Atrial inflammation and microvascular thrombogenicity are increased in deceased COVID-19 patients.

BACKGROUND: Histopathological studies have shown inflammation, cardiomyocyte injury, and microvascular thrombosis in the ventricular myocardium of patients with coronavirus disease 2019 (COVID-19). However, although atrial dysfunction is common in COVID-19, little is known about histopathological changes in the atria of the heart. We therefore analyzed inflammation, cardiomyocyte injury, and microvascular thrombogenicity in the atria of deceased patients with COVID-19. METHODS: Atrial tissue was obtained from autopsied COVID-19 (n=16) patients and control patients (n=10) and analyzed using immunohistochemistry. The infiltration of CD45+ leukocytes, CD3+ T lymphocytes, CD68+ macrophages, MPO+ neutrophils, and Tryptase+ mast cells were quantified as well as cardiomyocyte damage and microvascular thrombosis. In addition, Tissue Factor (TF) and Factor XII (FXII) were quantified as markers of microvascular thrombogenicity. RESULTS: The numbers of lymphocytes, macrophages, and neutrophils were significantly increased in the atrial myocardium and epicardial atrial adipose tissue of COVID-19 patients compared with the control group. This was accompanied by dispersed cardiomyocyte injury, the occasional presence of microvascular thrombosis, and an increased presence of TF and FXII in the microvascular endothelium. CONCLUSIONS: Severe COVID-19 induces inflammation, cardiomyocyte injury, and microvascular thrombosis in the atria of the heart.

The histopathological spectrum of myocardial inflammation in relation to circumstance of death: a retrospective cohort study in clinical and forensic autopsies.

Interpreting a myocardial inflammation as causal, contributory or as of no significance at all in the cause of death can be challenging, especially in cases where other pathologic and/or medico-legal findings are also present. To further evaluate the significance of myocardial inflammation as a cause of death we performed a retrospective cohort study of forensic and clinical autopsy cases. We revised the spectrum of histological inflammatory parameters in the myocardium of 79 adult autopsy cases and related these to the reported cause of death. Myocardial slides were reviewed for the distribution and intensity of inflammatory cell infiltrations, the predominant inflammatory cell type, and the presence of inflammation-associated myocyte injury, fibrosis, edema and hemorrhage. Next, the cases were divided over three groups, based on the reported cause of death. Group 1 (n = 27) consisted of all individuals with an obvious unnatural cause of death. Group 2 (n = 29) included all individuals in which myocarditis was interpreted to be one out of more possible causes of death. Group 3 (n = 23) consisted of all individuals in which myocarditis was reported to be the only significant finding at autopsy, and no other cause of death was found. Systematic application of our histological parameters showed that only a diffuse increase of inflammatory cells could discriminate between an incidental presence of inflammation (Group 1) or a potentially significant one (Groups 2 and 3). No other histological parameter showed significant differences between the groups. Our results suggest that generally used histological parameters are often insufficient to differentiate an incidental myocarditis from a (potentially) significant one.

The logic of forensic pathology opinion.

Evaluating evidence and providing opinions are at the heart of forensic science, and forensic experts are expected to provide opinions that are based on logically sound and transparent scientific reasoning, and that honour the boundaries of their area of expertise. In order to meet these objectives, many fields of science explicitly apply Bayes' theorem, which describes the logically correct way to update probabilities on the basis of observations. Making a distinction between 'investigative' and evaluative' modes of operating helps to implement the theorem into daily casework. Use of these principles promotes the logic and transparency of the reasoning that leads to expert's opinion and helps the expert to stay within her remit. Despite these important benefits, forensic pathology seems slow to adopt these principles. In this article, we explore this issue and suggest a way forward. We start with a short introduction to Bayes' theorem and its benefits, followed by a discussion of why its application is actually second nature to medical practitioners. We then discuss the difference between investigative and evaluative opinions, and how they enable the forensic pathologist to reconcile Bayes' theorem with the different phases of a forensic investigation. Throughout the text, practical examples illustrate the various ways in which the logically correct way of evidence interpretation can be implemented, and how it may help the forensic pathologist to provide an appropriate and relevant opinion.

Forensic issues arising in the assessment of chlorine-related deaths in a domestic setting.

A retrospective review of autopsy files at the Forensic Science South Australia, Australia, was undertaken over a 20-year period from January 2000 to December 2019 for all cases where chlorine had caused or contributed to death. Two cases were identified out of a total of 25,121 autopsies (0.008%): a 53-year-old man who committed suicide in a cellar with granulated chlorine, and a 49-year-old woman with asthma who died of acute bronchospasm due to exposure to chlorine gas while mixing swimming pool chemicals in her kitchen. Chlorine-related deaths are uncommon in domestic situations. However, the absence of biomarkers and non-specific findings at autopsy complicate the diagnosis, particularly as environmental levels are not stable. While accidents with cleaning agents or swimming pool reagents are the most common event in the literature in domestic settings (exclusive of industrial or transportation accidents), suicide may also very rarely occur. Individuals with asthma and chronic respiratory diseases are at higher risk of an adverse outcome upon exposure to chlorine gas, with inattention to proper storage conditions and handling protocols being additional risk factors.

Viral presence and immunopathology in patients with lethal COVID-19: a prospective autopsy cohort study.

BACKGROUND: Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) targets multiple organs and causes severe coagulopathy. Histopathological organ changes might not only be attributable to a direct virus-induced effect, but also the immune response. The aims of this study were to assess the duration of viral presence, identify the extent of inflammatory response, and investigate the underlying cause of coagulopathy. METHODS: This prospective autopsy cohort study was done at Amsterdam University Medical Centers (UMC), the Netherlands. With informed consent from relatives, full body autopsy was done on 21 patients with COVID-19 for whom autopsy was requested between March 9 and May 18, 2020. In addition to histopathological evaluation of organ damage, the presence of SARS-CoV-2 nucleocapsid protein and the composition of the immune infiltrate and thrombi were assessed, and all were linked to disease course. FINDINGS: Our cohort (n=21) included 16 (76%) men, and median age was 68 years (range 41-78). Median disease course (time from onset of symptoms to death) was 22 days (range 5-44 days). In 11 patients tested for SARS-CoV-2 tropism, SARS-CoV-2 infected cells were present in multiple organs, most abundantly in the lungs, but presence in the lungs became sporadic with increased disease course. Other SARS-CoV-2-positive organs included the upper respiratory tract, heart, kidneys, and gastrointestinal tract. In histological analyses of organs (sampled from nine to 21 patients per organ), an extensive inflammatory response was present in the lungs, heart, liver, kidneys, and brain. In the brain, extensive inflammation was seen in the olfactory bulbs and medulla oblongata. Thrombi and neutrophilic plugs were present in the lungs, heart, kidneys, liver, spleen, and brain and were most frequently observed late in the disease course (15 patients with thrombi, median disease course 22 days [5-44]; ten patients with neutrophilic plugs, 21 days [5-44]). Neutrophilic plugs were observed in two forms: solely composed of neutrophils with neutrophil extracellular traps (NETs), or as aggregates of NETs and platelets.. INTERPRETATION: In patients with lethal COVID-19, an extensive systemic inflammatory response was present, with a continued presence of neutrophils and NETs. However, SARS-CoV-2-infected cells were only sporadically present at late stages of COVID-19. This suggests a maladaptive immune response and substantiates the evidence for immunomodulation as a target in the treatment of severe COVID-19. FUNDING: Amsterdam UMC Corona Research Fund.

Fatal rooster attack.

A 76-year-old woman was attacked by a domestic rooster on her rural property while collecting eggs. The bird pecked her lower left leg causing significant hemorrhage with collapse and death. The decedent's past medical history included treated hypertension, hyperlipidemia, non-insulin dependent diabetes mellitus and varicose veins. At autopsy the major findings were limited to the lower left leg which was covered with adherent dried blood. Two small bleeding lacerations were present, one of which was located immediately over a perforated large varix. Death was therefore due to exsanguination from bleeding varicose veins following an attack by a rooster. This case demonstrates that even relatively small domestic animals may be able to inflict lethal injuries in individuals if there are specific vascular vulnerabilities present.

Post-mortem imaging compared with autopsy in trauma victims--A systematic review.

BACKGROUND: Post-mortem imaging or virtual autopsy is a rapidly advancing field of post-mortem investigations of trauma victims. In this review we evaluate the feasibility of complementation or replacement of conventional autopsy by post-mortem imaging in trauma victims. MATERIALS AND METHODS: A systematic review was performed in compliance with the PRISMA guidelines. MEDLINE, Embase and Cochrane databases were systematically searched for studies published between January 2008 and January 2014, in which post-mortem imaging was compared to conventional autopsy in trauma victims. Studies were included when two or more trauma victims were investigated. RESULTS: Twenty-six studies were included, with a total number of 563 trauma victims. Post-mortem computer tomography (PMCT) was performed in 22 studies, post-mortem magnetic resonance imaging (PMMRI) in five studies and conventional radiography in two studies. PMCT and PMMRI both demonstrate moderate to high-grade injuries and cause of death accurately. PMCT is more sensitive than conventional autopsy or PMMRI in detecting skeletal injuries. For detecting minor organ and soft tissue injuries, autopsy remains superior to imaging. Aortic injuries are missed frequently by PMCT and PMMRI and form their main limitation. CONCLUSION: PMCT should be considered as an essential supplement to conventional autopsy in trauma victims since it detects many additional injuries. Despite some major limitations, PMCT could be used as an alternative for conventional autopsy in situations where conventional autopsy is rejected or unavailable.

Analysis of morphological characteristics and expression levels of extracellular matrix proteins in skin wounds to determine wound age in living subjects in forensic medicine.

OBJECTIVE: Wound age determination in living subjects is important in routine diagnostics in forensic medicine. Macroscopical description of a wound to determine wound age however is inadequate. The aim of this study was to assess whether it would be feasible to determine wound age via analysis of morphological characteristics and extracellular matrix proteins in skin biopsies of living subjects referred to a forensic outpatient clinic. METHODS: Skin biopsies (n=101), representing the border area of the wound, were taken from skin injuries of known wound age (range: 4.5h-25 days) in living subjects. All biopsies were analyzed for 3 morphological features (ulceration, parakeratosis and hemorrhage) and 3 extracellular matrix markers (collagen III, collagen IV and α-SMA). For quantification, biopsies were subdivided in 4 different timeframes: 0.2-2 days, 2-4 days, 4-10 days and 10-25 days old wounds. Subsequently, a probability scoring system was developed. RESULTS: For hemorrhage, collagen III, collagen IV and α-SMA expression no relation with wound age was found. Ulceration was only found in wounds of 0.2-2, 2-4 and 4-10 days old, implying that the probability that a wound was more than 10 days old in case of ulceration is equal to 0%. Also parakeratosis was almost exclusively found in wounds of 0.2-2, 2-4 and 4-10 days old, except for one case with a wound age of 15 days old. The probability scoring system of all analyzed markers, as depicted above, however can be used to calculate individual wound age probabilities in biopsies of skin wounds of living subjects. CONCLUSIONS: We have developed a probability scoring system, analysing morphological characteristics and extracellular matrix proteins in superficial skin biopsies of wounds in living subjects that can be applied in forensic medicine for wound age determination.

Analysis of inflammatory cells and mediators in skin wound biopsies to determine wound age in living subjects in forensic medicine.

OBJECTIVE: In forensic medicine it is important to determine the age of skin wounds in living subjects. The aim of this study was to assess whether analysis of inflammatory cells and inflammatory mediators in skin biopsies of wounds from living subjects could improve wound age determination. METHODS: Biopsies (n=101), representing the superficial border area of a skin wound, were taken from skin injuries of known wound age (range: 4.5 hours to 25 days) of living subjects. All biopsies were analyzed for 3 inflammatory cell markers (MPO, CD45 and CD68) and 4 inflammatory mediators (MIP-1, IL-8, CML and vitronectin). For quantification, biopsies were subdivided in 4 different timeframes: 0.2-2 days, 2-4 days, 4-10 days and 10-25 days old wounds. Subsequently, a probability scoring system was developed. RESULTS: MPO, CD45, MIP-1, IL-8 (inflammatory cell markers) and N(epsilon)-(carboxymethyl)lysine (CML) positivity were maximal in wounds of 0.2-2 days old and then decreased in time. Remarkably, CD45, CD68 and CML showed a minor but non-significant increase again in 10-25 days old wounds. MPO and CD68 positivity was significantly lower in 4-25 days old wounds compared to 0.2-4 days old wounds. MPO positivity was also significantly lower in 10-25 days old wounds compared to 0.2-10 days old wounds. For CD45, MIP-1, IL-8 and CML no significant differences between the age groups were found. In case of vitronectin positivity in the extravasate or when the number of MIP-1 or IL-8-positive cells was more than 10 cells/mm(2) the probability that a wound was more than 10 days old was 0%. A probability scoring system of all analyzed markers can be used to calculate individual wound age probabilities in biopsies of skin wounds of living subjects. CONCLUSIONS: We have developed a probability scoring system of inflammatory cells and mediators that can be used to determine wound age in skin biopsies of living subjects.

A new method to determine wound age in early vital skin injuries: a probability scoring system using expression levels of Fibronectin, CD62p and Factor VIII in wound hemorrhage.

PURPOSE OF THE STUDY: In forensic autopsies it is important to determine the age of early vital skin wounds as accurate as possible. In addition to inflammation, coagulation is also induced in vital wounds. Analysis of blood coagulation markers in wound hemorrhage could therefore be an important additional discriminating factor in wound age determination. The aim of this study was to develop a wound age probability scoring system, based on the immunohistochemical expression levels of Fibronectin, CD62p and Factor VIII in wound hemorrhage. METHODS: Tissue samples of (A) non injured control skin (n=383), and samples of mechanically induced skin injuries of known wound age, (B) injuries inflicted shortly before death (up to a few minutes old) (n=382), and (C) injuries inflicted 15-30 min before death (n=42) were obtained at autopsy in order to validate wound age estimation. Tissue slides were stained for Fibronectin, CD62p and Factor VIII and were subsequently scored for staining intensity (IHC score) in wound hemorrhage (1=minor, 2=moderate, 3=strong positive). Finally, probability scores of these markers were calculated. RESULTS: In at most 14% of the non-injured control samples, hemorrhage was found, with mean±standard deviation IHC scores of 0.1±0.4, 0.2±0.4 and 0.2±0.5 for Fibronectin, CD62p, and Factor VIII, respectively. Expression of these markers significantly increased to mean IHC scores 1.4±0.8 (Fibronectin), 1.2±0.6 (CD62p), and 1.6±0.8 (Factor VIII) in wounds inflicted shortly before death (few minutes old) and to 2.6±0.5 (Fibronectin), 2.5±0.6 (CD62p), and 2.8±0.4 (Factor VIII) in 15-30 min old wounds. The probabilities that a wound was non vital in case of an IH score 0 were 87%, 88% and 90% for Fibronectin, CD62p, and Factor VIII, respectively. In case of an IHC score 1 or 2, the probabilities that a wound was a few minutes old were 82/90%, 82/83% and 72/93%. Finally, in case of an IHC score 3, the probabilities that a wound was 15-30 min old were 65%, 76% and 55%. CONCLUSIONS: Based on the expression of Fibronectin, CD62p and Factor VIII in wound hemorrhage, we developed a probability scoring system that can be used in forensic autopsies to improve wound age estimation in early skin injuries.

The value of autopsies in the era of high-tech medicine: discrepant findings persist.

AIMS: Although the autopsy is still the gold standard for quality assessment of clinical diagnoses, autopsy rates have been declining over the last decades to <10%. The aim of this study was to investigate the value of autopsies in the high-tech medicine era by determining the frequency of discrepancies between clinical and autopsy diagnoses. METHODS: We classified all adult autopsy cases (n=460), performed at Symbiant, Pathology Expert Centre, in 2007 and 2012/2013, as having major, or minor discrepancy or total concordance. The roles of possible contributory factors were analysed. Finally, we assessed the role of microscopic examination in identifying cause of death. RESULTS: Major and minor discrepancies were found in 23.5% and 32.6% of the classifiable autopsies, respectively. Most commonly observed major discrepancies were myocardial infarction, pulmonary embolism and pneumonia. Improper imaging and discontinuation of active treatment were significantly associated with a higher and a lower frequency of major discrepancies, respectively. Comparing 2007 and 2012/2013, the frequency of minor discrepancies significantly increased from 26.8% to 39.3%. Final admission length of >2 days was significantly associated with a lower frequency of class III minor discrepancies. Microscopic examination contributed to establishing cause of death in 19.6% of the cases. CONCLUSIONS: Discrepant findings persist at autopsy, even in the era of high-tech medicine. Therefore, autopsies still should serve as a very important part of quality control in clinical diagnosis and treatment. Learning from individual and system-related diagnostic errors can aid in improving patient safety.

The role of histology in forensic autopsies: is histological examination always necessary to determine a cause of death?

In England and Wales there is a conflict between the law and advice from regulatory bodies in relation to the sampling of human tissue for histological examination following medico-legal post-mortem examinations. Considering the results of previous publications, we performed a specific study to investigate the role of histology in determining the cause of death in cases at a forensic unit. A retrospective study of 500 adult forensic cases was performed. Cases were categorized by the role the histological examination played in determining a cause of death and its contributory factors. Furthermore, cause of death, manner of death, organ systems involved, and discrepancies were assessed. Of the 500 cases, histology was undertaken in 287 cases (58 %). Microscopic examination provided the cause of death in 2 % of cases where histology had been undertaken, and it added to the cause of death in 8 %. In 61 % of cases microscopy confirmed the macroscopic findings, and in 30 % it did not influence the medical cause of death. Histological examination of all organs in all forensic cases for the purpose of providing a medical cause of death is not supported. Practice guidance should be adjusted to reflect that, while histological examination is essential in certain circumstances, the decision to retain material for histology should be made on a case by case basis at the pathologist's discretion.