Usefulness of Heart-Type Fatty Acid-Binding Protein Measurement in Postmortem Urine Specimens.

ABSTRACT: Heart-type fatty acid-binding protein (HFABP) is a 15-kDa substance reported to pass through the renal tubules and be renally excreted. Therefore, it is possible that its concentration in the urine collected postmortem may reflect antemortem blood levels. We measured the postmortem urine concentration of HFABP in 94 forensic autopsy cases and compared it between acute myocardial infarction (AMI), sepsis, heat stroke cases, and asphyxia cases as control cases to examine its diagnostic validity. Kidney tissue collected at autopsy was immunostained with antibodies against HFABP to evaluate the correlation with the urinary measurements. Urinary HFABP was significantly higher in AMI, sepsis, and heat stroke cases than in asphyxia cases. Quantitative immunostaining results showed no significant differences between any 2 groups. The usefulness of kidney immunostaining for HFABP in elucidating the cause of death was low. Two reasons may explain the lack of significant differences in kidney immunostaining: nonspecific leakage of tubular epithelial HFABP into the tubules because of postmortem changes and oliguria due to dehydration caused by heat stroke. In conclusion, the measurement of urinary HFABP may be useful in elucidating the cause of death; however, the kidney HFABP immunostaining was not significantly different from AMI.

Ruptured Aortic Valve Aneurysm Caused by Infective Endocarditis in a SARS-CoV-2-Positive Autopsy Case.

ABSTRACT: Aortic valve aneurysm, an extremely rare complication secondary to infective endocarditis (IE), may cause heart failure due to rupture of the aneurysm. Coronavirus disease 2019 (COVID-19) has been reported to cause cardiovascular complications and alter susceptibility to secondary infections such as IE. Herein, we report a case of IE with a fatal outcome caused by rupture of an aortic valve aneurysm in a severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2)-positive autopsy case. The patient was a 67-year-old male diagnosed with heart failure after presenting with edema and weight gain. He was found dead in bed 2 weeks after initial symptom presentation. Autopsy revealed an aneurysmal sac in the center of the noncoronary cusp of the aortic valve with an opening of approximately 1 cm in the center of the aneurysmal wall. Histologically, aortic valve vegetation, destruction of the aortic valve cusp, rupture of the aneurysmal wall, and an abscess under the aortic intima were observed. Gram staining of the aneurysmal wall showed a gram-positive coccus. The reverse transcription quantitative polymerase chain reaction assay was positive for SARS-CoV-2. Because no defined risk factors for IE other than SARS-CoV-2 infection were observed, the association between IE and COVID-19 was highly likely.

Prominent black esophagus, Wischnewsky spots, and black duodenum in a fatal hypothermia case with an underlying diabetic ketoacidosis.

Hypothermia can occur in patients with diabetic ketoacidosis (DKA), and these two conditions can exacerbate each other. Fatal hypothermia and DKA have overlapping features and findings such as Wischnewsky spots (WS), black esophagus, basal subnuclear vacuolization in the renal tubule, dehydration, and increased acetone levels. Therefore, it may be challenging to differentiate or clarify the context of these two conditions. Herein, we report a case of a 49-year-old man with type 1 diabetes who was found lying in his house in mid-winter. He experienced cardiopulmonary arrest 10 h after the initial discovery and died at the hospital. On autopsy, florid left cardiac blood was observed. Black discoloration of the distal part of the esophageal mucosa, widespread WS in the gastric mucosa, and black discoloration of the duodenal mucosa were observed. Histologically, neutrophil infiltration in the esophageal mucosa, neutrophil infiltration and bleeding in the gastric mucosa, basal subnuclear vacuolization and Armanni-Ebstein lesion in the renal tubule epithelium in the kidney, and hyalinization of the islets of Langerhans were observed in the pancreas. Blood acetone and ß-hydroxybutyrate levels were 538 µg/mL and 8947 µmol/L, respectively. Glycated hemoglobin A1c and glucose levels were 16.2% and 883 mg/dL, respectively, while C-reactive protein level was 3.64 mg/dL. In conclusion, obnubilation due to DKA was assumed to be the underlying cause of hypothermia, and the combination of these two conditions led to the outcome of death. The concurrent presence of these conditions likely contributed to the conspicuous mucosal findings in the upper gastrointestinal tract.

Polymorphism and haplotype analysis of three novel short tandem repeat loci in the p11.4 region of human X chromosome.

X-chromosomal short tandem repeats (X-STRs) are useful for the identification of absent single parents and complex blood relations. In the present study, we aimed to identify novel STR loci for use as DNA markers by conducting polymorphism and haplotype analyses. We detected three novel STR loci (LC552061, LC552062, and LC552063, with repetitive structures of (GGAA)n(GGGA)m, (CCTT)n(CCCT)m, and (ATTT)n, respectively) in the p11.4 region of the X chromosome. For these X-STRs, the polymorphism information content values ranged from 0.5766 to 0.6377 and the power of discrimination in males and females ranged from 0.6269 to 0.6844 and from 0.8105 to 0.8537, respectively. The linkage disequilibrium analysis revealed p values of < 0.0001, < 0.0001, and 0.00909 between LC552061 and LC552062, LC552061 and LC552063, and LC552062 and LC552063, respectively. Additional linkage disequilibrium analysis including seven previously analyzed loci (LC149476, LC149479, LC149480, LC149484, LC317283, LC317284, and LC317285) revealed a p value of < 0.001 among each of the five loci (LC149476, LC149479, LC149480, LC149484, and LC317283) and between LC317284 and LC317285, indicating that they were a linked group. These results indicate that, in addition to the seven previously detected loci, the three novel X-STR loci identified in the present study might be useful DNA markers for complex kinship analysis and might support the Investigator® Argus X-12 kit.

Accuracy of Urea Nitrogen and Creatinine Measurements in Postmortem Serum and Pericardial Fluid Compared With Antemortem Data.

ABSTRACT: Although several studies have measured urea nitrogen (UN) and creatinine (Cr) concentrations in postmortem serum and pericardial fluid, no recent antemortem biochemical data have been available for forensic autopsy, thereby making the evaluation of the accuracy of postmortem data difficult. This study compared antemortem (from emergency room results before the declaration of death) and postmortem serum UN and Cr concentrations, as well as postmortem serum and pericardial fluid values, in 51 forensic autopsy cases (postmortem interval within 87 hours). Postmortem UN concentrations were strongly correlated with antemortem data. Moreover, no significant difference between pericardial fluid UN concentrations and antemortem data was observed. Postmortem serum and pericardial fluid Cr values were also correlated with antemortem data, although postmortem values were significantly higher than antemortem ones. Given our observation of early postmortem elevation in Cr concentrations, such an elevation was attributed to rigor mortis. In conclusion, the current study demonstrated the utility of postmortem UN and Cr concentrations, in particular of those measured in the pericardial fluid.

Comprehensive Severe Acute Respiratory Syndrome Coronavirus 2 Detection Using Polymerase Chain Reaction and Rapid Antigen Testing in Postmortem Specimens.

ABSTRACT: Polymerase chain reaction (PCR) is indispensable for diagnosing coronavirus disease 2019 (COVID-19) in autopsy cases. In this study, we performed comprehensive reverse transcription quantitative PCR (RT-qPCR) and rapid antigen tests for COVID-19 on forensic postmortem specimens, regardless of the antemortem symptoms and causes of death. Immediately before forensic external examination and autopsy, a wiping solution was collected from the nasopharynx with a dry swab, and rapid antigen testing and RT-qPCR were performed. Severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) was detected by RT-qPCR in 12 of the 487 cases; the infection rate was 2.46%. Of the RT-qPCR-positive cases, 7 were associated with COVID-19-related deaths. Cycle threshold values were not correlated with the cause of death or postmortem time. The sensitivity and specificity of the rapid antigen test were 91.67% and 100.00%, respectively. The RT-qPCR positivity rate of forensic cases was higher than the cumulative infection rate for the entire population. SARS-CoV-2 could be detected with the rapid antigen test and RT-qPCR within 216 hours of death. Because the rapid antigen test showed the same sensitivity and specificity as those observed in clinical practice, the test combined with RT-qPCR may be useful for diagnosing COVID-19 even in postmortem specimens.

Effectiveness of Rapid Antigen Testing in Forensic Cases of Severe Acute Respiratory Syndrome Coronavirus 2 Infection, Including Delta Variant.

ABSTRACT: The polymerase chain reaction is indispensable for diagnosing severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) infection in forensic cases. However, studies regarding the effectiveness of rapid antigen testing (RAT) in forensic cases remain limited. Therefore, we investigated the efficacy of RAT compared with reverse-transcription quantitative polymerase chain reaction (RT-qPCR) for confirming SARS-CoV-2 infection (including the delta variant). Before the external examination or autopsy, we collected samples from the nasopharyngeal mucosa, which were then assessed via RAT (QuickNavi COVID-19 Ag kit, QuickNavi-Flu+COVID-19 Ag kit) and RT-qPCR. Reverse-transcription quantitative polymerase chain reaction results were positive in 73 of 1255 cases, and 21 cases were identified as those of delta variants. Low RT-qPCR threshold cycle value cases and delta variant infections were more likely to result in coronavirus disease-related deaths. The sensitivity of the QuickNavi COVID-19 Ag kit was 76.32%, and that of the QuickNavi-Flu+COVID-19 Ag kit was 77.14%. The specificity of both RATs was 100%. In QuickNavi COVID-19 Ag kit cases, delta variant cases showed lower sensitivity than non-delta variant cases, even for a similar viral load. Thus, RAT in forensic cases is sufficiently useful as a screening test for SARS-CoV-2 infection. However, RAT carries a risk of false negatives, especially for delta variant cases.

Cerebral venous sinus thrombosis associated with COVID-19: an autopsy case report.

Coronavirus disease 2019 (COVID-19) is an infectious disease caused by the SARS-CoV-2 virus. COVID-19 has been reported to increase the propensity for systemic hypercoagulability and thromboembolism disorders such as cerebral venous sinus thrombosis (CVST). A 66-year-old woman was found dead at her home. She had symptoms of fever, dizziness, and malaise 2 weeks prior to her death. However, her fever declined 3 days before death. Postmortem computed tomography conducted before the autopsy suggested CVST. On autopsy, a massive thrombus was observed from the cortical veins to the superior sagittal sinus and transverse sinus accompanied by a small infarction region in the left parietal region. Although the rapid antigen test was negative, the reverse transcription-quantitative polymerase chain reaction test was positive for SARS-CoV-2, with a cycle threshold (Ct) value of 38.9. The serum C-reactive protein level was 0.532 mg/dL. COVID-19 was the only risk factor for CVST, and no other cause of death was determined. Therefore, the cause of death was determined as acute intracranial hypertension due to CVST associated with COVID-19. The patient died after the symptoms improved, the Ct value of RT-qPCR was 38.9, and the serum C-reactive protein level decreased. Therefore, CVST might have occurred in the convalescent phase of COVID-19 infection.

Relationship between N-terminal pro-brain natriuretic peptide concentration and heart-type fatty acid-binding protein in postmortem urine.

The clinical use of N-terminal pro-brain natriuretic peptide (NT-proBNP) and blood concentrations of heart-type fatty acid-binding protein (HFABP) is well-established in diagnosing heart conditions. However, their applicability in forensics is controversial due to postmortem changes. NT-proBNP and HFABP are excreted in the urine due to their small molecular weights and may be found in postmortem urine samples; however, their correlation has not been evaluated. In this study, we compared the concentrations of urinary NT-proBNP and HFABP in 386 forensic autopsy cases. The urinary NT-proBNP levels were significantly higher in acute myocardial infarction (AMI), congestive heart failure (CHF), sepsis, and hyperthermia cases, with the highest levels in CHF cases. Similarly, HFABP concentration was significantly higher in CHF, sepsis, and hyperthermia cases, with the highest level observed in hyperthermia cases. However, the difference in urinary HFABP levels between the AMI and control cases was not significant. Our analysis revealed a correlation between postmortem urine NT-proBNP and HFABP levels, and the NT-proBNP/HFABP ratio was high in patients with CHF and sepsis cases and low in those with hyperthermia. The difference between the ratios was possibly due to the combined release of ventricular myocardial cells in response to ventricular wall stress and myocardial injury for NT-proBNP, as well as myocardial and skeletal muscle injuries for HFABP. This study, for the first time, demonstrates the utility of postmortem measurements of urinary NT-proBNP and HFABP levels, offering valuable insights for improving the accuracy of postmortem diagnosis in forensic medicine.

Postmortem pericardial fluid sLOX-1 levels and LOX-1 immunostaining in forensic specimens: Relation to cause of death.

Lectin-like oxidized LDL receptor-1 (LOX-1) is the endothelial receptor for oxidized LDL. This receptor's extracellular domain is released into the blood as soluble LOX-1 (sLOX-1) and has been linked to ischemic heart disease (IHD), cerebrovascular diseases (CVDs), obesity, and diabetes. We recently reported that sLOX-1 fluid levels in postmortem pericardial fluid were comparable to clinical values in live patients and that significant increases in sLOX-1 were observed in patients with IHD. However, postmortem serum and urine sLOX-1 levels were higher than serum levels in living patients. Here, we conducted LOX-1 immunostaining in forensic specimens (aorta and heart) and evaluated pericardial fluid sLOX-1 in 221 medicolegal autopsy cases (67 IHD, 11 CVD, 17 inflammatory diseases, and 126 control cases) with a postmortem interval < 72 h to assess the diagnostic efficiency of postmortem pericardial fluid sLOX-1. Furthermore, we evaluated the relationships between pericardial fluid sLOX-1 and body mass index (BMI), blood HbA1c, serum C-reactive protein (CRP), high-density lipoprotein cholesterol (HDL-C), and low-density-lipoprotein cholesterol (LDL-C). LOX-1 immunostaining positivity was found in the aortic intima. Pericardial fluid sLOX-1 levels were considerably higher in patients with IHD and CVD. However, there were no significant differences in patients with inflammatory diseases and controls. No associations between pericardial fluid sLOX-1 and BMI, HbA1c, CRP, HDL-C, or LDL-C were found. These results indicate sLOX-1 utility in the postmortem diagnosis of IHD and CVD.

Fatal intoxication due to transrectal methamphetamine overdose: A case report.

Methamphetamine (MA) abuse is common in Asian countries. The major ways of abuse include intravenous injection, absorption, and ingestion. Although two cases of survival after transrectal MA administration have been reported in the clinical field, to the best of our knowledge, there is no report of death due to intentional transrectal MA overdose. This is the first report of such a death. A single, 42-year-old male with a history of habitual MA use was found dead at his home. The police found numerous unused injectors in his closet. A rapid test of his urine was positive for MA. The cause of death was unclear, so a medicolegal autopsy was performed. RESULTS: Autopsy findings revealed a tubular plastic container without a lid in the rectum at the time of excision, with a small, open plastic bag inside. MA was detected in both the plastic container and the plastic bag. The MA concentration in the femoral vein blood was enough to cause death, and the cause of death was considered transrectal MA overdose. MA was absorbed through the lower part of the rectum, so the absorbed MA bypassed the liver and was transported directly to the systemic circulation. Since MA is largely metabolized in the liver, the absorbed MA was unaffected by the hepatic first-pass effect and may have caused more rapid and serious intoxication.

Postmortem urine concentration of N-terminal pro-brain natriuretic peptide in relation to the cause of death.

The utility of biochemical marker analysis in forensic autopsy cases is still uncertain due to the postmortem changes which they undergo. Thus, research is required to elucidate alternative samples and biochemical markers which are less affected by postmortem changes. Levels of N-terminal pro-brain natriuretic peptide (NT-proBNP) are known to be elevated in congestive heart failure (CHF), acute myocardial infarction (AMI), and sepsis patients. Although NT-proBNP is reportedly excreted into the urine, no study has previously evaluated the diagnostic efficacy of urinary concentrations in a forensic setting. The aim of this study was to evaluate the diagnostic efficacy of NT-proBNP concentration in urine obtained postmortem in a series of forensic autopsy cases. METHODS: Urinary NT-proBNP was measured in 36 AMI, 10 CHF, and 19 sepsis cases, and in 124 control cases (all with postmortem interval [PMI]<72h). RESULTS: Urinary NT-proBNP was significantly higher in AMI, CHF, and sepsis cases than in control cases. Cut-off values for diagnosing AMI, CHF, and sepsis-related fatalities were 98 (sensitivity, 55.6 %; specificity, 73.4 %), 1050 (sensitivity, 80.0 %; specificity, 94.4 %), and 363pg/mL (sensitivity, 84.2 %; specificity, 85.5 %), respectively. Furthermore, we subdivided the control cases according to the death process as either acute death (87 cases) or prolonged death cases (37 cases). Although urine NT-proBNP of CHF and sepsis cases were significantly higher compared with both cases, the concentration in the AMI cases were significantly high only when compared with the acute death cases. CONCLUSION: This study is the first to elucidate the diagnostic utility of NT-proBNP measurement in urine obtained postmortem in a series of causes of death. This study suggests the diagnostic efficacy for AMI, CHF, and sepsis-related fatality in cases in which the PMI was within 72h.

Comparison of C-reactive protein levels between antemortem serum and postmortem serum and pericardial fluid.

Biochemical markers undergo postmortem changes that complicate diagnostic measurement. C-reactive protein (CRP) is one marker that is known to be useful in postmortem specimens, with high levels reported in forensic cases of sepsis, trauma, and ketoacidosis. In the present study, we included 30 cases (17 males and 13 females) that underwent forensic autopsy within 80 h of death and had a CRP result from two postmortem specimens (serum from cardiac blood and pericardial fluid) and an emergency room specimen. Antemortem results were taken at a time near to cardiopulmonary arrest and the declaration of death. CRP levels in postmortem serum and pericardial fluid correlated with those in antemortem serum. Although no significant difference was observed between the antemortem and postmortem serum levels, the pericardial level was significantly low and five false negatives were observed. We conclude that postmortem serum is suitable for use in CRP measurement, and in cases with high antemortem CRP levels, postmortem pericardial fluid may be an appropriate alternative.

Utility of soluble lectin-like oxidized low-density lipoprotein receptor-1 (sLOX-1) in the postmortem diagnosis of ischemic heart disease.

PURPOSE: Ischemic heart disease (IHD) is a major cause of death in developed countries. Postmortem IHD diagnosis using biochemical markers is difficult because of the postmortem changes. In the present study, we investigated the utility of soluble lectin-like low-density lipoprotein receptor-1 (sLOX-1) in body fluids obtained from forensic autopsy cases. METHODS: We measured pericardial fluid, urine, and serum sLOX-1 levels; these samples were obtained from medicolegal autopsy cases (n = 149, postmortem interval <72 h), and the utility of these biomarkers postmortem acute IHD diagnosis was evaluated. RESULTS: The pericardial fluid and urine of patients with acute IHD had higher sLOX-1 levels (p < .05) compared to the controls. No significant differences were found between the sLOX-1 level and the degree of coronary atherosclerosis, body mass index, and postmortem interval. CONCLUSION: sLOX-1 levels in pericardial fluid and urine samples obtained postmortem are useful markers of acute IHD.

Development of a dual test procedure for DNA typing and methamphetamine detection using a trace amount of stimulant-containing blood.

Investigation of drug-related crimes, such as violation of the Stimulant Drug Control Law, requires identifying the used drug (mainly stimulant drugs, methamphetamine hydrochloride) from a drug solution and the DNA type of the drug user from a trace of blood left in the syringe used to inject the drug. In current standard test procedures, DNA typing and methamphetamine detection are performed as independent tests that use two separate portions of a precious sample. The sample can be entirely used up by either analysis. Therefore, we developed a new procedure involving partial lysis of a stimulant-containing blood sample followed by separation of the lysate into a precipitate for DNA typing and a liquid-phase fraction for methamphetamine detection. The method enables these two tests to be run in parallel using a single portion of sample. Samples were prepared by adding methamphetamine hydrochloride water solution to blood. Samples were lysed with Proteinase K in PBS at 56°C for 20min, cooled at -20°C after adding methanol, and then centrifuged at 15,000rpm. Based on the biopolymer-precipitating ability of alcohol, the precipitate was used for DNA typing and the liquid-phase fraction for methamphetamine detection. For DNA typing, the precipitate was dissolved and DNA was extracted, quantified, and subjected to STR analysis using the AmpFℓSTR® Identifiler® Plus PCR Amplification Kit. For methamphetamine detection, the liquid-phase fraction was evaporated with N2 gas after adding 20µL acetic acid and passed through an extraction column; the substances captured in the column were eluted with a solvent, derivatized, and quantitatively detected using gas chromatograph/mass spectrometry. This method was simple and could be completed in approximately 2h. Both DNA typing and methamphetamine detection were possible, which suggests that this method may be valuable for use in criminal investigations.

Infant death associated with maternal methamphetamine use during pregnancy and delivery: A case report.

The case described in this report is of a male infant who was found dead in a closet. His mother delivered the infant in the kitchen, left him wrapped in a towel, and called emergency medical services 4days after the delivery. At the autopsy, the growth suggests a full-term delivery, significant pathological findings were not observed, and the infant was estimated to be stillborn. After the autopsy, the police investigation discovered that the mother used a stimulant during the pregnancy and shortly before the rupture of the membrane. Toxicological analysis showed 1.60mg/L of methamphetamine in the blood, strongly suggesting that the fetal death was associated with this acute intoxication. Thus far, only a few cases of infant deaths have been reported in association with methamphetamine intoxication. The present case showed the highest blood concentration of methamphetamine compared to the past infant cases with this intoxication.