are widespread, they fail to mimic the exact mechanisms in human TBI response. Therefore, more studies focusing on cadaveric samples need to follow to thoroughly define the mechanisms leading to cell destruction and eventually evaluate their forensic value.

Post-mortem in situ stability of serum markers of cerebral damage and acute phase response.

The aim of the given study was to test the in situ stability of biochemical markers of cerebral damage and acute phase response in the early post-mortem interval to assess their usability for forensic pathology. A monocentric, prospective study investigated post-mortem femoral venous blood samples at four time points obtained within 48 h post-mortem starting at the death of 20 deceased, using commercial immunoassays for the ten parameters: S100 calcium-binding protein B (S100B), glial fibrillary acidic protein (GFAP), neuron-specific enolase (NSE), brain-derived neurotrophic factor (BDNF), interleukin-6 (IL-6), C-reactive protein (CRP), procalcitonin (PCT), ferritin, soluble tumor necrosis factor receptor type 1 (sTNFR1), and lactate dehydrogenase (LDH). Significant changes in serum levels were observed only later than 2 h after death for all markers. Inter-laboratory comparability was high, and intra-assay precision was sufficient for most markers. Most of the biomarker levels depended on the severity of hemolysis and lipemia but were robust against freeze-thaw cycles. Serum levels increased with longer post-mortem intervals for S100B, NSE, ferritin, sTNFR1, and LDH (for all p < 0.001) but decreased over this period for CRP (p = 0.089) and PCT (p < 0.001). Largely unchanged median values were found for GFAP (p = 0.139), BDNF (p = 0.106), and IL-6 (p = 0.094). Serum levels of CRP (p = 0.059) and LDH (p = 0.109) did not differ significantly between the final ante-mortem (resuscitation) and the first post-mortem sample (moment of death). Collecting the post-mortem blood sample as soon as possible will reduce the influence of post-mortem blood changes. Serum GFAP for detection of cerebral damage as well as serum IL-6 and CRP as proof of acute phase response seemed to be preferable due to their in situ stability in the first 2 days after death.

Frequency and intensity of pulmonary bone marrow and fat embolism due to manual or automated chest compressions during cardiopulmonary resuscitation.

Iatrogenic consequences of cardiopulmonary resuscitation (CPR) include sternal or rib fractures, pulmonary bone marrow embolisms (BME) and fat embolisms (FE). This report aimed to analyze the frequency and intensity of pulmonary BME and FE in fatal cases receiving final CPR efforts with the use of automated chest compression devices (ACCD) or manual chest compressions (mCC). The study cohort (all cardiac causes of death, no ante-mortem fractures) consisted of 15 cases for each group 'ACCD', 'mCC' and 'no CPR'. Lung tissue samples were retrieved and stained with hematoxylin eosin (n = 4 each) and Sudan III (n = 2 each). Evaluation was conducted microscopically for any existence of BME or FE, the frequency of BME-positive vessels, vessel size for BME and the graduation according to Falzi for FE. The data were compared statistically using non-parametric analyses. All groups were matched except for CPR duration (ACCD > mCC) but this time interval was linked to the existence of pulmonary BME (p = 0.031). Both entities occur in less than 25% of all cases following unsuccessful CPR. BME was only detectable in CPR cases, but was similar between ACCD and mCC cases for BME frequency (p = 0.666), BME intensity (p = 0.857) and the size of BME-affected pulmonary vessels (p = 0.075). If any, only mild pulmonary FE (grade I) was diagnosed without differences in the CPR method (p = 0.624). There was a significant correlation between existence of BME and FE (p = 0.043). Given the frequency, intensity and size of pulmonary BME and FE following CPR, these conditions may unlikely be considered as causative for death in case of initial survival but can be found in lower frequencies in autopsy histology.

Proposals for best-quality immunohistochemical staining of paraffin-embedded brain tissue slides in forensics.

Immunohistochemistry (IHC) has become an integral part in forensic histopathology over the last decades. However, the underlying methods for IHC vary greatly depending on the institution, creating a lack of comparability. The aim of this study was to assess the optimal approach for different technical aspects of IHC, in order to improve and standardize this procedure. Therefore, qualitative results from manual and automatic IHC staining of brain samples were compared, as well as potential differences in suitability of common IHC glass slides. Further, possibilities of image digitalization and connected issues were investigated. In our study, automatic staining showed more consistent staining results, compared to manual staining procedures. Digitalization and digital post-processing facilitated direct analysis and analysis for reproducibility considerably. No differences were found for different commercially available microscopic glass slides regarding suitability of IHC brain researches, but a certain rate of tissue loss should be expected during the staining process.

Acute phase response after fatal traumatic brain injury.

An inflammatory response occurring after fatal traumatic brain injury (TBI) initiates time-dependent cascades of acute phase response. This may offer the potential to monitor postmortem biomarker levels of several pro-inflammatory cytokines to gain information about the cause of death and the trauma survival time. Cerebrospinal fluid (CSF) and serum samples were collected from forensic autopsies of 95 adult cadavers after postmortem intervals up to 6 days. The cases were divided according to their cause of death into fatal TBI (n = 46) with different survival times and age- and gender-matching non-TBI fatalities as controls (n = 49). Quantitative marker levels of interleukin-6 (IL-6), ferritin, soluble tumor necrosis factor receptor type 1, C-reactive protein, and lactate dehydrogenase were analyzed using immunoassays. Standardized statistical tests were performed to differentiate causes of death and survival time of TBI cases. The CSF IL-6, ferritin, and LDH levels after TBI were significantly higher than those in the controls (p < 0.001). Only serum IL-6 values showed comparable differences (p < 0.05). Both CSF and serum ferritin levels were discriminative between early and delayed death after TBI (p < 0.05). There were partly distinctive correlations between marker levels in both fluids with rising values after longer survival. There were up to moderate correlation between the marker levels and the postmortem interval due to postmortem hemolysis. However, neither CSF nor serum level ranges were affected by the age or gender of the subjects. This study is the first to measure all five proteins systematically in postmortem trauma cases. Ferritin and IL-6 proved themselves to be interesting postmortem biomarkers to provide specific information on the injury pattern and the survival time of traumatic fatalities. Such forensic investigations could serve as inexpensive and fast laboratory tests.

Chest compression-associated injuries in cardiac arrest patients treated with manual chest compressions versus automated chest compression devices (LUCAS II) - a forensic autopsy-based comparison.

The aim of this autopsy study was to investigate chest-compression associated injuries to the trunk in out-of-hospital and in-hospital non-traumatic cardiac arrest patients treated with automated external chest compression devices (ACCD; all with LUCAS II devices) versus exclusive manual chest compressions (mCC). In this retrospective single-center study, all forensic autopsies between 2011 and 2017 were included. Injuries following cardiopulmonary resuscitation (CPR) in patients treated with mCC or ACCD were investigated and statistically compared using a bivariate logistic regression. In the seven-year period with 4433 autopsies, 614 were analyzed following CPR (mCC vs. ACCD: n = 501 vs. n = 113). The presence of any type of trunk injury was correlated with longer resuscitation intervals (30 ± 15 vs. 44 ± 25 min, p < 0.05). In comparison with mCC, treatment with ACCD led to more frequent skin emphysema (5 vs 0%, p = 0.012), pneumothorax (6 vs. 1%, p = 0.008), lung lesions (19 vs. 4%, p = 0.008), hemopericardium (3 vs 1%, p = 0.025) and liver lesions (10 vs. 1%, p = 0.001), all irrespective of confounding aspects. Higher age and longer CPR durations statistically influenced frequency of sternal and rib fractures (p < 0.001). The mean number of fractured ribs did not vary significantly between the groups (6 ± 3 vs. 7 ± 2, p = 0.09). In this cohort with unsuccessful CPR, chest compression-related injuries were more frequent following ACCD application than in the mCC group, but with only minutely increased odds ratios. The severity of injuries did not differ between the groups, and no iatrogenic injury was declared by the forensic pathologist as being fatal. In the clinical routine after successful return of spontaneous circulation a computed tomography scan for CPR-associated injuries is recommended as soon as possible.

X-chromosomal 21-indel marker panel in German and Baltic populations.

In order to verify specific biallelic X-indels and their characteristic properties in distinct populations, one German and three Baltic population groups (Estonia, Latvia, and Lithuania) have been analyzed by a short amplicon method, which also enables detection of degraded DNA samples. To combine 21 indels in a single multiplex PCR, all products were arranged according to their expected amplicon length (~40-160 bp) on the basis of three different fluorochromes. Separation of PCR products was carried out in a single capillary electrophoresis. Data evaluating was performed including five further indel markers which have already been tested in identical samples, resulting in altogether 26 markers. The majority of the genetic material showed combinations of insertion elements (L-fragments). Combinations of deletion elements (S-fragments) in contrast occurred with significant lower ratios. Hardy-Weinberg equilibrium (HWE) was observed for all markers except for MID1361 and MID329. This was attributed to an insufficient number of samples. For two known linkage groups within the 26-indel set (MID357-MID356 and MID3690-MID3719-MID2089), haplotype data were determined. A pairwise comparison of German and Baltic allele frequencies did not show significant deviation. This result indicates a possible genetic association between all four population groups. The calculated biostatistical parameters show high forensic efficiency for this set of indel markers. In a segregation analysis investigating 194 meiosis, no mutations have been detected regarding expected transmission patterns.

Unusual planned complex suicide committed with a muzzle-loading pistol in combination with subsequent hanging.

In Germany, suicides by firearms are not very common in contrast to deaths by hanging and intoxications. The use of historical muzzle-loading firearms in the context of suicides is a rarity. Contact shots from muzzle loaders cause an unusual wound morphology with extensive soot soiling. We report the case of a 59-year-old man, who committed a planned complex suicide by shooting into his mouth with a replica percussion gun in combination with hanging. The gunshot injury showed strong explosive effects in the oral cavity with fractures of the facial bones and the skull associated with cerebral evisceration (so-called Krönlein shot). Due to the special constellation of the case with hanging immediately after the shot, external bleeding from the head injuries was only moderate. Therefore, the head injuries could be assessed and partially reconstructed already at the scene.

[On the forensic relevance of orphan diseases]. / Zur forensischen Relevanz seltener Erkrankungen.

A 40-year-old woman died shortly after complaining of non-specific symptoms after a pharmacist had accidentally given her the wrong medication. The woman's partner was not familiar with her medical history and the medical file had to be obtained from the family doctor. Autopsy findings and histological examination confirmed the clinically diagnosed autoimmune polyglandular syndrome without a tangible cause of death. Poisoning could not be demonstrated and no relation between the dosage error and death could be established. Laboratory tests revealed diabetic coma with ketoacidosis as the cause of death, which was probably caused by a prolonged lack of insulin administration. In addition to the clarification of legal issues, the complete post-mortem examination of orphan diseases is also relevant for achieving a better understanding of differential diagnostic aspects and complex pathophysiological contexts. Moreover, the genetic background often underlying such diseases should be a reason to inform the family of the deceased about the autopsy results. Only then can secondary preventive measures be taken in time.