Age-related cytokine imbalance in the thymus in sudden infant death syndrome (SIDS).

BACKGROUND: Sudden infant death syndrome (SIDS) has been considered to be triggered by a combination of underlying immune dysregulation and infections. The thymus is a crucial lymphatic organ responsible for T cell development in infancy. We hypothesized that an altered thymic immune status may be detectable by intrathymic cytokine profiling in SIDS. METHODS: 27 cytokines in protein lysates of thymus tissue and thymus weights were assessed in 26 SIDS cases and 16 infants who died of other reasons. RESULTS: Seventeen out of 27 cytokines were increased in thymic tissue of SIDS compared to controls without infections, and the most significant discrepancy was in infants younger than 20 weeks. The thymic cytokine profiles in SIDS cases were similar to those in controls with severe infection; however, the magnitude of the cytokine concentration elevation in SIDS was less pronounced, indicating sub-clinical infections in SIDS. In contrast to SIDS, intrathymic cytokine concentrations and thymus weight were increased with age in control children. CONCLUSIONS: Elevated thymic cytokine expression and thymus weight, as well as impaired age-related alterations in SIDS, may be influenced by subclinical infection, which may play a role in initiating SIDS in infants with a compromised immune response. IMPACT STATEMENT: Increased thymic weight and cytokine concentration may suggest possible subclinical infection in SIDS. Elevated thymic weight and cytokine concentration mainly in SIDS cases aged <20 weeks. Age-related impairment in the thymic weight and cytokine expression may be impaired by subclinical infection in SIDS.

Pulmonary immune profiling of SIDS: impaired immune maturation and age-related cytokine imbalance.

BACKGROUND: For sudden infant death syndrome (SIDS), an impaired immunocompetence has been discussed for a long time. Cytokines and chemokines are soluble immune mediators (SIM) whose balance is essential for the immune status. We hypothesized that an imbalanced immune response might contribute to the etiology of SIDS. METHODS: We investigated 27 cytokines, chemokines, and growth factors in protein lysates of lungs derived from 29 SIDS cases and 15 control children deceased for other reasons. RESULTS: Except for the CCL5, no significant differences were detected in the lungs between SIDS cases with and without mild upper respiratory tract infections. In contrast, IL-1RA, IL-7, IL-13, and G-CSF were decreased in the merged SIDS cases compared to control cases without evidence of infection. Plotting SIM concentrations against infant age resulted in increasing concentrations in control but not in SIDS lungs, indicating a disturbed immune maturation. Moreover, an age-dependent shift towards a Th2-related pattern was observed in SIDS. CONCLUSIONS: Our findings suggest that an impaired maturation of the immune system, an insufficient response to respiratory pathogens, and an immune response modulated by Th1/Th2 imbalance might play a possible role in triggering SIDS. These findings might in part be explained by chronic stress. IMPACT: Maturation of the cytokine and chemokine network may be impaired in SIDS. An imbalance between Th1- and Th2-related cytokines, which may reflect a state of chronic stress causing a more Th2 shift. An impaired immune maturation, an insufficient response to respiratory pathogens, and an immune response modulated by Th1/Th2 imbalance might play a possible role in SIDS.

Sudden infant death syndrome revisited: serotonin transporter gene, polymorphisms and promoter methylation.

BACKGROUND: Based on findings in the brain stems of SIDS victims, the serotonin transporter (5-HTT) gene has been discussed to be associated with SIDS. METHODS: In the largest study to date, we investigated the promoter length (5-HTTLPR) and intron 2 VNTR polymorphisms in 274 cases and 264 controls and the Ile425Val polymorphism in 65 cases and 64 controls. Moreover, the methylation of the internal promoter region was investigated in 35 cases and 14 controls. RESULTS: For 5-HTTLPR, we observed a trend towards an association of allele L (58.8% vs. 53.4%) with SIDS and significant results were observed after stratifying for age, season at death, and prone position. Nevertheless, when pooling all published data, a significant association of allele L with SIDS is confirmed (p: 0.001). For the intron 2 VNTR polymorphism, no significant differences were observed. After pooling, a significant accumulation of the rare allele 9 was observed in SIDS (2.1% vs. 0.6%; p: 0.018). For the Ile425Val polymorphism, no differences were observed. CONCLUSION: We conclude that genetic variation at this gene might be of some importance in SIDS. Epigenetic analysis of the internal promoter, however, revealed no influence on the relative risk to succumb to SIDS. IMPACT: This is the largest study published up to now on 5-HTT gene polymorphisms and SIDS. Polymorphisms in the 5-HTT gene appear to contribute (although to a small degree) to the risk to die from SIDS. There is no evidence that a methylation of the promoter region is of impact for the etiology of SIDS.

Fatal anogenital exenteration of the intestine.

A case of a sadistically motivated homicide with extraordinary injuries is reported. A 32-year-old woman was naked with signs of severe blunt trauma and oral, vaginal and anal penetration. At the crime scene, the intestine lay next to the woman without connection to the body. During the trial before the criminal court, the perpetrator admitted fisting and inserting several objects into the vagina, anus and oral cavity. Moreover, after anal and vaginal insertion of the hands, large parts of the intestine were torn and pulled out through the anus and the vagina. The results of the forensic pathological examination and additional investigation are discussed and compared with the pertinent literature. This extraordinary case of a sadistically motivated homicide ended with a final judgment that is extremely rare in German jurisdiction.

Just another railway fatality.

We report on a 56-year-old male who was found dead on railroad tracks, equipped with overhead high-voltage power lines (15,000 V AC). Apparently, the body had been hit by a passing train and completely severed at the mid-abdominal level. Based on this apparently unequivocal finding, the police initially assumed a fatal railway accident or suicide. However, close examination of the deceased's clothes revealed heat damage to at least four overlying layers of clothing in different locations. The rubber soles of his heavy leather shoes showed small holes in blackened areas underneath the toes. Furthermore, both socks revealed tears and fabric defects with burnt rims, again most prominent in the area of the toes. Skin burns, consistent with electric burns, and blistering were detected on the deceased's hands and feet. A broken fishing rod was found in the proximity. On autopsy, multiple injuries caused by severe blunt force with subsequent skull fracture and brain laceration as well as multiple injuries to the spinal column and rib fractures were found and visceral organs displayed multiple lacerations. However, the lack of relevant hematomas argues that these injuries were inflicted postmortem. Histological examination confirmed the presence of electric burns from electrocution. Based on the results of the forensic-pathological examination and additional investigations carried out at the scene of death, we could demonstrate that this highly unusual death was caused by an electrocution after contact of the fishing rod with the high-voltage power line and not by overrunning by the train.

Apparent versus true gene expression changes of three hypoxia-related genes in autopsy derived tissue and the importance of normalisation.

The aim of our work was to show how a chosen normal-isation strategy can affect the outcome of quantitative gene expression studies. As an example, we analysed the expression of three genes known to be upregulated under hypoxic conditions: HIF1A, VEGF and SLC2A1 (GLUT1). Raw RT-qPCR data were normalised using two different strategies: a straightforward normalisation against a single reference gene, GAPDH, using the 2(-ΔΔCt) algorithm and a more complex normalisation against a normalisation factor calculated from the quantitative raw data from four previously validated reference genes. We found that the two different normalisation strategies revealed contradicting results: normalising against a validated set of reference genes revealed an upregulation of the three genes of interest in three post-mortem tissue samples (cardiac muscle, skeletal muscle and brain) under hypoxic conditions. Interestingly, we found a statistically significant difference in the relative transcript abundance of VEGF in cardiac muscle between donors who died of asphyxia versus donors who died from cardiac death. Normalisation against GAPDH alone revealed no upregulation but, in some instances, a downregulation of the genes of interest. To further analyse this discrepancy, the stability of all reference genes used were reassessed and the very low expression stability of GAPDH was found to originate from the co-regulation of this gene under hypoxic conditions. We concluded that GAPDH is not a suitable reference gene for the quantitative analysis of gene expression in hypoxia and that validation of reference genes is a crucial step for generating biologically meaningful data.

Serial Killings and Attempted Serial Killings in Hospitals, Nursing Homes, and Nursing Care.

BACKGROUND: Serial killing by doctors or nurses is rare. When it occurs, it is generally only detected after multiple homicides by the same perpetrator have escaped detection in the past. The persons at greatest risk are multimorbid elderly patients whose sudden death for natural reasons would not come as a surprise. However, patients' risk of falling victim to homicide is increased only if such vulnerable patients are exposed to perpetrators with certain personality traits. In this situation, homicides can be committed in which little or no evidence of the crime is left behind. In this review, we address the frequency, nature, and circumstances of serial killings and attempted serial killings in hospitals, nursing homes, and nursing care. METHODS: This review is based on publications retrieved by a selective review of the literature in monographs, medical databases, specialty journals, general-interest media, and the Internet. RESULTS: An evaluation of searchable, published case descriptions of serial killings and attempted serial killings in hospitals, nursing homes, and nursing care, mainly from Europe and the English-speaking countries, enables identification of the type of patients at risk, the modes of homicide, and the personality traits of the perpetrators. Multimorbid, care-dependent and nursing-dependent persons are the main victims. The perpetrators (men and women) generally act alone and have often been working in patient care for many years. The most common method of homicide is by drug injection; violent physical homicide is rarer. In many cases, irregularities in drug stocks, erratic behavior of a staff member, and/or a cluster of sudden deaths are indeed noticed, but are too slowly acted upon. CONCLUSION: Irregularities in drug stocks, inexplicably empty drug packages and used syringes, erratic behavior of a staff member before and after a patient's death, or a cluster of unexpected deaths mainly involving elderly, multimorbid patients (detectable from internal mortality statistics) should always lead to further questioning and investigation.

RNA integrity in post-mortem samples: influencing parameters and implications on RT-qPCR assays.

Messenger RNA (mRNA) profiling in post-mortem human tissue might reveal information about gene expression at the time point of death or close to it. When working with post-mortem human tissue, one is confronted with a natural RNA degradation caused by several parameters which are not yet fully understood. The aims of the present study were to analyse the influence of impaired RNA integrity on the reliability of quantitative gene expression data and to identify ante- and post-mortem parameters that might lead to reduced RNA integrities in post-mortem human brain, cardiac muscle and skeletal muscle tissues. Furthermore, this study determined the impact of several parameters like type of tissue, age at death, gender and body mass index (BMI), as well as duration of agony, cause of death and post-mortem interval on the RNA integrity. The influence of RNA integrity on the reliability of quantitative gene expression data was analysed by generating degradation profiles for three gene transcripts. Based on the deduced cycle of quantification data, this study shows that reverse transcription quantitative polymerase chain reaction (RT-qPCR) performance is affected by impaired RNA integrity. Depending on the transcript and tissue type, a shift in cycle threshold values of up to two cycles was observed. Determining RNA integrity number of 136 post-mortem samples revealed significantly different RNA qualities among the three tissue types with brain revealing significantly lower integrities compared to skeletal and cardiac muscle. The body mass index was found to influence RNA integrity in skeletal muscle tissue (M. iliopsoas). Samples originating from deceased with a BMI > 25 were of significantly lower integrity compared to samples from normal weight donors. Correct data normalisation was found to partly diminish the effects caused by impaired RNA quality. Nevertheless, it can be concluded that in post-mortem tissue with low RNA integrity numbers, the detection of large differences in gene expression activities might still be possible, whereas small expression differences are prone to misinterpretation due to degradation. Thus, when working with post-mortem samples, we recommend generating degradation profiles for all transcripts of interest in order to reveal detection limits of RT-qPCR assays.

Determination of drugs in exhumed liver and brain tissue after over 9 years of burial by liquid chromatography-tandem mass spectrometry-Part 2: Benzodiazepines, opioids, and further drugs.

In this publication, benzodiazepines, opioids, and further drugs were analyzed in exhumed brain and liver tissue samples in 116 cases (total) after 9.5-16.5 years of burial. Solid phase extraction followed by liquid chromatography-tandem mass spectrometry was applied. Data from literature is listed summarizing the detectability of the presented analytes after a certain time of burial. In our study, 60% of the analyzed benzodiazepines, 100% of the opioids, and 82% of further drugs were detectable. Only the benzodiazepines lorazepam, nitrazepam, flunitrazepam, and its metabolite norflunitrazepam, and the drugs butylscopolamine, metronidazole, and omeprazole were not detectable at all. Percentage of positive findings (total, and separately for brain and liver tissue) and postmortem period are listed for each analyte. Correlation of detectability depending on postmortem period and condition of tissue are presented exemplarily for midazolam. No substantial correlation was observed. Despite a long time of burial, most benzodiazepines, opioids, and further drugs were detectable in the examined tissue samples. Our results may be a good support for future exhumations in which toxicological analyses are relevant.

Determination of drugs in exhumed liver and brain tissue after over 9 years of burial by liquid chromatography-tandem mass spectrometry-Part 1: Cardiovascular drugs.

This paper should serve as support for future exhumations in which an analysis of cardiovascular drugs is issued after over 9 years of burial. Amiodarone, amlodipine, atropine, bisoprolol, cafedrine, clonidine, esmolol, furosemide, hydrochlorothiazide, lisinopril, nifedipine, nitrendipine, phenprocoumon, torsemide verapamil, and xipamide were determined in liver and brain tissue of over 100 cases in which exhumation was performed after over 9 years of burial. Diagrams, showing the detectability depending on postmortem period as well as condition of tissues, are presented for furosemide.

Validation of adequate endogenous reference genes for the normalisation of qPCR gene expression data in human post mortem tissue.

Gene expression analyses based on messenger RNA (mRNA) profiling require accurate data normalisation. When using endogenous reference genes, these have to be validated carefully. Therefore, we examined the transcript stability of 10 potential reference genes using quantitative real-time polymerase chain reaction: beta actin, 18S rRNA, glyceraldehyde-3-phosphate dehydrogenase, TATA box-binding protein, hypoxanthine phosphoribosyl-transferase I, beta-2-microglobulin, hydroxymethylbilane synthase, succinate dehydrogenase complex, subunit A, cyclophilin A and ubiquitin C. The aim of the current study was to assess which reference genes show stable mRNA levels in human post mortem cardiac muscle, skeletal muscle and brain tissue. Considering cardiac muscle tissue, CYCA and TBP were identified as the most stable while in skeletal muscle tissue, SDHA and TBP, and in brain tissue, SDHA and HMBS turned out to be the most stable. Furthermore, we recommend a minimum of four carefully validated endogenous control genes for reliable data normalisation in human post mortem tissue. Parameters influencing the stability of transcript amounts were found to be mainly the post mortem interval in cardiac muscle and skeletal muscle tissue and the donor's cause of death in skeletal muscle and brain samples. Further parameters like gender, age at death and body mass index were found to influence mRNA quantities in skeletal muscle only. The set of stable control genes identified in this study may be used in further studies if the composition of the samples is similar to the one used here.

A survey of warning colours of pesticides.

Pesticides are used to protect plants all over the world. Their increasing specificity has been due to utilization of differences in biochemical processes, and has been accompanied by lower human toxicity. Nevertheless cases of poisoning are still observed. While certain toxic substances are provided with characteristic dyes or pigments to facilitate easy identification, no overview of pesticide colors exists. The lack of available product information prompted us to explore the colors and dyes of pesticides registered in Germany, most of which are commercially available worldwide. A compilation of the colors and odors of 207 pesticide products is presented. While some of the substances can be identified by their physical characteristics, in other cases, the range of possibilities can be narrowed by their nature and color.

Postmortem "delivery" in a pregnant fire victim.

A 23-year-old woman traveling in the front passenger seat of a car was killed in a traffic accident involving two rear-end collisions. After the second rear-end collision, the car caught fire. While the driver could be rescued, the pregnant passenger remained inside the car. After the fire had been extinguished, a charred body was found in a pugilistic attitude with an exposed abdominal cavity and a ruptured anterior wall of the gravid uterus. Between the woman's legs, an almost unburned fetus was found with body measurements corresponding to a gestational age of 28 weeks. The placenta had remained inside the uterine cavity and was still connected with the fetus via the umbilical cord. In conclusion, the rupture of the pregnant uterus with expulsion of the fetus had to be considered a secondary event due to the initial heating and subsequent shrinking of the uterine wall.