Interpretation of lesions of the cardiac conduction system in cocaine-related fatalities.

This study examines cases of chronic drug users who died suddenly after drug administration. Victims were young subjects, aged from 19 to 35 from Switzerland and known to the police as long-term drug users. The circumstances of death suggested the occurrence of a sudden, unexpected death. Some victims were undergoing methadone treatment. In each case, a forensic autopsy and toxicological analyses were performed at the Institute of Forensic Medicine in Lausanne in Switzerland between 2002 and 2004, including hair analysis as a means to establish chronic drug use in general, and cocaine use in particular. The conduction system was examined histologically and cases showing potentially lethal changes were chosen for this report. The most frequent lesions found were severe thickening of the atrioventricular node artery, intranodal and perinodal fibrosis, and microscopic foci of chronic inflammatory infiltration. The authors conclude that pathological lesions in the conduction tissue may play a role in the occurrence of death attributed to intoxication consecutive to cocaine ingestion.

Deposition rates of sidestream tobacco smoke particles in an experimental chamber

The natural dissipation rates of sidestream smoke (SS) particles dispersed in a chamber were studied from the standpoint of a static atmosphere and were expressed as half-lives of residence in the air. The half-lives for particles less than 0.3 micron, 0.3-0.5 micron and 0.5-1 micron were found to be 25.5, 12.8 and 4.9 h, respectively. Total particulate matter (TPM) decreases by half after 6.2 h. Other data on diluted SS in the indoor air were also reported.

Ochratoxin A at nanomolar concentration perturbs the homeostasis of neural stem cells in highly differentiated but not in immature three-dimensional brain cell cultures.

Ochratoxin A (OTA), a fungal contaminant of basic food commodities, is known to be highly cytotoxic, but the pathways underlying adverse effects at subcytotoxic concentrations remain to be elucidated. Recent reports indicate that OTA affects cell cycle regulation. Therefore, 3D brain cell cultures were used to study OTA effects on mitotically active neural stem/progenitor cells, comparing highly differentiated cultures with their immature counterparts. Changes in the rate of DNA synthesis were related to early changes in the mRNA expression of neural stem/progenitor cell markers. OTA at 10nM, a concentration below the cytotoxic level, was ineffective in immature cultures, whereas in mature cultures it significantly decreased the rate of DNA synthesis together with the mRNA expression of key transcriptional regulators such as Sox2, Mash1, Hes5, and Gli1; the cell cycle activator cyclin D2; the phenotypic markers nestin, doublecortin, and PDGFRα. These effects were largely prevented by Sonic hedgehog (Shh) peptide (500ngml(-1)) administration, indicating that OTA impaired the Shh pathway and the Sox2 regulatory transcription factor critical for stem cell self-renewal. Similar adverse effects of OTA in vivo might perturb the regulation of stem cell proliferation in the adult brain and in other organs exhibiting homeostatic and/or regenerative cell proliferation.

Fast gas chromatography and negative-ion chemical ionization tandem mass spectrometry for forensic analysis of cannabinoids in whole blood.

The present work describes a fast gas chromatography/negative-ion chemical ionization tandem mass spectrometric assay (Fast GC/NICI-MS/MS) for analysis of tetrahydrocannabinol (THC), 11-hydroxy-tetrahydrocannabinol (THC-OH) and 11-nor-9-carboxy-tetrahydrocannabinol (THC-COOH) in whole blood. The cannabinoids were extracted from 500 microL of whole blood by a simple liquid-liquid extraction (LLE) and then derivatized by using trifluoroacetic anhydride (TFAA) and hexafluoro-2-propanol (HFIP) as fluorinated agents. Mass spectrometric detection of the analytes was performed in the selected reaction-monitoring mode on a triple quadrupole instrument after negative-ion chemical ionization. The assay was found to be linear in the concentration range of 0.5-20 ng/mL for THC and THC-OH, and of 2.5-100 ng/mL for THC-COOH. Repeatability and intermediate precision were found less than 12% for all concentrations tested. Under standard chromatographic conditions, the run cycle time would have been 15 min. By using fast conditions of separation, the assay analysis time has been reduced to 5 min, without compromising the chromatographic resolution. Finally, a simple approach for estimating the uncertainty measurement is presented.

Predictive models for nanotoxicology: current challenges and future opportunities.

Characterizing the risks posed by nanomaterials is extraordinarily complex because these materials can have a wide range of sizes, shapes, chemical compositions and surface modifications, all of which may affect toxicity. There is an urgent need for a testing strategy that can rapidly and efficiently provide a screening approach for evaluating the potential hazard of nanomaterials and inform the prioritization of additional toxicological testing where necessary. Predictive toxicity models could form an integral component of such an approach by predicting which nanomaterials, as a result of their physico-chemical characteristics, have potentially hazardous properties. Strategies for directing research towards predictive models and the ancillary benefits of such research are presented here.

Naphthylamine, 2-

2-Naphthylamine (2NA, CAS # 91-59-8) was used in as an intermediate in the dye industry and in the rubber industry. 2NA is frequently produced when nitrogenous organic materials are burned or heated. Tobacco smoke, heated cooking oil, and many other smokes contain 2NA as well as other aromatic amines. 2NA is among the most potent human urinary bladder carcinogens. It is well absorbed by all routes.

Sensitivity analysis, dominant factors, and robustness of the ECETOC TRA v3, Stoffenmanager 4.5, and ART 1.5 occupational exposure models

Occupational exposure modeling is widely used in the context of the E.U. regulation on the registration, evaluation, authorization, and restriction of chemicals (REACH). First tier tools, such as European Centre for Ecotoxicology and TOxicology of Chemicals (ECETOC) targeted risk assessment (TRA) or Stoffenmanager, are used to screen a wide range of substances. Those of concern are investigated further using second tier tools, e.g., Advanced REACH Tool (ART). Local sensitivity analysis (SA) methods are used here to determine dominant factors for three models commonly used within the REACH framework: ECETOC TRA v3, Stoffenmanager 4.5, and ART 1.5. Based on the results of the SA, the robustness of the models is assessed. For ECETOC, the process category (PROC) is the most important factor. A failure to identify the correct PROC has severe consequences for the exposure estimate. Stoffenmanager is the most balanced model and decision making uncertainties in one modifying factor are less severe in Stoffenmanager. ART requires a careful evaluation of the decisions in the source compartment since it constitutes ∼75% of the total exposure range, which corresponds to an exposure estimate of 20-22 orders of magnitude. Our results indicate that there is a trade off between accuracy and precision of the models. Previous studies suggested that ART may lead to more accurate results in well-documented exposure situations. However, the choice of the adequate model should ultimately be determined by the quality of the available exposure data: if the practitioner is uncertain concerning two or more decisions in the entry parameters, Stoffenmanager may be more robust than ART.

Assessment of the chemosensitizing activity of TAT-RasGAP317-326 in childhood cancers.

Although current anti-cancer protocols are reasonably effective, treatment-associated long-term side effects, induced by lack of specificity of the anti-cancer procedures, remain a challenging problem in pediatric oncology. TAT-RasGAP317-326 is a RasGAP-derived cell-permeable peptide that acts as a sensitizer to various anti-cancer treatments in adult tumor cells. In the present study, we assessed the effect of TAT-RasGAP317-326 in several childhood cancer cell lines. The RasGAP-derived peptide-induced cell death was analyzed in several neuroblastoma, Ewing sarcoma and leukemia cell lines (as well as in normal lymphocytes). Cell death was evaluated using flow cytometry methods in the absence or in the presence of the peptide in combination with various genotoxins used in the clinics (4-hydroperoxycyclophosphamide, etoposide, vincristine and doxorubicin). All tested pediatric tumors, in response to at least one genotoxin, were sensitized by TAT-RasGAP317-326. The RasGAP-derived peptide did not increase cell death of normal lymphocytes, alone or in combination with the majority of the tested chemotherapies. Consequently, TAT-RasGAP317-326 may benefit children with tumors by increasing the efficacy of anti-cancer therapies notably by allowing reductions in anti-cancer drug dosage and the associated drug-induced side effects.

The anticancer drug metabolites endoxifen and 4-hydroxy-tamoxifen induce toxic effects on Daphnia pulex in a two-generation study.

Although pharmaceutical metabolites are found in the aquatic environment, their toxicity on living organisms is poorly studied in general. Endoxifen and 4-hydroxy-tamoxifen (4OHTam) are two metabolites of the widely used anticancer drug tamoxifen for the prevention and treatment of breast cancers. Both metabolites have a high pharmacological potency in vertebrates, attributing prodrug characteristics to tamoxifen. Tamoxifen and its metabolites are body-excreted by patients, and the parent compound is found in sewage treatment plan effluents and natural waters. The toxicity of these potent metabolites on non-target aquatic species is unknown, which forces environmental risk assessors to predict their toxicity on aquatic species using knowledge on the parent compounds. Therefore, the aim of this study was to assess the sensitivity of two generations of the freshwater microcrustacean Daphnia pulex towards 4OHTam and endoxifen. Two chronic tests of 4OHTam and endoxifen were run in parallel and several endpoints were assessed. The results show that the metabolites 4OHTam and endoxifen induced reproductive and survival effects. For both metabolites, the sensitivity of D. pulex increased in the second generation. The intrinsic rate of natural increase (r) decreased with increasing 4OHTam and endoxifen concentrations. The No-Observed Effect Concentrations (NOECs) calculated for the reproduction of the second generation exposed to 4OHTam and endoxifen were <1.8 and 4.3μg/L, respectively, whereas the NOECs that were calculated for the intrinsic rate of natural increase were <1.8 and 0.4μg/L, respectively. Our study raises questions about prodrug and active metabolites in environmental toxicology assessments of pharmaceuticals. Our findings also emphasize the importance of performing long-term experiments and considering multi-endpoints instead of the standard reproduction outcome.

Cardioinhibitory reflex due to a karate kick: a case report.

This article describes the case of a 17-year-old adolescent boy who received a foot kick in the trunk area from an expert in karate. He presented with immediate cardiocirculatory arrest. After a prolonged resuscitation, he was transferred to a hospital where he died 5 days later without ever regaining consciousness. Postmortem investigations including autopsy, radiology, histology, toxicology, and postmortem chemistry were performed that showed signs of multiple organ failure, an acute hemorrhage in the region of the celiac plexus, and signs of medical resuscitation. No preexisting disease, particularly those concerning the heart, was objectified. The cause of death was attributed to multiple organ failure after a prolonged cardiocirculatory arrest. Concerning the origin of the cardiac arrest, 2 hypotheses were considered-a cardioinhibitory reflex and a cardiac contusion (commotio cordis). Because of the presence of traumatic lesions in the celiac plexus, the first hypothesis was finally submitted. This case is reported because rare cases of sudden death from celiac reflex are described in the literature where it is almost impossible to find references with accurate documentation. The presented case confirms the importance of detailed documentation of the circumstances and postmortem investigations to establish a diagnosis of death due to cardioinhibitory reflex.

Pathological changes in anabolic androgenic steroid users.

Several classes of recreational and prescription drugs have additional effects on the heart and vasculature, which may significantly contribute to morbidity and mortality in chronic users. The study presented herein focuses on pathological changes involving the heart possibly due to anabolic androgenic steroid use. The role these hormones may play in their occurrence of sudden cardiac death is also investigated. 98 medico-legal cases including 6 anabolic androgenic steroid users were retrospectively reviewed. Autopsies, histology, immunohistochemistry, biochemistry and toxicology were performed in all cases. Pathological changes consisted of various degrees of interstitial and perivascular fibrosis as well as fibroadipous metaplasia and perineural fibrosis within the myocardium of the left ventricle. Within the limits of the small number of investigated cases, our results appear to confirm former observations on this topic and suggest anabolic androgenic steroid's potential causative role in the pathogenesis of sudden cardiac deaths in chronic users.

Postmortem angiography using femoral cannulation and postmortem microbiology.

Despite the undeniable advantages of postmortem angiography, numerous questions have arisen concerning the influence that the injected contrast media may exercise on biological fluids and tissues collected for toxicological and biochemical investigations. Moreover, cardiac blood for microbiological investigations cannot be obtained post-angiography. In this study, we examined whether the peripheral blood collected prior to postmortem angiography, using percutaneous access to femoral vessels after skin surface disinfection, could be suitable for microbiological investigations when postmortem angiography with femoral vessel cannulation is also performed. A total of 66 cases were included in the study and were divided into two subgroups (angiography and bacteriology group, 33 cases and control group, 33 cases). Autopsies, histology, toxicology, bacteriology, and biochemical investigations (procalcitonin, C-reactive protein, interleukin-6, and soluble triggering receptors expressed on myeloid cells type 1) were performed in all cases. No statistically significant differences between the two groups were noted, and identified category distribution (death unrelated to infection, true infection, false positive, and undetermined) was rather similar in both studied populations. These preliminary results suggest that postmortem angiography using a femoral approach does not constitute an impediment to the collection of peripheral blood for microbiology and vice versa. Moreover, the use of femoral blood for microbiology does not lead to an increased risk of doubtful results.

Ewing sarcoma cancer stem cells : from mechanisms to therapeutic targeting

Le sarcome d'Ewing (SE) est la 2ème tumeur des os la plus fréquente chez les enfants, et le pronostic est sombre au stade métastatique. La pathogenèse du SE repose sur une translocation, provocant la fusion du domaine activateur du facteur de transcription EWS, avec la partie liant l'ADN de la protéine FLI-1. Les cellules souches cancéreuses (CSC) sont supposées être les moteurs de la croissance tumorale, et représente de ce fait des cibles thérapeutiques préférentielles. Dans ce travail nous nous sommes efforcés de comprendre, ainsi que de cibler les mécanismes liés à l'émergence des CSC dans le sarcome d'Ewing. La formation des CSC du ES est liée à un défaut de maturation des miRNAs provoqué par une sous-expression d'un gène, TARBP2, dans les CSC. Ce défaut de maturation peut être corrigé par un traitement des cellules avec de l'enoxacine, une fluoroquinolone utilisée pour traiter les infections urinaires. L'enoxacine seule n'étant pas suffisante pour éradiquer les tumeurs in vivo, nous avons testé la combinaison d'une thérapie ciblée sur les CSC avec une chimiothérapie classique, la doxorubicine, ciblant les cellules différentiées. In vitro l'enoxacine induit l'apoptose dans les CCS sans affecter les cellules différentiées, alors que à l'inverse, la doxorubicine n'affecte que les cellules de la « masse » tumorale. In vivo la combinaison de ces deux drogues inhibe la croissance de tumeurs provenant de cellules primaires xenotranplantées et éradique les CSCs. Nos résultats mettent en lumière une nouvelle approche thérapeutique directement applicable pour le sarcome d'Ewing, et pourraient ainsi rapidement déboucher sur des essais cliniques. Dans la deuxième partie de ce travail nous avons essayé de comprendre comment EWS-FLI1, la protéine de fusion issue de la translocation chromosomique du sarcome d'Ewing conduit à la génération des CSC. Pour cela nous avons effectué des ChIPseq (immunoprecipitation de la chromatine suivi de séquençage) pour EWS-FLI1 ainsi que pour certaines modifications histoniques. -- Ewing sarcoma family tumors (ESFT) are the second most frequent bone tumors in children and have a high rate of recurrence when metastatic at presentation. The pathogenesis of Ewing sarcoma is underlayed by a translocation, leading to the fusion of the trans-activating domain of EWS with the FLU DNA binding domain. Cancer stem cells (CSCs) are thought to be the driving force of tumor growth. In this work we focused on understanding the mechanisms underlying ESFT CSC emergence as well as defining targeted therapeutic strategies. Emergence of CSCs in ESFT has been shown to arise from a defect in TARBP2-dependent microRNA maturation, which can be corrected by exposure to the fluoroquinolone enoxacin. As enoxacin alone is not sufficient to reverse tumor growth in vivo, we assessed the effect of combining a drug that abrogates CSC properties with doxorubicin, a standard-of-care therapy in ESFT. Primary ESFT CSCs and bulk tumor cells were treated with different concentration of drugs and displayed divergent responses to doxorubicin and enoxacin. Doxorubicin, which targets the tumor bulk, displayed toxicity toward primary adherent ESFT cells in culture but not to CSC-enriched ESFT spheres. Conversely, enoxacin induced apoptosis but only in ESFT spheres and specifically on the CD133+ population. In combination, the two drugs markedly depleted CSC and strongly reduced primary growth in xenograft assays of two primary ESFT. Our results identify a potentially attractive therapeutic strategy for ESFT that combines mechanism-based targeting of CSC using a low toxicity antibiotic with a standard-of-care cytotoxic drug, offering immediate applications for clinical evaluation. In the second part of this work we performed chromatin immunopercipitation on CSCs and bulk cells for EWS-FLI1 binding as well as some chromatin modifications, and concluded that EWS-FLI1 shows cell context dependent binding.

Innate Immune Sensing of Fusarium culmorum by Mouse Dendritic Cells.

Chronic inhalation of grain dust is associated with asthma and chronic bronchitis in grain worker populations. Exposure to fungal particles was postulated to be an important etiologic agent of these pathologies. Fusarium species frequently colonize grain and straw and produce a wide array of mycotoxins that impact human health, necessitating an evaluation of risk exposure by inhalation of Fusarium and its consequences on immune responses. Data showed that Fusarium culmorum is a frequent constituent of aerosols sampled during wheat harvesting in the Vaud region of Switzerland. The aim of this study was to examine cytokine/chemokine responses and innate immune sensing of F. culmorum in bone-marrow-derived dendritic cells and macrophages. Overall, dendritic cells and macrophages responded to F. culmorum spores but not to its secreted components (i.e., mycotoxins) by releasing large amounts of macrophage inflammatory protein (MIP)-1α, MIP-1β, MIP-2, monocyte chemoattractant protein (MCP)-1, RANTES, and interleukin (IL)-12p40, intermediate amounts of tumor necrosis factor (TNF), IL-6, IL-12p70, IL-33, granulocyte colony-stimulating factor (G-CSF), and interferon gamma-induced protein (IP-10), but no detectable amounts of IL-4 and IL-10, a pattern of mediators compatible with generation of Th1 or Th17 antifungal protective immune responses rather than with Th2-dependent allergic responses. The sensing of F. culmorum spores by dendritic cells required dectin-1, the main pattern recognition receptor involved in β-glucans detection, but likely not MyD88 and TRIF-dependent Toll-like receptors. Taken together, our results indicate that F. culmorum stimulates potently innate immune cells in a dectin-1-dependent manner, suggesting that inhalation of F. culmorum from grain dust may promote immune-related airway diseases in exposed worker populations.