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Sudden death is one of the major concerns in forensic medicine. Especially when the deceased is a young subject without significant history, the case will be of major interest to the authorities. Sudden unexplained cardiac death has been known as "Pokkuri Death Syndrome" (PDS) in Japan, "Lai Tai" in Thailand, "Bangungut" in the Philippines, "Dream Disease" in Hawaii, and "Sudden Unexpected Nocturnal Death Syndrome" among South Asian immigrants in the USA. However, the clinical and pathological features of these sudden death cases, especially the characteristics of no coronary atherosclerosis, are surprisingly similar and mainly occur among Southeast Asian young males during sleep in the midnight. In this manuscript, we have reviewed the pathological characteristics and the possible mechanism of death in PDS cases, which were associated with significantly elevated remnant lipoproteins in plasma as revealed from our studies during the past 15 years in Japan. Although elevated plasma remnant lipoproteins have been known to be strongly atherogenic, coronary atherosclerosis was not observed in PDS cases. PDS cases were shown to be an interesting cardiovascular disease death discovered in forensic medicine research, which may suggest the difference between the occurrence of cardiovascular events and the severity of coronary atherosclerosis as separate factors. These observations in PDS cases suggest the possibility that the intervention could be more targeted to suppress the cardiovascular events rather than to slow down the progression of atherosclerosis, which is now most extensively targeted for the therapy of cardiovascular disease in Western countries.

Background ApoB-48 is a major apolipoprotein secreted by the small intestine and is the main constitutive apolipoprotein in chylomicrons (CM). In the past, presence of apoB-48 in human aortic atherosclerotic plaques has not been detected. Methods A newly developed apoB-48 ELISA together with an HPLC fractionation technique, were applied to investigate the presence of apoB-48 (CM) in aortic atherosclerotic plaques. The atherosclerotic plaques were obtained from aortae of sudden cardiac death cases. Total cholesterol, triglycerides (TG), apoB-100 and apoB-48 were measured in the aortic plaques extracts. Results HPLC analysis of plaques extracts monitored by cholesterol revealed mainly particle sizes of CM and very low density lipoproteins (VLDL) in the d > 1.006 fractions. The plaques extracts were monitored by apoB-48 and apoB-100 ELISA. There were no TG peaks in any lipoprotein fraction extracted from the plaques except as free glycerol. ApoB-100 was detected in VLDL particles and in LDL sizes. In contrast, apoB-48 was detected in particles of CM, VLDL and LDL sizes. Further, in postmortem plasma, apo B-48 was detected in particles sizes of HDL or smaller and the Western blot analysis could not show any 250 kDa molecular weight (MW) protein in the plaque extracts, but smaller and broader MW staining were observed at 20–150 kDa. Conclusion Hitherto there has been lack of an appropriate assay to measure apoB-48 in plaques. Our investigations show that apoB-48 is present in atherosclerotic plaques with denatured or degraded structure. This is the first report describing presence of apoB-48 in human atherosclerotic plaques.

We have previously reported that plasma levels of remnant-like lipoprotein particles (RLP) significantly increased in sudden cardiac death cases with and without coronary atherosclerosis. In this study we have elucidated the major subset of proatherogenic RLP, containing both apoB-48 and apoB-100-carrying remnants, in plasma of SCD and control death cases. One hundred and sixty seven Japanese cases of sudden cardiac death and 78 cases of control death underwent autopsy within 12h after death were studied. Heart weight was 9.2% higher in SCD cases than controls (P

This paper presents a simple and sensitive chromatographic procedure for the simultaneous determination and quantification of ephedrines, amphetamines, cocaine, cocaine metabolites, and opiates in human urine by gas chromatography-mass spectrometry. This method involves enzyme hydrolysis in the presence of a deuterated internal standard, liquid-liquid extraction, and derivatization with pentafluoropropionic anhydride and pentafluoropropanol. The recovery of each compound averaged at 65.8% or more. The limits of detection determined for each compound by using a 2-mL sample volume ranged from 5 to 50 ng/mL. The calibration curves were linear to 100 ng/mL for all compounds when determined using methamphetamine-d4 and MDMA-d5 as internal standards. This method was successfully applied for the analysis of urine samples suspected to contain intoxicants such as methamphetamine and heroin.

We report an autopsy case of sudden death in a 36-year-old craftsman with idiopathic scoliosis. The doctor identified his scoliosis at the age of thirteen, and he was under medical care for three years until he stopped consulting the doctor. He collapsed while walking at the station and was sent to an emergency room in cardiopulmonary arrest state, where he was declared dead in spite of more than an hour of CPR. Numbers of petechiae were seen on the bilateral palpebral conjunctivae and the lips were cyanotic. There were no particular injuries except for small abrasions observed on the face. The back showed right rib hump owing to midthoracic scoliosis (with 73 degrees of Cobb's angle) and right hemithorax was deformed showing an appearance of pectus excavatum in the front. The volume of the right thoracic cavity was significantly decreased. In the right lung, there was extensive stromal fibrosis, leaving almost no normal alveolar structures, and medial hypertrophy of pulmonary arteriolar walls. Hypertrophy of the right heart ventricle due to these pulmonary changes and the congestion of other organs suggested that the cause of death in this case was cor pulmonale due to pulmonary hypertension. This was a rare case of fatal outcome of advanced idiopathic scoliosis without medical care in spite of early detection through mass screening.

Thinner is mainly composed of toluene and xylenes, and we studied the incorporation of the main metabolites of toluene and xylenes, hippuric acid (HA) and o-, m-, and p-methyl hippuric acids (o-, m-, p-MHA), in dark agouti rats' hair. Rat black hair was shaved before any exposure with an electric shaver designed for animals. Studies were performed in vivo with exposures of 30 min per day at three different concentrations (100, 300, and 1000 ppm) of toluene and o-, m-, and p-xylene for a total of 10 times over 2 weeks. Newly grown hair was tweezed out from the root with tweezers at seventh of the last exposure. Hair samples were then washed, extracted, derivatized, and analyzed by gas chromatography-mass spectrometry (GC-MS). HA and o-, m-, and p-MHA were not detected (ND) in the unexposed rat hair. After exposure, the metabolite concentration in the hair changed depending on the exposure concentration. Mean concentrations ranged from ND to 7.6 ng/mg, from ND to 13.8 ng/mg, from ND to 10.1 ng/mg, and from ND to 9.2 ng/ml hair for HA, o-, m-, and p-MHA, respectively. These results indicate that the metabolites concentrations in hair are effective indices of thinner exposure.

The ever-increasing presence of environmental toxicants and their disruptive effect on the reproductive systems of wildlife raises the question of possible damage to the human reproductive system. Using medicolegal data from over 20,000 Japanese men subjected to necropsy from 1948 to 1998, we investigated temporal changes in testis weight to find possible evidence of male reproductive disorders. We also carried out a histological examination of 747 testes collected from 1978 to 1998. Our detailed analyses of the development of testis weight over the past 50 years have revealed four clear phenomena: (i) the age at which testis weight reaches its maximum has decreased; (ii) peak weight showed a general increase until it started to decline in boys born after 1960; (iii) the decline-rate at which testis weight decreases after its peak has greatly accelerated; and (iv) the onset of increasing testis weight of boys has occurred at a progressively younger age. Our quantitative analyses of testis weight indicate the possibility of a subtle reproductive disorder in Japanese men, especially in those born after 1960. Together with the accelerated development and decline in testis weight during the past 50 years, the decline in peak weight might be indicative of a subtle interference of environmental toxicants with male reproductive organogenesis.

The effects of baclofen and pancuronium bromide on evoked electromyogram (EMG), cortical electroencephalogram (EEG) and auditory brainstem responses (ABR) were studied in pentobarbital anesthetized normal rabbits. Evoked EMG was measured in the gastrocnemius muscle by electrical stimulation of the sciatic nerve. Intravenous injection of baclofen decreased EEG and arterial blood pressure and light reflex, however, it had no significant influence on EMG or ABR at doses of 10 and 20 mg/kg/h. Pancuronium bromide immediately inhibited respiration, decreased EEG and EMG, however, it had no significant influence on arterial blood pressure, ABR, or light reflex, at doses of 0.4 and 1.0 mg/kg/h in anesthetized rabbits. ABR waves were observed until just before cardiac arrest with both of the muscle relaxants. It is suggested that ABR are not influenced by central or peripheral muscle relaxants, or by pentobarbital.

Summary: Gc globulin, also called vitamin D-binding protein, is a human 56 kD plasma protein. Antigenic epitopes of the protein recognized by 12 different monoclonal antibodies (MoAbs), including the previously prepared MoAb El2 of which binding to Gc is inhibited by actin, were analyzed in relation to the functional domains of the protein. To map the epitopes recognized by the respective antibody, the reactivities of the MoAbs were tested by immunoblotting against the recombinant Gc fragments expressed in E. coli cells, and then a competitive ELISA was performed. The results showed that the antibodies were classified into at least six groups. Furthermore, in addition to El2, actin inhibited the reactivity of MoAb 21, of which the epitope was located within residues Asp32O to Glu 379, in an ELISA in the presence of the ligand. This result must indicate that this antibody-binding site is near the site that interacts with actin. In contrast, no inhibitory effect by 25-hydroxyvitamin D to any antibodies was observed. Furthermore, all antibodies, including El2 and 21, reacted with membrane-bound Gc of lymphocytes by an immunofluorescence study, suggesting that Gc is unlikely to bind to the plasma membrane in its interaction with actin.

A polyclonal bispecific (bifunctional) antibody was prepared to develop a hemagglutination immunoassay for beta-microseminoprotein (beta-MSP), a predominant seminal protein. Three types of F(ab')2 fragments of rabbit IgG, affinity-purified anti-human red blood cell (RBC) F(ab')2 nonaffinity-purified anti-beta-MSP F(ab')2 and nonspecific (nonimmunized) F(ab')2, were mixed to obtain a F(ab')2 mixture containing 10% anti-RBC molecules and 10% anti-beta-MSP molecules. Fab' was obtained from the F(ab')2 mixture, and then reacted with maleimide-activated bovine serum albumin (BSA) at a molar ratio of 10:1. As estimated by the decrease in the maleimide content, approximately 7 Fab' molecules were introduced per one BSA molecule. The bispecific (anti-beta-MSP and anti-RBC) Fab'-BSA conjugate thus prepared was incubated successively with a human RBC suspension and with samples. In the presence of beta-MSP, RBCs become agglutinated, providing a test simple for forensic semen identification.

Methemoglobin formation caused by a liniment solution containing sodium nitrite (30 g/L and 140 g/L) was studied in rats with normal or abraded skin, by measuring the methemoglobin concentration before and after application of liniment solutions with differing nitrite concentration.Each liniment solution (120 microL) was applied. Methemoglobin was measured for 180 minutes using a hemoximeter. Simultaneously, arterial blood pressure and cutaneous blood flow was measured by laser Doppler flowmetry and a pressure transducer.After the application of each liniment solution to normal skin, the methemoglobin concentration was not significantly modified depending on the time after application. Application of liniment solution to abraded skin (140 g/L) resulted in a marked increase in methemoglobin concentration. A remarkable decrease in arterial blood pressure and subcutaneous blood flow were observed after application of liniment solution to abraded skin (140 g/L).Each of these findings are characteristic of nitrite and they imply the percutaneous absorption of nitrite. Regardless of the nitrite concentration, the methemoglobin concentration was consistently higher in abraded skin than in normal skin.

The purpose of this study was to determine whether the toxicity of a lithographic developer solution which contains hydroquinone is caused by hydroquinone or the alkaline lithographic developer solution. Male Wistar rats were divided into seven groups. In four groups, rats were dosed orally with 3% hydroquinone or 3% hydroquinone in 3% lithographic developer solution. Hydroquinone levels were measured after one and 24 hours. In two groups, rats were dosed orally with 6% hydroquinone or 6% hydroquinone in lithographic developer solution. In the seventh group, rats received the alkaline solution only. Hydroquinone measurement was made using gas chromatography-mass spectrometry. Hydroquinone was rapidly absorbed from the gastrointestinal tract and subsequently distributed throughout the body. Nearly all hydroquinone was excreted in the urine as either a glucuronide or a sulfate (78-82%) within 24 hours. All rats administered 6% hydroquinone in non-alkaline vehicle died, but the mortality rate of rats administered 6% hydroquinone in lithographic developer solution was 60%. Tissue hydroquinone was lower at one hour and 24 hours after administration in lithographic developer solution than in equal dose of hydroquinone in non-alkaline vehicle suggesting decreased absorption in an alkaline pH. Hydroquinone was not associated with gross pathologic changes of the intestine but all animals treated with lithographic developer solution or alkaline solution had congestion, hemorrhagic petechiae and purple-brown discoloration throughout the small intestine. The combination of alkaline/formaldehyde diluent with hydroquinone may delay hydroquinone absorption but increase the risk of intestinal necrosis.

To the Editor: The chambers of the great veins and the right side of the heart are frequently engorged not only in asphyxial deaths, but also in unquestioned natural deaths’)*). For the traditional reasons, in many Japanese forensic textbooks, however, the engorgement is described in a section of asphyxia with insufficient description of the association of engorgement with other types of deaths. Because of this, our students tend to assume that engorgement is a specific sign of asphyxia, and they often “comment upon” the engorgement whilst observing our postmortems (Fig. 1). To such students, we have been explaining that sudden cardiac arrest may be of itself sufficient to cause significant engorgement. In this letter, we

Pathological characteristics of Pokkuri Death Syndrome; narrow circumferences of the coronary arteries in Pokkuri Death Syndrome cases S. Takeichi a, K. Nakajima b,∗, Y. Nakajima b, M.Q. Fujita b a Department of Forensic Medicine, Tokai University School of Medicine, Isehara, Kanagawa 259-1193, Japan b Department of Legal Medicine, Keio University School of Medicine, 35 Shinanomachi, Shinjuku-ku, Tokyo 160-8582, Japan

We have previously reported that the majority of sudden cardiac death (SCD) events were associated with postprandial hyperlipidemia in Japanese subjects. In this investigation, we have compared LDL-cholesterol (LDL-C) and remnant-like lipoprotein particles (RLP) as cardiovascular risk factors in SCD cases, especially in Pokkuri death syndrome (PDS) cases who had nearly normal coronary arteries. To predict the risk of plasma RLP-cholesterol, triglyceride (RLP-C, RLP-TG) and LDL-C in fatal clinical events associated with SCD cases with or without atherosclerosis (PDS), we calculated the cut-off values and likelihood ratio of these lipoproteins from ROC analysis. Sixty-eight percent of SCD cases were above cut-off value of RLP-C (12.8 mg/dL) versus 32% for control death cases (P0.0001) and the likelihood ratio of RLP-C was 2.12. Significantly higher incidence of RLP-C above cut-off value (10.1 mg/dL) was seen in PDS compared to controls (P0.0001) and the likelihood ratio was 3.13. Similarly, significantly higher incidence of RLP-TG above cut-off values, SCD53 mg/dL and PDS67 mg/dL, was seen compared to controls (P0.0001) and the likelihood ratio was 1.86 and 2.73, respectively. Further, significantly higher incidence of LDL-C above cut-off value (93 mg/dL) was seen in SCD compared to controls (P0.0001) and the likelihood ratio was 1.68. However, the incidence of LDL-C above cut-off value (LDL-C106 mg/dL) was not significantly different between PDS and controls and the likelihood ratio was 1.52. In conclusion, this study has shown high levels of plasma remnant lipoproteins in PDS and that PDS cases did not present with atherosclerotic lesions or elevated LDL-C. In contrast, SCD cases showed high levels of plasma remnant lipoproteins together with elevated plasma LDL-C. Accordingly, we believe that plasma remnant lipoproteins level rather than plasma LDL-C is a major pathologic factor in cardiovascular events.

Accidental or suicidal poisonings due to benzodiazepines have been previously reported. A case of fatal, homicidal poisoning with benzodiazepines, antipyretic analgesics (anti-inflammatory drugs), and beer is described here. In this homicidal poisoning, the drugs and beer were given to the decedent by his wife. Autopsy findings showed no clinically significant macroscopic findings except for slight postmortem change. Capillary gas chromatography coupled to mass spectrometry was employed to quantitate the drugs in biological fluids and stomach contents. Six drugs (brotizolam, triazolam, ibuprofen, dihydrocodeine, phenylpropanolamine, and chlorpheniramine) were identified and quantitated in blood, urine, and stomach contents. Although each drug was present in a very small quantity, the cause of death was determined to be the combination of these drugs and alcohol poisoning. Language: en

CO-oximeters are specialized spectrophotometers that automatically determine hemoglobin (Hb) derivatives by measuring absorbance at selected wavelengths (1). We believe that a good understanding of the relevant theory may allow users to avoid many pitfalls during operation of these instruments. The mathematical basis of their operation has not, however, been fully explained by the manufacturers apart from Instrumentation Laboratory (Lexington, MA) at the introduction of their first CO-Oximeter™ (2). Here, we discuss what mathematical methods for data processing might be used in commercial CO-oximeters, particularly in those models that use an “overdetermined” system. CO-oximeters depend on the observation that Hb solutions obey the Lambert–Beer Law; thus, the absorbance measured at a given wavelength is the sum of the absorbance of each Hb derivative at the same wavelength (2). When we measure n wavelengths to determine the m Hb derivatives χ i , we get n equations: \batchmode \documentclass[fleqn,10pt,legalpaper]{article} \usepackage{amssymb} \usepackage{amsfonts} \usepackage{amsmath} \pagestyle{empty} \begin{document} \[\ A\_{j}{=}\ {{\sum}\_{i{=}1}^{m}}{\epsilon}\_{ji}C\_{i}l\mathrm{\ for\ 1}{\leq}j{\leq}n,\] \end{document} where A j is the absorbance at wavelength λ j , C i is the concentration of derivative χ i , and l is the pathlength. e ji is the molar absorptivity at wavelength λ j for derivative χ i . When n = m , we can solve Eq. 1 to get C i . This is termed an “exactly determined” system (3) and has been implemented in the IL 482 CO-Oximeter (Instrumentation Laboratory), the IL 282 (its predecessor), and the Radiometer OSM3 Hemoximeter™ (Radiometer, Copenhagen, Denmark). The IL 482 uses four wavelengths for four Hb derivatives, whereas the OSM3 uses six wavelengths for six unknowns: five Hb derivatives plus one for noise (attributed to “turbidity”). The report by Steinke and Shepherd (4) illustrates how the full exposition of the algorithms used in specific instruments is useful not only to users but also to manufacturers. After measuring absorptive spectra of Hb derivatives … [↵][1]bAuthor for correspondence. [1]: #xref-corresp-2-1

A fatal case of methamphetamine poisoning whilst in police custody is described. Drug identification and quantification in postmortem blood, urine, brain and gastric contents was achieved using gas chromatography-mass spectrometry. The blood concentration of methamphetamine was 4.92 micromol/100 g, which is in the lethal range. It is necessary for police officers to have a good knowledge of the toxic effects of abused drugs.

Background: A sensitive gas chromatography–negative ion chemical ionization mass spectrometry (GC/MS-NICI) method was developed and validated for the measurement of Δ9-tetrahydrocannabinol (THC) in human sweat patches. Methods: THC-d0 and THC-d3 were added to worn blank sweat patches (PharmChek™; PharmChem Incorporated) and extracted with 3 mL of methanol–0.2 mol/L sodium acetate buffer (pH 5.0, 3:1 by volume) on a reciprocating shaker at ambient temperature for 30 min. Extracted solution (2 mL) was diluted with 8 mL of 0.1 mol/L sodium acetate buffer (pH 4.5) and extracted by use of solid-phase extraction columns (CleanScreen®; United Chemical Technologies). Dried extracts were derivatized with trifluoroacetic acid and analyzed with an Agilent 6890 gas chromatograph interfaced with an Agilent 5973 mass selective detector operated in NICI–selected ion-monitoring mode. Results: The lower limits of detection and quantification for THC in human sweat were 0.2 and 0.4 ng/patch, respectively. The calibration curve was linear from 0.4 to 10 ng/patch (R2 >0.995). Overall recovery of THC from blank worn patches to which 0.6, 4.0, and 8.0 ng of THC had been added was 44–46%. Assay imprecision, expressed as CV, was Conclusions: This GC/MS-NICI assay for THC in human sweat provides adequate sensitivity and performance characteristics for analyzing THC in sweat patches and meets the requirements of the proposed Substance Abuse and Mental Health Administration’s guidelines for sweat testing.

Objective— Sudden cardiac death (SCD) still remains a serious problem. We have previously shown that remnant-like particles (RLP) are the major risk factor for SCD and that Rho-kinase plays a central role in the molecular mechanism of coronary vasospasm. In this study, we examined whether RLP from patients with SCD upregulate Rho-kinase associated with an enhanced coronary vasospastic activity. Methods and Results— We isolated RLP and non-RLP in very-low-density lipoprotein (VLDL) fraction from SCD patients without coronary stenosis. We performed in vivo study in which we treated the coronary artery with RLP or non-RLP fraction at the adventitia in pigs. After 1 week, intracoronary serotonin caused marked coronary hyperconstriction at the segment treated with RLP fraction but not with non-RLP fraction ( P P P Conclusions— These results indicate that RLP from SCD patients upregulate Rho-kinase in coronary VSMC and markedly enhance coronary vasospastic activity.

Background: During pregnancy, serum cholesterol (TC) and triglyceride (TG) concentrations are known to increase significantly, but whether remnant lipoprotein particles (RLP) increase has not been shown. Methods: We compared lipid profiles in 22 healthy pregnant women to 31 healthy nonpregnant women and 24 patients with diabetes mellitus (DM), by measuring cholesterol and TG concentrations in major lipoprotein classes after HPLC separation and immunoseparation of RLP. Results: Serum TG and TC concentrations were significantly higher in the pregnant group than in the healthy control or DM groups. Cholesterol and TG concentrations of all major lipoprotein classes were also significantly higher in the pregnant group than the control and DM groups, except for VLDL-TG in the DM group. RLP-C and RLP-TG concentrations were significantly higher in the pregnant group (8.7 mg/dl and 25.4 mg/dl on average) than the control group (2.4 mg/dl and 5.7 mg/dl), but not different from the DM group (8.8 mg/dl and 24.1 mg/dl). RLP-TG to RLP-C ratios were similar among the three groups and correlated with the VLDL-TG to VLDL-C ratio. The percentages of RLP-C in VLDL-C and RLP-TG in VLDL-TG in the pregnant group (15.9% and 15.7%) were significantly lower than those of the control (48.5% and 35.6%) and the DM (32.7% and 20.8%) groups. Conclusions: RLP increased moderately during gestation with the increase in VLDL and TG, but the percentage of RLP in VLDL was significantly lower in the pregnant women compared with the control and DM patients, suggesting that hypertriglyceridemia in pregnancy is not primarily due to an increase in the atherogenic RLP.

Background : Plasma levels of remnant-like particles (RLP) is one of the predictive markers for coronary artery disease (CAD), and the inhibition by RLP of endothelium-dependent vasorelaxation has been reported. We attempted to clarify whether or not RLP, which inhibits endothelium-dependent vasorelaxation, affects nitric oxide (NO) production and NO synthase (eNOS) levels in cultured endothelial cells. Methods : RLP were obtained from postmortem blood of subjects who died of CAD. Modification by RLP of acetylcholine-induced relaxation of rabbit aorta, and changes in NO production and (eNOS) in cultured bovine endothelial cells were examined. Results : RLP at 750 and 1500 μg triglyceride/ml inhibited vasorelaxation, and at 5–160 μg triglyceride/ml, concentration-dependently inhibited NO production. However, (eNOS) did not decrease after incubation with RLP. Conclusion : Postmortem RLP from subjects who died of CAD do not change the amount of (eNOS), but rather, inhibits its activity and attenuates endothelium-dependent vasorelaxation.

The present study analyses the human Y-chromosome minisatellite locus, MSY1 (DYF155S1), in 205 Japanese males of 191 pedigrees using the minisatellite variant repeat (MVR) mapping system. The internal haploid structures of the detected alleles considerably varied and consisted of three major repeat units: types 2, 3 and 4. A comparison of the haploid profiles of the MVR codes identified 185 distinct alleles, of which only five were shared. We did not detect a type 1 repeat unit, and variations were frequent at the 5' end of the minisatellite locus. Within an analysis of 24 paternally linked DNA samples donated by ten families, no mutational events were identified even over two generation gaps. Furthermore, we applied this mapping system to a paternity test in which the alleged father was missing.

In our recent report, it remained unclear whether or not triglyceride-rich lipoprotein remnants (RLP) were associated with the risk of sudden coronary death in younger cases without coronary atherosclerosis that were detected in about 10% of all sudden coronary death cases in Japan. These cases were categorized as 'origin unknown, but suspected to be due to coronary spasm', the so called 'pokkuri disease' in Japan. The present study population consisted of 108 sudden death cases without coronary atherosclerosis [(pokkuri disease n=57) and non-cardiac sudden death (control n=51)] aged 20-69 years from Kanagawa prefecture in Japan. All individuals had died suddenly and unexpectedly, most had no significant history of medical conditions including cardiac symptoms and had not taken medications prior to death according to their medical records. All the autopsies were performed within 12 h after death. Plasma total cholesterol (TC), triglycerides (TG), phospholipids, RLP-C and RLP-TG, VLDL-C, LDL-C, HDL-C, apolipoproteins A-I, A-II, B, C-II, C-III, E, Lp (a) and homocysteine were measured in postmortem plasma samples. The TG-rich lipoprotein remnants measured as RLP-C and RLP-TG were significantly higher in pokkuri disease compared with controls both in fasting and postprandial states (P

Hitherto triglycerides (TG) and TG-rich lipoproteins were been of limited value as surrogates for antemortem levels. We measured TG levels in postmortem plasma from sudden coronary death cases (SCD, n=91) by using two TG assays, Dry Chem TG (free glycerol was added) and the Determiner L-TG (without added free glycerol) that measured net TG. TG levels were markedly higher by the Dry Chem TG (y) vs. Determiner L-TG (x), y=1.03x+229 mg/dl. HPLC showed large amounts of free glycerol in postmortem plasma and in TG-rich lipoprotein remnants (RLP). These results were verified in a rabbit model of SCD. Further, RLP from SCD were found to be biophysically similar to those from living patients with coronary artery disease (CAD). In conclusion, postmortem plasma sampled up to 12 h after death is appropriate for measuring lipid and lipoproteins, TG and RLP-TG as surrogates for antemortem levels when a TG assay without added free glycerol is used.

A method is described for the determination of hippuric acid (HA) and o-, m-, and p-methylhippuric acids (o-, m-, p-MHAs) in urine using solid-phase extraction and gas chromatography–mass spectrometry (GC–MS). The extraction procedure uses an Empore™ disk, derivatized into the respective trimethyl silyl derivatives. All metabolites including the internal standard (I.S.) were clearly able to be analyzed by the DB-17 column. The calibration curves for the four acids show linearity in the range of 5–70 μg/ml. The detection limit of each acid was 1.0–2.5 μg/ml.

A simple and rapid extraction method for the analysis of p,p'-DDE from rat whole blood and tissues was developed using headspace solid-phase microextraction (SPME). A vial containing a sample of p,p'-DDE, sodium hydroxide, and benzophenone as internal standard was heated at 120 degrees C. A polydimethylsiloxane-coated SPME fiber was exposed for 15 min in the headspace of the vial. The fiber needle was then injected and desorbed by exposing the fiber in the injection port of a capillary gas chromatography-mass spectrometry system. The calibration curve demonstrated good linearity throughout the concentration range from 0.02 to 50 microg/g for rat whole blood and liver samples. The limit of detection for p,p'-DDE was 0.020 microg/g using 0.5 g rat whole blood and liver samples. Coefficients of variation ranged from 7.0 to 7.9%. This method was used to analyze a rat whole blood sample after administration of p,p'-DDE.

A 30-year-old male truck driver, known to be asthmatic, was found dead at the roadside lying near his car. A nebulizer bottle of Berotec (fenoterol hydrobromide) was found near his hand. The anatomic cause of death was suspected to be asthma. Toxicological screening of urine using Triage demonstrated the presence of methamphetamine. The blood concentration of methamphetamine was 0.4 microg/ml, and fenoterol was not detected. Hair analysis clearly indicated chronic methamphetamine abuse and medium dependency during the 2 months before death. We conclude that death might have been induced by the interaction of fenoterol and methamphetamine.

A case of fatal methemoglobinemia (MetHb-emia) which was presumably due to ingestion of nitrate is presented. An unidentified man was taken to a local emergency hospital suffering from deep cyanosis and 7 h later he was found dead in the hospital bed. The post-mortem examination of the blood revealed a methemoglobin (MetHb) concentration of 78% and the concentrations of nitrate and nitrite were 1.50 and 0.76 microg/mL, respectively. Capillary gas chromatography coupled to mass spectrometry (GC-MS) and capillary gas chromatography with a nitrogen-phosphorus detector (NPD) were used to detect nitrates and nitrites in the blood.

Among the risk factors for coronary atherosclerosis, elevated LDL-C level is best known. The action of lipoprotein lipase on triglyceride-rich lipoproteins produces remnant lipoprotein particles enriched in cholesterol and apolipoprotein E (apo E). Apo E serves as the ligand for uptake of remnant lipoproteins via the LDL-receptor or the remnant receptor. In this study, postmortem plasma total cholesterol, triglycerides (TG), VLDL-C, HDL-C, lipoprotein (a) [Lp(a)] and remnant-like lipoprotein particles (RLP)-cholesterol, RLP-TG, apolipoproteins B, C III and E were measured, together with LDL-C to assess their potential contribution to the severity of coronary and aortic atherosclerosis of the 197 cases of sudden death (132 cardiac death and 65 non-cardiac death). In all cases, the severity of coronary atherosclerosis was determined at postmortem pathological examination. RLP-cholesterol (RLP-C) and LDL-C concentrations were significantly higher in cases with advanced coronary atherosclerosis compared with those without coronary atherosclerosis; respective median values were 13.5 vs 8.4 mg/dl (P < 0.001) and 140 vs 115 mg/dl (P < 0.05). RLP-C levels were more strongly correlated with the severity score of coronary atherosclerosis than LDL-C.

Gc globulin binds actin at an equimolar ratio with a high affinity. It has been revealed that complexes of Gc with actin are present in the plasma of human cadavers. In the present study, an immunoassay which we developed using a specific anti-Gc monoclonal antibody was applied to evaluate the complexed Gc in cadaver plasma. The percentage of Gc in the complexed form was found to be 38.3 +/- 19.8% (n = 45), and a significantly higher percentage was observed in the plasma of cadavers from burn death. However, the plot of the complex percentage against the time since death showed a relatively low correlation, with a coefficient (r) of 0.51. This was supposed to be due to different environmental conditions after death. Since the complex formation represents the release of cytoplasmic actin into plasma from collapsed cells, the phenomenon should be noted in forensic postmortem chemistry.

Alpha 2 -HS glycoprotein (AHSG) is a human plasma glycoprotein and fetuin is the homologue in the calf. In this report, we present the structure and organization of the AHSG gene. Introns and the 5′ and 3′-flanking regions were obtained by polymerase chain reaction (PCR) and the inverted PCR, respectively, from genomic DNA using AHSG cDNA-specific oligonucleotide primers. The sequence of the PCR products shows that the coding region spans approximately 8.2 kb and is composed of seven exons interrupted by six introns. The exon–intron splice junctions agree with the consensus sequence, and the positions interrupted by introns are precisely identical to those of the rat insulin receptor tyrosine kinase inhibitor (fetuin) gene. The 5′-promoter region contains several characteristic sequences such as an A+T-rich sequence of TAAATAA, C/EBP-binding site, and hepatocyte nuclear factor-5 (HNF-5) and serum response factor (SRF) sites.

Postmortem plasma lipid and lipoprotein levels were analyzed in two groups of Japanese subjects who died suddenly and unexpectedly due to cardiac (n = 93) or non-cardiac (n = 26) causes. No individuals in either group had a significant medical or cardiac history. In this study, we measured plasma total cholesterol, triglycerides, VLDL-cholesterol, LDL-cholesterol, HDL-cholesterol, and especially triglyceride-rich lipoprotein remnants. Triglyceride and apo E-rich remnant-like particles (RLP) were studied as a possible risk factor for sudden cardiac death in relation to the progression of coronary atherosclerosis. The receiver-operating characteristic curve (ROC) analysis showed that RLP-TG was the most significant risk factor for sudden cardiac death among the lipids and lipoproteins and RLP-C was the best predictor for coronary atherosclerosis. HDL-C and LDL-C levels were within normal limits in the majority of the cases and did not appear to relate to the sudden cardiac death. Apo E phenotyping was performed for the detection of the genetic background in the lipid metabolism. The frequency of the Apo E3/3 (wild type) phenotype, which closely relates with the remnant metabolism, was significantly reduced in the sudden cardiac death group. Our study on the postmortem plasma lipid analysis suggested that RLP-C and RLP-TG are the best risk predictor for coronary atherosclerosis and sudden cardiac death, respectively.

Alpha 2-HS glycoprotein (AHSG) is a human plasma glycoprotein that exhibits genetic polymorphism on isoelectric focusing (IEF). To identify the origin of two common alleles, AHSG*1 and *2, we examined nucleotide exchanges in the gene. AHSG cDNA was obtained by RT-PCR from poly(A) RNA of seven liver tissue samples and subcloned into a plasmid vector. After sequencing, we found six single nucleotide differences in comparison with the originally reported sequence. In particular, the nucleotide substitutions of C to T at amino acid position 230 and C to G at position 238 were common among the samples exhibiting phenotype 2-1 or 2. Since these substitutions might give rise to a NlaIII site and a SacI site, respectively, for the potential AHSG*2, we analyzed these substitutions by PCR-RFLP using genomic DNA of 68 individuals. The result was consistent with the IEF analysis of the corresponding serum, indicating that AHSG*1 was characterized by ACG (Thr) at position 230 in exon 6 and ACC (Thr) at position 238 in exon 7, and that AHSG*2 was characterized by ATG (Met) at position 230 and AGC (Ser) at position 238.

A new method is described for isolating the 20-kDa protein, which used to be purified using an actin-immobilized column from human seminal plasma. This method employed ion-exchange column chromatography and isoelectric focusing separation. The level of seminal protein was determined by sandwich ELISA to be 1.06 +/- 0.27 mg/mL (mean +/- SD, n = 8). Its cDNA was cloned from a human salivary gland cDNA library by immunoscreening. The 553-nucleotide sequence included the 5' untranslated region and extended to the poly(A) tail. It encoded a protein of 118 amino acid residues in addition to a signal sequence of 28 residues. This sequence was identical to those of gross cystic disease fluid protein 15 and prolactin-inducible protein cDNAs. Northern blotting revealed the common expression in the submandibular gland and seminal vesicle. An immunohistochemical study in paraffin-embedded tissues from human male sex organs also indicated that the 20-kDa protein is mainly produced in the seminal vesicle.

We describe a case of fatal methemoglobinemia (MetHb-emia) resulting from application of liniment solution containing large quantities of sodium nitrite. As a remedial treatment of atopic dermatitis, the liniment solution was applied all over the boy's body. Autopsy findings showed no significant macroscopic or microscopic findings except blood tinted chocolate brown color and chronic atopic dermatitis over the whole surface of the body. Quantitation of the methemoglobin (MetHb) in the blood was performed using spectrophotometer; MetHb concentration of the blood was 76%. Ion chromatographic determination revealed a nitrite concentration of 1 mg/L in the serum. Such a liniment solution is not authorized by the Ministry of Public Welfare.

The present study was designed to measure the ethanol and its toxic metabolite acetaldehyde concentrations in post-mortem tissue specimens of corpse from whom no quantity of the blood could be collected. Another purpose was to discuss the toxic effects of ethanol and/or acetaldehyde on the various organs.

A 38-year-old male slipped accidentally and fell from a height of 4.6 m, and was admitted to a hospital. A radiograph indicated a linear fracture of the parietal bone across the sagittal sutura, and a computed tomography scan of the head showed severe subdural hematoma on both parietal sides, so an operation was performed to remove the hematoma. Five hours after the operation, brain death due to a cerebral herniation occurred, and he died six days later. Autopsy revealed a linear fracture of the parietal bones, which crossed obliquely the sagittal sutura. A slightly lacerated wound of the dura mater was seen on the sagittal sutura, from where the latter half of that point to the left and right transverse sinuses were occluded completely with thrombosis. The brain was markedly swollen. From these findings, the victim was considered to have died from acute cerebral swelling due to traumatic thrombosis of the superior sagittal sinus.

A fatality due to the ingestion of photographic developer solution containing hydroquinone is described. Hydroquinone was extracted from autopsy materials and identified using gas chromatography-mass spectrometry (GC-MS). The concentration of hydroquinone in the urine, liver, and kidney were 3.4 micrograms/mL, 0.5 and 0.2 microgram/g, respectively. Language: en

The ever increasing presence of environmental toxicants and their disrupting effects on the reproductive systems of wildlife, raise the question of their possible damaging effects on the human reproductive system. Using medicolegal data from over 20,000 Japanese men subjected to necropsy from 1948 to 1998, we investigated temporal changes of testis-weight to find possible evidence of male reproductive disorders. We also made histological observations of 747 testes collected from 1978 to 1998. Our detailed analyses indicates that the development of testis-weight over the last fifty years has followed four marked phenomena: (a) the age at which testis-weight reaches its maximum has become younger, (b) the weight at peak had become generally higher, until a decline in peak-weight beginning after 1960 birth years, (c) the decline-rate at which testis-weight decreases after its peak has greatly accelerated, and (d) the onset of testis-weight of boys has started progressively younger. Our quantitative analyses of testis-weight indicate some possibility of a subtle reproductive disorder in Japanese men, especially those born after 1960. Together with the accelerated development and decline of testis-weight during the last fifty years, the decline in peak-weight might be indicative of a subtle interference of environmental toxicants with male reproductive organogenesis.

To understand the mechanism in postmortem autolysis better, processes in the postmortem degradation of myofibril proteins in the presence of protease inhibitors were studied. Male Wistar rats were given injections of the carboxyl-, thiol-, and serine-protease inhibitors, pepstatin, Ep-475[L-transepoxysuccinyl-leucylamide(3-methyl) butane; E-64-C], and chymostatin, via the femoral vein. Control rats were similarly treated with saline. Then, myofibril proteins were isolated from their cardiac and femoral muscles and from those of control animals at various times after death, and degradation of these myofibril proteins with time was examined by sodium dodecyl sulfate-polyacrylamide gel electrophoresis. In cardiac muscle, alpha-actinin was degraded rapidly, followed by the heavy chain of myosin and light chain of myosin (L2). Actin and the light chain of myosin (L1) were degraded slowly. the degradations of the heavy chain of myosin, alpha-actinin, tropomyosin and L2 after 14 days were not inhibited by pepstatin, but were inhibited by Ep-475 and chymostatin. In skeletal muscle, L1 and L2 were degraded rapidly, followed by the heavy chain of myosin and alpha-actinin. Actin was degraded slowly and was still unchanged 2 weeks after death. The degradations of protein components were inhibited by pepstatin, Ep-475 and chymostatin. These results indicated that after death the components of myofibrils are degraded by various proteases at various rates depending on their properties or structures. This degradation is fundamentally the same in cardiac and skeletal muscles, but inhibitors have somewhat different effects on the postmortem degradation processes after death in the two types of muscle.

In rats, fibrinolytic activity and plasma catecholamine levels increased rapidly after death. Postmortem fibrinolysis was affected by the method of sacrifice, but catecholamine level was not always affected. Immediately after death, the plasma adrenalin level was higher than the noradrenalin level, but then adrenalin remained unchanged or tended to decrease while noradrenalin tended to increase gradually with time. In human subjects after rapid death, the plasma catecholamine and fibrinolytic activity levels were high. However, in cases after slow death they were low.

Experiments by the vascular perfusion technique in isolated hind leg of the dog showed that after addition of various vasoactive drugs to the perfusate, fibrinolytic activity (plasminogen activator level) of the effluent was elevated. The effects of the drugs were completely inhibited only by the specific blocker of their receptor on the vascular wall. High K+ level of the perfusate also induced abundant release of plasminogen activator from the vascular wall.

In rats, fibrinolytic activity and acidosis increased rapidly after death. Postmortem fibrinolysis and the pH, base excess (BE), and [HCO3-] levels were affected by the method of sacrifice: the lower the pH, BE, and [HCO3-] levels, the higher the fibrinolytic activity. Conversely, in experiments using the vascular perfusion technique, low pH, BE, and [HCO3-] levels of the perfusate induced abundant release of plasminogen activator from the vascular wall.

Male Wistar rats were treated with the carboxyl, thiol, and serine protease inhibitors, pepstatin, Ep-475[L-trans-epoxysuccinyl-leucylamide(3-methyl) butane; E-64-c], and chymostatin. Then the femoral muscles of these rats and control animals were used for preparation of myofibril proteins. Sodium dodecyl sulfate-polyacrylamide gel electrophoresis was used to analyze the degradation of these myofibril proteins with time (day) after death. The protease activities of the muscle were also measured. Tropomyosin was degraded most rapidly, followed by the heavy chain of myosin, alpha-actinin, and light chains of myosin (L1 and L2). Actin and troponin-T were degraded slowly, still remaining unchanged 2 weeks after death. The degradation of protein was not inhibited by pepstatin but was inhibited strongly by Ep-475 and very strongly by chymostatin. Chymostatin inhibited degradation of all components except alpha-actinin more strongly than Ep-475. Data on enzyme activities were consistent with these findings. These results suggest that after death the components of myofibrils are degraded with various proteases at various rates depending on their properties or their structure and that the proteases involved in the degradation show some specificity.

Studies were made on the acid-base balance, blood gases, and carbon monoxide (CO), cyanide, and sulfur dioxide concentrations in the blood of albino rabbits that died from automobile exhaust gas poisoning (group I) or fires in cars (complete combustion, group II; incomplete combustion, group III). In group I, the temperature and CO concentration increased gradually to 35 degrees C and 5.2% in 70 min. The animals died after 9 min, when the values were 20 degrees C and 5.2%, respectively. In group II the animals died after 9 min, when the values were 55 degrees C and 1.95%, respectively. In group III, the temperature was very high (870 degrees C), but the CO concentration was not (0.6-1.3%) after 4 min. The animals died after 5 min. In all experimental groups, marked acidosis and hypoxemia were seen, but the CO2 tension (PCO2) was high, in contrast to previous studies on pure CO poisoning. In group I, the level of carboxyhemoglobin (CO-Hb) was significantly higher (91.2 +/- 3.4% in arterial blood, 87.5 +/- 8.1% in venous blood; p less than 0.01) than in groups II and III. Although the O2 tensions of venous and arterial blood (PvO2, PaO2) were very low, that of arterial blood was higher, suggesting that O2 was still being utilized in the tissues at the time of death. In group II, CO-Hb was high (57.7 +/- 16.0% in arterial blood, 61.2 +/- 20.6% in venous blood) and the acid-base balance indicated marked acidosis. In group III, the CO-Hb, PCO2 and cyanide levels in the blood were very high.(ABSTRACT TRUNCATED AT 250 WORDS) Language: en

Autopsy findings on a man who fell from a height due to acute toluene poisoning while painting are described. Gas chromatographic examination revealed that the toluene concentrations of his blood, lung, liver and brain were 48, 35, 65 and 80 micrograms/g, respectively. These toluene levels were not enough to be definitely lethal, but were high enough to anesthetize the central nervous system.