Intraoperative Carotid Artery Injuries. Review of the literature, analysis of the material of one centre.

<b>Introduction:</b> Injuries of the carotid artery constitute a rare group of injuries. This study presents results of the treatment of 44 patients with iatrogenic carotid artery injuries for over 20 years. The patients were treated by the team of doctors of the Department of Vascular, General and Transplant Surgery in Wroclaw in the years 1997-2017 (Head of the Department Prof. Klemens Skóra, MD, and Prof. Piotr Szyber, MD, PhD - material used with permission). </br></br> <b>Aim:</b> Aims of the analysis are: to estimate the frequency of different forms of iatrogenic injuries to the common and internal carotid artery, to evaluate the results of treatment, to assess the most effective surgical method depending on the type of injury, and develop an effective preoperative, intraoperative and postoperative regimen.</br></br> <b>Discussion and results:</b> The frequency of various carotid artery injuries (blunt, acute, traffic) was constant between years, but the number of iatrogenic injuries definitely increased over time. The prognosis for patients with carotid artery injury, especially when combined with multi-organ trauma, is the gravest. Significantly better treatment results were achieved with both acute and iatrogenic injuries. This is mainly due to easier and quicker diagnosis and better conditions for assisting patients.</br></br> <b>Conclusions:</b> In iatrogenic injuries, a well-designed surgical scheme, i.e. primarily the administration of UTH and placing a temporary flow drain by the first operating team, reduces the risk of neurological complications.

Serum very long-chain fatty acid-containing lipids predict response to immune checkpoint inhibitors in urological cancers.

Checkpoint inhibitors (CPI) have significantly changed the therapeutic landscape of oncology. We adopted a non-invasive metabolomic approach to understand immunotherapy response and failure in 28 urological cancer patients. In total, 134 metabolites were quantified in patient sera before the first, second, and third CPI doses. Modeling the association between metabolites and CPI response and patient characteristics revealed that one predictive metabolite class  (n = 9/10) were very long-chain fatty acid-containing lipids (VLCFA-containing lipids). The best predictive performance was achieved through a multivariate model, including age and a centroid of VLCFA-containing lipids prior to first immunotherapy (sensitivity: 0.850, specificity: 0.825, ROC: 0.935). We hypothesize that the association of VLCFA-containing lipids with CPI response is based on enhanced peroxisome signaling in T cells, which results in a switch to fatty acid catabolism. Beyond use as a novel predictive non-invasive biomarker, we envision that nutritional supplementation with VLCFA-containing lipids might serve as an immuno sensitizer.

A plasma metabolite panel as biomarkers for early primary breast cancer detection.

In recent years, metabolites have attracted substantial attention as promising novel biomarkers of various diseases. However, breast cancer plasma metabolite studies are still in their infancy. Here, we investigated the potential of metabolites to serve as minimally invasive, early detection markers of primary breast cancer. We profiled metabolites extracted from the plasma of primary breast cancer patients and healthy controls using tandem mass spectrometry (UHPLC-MS/MS and FIA-MS/MS). Two metabolites were found to be upregulated, while 16 metabolites were downregulated in primary breast cancer patients compared to healthy controls in both the training and validation cohorts. A panel of seven metabolites was selected by LASSO regression analysis. This panel could differentiate primary breast cancer patients from healthy controls, with an AUC of 0.87 (95% CI: 0.81 ~ 0.92) in the training cohort and an AUC of 0.80 (95% CI: 0.71 ~ 0.87) in the validation cohort. These significantly differentiated metabolites are mainly involved in the amino acid metabolism and breast cancer cell growth pathways. In conclusion, using a metabolomics approach, we identified metabolites that have potential value for development of a multimarker blood-based test to complement and improve early breast cancer detection. The panel identified herein might be part of a prescreening tool, especially for younger women or for closely observing women with certain risks, to facilitate decision making regarding which individuals should undergo further diagnostic tests. In the future, the combination of metabolites and other blood-based molecular marker sets, such as DNA methylation, microRNA, and cell-free DNA mutation markers, will be an attractive option.

Reverse Genetics for Type I Feline Coronavirus Field Isolate To Study the Molecular Pathogenesis of Feline Infectious Peritonitis.

Feline infectious peritonitis (FIP), one of the most important lethal infections of cats, is caused by feline infectious peritonitis virus (FIPV), the high-virulence biotype of feline coronaviruses (FCoVs). FIPVs are suggested to emerge from feline enteric coronaviruses (FECVs) by acquiring mutations in specific genes in the course of persistent infections. Although numerous studies identified mutations predicted to be responsible for the FECV-FIPV biotype switch, the presumed roles of specific genetic changes in FIP pathogenesis have not been confirmed experimentally. Reverse genetics systems established previously for serotype I and the less common serotype II FCoVs were based on cell culture-adapted FIPV strains which, however, were shown to be unsuitable for FIP pathogenesis studies in vivo To date, systems to produce and manipulate recombinant serotype I field viruses have not been developed, mainly because these viruses cannot be grown in vitro Here, we report the first reverse genetics system based on a serotype I FECV field isolate that is suitable to produce high-titer stocks of recombinant FECVs. We demonstrate that these recombinant viruses cause productive persistent infections in cats that are similar to what is observed in natural infections. The system provides an excellent tool for studying FCoVs that do not grow in standard cell culture systems and will greatly facilitate studies into the molecular pathogenesis of FIP. Importantly, the system could also be adapted for studies of other RNA viruses with large genomes whose production and characterization in vivo are currently hampered by the lack of in vitro propagation systems.IMPORTANCE The availability of recombinant serotype I FCoV field isolates that are amenable to genetic manipulation is key to studying the molecular pathogenesis of FIP, especially since previous studies using cell culture-adapted FIPVs had proven unsuccessful. To our knowledge, we report the first serotype I FECV field isolate-based reverse genetics system that allows the production of high-titer recombinant virus stocks that can be used for subsequent in vivo studies in cats. The system represents a milestone in FCoV research. It provides an essential tool for studying the molecular pathogenesis of FIP and, more specifically, the functions of specific gene products in causing a fundamentally different progression of disease following acquisition of specific mutations. The system developed in this study will also be useful for studying other coronaviruses or more distantly related RNA viruses with large genomes for which suitable in vitro culture systems are not available.

Reduced mRNA expression levels of NFE2L2 are associated with poor outcome in breast cancer patients.

BACKGROUND: The transcription factor nuclear factor erythroid 2-related factor 2 (NFE2L2; previously known as NRF2) is a crucial regulator of the intracellular antioxidant response. It controls the expression of genes involved in the detoxification and elimination of reactive oxidants and electrophilic agents. The role of NFE2L2 in cancer is subject of controversial discussion, as it has been reported to have both pro-and anti-tumourigenic functions. To shed some light on this paradox, we analysed the NFE2L2 mRNA expression levels in breast cancer and its association with clinicopathological features and survival. METHODS: We retrospectively evaluated the NFE2L2 mRNA expression levels in tumour tissue of two independent breast cancer patient cohorts. In the training set we analysed data from the Molecular Taxonomy of Breast Cancer International Consortium (METABRIC). In the test set we measured the NFE2L2 mRNA expression levels in 176 breast tumour tissues by quantitative real-time reverse transcription PCR (qRT-PCR). Group differences were analysed using Mann-Whitney U-test, and associations between NFE2L2 mRNA expression levels and clinicopathological features were examined by means of univariate and multivariate survival analyses. Furthermore, we compared NFE2L2 mRNA expression levels between tumour and normal breast tissue samples by means of 108 paired samples from the The Cancer Genome Atlas (TCGA) dataset. RESULTS: In the training set we identified an independent predictive value for high NFE2L2 mRNA expression levels [HRdisease specific death 0.8 (0.6-1.0), P = 0.041; HRdeath 0.8 (0.6-1.0), P = 0.023] especially in the subgroup of oestrogen receptor (ER) positive tumours [HRdisease specific death 0.6 (0.4-0.9), P = 0.008; HRdeath 0.6 (0.4-0.8), P = 0.001]. Similarly, we found this association also in the test set [HRrelapse 0.4 (0.2-0.9), P = 0.031] and again, more pronounced in patients with ER positive tumours [HRrelapse 0.2 (0.1-0.7), P = 0.012]. In addition, we observed generally lower NFE2L2 expression levels in tumour tissues than in normal breast tissues. CONCLUSION: We concluded that reduced NFE2L2 mRNA expression in tumour tissues is an independent predictor of shortened survival in breast cancer patients.

High prevalence of side population in human cancer cell lines.

Cancer cell lines are essential platforms for performing cancer research on human cells. We here demonstrate that, across tumor entities, human cancer cell lines harbor minority populations of putative stem-like cells, molecularly defined by dye extrusion resulting in the side population phenotype. These findings establish a heterogeneous nature of human cancer cell lines and argue for their stem cell origin. This should be considered when interpreting research involving these model systems.

The Phenomenon of the Urban Mummy.

The environmental mummification of a body in an urban setting is an unusual event. Florida is known for its high temperatures and humidity, which typically result in the rapid putrefaction of bodies. However, under certain circumstances, Florida's climate provides an environment suitable for mummification, typically in isolated areas. We previously reported a case of a homicide/suicide in a residential neighborhood in which the mummified remains were undetected for several years. We have subsequently encountered two additional cases in which mummified bodies were discovered in urban settings. Presented in the historical context of the environmental conditions and postmortem processes involved in mummification, these cases illustrate the particular constellation of human behaviors and scenarios, as well as environmental conditions, which must coincide for this phenomenon to occur and highlight the necessity of the multidisciplinary approach to the medicolegal investigation of such deaths and the determination of the cause and manner of death.

Phylogenetic analysis of hepatitis E virus in domestic swine and wild boar in Germany.

Hepatitis E virus (HEV) is an emerging non-enveloped positive strand RNA virus with worldwide distribution that can cause acute liver disease in humans. The virus has also been detected in both domestic and wild animals. In this study we investigated the presence of HEV in free-living wild boar as well as in domestic swine. A total of 105 domestic swine fecal samples and 124 wild boar sera were tested for the presence of HEV RNA by RT-PCR. A 241 nucleotide (nt) fragment from the capsid gene of HEV from one domestic swine and from 18 wild boars were amplified and sequenced. In addition, the complete capsid of three HEV sequences found in wild boar and the complete genomic sequence of the domestic swine HEV were obtained. Phylogenetic analyses based on both the 241 nt fragments as well as four complete capsid gene sequences demonstrated that all sequences belong to genotype HEV-3.

Mutations of 3c and spike protein genes correlate with the occurrence of feline infectious peritonitis.

The genes encoding accessory proteins 3a, 3b, 3c, 7a and 7b, the S2 domain of the spike (S) protein gene and the membrane (M) protein gene of feline infectious peritonitis virus (FIPV) and feline enteric coronavirus (FECV) samples were amplified, cloned and sequenced. For this faeces and/or ascites samples from 19 cats suffering from feline infectious peritonitis (FIP) as well as from 20 FECV-infected healthy cats were used. Sequence comparisons revealed that 3c genes of animals with FIP were heavily affected by nucleotide deletions and point mutations compared to animals infected with FECV; these alterations resulted either in early termination or destruction of the translation initiation codon. Two ascites-derived samples of cats with FIP which displayed no alterations of ORF3c harboured mutations in the S2 domain of the S protein gene which resulted in amino acid exchanges or deletions. Moreover, changes in 3c were often accompanied by mutations in S2. In contrast, in samples obtained from faeces of healthy cats, the ORF3c was never affected by such mutations. Similarly ORF3c from faecal samples of the cats with FIP was mostly intact and showed only in a few cases the same mutations found in the respective ascites samples. The genes encoding 3a, 3b, 7a and 7b displayed no mutations linked to the feline coronavirus (FCoV) biotype. The M protein gene was found to be conserved between FECV and FIPV samples. Our findings suggest that mutations of 3c and spike protein genes correlate with the occurrence of FIP.

Association of TNFRSF10D DNA-methylation with the survival of melanoma patients.

In this retrospective pilot study, the DNA-methylation status of genes that have been demonstrated to be involved in melanoma carcinogenesis was analyzed in order to identify novel biomarkers for the risk assessment of melanoma patients. We analyzed DNA extracted from punch-biopsies from 68 formalin-fixed paraffin-embedded (FFPE) melanoma specimens. Using MethyLight PCR, we examined 20 genes in specimens from a training set comprising 36 melanoma patients. Selected candidate genes were validated in a test set using FFPE tissue samples from 32 melanoma patients. First, we identified the TNFRSF10D DNA-methylation status (TNFRSF10D methylated vs. unmethylated) as a prognostic marker for overall (p = 0.001) and for relapse-free survival (p = 0.008) in the training set. This finding was confirmed in the independent test set (n = 32; overall survival p = 0.041; relapse-free survival p = 0.012). In a multivariate Cox-regression analysis including all patients, the TNFRSF10D DNA-methylation status remained as the most significant prognostic parameter for overall and relapse-free survival (relative-risk (RR) of death, 4.6 (95% CI: 2.0-11.0; p < 0.001), RR of relapse, 7.2 (95% CI: 2.8-18.3; p < 0.001)). In this study, we demonstrate that TNFRSF10D DNA-methylation analysis of a small tissue-punch from archival FFPE melanoma tissue is a promising approach to provide prognostic information in patients with melanoma.

Fatalities due to indigenous and exotic species in Florida.

Florida's climate is suitable for many potentially hazardous animals, including both indigenous and exotic species, which are frequently kept as in zoos or as pets. This has resulted in many unforeseen fatal encounters between animals and the ever expanding human population. While the literature and knowledge pool for more common types of deaths referred to medical examiner/coroner's offices is abundant, the appreciation of wildlife and exotic pet-related deaths is far less widespread. We report seven animal attack-related deaths that occurred in Florida. The inflicted injuries included blunt and sharp force injuries, asphyxia, drowning, and envenomation. The underlying cause of death, however, was always a result of the human/animal interaction and in many cases related to human error and failure to appreciate the potentially dangerous behavior of nondomesticated species. These cases illustrate the varied circumstances and pathophysiologies associated with deaths due to indigenous and exotic species and the importance of the multidisciplinary approach in the medicolegal investigation of these cases.

A reverse genetics approach to study feline infectious peritonitis.

Feline infectious peritonitis (FIP) is a lethal immunopathological disease caused by feline coronaviruses (FCoVs). Here, we describe a reverse genetics approach to study FIP by assessing the pathogenicity of recombinant type I and type II and chimeric type I/type II FCoVs. All recombinant FCoVs established productive infection in cats, and recombinant type II FCoV (strain 79-1146) induced FIP. Virus sequence analyses from FIP-diseased cats revealed that the 3c gene stop codon of strain 79-1146 has changed to restore a full-length open reading frame (ORF).

Multidisciplinary investigation of an unusual apparent homicide/suicide.

The investigation of deaths of individuals whose bodies are decomposed, mummified, or skeletonized is particularly difficult for medical examiners and medicolegal death investigators. Determination of the cause and manner of death in such cases frequently requires consultation with experts in a variety of disciplines in the forensic sciences and necessitates correlation of the autopsy results, scene investigation, medical and social history of the deceased, and laboratory studies. The authors report an unusual case of an apparent homicide/suicide involving 2 individuals and a canine that went undetected for almost 4 years. Determination of the cause and manner of death in these cases involved a multidisciplinary, intercontinental investigation and necessitated the performance of toxicologic studies on specimens not commonly analyzed. These cases illustrate the importance of the multidisciplinary approach to medicolegal death investigations and the necessity of considering nontraditional sources of potential information and specimens for laboratory analysis in selected cases.

Household furniture tip-over deaths of young children.

The potential for the injury or death of a child resulting from the tip-over of a piece of household furniture or a domestic appliance has not been previously well recognized. We reviewed nine accidental deaths of young children that resulted from avoidable residential hazards and/or lapses in supervision of the children by their caregivers. The offending household items included televisions, bedroom dressers, a kitchen stove, and a lounge chair. The causes of death were mechanical asphyxia, blunt trauma, and combined blunt head trauma and asphyxia. All of the deaths could have been prevented by appropriate anchoring of the piece of furniture and/or closer supervision of the child. A thorough multidisciplinary investigation is essential in establishing the cause and manner of death in such cases and in identifying risk factors that may aid in the prevention of future childhood deaths.

Chimeric feline coronaviruses that encode type II spike protein on type I genetic background display accelerated viral growth and altered receptor usage.

Persistent infection of domestic cats with feline coronaviruses (FCoVs) can lead to a highly lethal, immunopathological disease termed feline infectious peritonitis (FIP). Interestingly, there are two serotypes, type I and type II FCoVs, that can cause both persistent infection and FIP, even though their main determinant of host cell tropism, the spike (S) protein, is of different phylogeny and displays limited sequence identity. In cell culture, however, there are apparent differences. Type II FCoVs can be propagated to high titers by employing feline aminopeptidase N (fAPN) as a cellular receptor, whereas the propagation of type I FCoVs is usually difficult, and the involvement of fAPN as a receptor is controversial. In this study we have analyzed the phenotypes of recombinant FCoVs that are based on the genetic background of type I FCoV strain Black but encode the type II FCoV strain 79-1146 S protein. Our data demonstrate that recombinant FCoVs expressing a type II FCoV S protein acquire the ability to efficiently use fAPN for host cell entry and corroborate the notion that type I FCoVs use another main host cell receptor. We also observed that recombinant FCoVs display a large-plaque phenotype and, unexpectedly, accelerated growth kinetics indistinguishable from that of type II FCoV strain 79-1146. Thus, the main phenotypic differences for type I and type II FCoVs in cell culture, namely, the growth kinetics and the efficient usage of fAPN as a cellular receptor, can be attributed solely to the FCoV S protein.

Zoonotic aspects of infections with noroviruses and sapoviruses.

The close genetic relationship of noroviruses and sapoviruses found in animals and humans has raised the question whether these viruses have a zoonotic potential. Transmission from animals to humans and vice versa would have far-reaching consequences for epidemiology and food safety. So far animal noro- and sapoviruses have not been found in humans. However detection of human noroviruses in animals as well as simultaneous presence of animal and human viruses in bivalve molluscs suggest a risk of transmission. Furthermore, antibodies against animal noroviruses were detected in humans as well as antibodies against human noroviruses in swine. Experimental infection of gnotobiotic calves and pigs with human noroviruses demonstrated that virus replication and seroconversion can occur. Accordingly the possible role of noro- and sapoviruses as zoonotic agents needs to be further investigated.

Parasailing fatalities in southwest Florida.

Parasailing is a recreational sport that is generally considered to be of little risk to the participants. Typically, the passenger launches from a motorboat with a specially designed winch that pulls him or her back to the boat at the end of the ride. The sport is not regulated at the federal, state, or county level. There have been few reports of injuries to parasailors. Additionally, there have been only 2 fatalities reported to the United States Coast Guard in a 10-year review. We report the details of these 2 deaths, those of a mother and daughter riding in a tandem parasail, which occurred on Fort Myers Beach in 2001, as well as an additional case of a parasailing fatality that occurred in southwest Florida in 1999. These cases illustrate the injuries seen in such fatalities and the hazards posed by adverse weather conditions and faulty equipment, as well as the impairment of passenger judgment by drugs and/or alcohol.

Investigative and autopsy findings in sport aircraft-related deaths in southwest Florida.

Aircraft designated for sport or recreational use only, including ultralights, experimental aircraft and light-sport aircraft, have become increasingly popular. Because of their relative safety and the rarity of fatalities resulting from crashes of these aircraft, the forensic literature contains little information concerning the pathologic findings in such deaths. We report 9 deaths resulting from 6 sport aircraft crashes in southwest Florida, 6 pilots and 3 passengers. The vehicles involved 3 experimental aircraft, 1 ultralight and 2 "ultralight-like" aircraft. The patterns of injuries included trauma predominantly to the chest (3 cases), abdomen (1) or head (1), as well as multiple blunt force injuries involving the chest and abdomen (1) or the head and torso (3). Extremity fractures were found in only 2 cases, whereas injuries to the symphysis pubis were found in six. No "control-type" injuries were identified. These cases illustrate the varied pathologies associated with deaths due to crashes of sport aircraft and reveal the lack of uniformity associated with the investigations of such deaths.

Case report of suicide by inhalation of nitrogen gas.

Nitrogen is an inert gas that is a normal constituent of the air that we breathe. It is a suffocating gas that does not support life and that can be a cause of death by the displacement of oxygen in the atmosphere. The majority of deaths associated with nitrogen have occurred in the setting of scuba diving. Although other suffocating gases have been used as a means of committing suicide, the literature contains little information about the use of nitrogen as a suicidal agent. A case of a 50-year-old man who committed suicide using a homemade suicide device and nitrogen gas is presented.

Sudden unexpected death due to a previously undiagnosed plasma cell dyscrasia.

The plasma cell dyscrasias are a diverse group of disorders characterized by the production of a clonal paraprotein. Sudden death is a recognized complication of the plasma cell dyscrasias, most commonly in individuals with cardiac involvement by amyloidosis. However, the current forensic literature has no reported cases in which sudden death resulted from complications of a plasma cell dyscrasia that was first diagnosed by postmortem histologic examination. We present the case of a woman whose sudden and unexpected death resulted from a seizure. Postmortem examination revealed no evidence of trauma or a grossly identifiable natural disease process that would have accounted for her death. However, microscopic and immunohistologic studies revealed a previously undiagnosed plasma cell dyscrasia, the clonality of which was determined by immunohistochemical studies for immunoglobulin light chains, that was not associated with amyloid deposition. This case elucidates a previously unrecognized cause of sudden unexpected death and illustrates the importance of microscopic studies in selected cases examined in medical examiner/coroner offices.