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In forensic practice, determining the postmortem submersion interval (PMSI) and cause-of-death of cadavers in aquatic ecosystems has always been challenging task. Traditional approaches are not yet able to address these issues effectively and adequately. Our previous study proposed novel models to predict the PMSI and cause-of-death based on metabolites of blood from rats immersed in freshwater. However, with the advance of putrefaction, it is hardly to obtain blood samples beyond 3 days postmortem. To further assess the feasibility of PMSI estimation and drowning diagnosis in the later postmortem phase, gastrocnemius, the more degradation-resistant tissue, was collected from drowned rats and postmortem submersion model in freshwater immediately after death, and at 1 day, 3 days, 5 days, 7 days, and 10 days postmortem respectively. Then the samples were analyzed with liquid chromatography-tandem mass spectrometry (LC-MS/MS) to investigate the dynamic changes of the metabolites. A total of 924 metabolites were identified. Similar chronological changes of gastrocnemius metabolites were observed in the drowning and postmortem submersion groups. The difference in metabolic profiles between drowning and postmortem submersion groups was only evident in the initial 1 day postmortem, which was faded as the PMSI extension. Nineteen metabolites representing temporally-dynamic patterns were selected as biomarkers for PMSI estimation. A regression model was built based on these biomarkers with random forest algorithm, which yielded a mean absolute error (± SE) of 5.856 (± 1.296) h on validation samples from an independent experiment. These findings added to our knowledge of chronological changes in muscle metabolites from submerged vertebrate remains during decomposition, which provided a new perspective for PMSI estimation., (© 2024. The Author(s), under exclusive licence to Springer-Verlag GmbH Germany, part of Springer Nature.)

Seawater-drowning-induced acute lung injury (SD-ALI) is a life-threatening disorder characterized by increased alveolar-capillary permeability, an excessive inflammatory response, and refractory hypoxemia. Perfluorocarbons (PFCs) are biocompatible compounds that are chemically and biologically inert and lack toxicity as oxygen carriers, which could reduce lung injury in vitro and in vivo . The aim of our study was to explore whether the vaporization of PFCs could reduce the severity of SD-ALI in canines and investigate the underlying mechanisms. Eighteen beagle dogs were randomly divided into three groups: the seawater drowning (SW), perfluorocarbon (PFC), and control groups. The dogs in the SW group were intratracheally administered seawater to establish the animal model. The dogs in the PFC group were treated with vaporized PFCs. Probe-based confocal laser endomicroscopy (pCLE) was performed at 3 h. The blood gas, volume air index (VAI), pathological changes, and wet-to-dry (W/D) lung tissue ratios were assessed. The expression of heme oxygenase-1 (HO-1), nuclear respiratory factor-1 (NRF1), and NOD-like receptor family pyrin domain containing-3 (NLRP3) inflammasomes was determined by means of quantitative real-time polymerase chain reaction (qRT-PCR) and immunological histological chemistry. The SW group showed higher lung injury scores and W/D ratios, and lower VAI compared to the control group, and treatment with PFCs could reverse the change of lung injury score, W/D ratio and VAI. PFCs deactivated NLRP3 inflammasomes and reduced the release of caspase-1, interleukin-1β (IL-1β), and interleukin-18 (IL-18) by enhancing the expression of HO-1 and NRF1. Our results suggest that the vaporization of PFCs could attenuate SD-ALI by deactivating NLRP3 inflammasomes via the HO-1/NRF1 pathway., Competing Interests: The authors declare that the research was conducted in the absence of any commercial or financial relationships that could be construed as a potential conflict of interest., (Copyright © 2024 Su, Zhang, Liu, Meng, Ma, Mo, Ren, Liang, Yang, Li and Chen.)

Background: Ferroptosis is a new type of nonapoptotic cell death model that was closely related to reactive oxygen species (ROS) accumulation. Seawater drowning-induced acute lung injury (ALI) which is caused by severe oxidative stress injury, has been a major cause of accidental death worldwide. The latest evidences indicate nuclear factor (erythroid-derived 2)-like 2 (Nrf2) suppress ferroptosis and maintain cellular redox balance. Here, we test the hypothesis that activation of Nrf2 pathway attenuates seawater drowning-induced ALI via inhibiting ferroptosis., Methods: we performed studies using Nrf2-specific agonist (dimethyl fumarate), Nrf2 inhibitor (ML385), Nrf2-knockout mice and ferroptosis inhibitor (Ferrostatin-1) to investigate the potential roles of Nrf2 on seawater drowning-induced ALI and the underlying mechanisms., Results: Our data shows that Nrf2 activator dimethyl fumarate could increase cell viability, reduced the levels of intracellular ROS and lipid ROS, prevented glutathione depletion and lipid peroxide accumulation, increased FTH1 and GPX4 mRNA expression, and maintained mitochondrial membrane potential in MLE-12 cells. However, ML385 promoted cell death and lipid ROS production in MLE-12 cells. Furthermore, the lung injury became more aggravated in the Nrf2-knockout mice than that in WT mice after seawater drowning., Conclusions: These results suggested that Nrf2 can inhibit ferroptosis and therefore alleviate ALI induced by seawater drowning. The effectiveness of ferroptosis inhibition by Nrf2 provides a novel therapeutic target for seawater drowning-induced ALI.

Background: Drowning is the leading cause of unintentional death in the world, with about 372,000 drowning deaths reported annually. Forensic opinion of drowning is considered one of the most difficult tasks in confirming the exact cause of death. Drowning is now possibly patronized to cover up intentional crimes. This study was aimed at utilizing concentrations of vitreous proteins and lipids in making inroad into the inquest of true drowning. Twelve albino rabbits constituted the sample size. The rabbits were divided into three groups; control death (CD), post-mortem submersion death (PSD) and truly drowned death (TDD). Vitreous humor was extracted from the eyes of all the rabbits and centrifuged for biochemical analysis. The biochemical parameters analyzed included vitreous total protein (TP), albumin (ALB), globulin (GLO), albumin/globulin (A/G) ratio, immunoglobulin G (IgG), immunoglobin M (IgM) total cholesterol (TC), triacylglycerol (TG), high density lipoprotein (HDL), low density lipoprotein (LDL), very low density lipoprotein (VLDL) and glucose (GLU) using standard methods., Result: The mean concentrations of vitreous parameters amongst the groups were compared using one-way anova (pos hoc-LSD). The findings revealed that the concentrations of vitreous TP, GLO, IgG, IgM, TG, TC, TG, HDL, VLDL and GLU were significantly elevated (p < 0.05), whereas ALB and A/G ratio decreased significantly amongst the drowned death group compared to that either CD or PSD., Conclusion: The findings have indicated that some vitreous protein and lipid parameters could aid in discriminating deaths that truly emanated from true drowning from that of post-mortem drowning and suffocation., Competing Interests: Declaration of competing interest All the three authors fund the project without grant or finance from any organization. It was a totally self-help research wholly financed by the authors. Interpersonal and intrapersonal relationship did not in any way influence the result or direction of the research., (Copyright © 2020 Elsevier Ltd and Faculty of Forensic and Legal Medicine. All rights reserved.)

Tidal volume V T required for mouth-to-mouth (MTM) and bag-valve-mask (BVM) rescue ventilation remains debatable owing to differences in physiology and end-point objectives. Analysis of gas transport may clarify minimum necessary V T and its determinants. Alveolar and arterial O 2 and CO 2 responses to MTM and air BVM ventilation for V T between 0.4 and 1.2 liters were computed using a model of gas exchange that incorporates inspired gas concentrations, airway dead space, cardiac output, pulmonary shunt, blood gas dissociation curves, tissue compartments, and metabolic rate. Parameters were adjusted to match published human data. Steady state arterial oxygen saturation reached plateaus at V T above 0.7 liters with MTM and 0.6 liters with air ventilation at 12 breaths per minute. Increasing shunt shifted oxygenation plateaus downward, but larger tidal volumes did not improve oxygen saturation. Carbon dioxide retention occurred at V T below 2.3 liters for MTM ventilation and 0.6 liters for air ventilation. Results establish a physiological foundation for tidal volume requirements during resuscitation., (Copyright © 2019 Elsevier B.V. All rights reserved.)

Objective: Heme oxygenase-1 (HO-1) plays a critical protective role in various insults-induced acute lung injury (ALI) through its strong anti-inflammatory, anti-oxidant, and anti-apoptotic properties, but its protective role and mechanism on seawater aspiration-induced acute lung injury remains unclear. This study aimed to explore the therapeutic potential and mechanism of HO-1 to attenuate seawater aspiration-induced ALI in vivo and in vitro., Methods: The viability and invasion of A549 cell were analyzed through cell counting kit-8 and lactate dehydrogenase release assay; the transcriptional level of inflammatory cytokines (TNF-α, IL-6, IL-8 and MCP-1) and cell proliferation-related cytokines (FoxM1, Ccnb1 and Cdc25C) in seawater-treated A549 cell were tested by qPCR; apoptotic cells were analyzed by flow cytometryd; HO-1mRNA and protein were determined by qPCR and western blotting; the fluorescent indicators (DCFH-DA, dihydroethidium, MitoSox Red and Fluo-4) were used to monitor generation of ROS and mitochondrial function. The lung wet/dry weight radio and lactate dehydrogenase activity, Sirius red staining, TUNEL assay and immunohistochemical staining with anti-pan Cytokeratin antibody were analyzed in seawater-drowning mice. The role of HO-1 on seawater-drowning pulmonary injury was explored via HO-1 activity inhibitors (Zinc protoporphyrin) in vitro and in vivo., Results: Seawater exposure decreased the cellular viability, increased the production of pro-inflammatory cytokines (IL-6, IL-8 and TNF-α), induced cellular apoptosis and inhibited the expression of cell proliferation-related cytokines (FoxM1, Ccnb1 and Cdc25C). Moreover, seawater exposure led to mitochondrial dysfunction in A549 cells. Supplement of HO-1 sepcific inducer (heme) or its catalytic product (biliverdin) significantly attenuated seawater-induced A549 damage and promoted cell proliferation. However, Zinc protoporphyrin abolished the beneficial effects of HO-1 on seawater drowning-induced pulmonary tissue injury., Conclusion: HO-1 attenuates seawater drowning-induced lung injury by its anti-inflammatory, anti-oxidative, and anti-apoptosis function., (Copyright © 2019 Elsevier B.V. All rights reserved.)

The water contents of individual organs are maintained in a narrow range, but altered in morbidity owing to a disturbance of water equilibrium. The present study investigated the tissue water contents of major organs with regard to the cause of death in serial autopsy cases within 3 days postmortem (n = 329; 223 males, 106 females; age range, 1-100 years). Individual tissue water contents differed markedly across organs, but no significant postmortem or survival-period dependence, gender-related difference, or age dependence was observed. However, the lung water contents were higher in drowning cases, especially in saltwater cases (p < 0.05), and in strangulation among the acute mechanical asphyxiation cases. The brain water contents were higher in hypothermia cases (cold exposure) and tended to be higher in hyperthermia cases (heatstroke). The kidney water contents were higher in drowning and acute cardiac cases than in fatal intoxication and fire fatality cases, but tended to be higher in fresh- and bathwater drowning cases than in saltwater cases (p > 0.05). The spleen water contents were higher in bathwater drowning than in saltwater and freshwater cases, but did not differ among other the causes of death. These findings suggest that the postmortem tissue water content of individual organs, especially the lungs and/or kidney, depends on the cause of death and particularly contributes to differentiation between saltwater and freshwater drowning, respectively. This work therefore provides insight into the investigation of varied tissue water imbalances during the death process. In conclusion, we recommend the measurement of tissue water content because it is easy to perform and appears to be useful for evaluating the pathophysiology of systemic circulatory failure., (Copyright © 2019. Published by Elsevier Ltd.)

Background: Elevation in postmortem vitreous humor sodium and chloride (PMVSC) in salt water drowning (SWD) when the immersion time is less than 1 hour (SWD1) is hypothesized to result from electrolyte changes in blood from salt water inhalation/ingestion during drowning. After approximately 1 hour after death, electrolytes may diffuse into the vitreous humor via the eye coverings. Another abundant element in salt water is magnesium, which is approximately 50 times higher in concentration than the blood and vitreous humor magnesium levels. Magnesium is able to diffuse across the eye coverings but not as easily through the blood-ocular barrier. With these properties, we hypothesize that postmortem vitreous magnesium (PMVM) would not be elevated in SWD1 but become elevated in SWD with immersion times greater than 1 hour (SWD>1)., Aim: The aim of this article was to investigate the differences in PMVM and PMVSC between nonimmersion deaths, SWD1, and SWD>1., Methods: This is a 1-year retrospective study comparing PMVM and PMVSC in nonimmersion deaths, SWD1, and SWD>1., Results: Postmortem vitreous magnesium is significantly higher in SWD>1 than SWD1 and nonimmersion deaths, with no significant difference between SWD1 and nonimmersion deaths. Postmortem vitreous humor sodium chloride is statistically higher in SWD1 and SWD>1 than nonimmersion deaths., Conclusions: As a conclusion, PMVSC elevates and PMVM does not elevate in SWD1.

Many terrestrial insects live in environments that flood intermittently, and some life stages may spend days underwater without access to oxygen. We tested the hypothesis that terrestrial insects with underground pupae show respiratory adaptations for surviving anoxia and subsequently reestablishing normal patterns of respiration. Pupae of Manduca sexta were experimentally immersed in water for between 0 and 13 days. All pupae survived up to 5 days of immersion regardless of whether the water was aerated or anoxic. By contrast, fifth-instar larvae survived a maximum of 4 h of immersion. There were no effects of immersion during the pupal period on adult size and morphology. After immersion, pupae initially emitted large pulses of CO 2 After a subsequent trough in CO 2 emission, spiracular activity resumed and average levels of CO 2 emission were then elevated for approximately 1 day in the group immersed for 1 day and for at least 2 days in the 3- and 5-day immersion treatments. Although patterns of CO 2 emission were diverse, most pupae went through a period during which they emitted CO 2 in a cyclic pattern with periods of 0.78-2.2 min. These high-frequency cycles are not predicted by the recent models of Förster and Hetz (2010) and Grieshaber and Terblanche (2015), and we suggest several potential ways to reconcile the models with our observations. During immersion, pupae accumulated lactate, which then declined to low levels over 12-48 h. Pupae in the 3- and 5-day immersion groups still had elevated rates of CO 2 emission after 48 h, suggesting that they continued to spend energy on reestablishing homeostasis even after lactate had returned to low levels. Despite their status as terrestrial insects, pupae of M. sexta can withstand long periods of immersion and anoxia and can reestablish homeostasis subsequently., (© 2016. Published by The Company of Biologists Ltd.)

Post-mortem determination of biochemical parameters, especially for obscure cases, has been recognized useful in diagnosis of the underlying causes of death. Procalcitonin (PCT) is known to rise in a response to any proinflammatory stimulus. The present study aims to estimate postmortem PCT levels in serum and kidney, liver, brain; and whether it is similar in different causes of death models (trauma, drowning and freezing) models or not. The study was performed on 60 male rabbits. Rabbits were divided into four different death induced models (15 rabbit each): trauma, infection, drowning and freezing models. At the end of the study, all rabbits were sacrificed; blood samples, kidneys, livers and brains were collected. PCT was measured using ELISA assay. Results showed highly significant increase in PCT levels in all tested samples in different models of death. The infection induced model showed the highest levels in all tested samples compared to other groups mainly in liver; followed by trauma model and drowning model which were increased mainly in brain's samples. The least model which showed increased PCT levels was the freezing model mainly in liver samples. Post Hoc multiple comparisons test showed significant differences between groups in most of liver, brain and kidney samples, while PCT serum blood samples were significant only between trauma and infection groups. It was concluded that PCT can differentiate between sepsis and non-sepsis related deaths and that organs like liver, kidney and brain PCT levels could be an alternative to serum PCT for the diagnosis of postmortem sepsis., (Copyright © 2015 Elsevier Ltd and Faculty of Forensic and Legal Medicine. All rights reserved.)

Drowning is an important public health problem, but the mechanism of acute lung injury induced by near-drowning is rarely reported. The aim of this study is to investigate the role of hypertonicity and HIF-1α in seawater aspiration-induced lung injury. Diverse solutions were used to study the effect of hypertonicity on hypoxia, inflammation, vascular leakage, edema, and HIF-1α expression in lungs of rats. The relationship between hypertonicity and hypoxia, when they induced HIF-1α, was studied and the roles of ATM, PI3K, and p38 in the course of hypertonicity inducing HIF-1α were investigated. At last, our conclusion was verified with HIF-1α inhibitor and inducer in seawater aspiration rats. The results showed that hypertonicity, but not isotonicity and hypotonicity, promoted hypoxia, inflammation, vascular leakage, edema, and HIF-1α expression in lungs. Hypertonicity not only induced HIF-1α in a time- and dose-dependent manner but also could increase HIF-1α synergistically with hypoxia in AEC. Furthermore, hypertonicity increased HIF-1α by promoting its mRNA expression through both ATM and PI3K activation and by suppressing its protein degradation through p38 activation. During hyperosmotic stress, the increased HIF-1α promoted the production of the inflammatory cytokines in NR8383 and elevated monolayer permeability through increasing VEGF in RLMVEC. In conclusion, hypertonicity induced by aspirated seawater aggravated lung injury through increasing HIF-1α which promoted inflammation and edema in lung tissues in rats.

Although electrolyte analysis of pleural effusion at autopsy is useful for the diagnosis of water aspiration (i.e., drowning), the method of comparing each level of sodium (Na(+)), potassium (K(+)), and chloride (Cl(-)) ions does not clearly differentiate between freshwater drowning, seawater drowning, and non-drowning. Therefore, here we introduce the summation of Na(+), K(+), and Cl(-) levels, that is SUM(Na+K+Cl), as a modified diagnostic indicator. In 21 autopsy cases of freshwater drowning, 32 cases of seawater drowning, and 43 non-drowning controls (with pleural effusion), mean SUM(Na+K+Cl) differed significantly between the groups (188.8±33.2, 403.5±107.9, and 239.3±21.7 mEq/L, respectively). We defined a SUM(Na+K+Cl) cut-off value of <195.9 mEq/L as strongly suggestive of freshwater aspiration and that of >282.7 mEq/L as strongly suggestive of seawater aspiration. When these values were applied to the two drowning groups, 15 cases (71%) of freshwater drowning and 29 cases (91%) of seawater drowning were diagnosed correctly. This new approach may be more valid than previous methods in cases found >2 days after death or those with substantial pleural effusion (>100 mL). For an additional 15 bathtub deaths, mean SUM(Na+K+Cl) was 198.8±40.0 mEq/L, and in 14 of these cases (93%) the relationship between cause of death and SUM(Na+K+Cl) could be explained using this method. Forensic pathologists should not depend exclusively on chemical findings and should consider also typical pathological indicators of drowning. This new method may be useful as a supplementary diagnostic tool when used alongside consideration of the pathological findings., (Copyright © 2013 Elsevier Ireland Ltd. All rights reserved.)

Resveratrol is a phytoalexin synthesized by a wide variety of plants, which has been proven to be effective in suppressing oxidative stress and inflammation. The aim of the present study was to investigate the effect of Resveratrol's prodrug: 3,5,4'-tri-O-acetylresveratrol, on seawater drowning-induced acute lung injury (SWD-ALI). Histological changes were assessed to study lung injuries; cytokines in lung samples were monitored by ELISA to reflect inflammation; T-SOD and MDA activity were detected to examine oxidative stress in lung tissues. Besides, we also tested the expression of NF-κB and HIF-1α to probe the possible protecting mechanism of 3,5,4'-tri-O-acetylresveratrol on AWD-ALI. The results showed that pretreatment with different doses of 3,5,4'-tri-O-acetylresveratrol improved seawater-induced lung histopathologic changes, alleviated lung edema, reduced the production of inflammatory mediators including TNF-α and IL-1β, inhibited MDA activity, and enhanced T-SOD activity, which was possibly associated with inhibition of NF-κB and HIF-1α. In conclusion, the current study demonstrated that 3,5,4'-tri-O-acetylresveratrol exhibited a protective effect on SWD-ALI by inhibiting of the inflammatory response, which may also involve the suppression of oxidative stress in lung tissues., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Our previous study showed that seawater can cause lung tissue cell apoptosis; in the present study, the immunohistochemistry and Western blot analysis results demonstrated that Fas, FasL, and cleaved caspase-8 and caspase-3 were up-regulated in the rat lungs exposed to seawater. We found that seawater-induced human lung alveolar epithelial A549 cell apoptosis was concentration and time dependent. Moreover, seawater increased the expression of Fas, FasL, and cleaved caspase-8 and caspase-3 in A549 cells. The incubation of A549 cells in the presence of FasL-neutralising antibody (NOK-2) or caspase-8 inhibitor (Z-IETD-FMK) resulted in a decrease of seawater-induced cell apoptosis. NOK-2 inhibited Fas/FasL interaction and reduced the cleavage of caspase-8 and caspase-3, and Z-IETD-FMK blocked caspase-8 and caspase-3 activation. Seawater similarly produced a significant increase in rat alveolar type II cell apoptosis and expression of Fas and cleaved caspase-8. In summary, the Fas/FasL pathway involved in alveolar epithelial cell (AEC) apoptosis could be important in the pathogenesis of seawater-induced acute lung injury (SW-ALI)., (Copyright © 2012 Elsevier B.V. All rights reserved.)

Drowning involves complex fatal factors, including asphyxiation and electrolyte/osmotic disturbances, as well as hypothermia in cold water. The present study investigated the molecular pathology of pulmonary injury due to drowning, using lung specimens from forensic autopsy cases of drowning (n = 21), acute mechanical asphyxia due to neck compression and smothering (n = 24), and hypothermia (cold exposure, n = 11), as well as those of injury (n = 23), intoxication (n = 13), fire fatality (n = 18), and acute cardiac death (n = 9) for comparison. TaqMan real-time reverse transcription polymerase chain reaction was used to quantify messenger RNA (mRNA) expressions of pulmonary surfactant-associated proteins A and D (SP-A and SP-D), tumor necrosis factor (TNF)-α, interleukin (IL)-1β, and IL-10. SP-A and SP-D mRNA levels were lower for drowning, mechanical asphyxiation, fire fatality, and acute cardiac deaths than for hypothermia and injury. TNF-α, IL-1β, and IL-10 mRNA levels were higher for drowning or for drowning and injury than for other groups; there was no significant difference between fire fatality, involving airway injury due to inhalation of hot/irritant gases, and other control groups. These observations suggest characteristic molecular biological patterns of pulmonary injury involving suppression of pulmonary surfactants and activation of early-phase mediators of inflammation in drowning, with high mRNA expression levels of pulmonary surfactants in fatal hypothermia; however, there was no significant difference among these markers in immunohistochemical detection, except for SP-A. These mRNA expressions can be used as markers of pulmonary injury to assist in investigations of the pathophysiology of drowning and fatal hypothermia in combination with other biochemical and biological markers.

In investigating death due to mechanical asphyxiation and drowning, a cardiac attack is important for discriminating between possible causes of death and as a contributory factor in death processes; however, general pathologies involving visceral congestion are often similar. The present study compared terminal cardiac dysfunction in these fatalities using the molecular pathology of atrial and brain natriuretic peptides (ANP and BNP) in the myocardium as markers of cardiac strain. Both mechanical asphyxiation (n=27) and drowning (n=23) showed significantly lower ANP and BNP mRNA expressions in bilateral ventricular walls than sudden cardiac deaths (n=36). In addition, right atrial wall BNP mRNA expression was lower in asphyxiation; however, immunostaining did not demonstrate any difference among these fatalities. Differences among the subtypes of asphyxiation or between fresh- and saltwater drowning were insignificant. These observations suggest a difference between primary heart failure in sudden cardiac death and terminal cardiac dysfunction secondary to fatal asphyxiation or drowning., (Copyright © 2012 Elsevier Ireland Ltd. All rights reserved.)

Aquaporins (AQPs), a family of transmembrane water channels, mediate physiological response to changes of fluid volume and osmolarity. It is still unknown what role of AQPs plays in seawater drowning-induced acute lung injury (ALI) and whether pharmacologic modulation of AQPs could alleviate the severity of ALI caused by seawater aspiration. In our study, the results from RT-PCR and Western blotting showed that intratracheal installation of seawater up-regulated the mRNA and protein levels of AQP1 and AQP5 in lung tissues. Furthermore, we found that treatment of tanshinone IIA (TIIA, one of the main active components from Chinese herb Danshen) significantly reduced the elevation of AQP1 and AQP5 expression induced by seawater in rats, A549 cells and primary alveolar type II cells. Treatment of TIIA also improved lung histopathologic changes and blood-gas indices, and reduced lung edema and vascular leakage. These findings demonstrated that AQP1 and AQP5 might play an important role in the development of lung edema and lung injury, and that treatment with TIIA could significantly alleviate seawater exposure-induced ALI, which was probably through the inhibition of AQP1 and AQP5 over-expression in lungs., (Copyright © 2011 Elsevier B.V. All rights reserved.)

Pericardial fluid is a kind of serous fluid in pericardial cavity. Because blood undergoes postmortem changes such as autolysis and putrefaction, vitreous humor is limited,cerebrospinal fluid is easily mixed with blood, pericardial fluid, on the other hand, exists in a closed cavity and can be hardly contaminated by postmortem changes, and also is easily obtained. Pericardial fluid not only plays an important role in clinic practice, but also is widely applicable in forensic practice. This paper briefly presented the properties of pericardial fluid and its clinical significance. It reviewed biochemical changes in decedents died of heart diseases, drowning and asphyxia, and explored the significance in medico-legal investigation. Moreover, application of pericardial fluid in forensic serology, forensic toxicological analysis and other fields were also discussed. Pericardial fluid analysis may provide important information for determination of the cause of death with further investigation concerning forensic applicability of pericardial fluid.

Primary Objective: To report Magnetic Resonance Imaging (MRI) and/or Magnetic Resonance Spectroscopy (MRS) findings in subjects with hypoxic encephalopathy caused by drowning in recent literature and to compare them with non-specific hypoxic encephalopathy., Method: Systematic review of the Medline Database for bibliographic citations from 1996 to 2008., Results: The studies included in this review described a total of 68 victims of drowning. From those, 58 performed MRS with a decrease of N-Acetyl-Aspartate/Creatine ratio in 75.86% (n = 44), and presence of lactate in 65.52% (n = 38) of the cases. MRI data was available in 46 cases. The main finding was brain edema in 78.26% (n = 36) and abnormalities of MRI signal in basal ganglia in 75% (n = 27) of the cases. Worse clinical outcomes were reported in conjunction with degree of MRI and MRS alterations. The findings were more consistent in the latter. Comparing these results with literature from non-specific hypoxic brain injury, the drowning process is apparently more variable., Conclusions: We found a trend to a more variable pattern of brain injury as seen by MRI/MRS in victims of drowning, which may reflect the nature of the aggression. Possibly there are different mechanisms involved in aquatic submersion, such as temperature, not present in pure hypoxic injury.

It is important for forensic pathologists to determine the diagnosis of drowning as well as the site of drowning. In a previous study, we propose that analysis of electrolytes in pleural effusion from rats may be useful for determining whether drowning has occurred in seawater or freshwater. To test this proposal, we measured the concentration of sodium, potassium and chloride ions and total protein in pleural effusion from 40 autopsy cases: 24 involving seawater drowning, 9 freshwater drowning and 7 no drowning. The concentrations of sodium and chloride ions in pleural effusion showed a significant difference between seawater drowning and freshwater drowning. The concentration of potassium ions and total protein showed no difference between each group, although they increased in proportion to the postmortem interval in cases of both seawater and freshwater drowning. These results are almost same as our previous study and, thus, the quantitative analysis of electrolytes in pleural effusion may be useful for determining whether drowning has occurred in seawater or freshwater.

Objective: To investigate the differences between the lung injuries induced by seawater and freshwater drowning in a rabbit model., Methods: Forty-two New Zealand rabbits were divided randomly into three groups: control group (C, n = 18), freshwater drowning group (F, n = 12), seawater drowning group (S, n = 12). The drowning model was established by pouring seawater or freshwater (2 ml/kg) into the respiratory tract through a tracheal catheter. Mean arterial pressure (MAP) and heart rate (HR) were monitored continually. Respiratory rate (RR), blood gas analysis and electrolyte contents of every rabbit were observed at different time. The lung wet to dry weight (W/D) ratio and lung permeability index (LPI) were calculated. The contents of malondialdehyde (MDA), myeloperoxidase (MPO) and superoxide dismutase (SOD) were measured by biochemical method. The expressions of tumor necrosis factor-alpha (TNF-alpha) and interleukin-1 beta (IL-1 beta) were detected by enzyme linked immunosorbent assay (ELISA). At the same time, the changes in pathology were studied with by hematoxylin eosin (HE) staining, and lung pathologic score (LPS) was calculated., Results: There was no significant difference in blood electrolyte contents and HR among the three groups (all P > 0.05). In freshwater drowning, there was a temporary increase of MAP at 5 minutes, RR increased immediately, and partial pressure of carbon dioxide in artery (PaCO(2)) and base excess (BE) were persistently decreased (P < 0.05 or P < 0.01). Oxygenation index (PaO(2)/FiO(2)) fell to (297.8+/-81.3) mm Hg (1 mm Hg = 0.133 kPa) at 0.5 hour, then elevated to over 300 mm Hg rapidly, and then reverted to initiative level in around 2 hours. There were several edematous and petechial areas on the dependent region of the lung. Alveolar collapse and parenchymal congestion were the main pathological features. W/D ratio and LPI showed no remarkable change. In lung tissue, the level of LPS, MPO, MDA, TNF-alpha and IL-1 beta had a significant increase, while SOD had a significant decrease (P < 0.05 or P <0.01). In S group, respiratory symptoms were more serious; edematous and congestive areas of the lung were more extensive, and the dependent region showed hepatization changes. W/D ratio and LPI consisted of elevated significantly (all P < 0.01). The pathological characteristics were massive inflammatory cell infiltration and more serious alveolar edema. Compared with F group, the extent of up- or down-regulation of RR, MAP, PaO(2)/FiO(2), PaCO(2), BE, inflammatory media and cytokines in S group was more prominent and steady, and S 6-hour group had a higher pathological score than S 3-hour group and F group (P < 0.05 or P < 0.01)., Conclusion: Seawater and freshwater drowning could not only injure pulmonary parenchymal cells directly, but also induce acute inflammatory reaction. Lung injury induced by seawater is severer than that by freshwater.

To clarify drowning death, positive evidence for aspiration of the immersion medium and the subsequent fatal mechanism is necessary. This study investigated biochemical findings with regard to lung weight in drowning cases of adults (n=56, >18 years of age, <48 h postmortem: salt water, n=19; fresh water, n=21; brackish water, n=16), using acute cardiac death cases (n=240) as controls. The biochemical markers used in this study were urea nitrogen (UN), sodium (Na), chloride (Cl), calcium (Ca) and magnesium (Mg) in the blood and pericardial fluid (PCF). The left-to-right ratio of cardiac blood UN levels was lower for drowning, showing an inverse correlation to the total lung weight. There was a mild postmortem decrease in serum and PCF Na and Cl levels; however, left cardiac serum and PCF Na, Cl, Ca and Mg levels were higher for saltwater drowning, and left cardiac serum Na and Cl levels were lower for fresh water drowning. Correlation of the left cardiac serum level with lung weight was positive for Na, Cl and Mg in saltwater and brackish water drowning, and was also positive for Ca in saltwater drowning. There was an inverse correlation with lung weight for PCF Na and Cl levels in freshwater drowning. These findings suggest that analyses of serum and pericardial markers in relation to lung weight are useful for evaluating the composition and amount of aspirated medium when investigating drowning death.

Basic fibroblast growth factor (bFGF) is a highly conserved and ubiquitously distributed mitogen, and much is known at the molecular level. However the available information about in vivo distribution in human tissues and expression changes in relation to causes of death is not sufficient. The present study investigated 35 autopsy cases, comprising five cases for each cause of death: acute myocardial infarction/ischemia (AMI), mechanical asphyxiation, blunt brain injury, drowning, hypothermia, intoxication and sharp instrument injury. The bFGF immunopositivity was detected mainly in interstitial cells and sporadically in cardiomyocytes in the heart, in macrophages in the lung, astrocytes in the brain, mainly in the sinusoidal Kupffer cells and partly in the hepatocytes in the liver, red pulp of the spleen, pancreatic islets and proximal convoluted tubules and corpuscles in the kidney. Immunopositivity was frequently detected in the lung and liver for AMI and hypothermia, and in the kidney for AMI, mechanical asphyxiation, drowning and injuries, but was not evident in the kidney for hypothermia. Positivity in these tissues varied by case in other causes of death. High positivity in the brain was seen for intoxication, but AMI, mechanical asphyxiation and drowning showed lower positivity. For the heart, spleen and pancreas, there was no evident difference among the causes of death. These findings suggested that bFGF expression in the lung, liver, kidney and brain varies depending on the cause of death, and is useful for investigating deaths.

The histopathological alterations that permit the diagnosis of death by asphyxia are very unspecific, although pulmonary alterations are of great importance in this respect. The postmortem diagnosis of drowning, particularly, continues to be one of the most difficult in forensic pathology. The aim of this study is to jointly evaluate microscopic findings and immunohistochemical surfactant protein A (SP-A) expression in the upper and lower lobes of lungs in different causes of death, and their possible application to the diagnosis of drowning. We studied 120 cadavers from subjects with a mean age of 48.73 years (SD 19.45; range 2-86 years), and with a mean post-mortem interval of 30 hours (SD 39.59; range 3-216 hours). According to the scene, cause and circumstances of death, and autopsy findings, cases were classified into groups as follows: (a) drowning (n=47); (b) other asphyxia (n=44) and (c) other causes (n=29). In the upper and lower lobes of lungs, histological studies of H&E staining and immunohistochemical surfactant protein A expression were made. The presence and severity of congestion, haemorrhage and oedema, together with immunohistochemical SP-A expression, may have a diagnostic value in differentiating asphyxia and drowning from other causes of death, and drowning from other types of asphyxia. Our findings suggest that both lobes should be investigated to establish the diagnosis, although the findings in the upper lobe might be the most important for differentiating the exact cause of death.

The present study investigated cardiac troponin I (cTnI) and creatine kinase MB (CK-MB) in the blood and pericardial fluid from medicolegal autopsy cases (n=234, within 48h postmortem) with regard to the cause of death. The cTnI and CK-MB levels in cardiac, peripheral blood and pericardial fluid generally showed a mild and gradual postmortem time-dependent elevation (r=0.231-0.449, P<0.05-0.001). However, postmortem elevation of cTnI was larger for specific causes of death including acute myocardial infarction (AMI), cerebrovascular diseases (CVD), hyperthermia, fatal methamphetamine (MA) abuse and carbon monoxide (CO) intoxication and insignificant for recurrent myocardial infarction (RMI), chronic congestive heart diseases (CHD) and drowning, while that of CK-MB was greater for CO intoxication and insignificant for drowning. Cardiac blood and pericardial cTnI levels were relatively high for AMI, RMI, hyperthermia, MA abuse and CO intoxication, and was low for drowning. Elevated CK-MB level was observed for cardiac blood in asphyxiation and MA abuse cases and for peripheral blood in hyperthermia and MA abuse cases. When the cTnI/CK-MB ratio was estimated, it was independent of postmortem time, and the ratios for cardiac blood and pericardial fluid were significantly higher in cases of AMI, RMI, hyperthermia and CO intoxication but lower in cases of drowning. Elevations of cTnI levels in cardiac blood and pericardial fluid were related to the morphological severity of myocardial damage. These findings suggest that elevated cTnI and CK-MB levels in blood and pericardial fluid are related to ischemic, hypoxic and/or cytotoxic myocardial damage, which are characteristic of the cause of death, although the levels increase after death depending on myocardial damage at the time of death.

Previous studies showed significant differences in postmortem urea nitrogen (UN), creatinine (Cr) and uric acid (UA) levels in heart blood depending on the causes of death, including acute death. In addition, the levels in pericardial fluid approximated the clinical serum reference ranges, and their elevations may be assessed based on clinical criteria. The present study investigated difference between blood and pericardial levels of these markers. Medicolegal autopsy cases (n=556, within 48h postmortem) of the following causes of death were examined: injury (n=136), asphyxiation (n=50), drowning (n=39), fire fatalities (n=99), hyperthermia (n=11), hypothermia (n=8), poisoning (n=26), delayed traumatic death (n=44) and natural diseases (n=143). When serum UN, Cr and UA levels were compared with the pericardial levels, there was an equivalency for delayed traumatic death and chronic renal failure, although each level was markedly elevated. Parallel increases in serum and pericardial UA and/or Cr levels were also observed for hypothermia and gastrointestinal bleeding. However, in drowning cases, the left cardiac and pericardial UN levels were lower than the right cardiac and peripheral levels, suggesting the influence of water aspiration. Significant elevations in serum and pericardial Cr and UA levels with a higher serum/pericardial UA ratio for fatal methamphetamine intoxication suggest progressive skeletal muscle damage due to advanced hypoxia/acidosis. Similar findings were often observed for other acute and subacute deaths. These findings suggest that a comparison between blood and pericardial nitrogenous compounds would be useful for investigating the cause and process of death.

Accumulating studies demonstrate that the expressions of hypoxia-inducible factor 1 (HIF-1), erythropoietin (EPO) and vascular endothelial growth factor (VEGF) depend on cellular oxygen tension, which is involved in the pathological process of tissue hypoxia and/or ischemia. The present study investigated hypoxia-inducible factor-1alpha (HIF-1alpha), EPO and VEGF mRNA expressions in the kidney with regard to the cause of death in medicolegal autopsy. Relative quantifications of HIF-1alpha, EPO and VEGF mRNAs, based on real-time TaqMan reverse transcription-polymerase chain reaction (RT-PCR), were performed on tissue specimens obtained from consistent sites of the bilateral renal cortices. The cases (total, n=245, 6-48h postmortem) included fatal blunt/sharp instrument injuries (n=53/31), asphyxia (n=28: aspiration, n=8; strangulation/hanging, n=20), drowning (n=27), fire fatality (n=62), acute myocardial infarction/ischemia (AMI, n=39), and gastrointestinal hemorrhage (n=5). Both HIF-1alpha and EPO mRNA levels were significantly lower in drowning cases. More characteristic findings were found for VEGF mRNA: it showed higher expression levels for AMI, acute blunt/sharp instrument injury, and aspiration, whereas it was lower for neck compression (strangulation/hanging), drowning, fire fatality with higher blood carboxyhemoglobin (COHb) levels (>60%), peracute blunt injury, and gastrointestinal hemorrhage. Quantitative assays of renal HIF-1alpha, EPO and VEGF mRNA transcripts are potentially useful for investigating the pathophysiology of death, and VEGF mRNA may be especially useful as an indication of acute circulatory failure.

In forensic pathology, previous studies have suggested the possible application of cardiac troponins in the diagnosis of myocardial infarction. However, there appears to be insufficient practical data on other causes of death. The present study was a comprehensive analysis of the cardiac, peripheral blood and pericardial levels of cardiac troponin T (cTnT) in serial medicolegal autopsy cases (n = 405) with a survival time <24 h and within 48 h postmortem to assess the validity of investigating myocardial damage with special regard to traumatic causes of death. These included blunt and sharp instrument injury (n = 122 and 21, respectively), asphyxiation (n = 35), drowning (n = 27), fire fatalities (n = 94), hyperthermia (n = 13), hypothermia (n = 6), fatal methamphetamine (MA) abuse (n = 12) and carbon monoxide (CO) poisoning (n = 5) in comparison with myocardial infarction (MI, n = 57) and cerebrovascular diseases (n = 13). Cases within 12h postmortem usually showed lower cardiac and pericardial cTnT levels than did those of longer postmortem time of 12-48 h. In the early postmortem period of <12 h, significantly elevated serum cTnT levels were observed for hyperthermia. Thereafter, fatal MA abuse, CO poisoning and MI cases also showed higher levels. However, cTnT remained at lower levels for hypothermia and drowning. The elevation of cTnT was associated with the pathology of advanced myocardial damage involving swelling and liquefactive necrosis. The above-mentioned differences were the smallest for peripheral blood. These findings suggest that elevations in postmortem serum and pericardial cTnT levels depend on the severity of myocardial damage at the time of death and are related to the pathological findings, although postmortem interference should be taken into consideration.

Although previous forensic pathological studies have suggested the possible application of cardiac troponins in the diagnosis of myocardial infarction, there appears to be insufficient data with regard to its cardiac pathology. The present study analyzed the heart blood, peripheral blood and pericardial fluid levels of cardiac troponin T (cTnT) in sudden cardiac deaths (n = 96) within 48h postmortem in relation to pathological findings of acute myocardial infarction (AMI, n = 34), recurrent myocardial infarction (RMI, n = 23), ischemic heart disease without any pathological evidence of infarction (IHD, n = 24) and other heart diseases (OHD, n = 15). Control groups (n = 75, survival time <24 h) within 48 h postmortem consisted of asphyxiation (n = 35), drowning (n = 27) and cerebrovascular diseases (n = 13). There was a marked correlation in the cTnT levels between right and left heart blood samples. The pericardial level was usually higher than either heart blood level, and the external iliac venous blood level was the lowest. Although postmortem time-dependent increases in heart and pericardial blood cTnT levels were observed in most groups, they were most evident for AMI and asphyxiation. In the early postmortem period (<12 h) there was no significant difference between AMI or RMI and the other groups except for drowning. After 12 h postmortem, significantly elevated heart blood and pericardial cTnT levels were observed for AMI and RMI showing multiple interstitial hemorrhages and necrosis compared to those with localized eosinophilic changes or patchy interstitial hemorrhages, IHD and OHD. These differences were the smallest for peripheral blood. For sudden cardiac death cases, the difference in cTnT level at each site among the causes of death was independent of gender, age, heart or lung weight and pathologies of affected coronary artery and severity of coronary stenosis. These observations suggest that the elevation in postmortem blood and pericardial cTnT levels in sudden cardiac death may depend on the severity of ischemic myocardial damage including the size and intensity of myocardial lesions involving multiple interstitial hemorrhages and necrosis, and also the postmortem period for heart and pericardial levels.

S-100 protein (S100) is an acidic calcium-binding protein, which is abundantly found in the brain. The aim of the present study was to investigate the immunohistochemical distribution of S100 in the cerebral cortex in forensic autopsy cases with regard to the cause of death and relationship to its serum levels. The cases (n = 286, 3-48 h postmortem) included fatal head injuries (n = 89), acute death from other blunt injuries (n = 29), sharp instrument injuries (n = 20), asphyxiation (n = 29: strangulation/hanging, n = 22; aspiration, n = 7), drownings (n = 22), fire fatalities (n = 68), cerebrovascular diseases (n = 9) and acute myocardial infarction/ischemia (AMI, n = 20). S100-immunopositivity was mainly observed in the gliacytes, in part, in the neurons and myelins. For S100B-immunostaining, the gliacytes and myelins were positive, whereas the neurons were negative. Positivity in astrocytes was dependent on the cause of death, and was significantly lower in acute deaths due to strangulation/hanging and drowning, and mildly low in those due to injuries. Positivity in neurons and myelins was frequently observed in delayed head injury deaths and fire fatalities, showing an inverse relationship with the positivity in astrocytes in head injury cases. For cases of acute death, there was an inverse relationship between S100-positivity in the astrocytes and the serum S100B level. These observations suggest that astrocytes are more rapidly and severely injured than neurons during fatal brain damage, thereby causing an elevation in the serum S100B level.

The aim of the present study was to examine Ub-immunoreactivity in the midbrain periaqueductal gray matter (PGM), which is involved in pain processing and modulation, in forensic autopsy cases (n=273) in relation to the causes of death: acute deaths from blunt injuries (n=75), sharp weapon injuries (n=36), fatal asphyxiation (n=22), drownings (n=16: freshwater, n=9; saltwater, n=7), fire fatalities (n=64), poisoning (n=12), hyperthermia (n=5), hypothermia (n=5), delayed deaths from blunt head injury (n=8), acute cardiac deaths (n=24), and acute cerebrovascular strokes (n=6). The Ub-immunoreactivity was clearly observed in the nuclei of the PGM neurons, showing no postmortem interference or age-dependency. A higher value was observed in blunt injuries, fire fatalities and also in saltwater drowning, hyperthermia and delayed head injury deaths. These findings suggest a complicated mechanism for the ubiquitination of PGM neurons, to which multiple factors including the intensity and duration of pains possibly under alert consciousness, traumatic and metabolic neurodegeneration may contribute.

Objective: To perform a molecular autopsy involving the RyR2-encoded cardiac ryanodine receptor/calcium release channel to determine whether mutations responsible for catecholaminergic polymorphic ventricular tachycardia (CPVT) represent a novel pathogenic basis for unexplained drownings., Methods: A cardiac channel molecular autopsy was performed on 2 individuals who died of unexplained drowning and whose cases were referred to the Sudden Death Genomics Laboratory at the Mayo Clinic in Rochester, Minn. Comprehensive mutational analysis of all 60 protein-encoded exons of the 5 long QT syndrome-causing cardiac channel genes and a targeted analysis of 18 RyR2 exons known to host RyR2-mediated CPVT-causing mutations (CPVT1) was performed using polymerase chain reaction, denaturing high-performance liquid chromatography, and DNA sequencing., Results: Both individuals harbored novel mutations in RyR2. Postmortem mutational analysis revealed a familial missense mutation in exon 14, R414C, in a 16-year-old girl. A 9-year-old boy possessed a sporadic missense mutation in exon 49, V2475F. Both amino acid positions involve highly conserved residues that localize to critical functional domains in the calcium release channel. Neither substitution was present in 1000 reference alleles., Conclusions: This molecular autopsy study provides proof of principle that RyR2 mutations can underlie some unexplained drownings. A population-based genetic epidemiology study that involves molecular autopsies of individuals who die of unexplained drowning is needed to determine the prevalence and spectrum of KCNQ1 and now RyR2 mutations as potential pathogenic mechanisms for drowning.

In medico-legal autopsies for drowned bodies, the location of drowning needs to be determined. To investigate the usefulness of electrolyte analysis in pleural effusion as an indicator of the location where the deceased has drowned, we determined the concentrations of electrolytes in the pleural effusion of rats drowned in four kinds of water. The concentrations of sodium and chloride ions in the pleural effusion of rats that drowned in seawater were significantly greater than those of rats that drowned in freshwater at both 1 day and 3 days after drowning. The concentration of potassium ions in pleural effusion 1 day after drowning showed no difference between each group, although it then increased from 1 to 3 days after seawater drowning, whereas it decreased from 1 to 3 days after freshwater drowning. The concentration of total protein in pleural effusion increased from 1 to 3 days after drowning, however, there was no significant difference in the concentration of total protein in pleural effusion between each group at either 1 day or 3 days after drowning. These results suggest that analysis of electrolytes in pleural effusion may be useful for determining whether drowning has occurred in seawater or in freshwater.

In the central nervous system (CNS), a variety of ubiquitinated structures have been reported, usually as pathological alterations of the brain related to degenerative diseases or aging. However, previous studies showed an increase in the ubiquitin (Ub)-immunoreactive intranuclear inclusion of the pigmented neurons of the substantia nigra in the midbrain in asphyxiation and fire fatalities in the adult subjects. The aim of the present study was to examine granular 'dot-like' Ub-immunoreactivity in the crus cerebri (cortico-spinal tracts) in related fatalities (over 35 years of age, n=169), including fatal asphyxiation (n=27), drownings (n=14), fire fatalities (n=60), and control groups (n=68). Dot-like Ub-immunoreactivity was clearly observed in the descending tract of the crus cerebri. Morphometric analysis of the positive granular area (dot-like Ub-area) showed a higher value in strangulation and fire fatalities and a lower value in hemorrhagic and head injury deaths, as was observed for the inclusion-type neuronal Ub-positivity. However, there was a difference between those markers: a low value was seen for the inclusion-type neuronal Ub-positivity in hanging and drownings, and a difference in the dot-like Ub-area was detected between fire fatalities with lower and higher COHb levels. Our findings suggested the possible usefulness of these markers for examination of CNS stress responses in traumas, at least in middle-aged and elderly victims and a partial difference in stress reaction between the cortico-spinal tracts and dopaminergic neurons.

Background: Nitric oxide is a signal molecule regulating the organism functions in living bodies. The aim of this study was to investigate the NO levels of striated muscles after different types of death in rats., Methods: Nitric oxide levels in the muscles of masseter, triceps, and quadriceps obtained from right and left sides of 24 Spraque-Dawley rats following death were investigated. The rats were divided into three groups as cervical dislocation (control) group, electric shock group, and drowning group. After applying a light anesthesia, the rats were killed by cervical dislocation, electric shock and drowning. The samples were taken immediately and 120 minutes after death., Results: In all muscle types of all groups, NO concentrations were lower in samples obtained 120 minutes after death than in those obtained immediately after death. NO concentrations were lower in the electric shock and drowning group than in the control group for both times., Conclusion: We can conclude that the type of death may affect the occurrence of rigor mortis and NO measurement may give an important clue in evaluation the mode of death.

Objective: To observe water channel 1 protein in the lungs of the drown rat., Methods: Immunohistochemical method and Colloidal gold of the electron microscopy were used to observe the location of aquaporin1(AQP1) on the rat lung tissue between the experiment group and control group., Results: The positive expression of AQP1 was seen in the capillary endotheliocyte of the interstitium and around bronchi and in the alveolar endothelial cells. The difference in the value of integrated optical density between the two groups was statistically significant. AQP1 was located in the type I alveolar cells, and the difference in the semi-quantity of Colloidal gold of the electron microscopy between the two groups was statistically significant as well., Conclusion: The above data suggest that AQP1 positive reaction is one of the sensitive signs of drowning and it is located in the type I alveolar cells.

Objective: To determine the expression of water channel 1 protein in the lung of drown rat and that of after death thrown into the water., Methods: Immunohistochemical method was used and the computer image analysis was conducted to detect the distribution of AQP1., Results: The positive expression of AQP1 was seen in the capillary endotheliocyte of the interstitial and around bronchi and in the alveolar endothelial cells. The value of intergrated optical density was statistical significant., Conclusion: It suggests that AQP1 is one of sensitive signs to distinguish ante-mortem and postmortem immersion.

Endogenous alcohol production can increase the blood alcohol concentration (BAC) of drowning victims following submersion and confound epidemiological studies of the role of alcohol. This study seeks to determine how soon after a drowning death a victim's BAC is influenced by post-mortem alcohol production. The drop in mean lung weight that occurs over time in the water was hypothesized to serve as a proxy for the time course of decomposition, and thus provide an empirical measure to determine how soon after death to first suspect endogenous alcohol. The autopsy lung weights of 562 previously healthy males who drowned were compared across six submersion time groups (0-11.9, 12-23.9, 24-47.9, 48-95.9, 96-167.9 and >or=168 h) and two times of year (winter and non-winter). The hypothesis that a drop in lung weight is sensitive to the time course of decomposition was supported by (1). a statistically significant drop in mean lung weight that occurred 12-23.9 h post-submersion in the non-winter months, but not until 96-167.9 h in the colder winter months; and (2). a significant drop in lung weight was not observed in the group of cases with zero BAC. With a parallel finding that an increase in the proportion of cases with a positive BAC first occurred at the 12-23.9 h submersion group during the warmer non-winter months, we concluded that production of alcohol can occur in bodies recovered from the water as early as 12 h after death. Because excluding drownings with submersion durations greater than 12 h would exclude almost half of our cases from epidemiological studies of alcohol and drowning, additional evidence from the forensic literature was used to develop an adjustment procedure to account for endogenous alcohol production for submersion times of up to 1 week.

In this study, adenosine triphosphate (ATP), adenosine diphosphate (ADP), adenosine monophosphate (AMP) and lactic acid in the muscles of masseter, triceps, and quadriceps obtained from right and left sides of Spraque-Dawley rats following death were investigated. The samples were taken immediately and 120 minutes after death occurred. The rats were killed either by cervical dislocation or drowning. ATP concentrations in the muscles of masseter, triceps, and quadriceps were lower in samples obtained 120 minutes after death than in those obtained immediately after death. ADP, AMP, and lactic acid concentrations in these muscles were higher in samples obtained 120 minutes after death than those obtained immediately after death. A positive linear correlation was determined between ATP and ADP concentrations in quadriceps muscles of the rats killed with cervical dislocation and in triceps muscles of the rats killed with drowning. When rats killed with cervical dislocation and with drowning were compared, ADP, AMP, and lactic acid concentrations were lower in the former than in the latter for both times (immediately and 120 minutes after death occurred). In the case of drowning, ATP is consumed faster because of hard exercise or severe physical activity, resulting in a faster rigor mortis. Higher lactic acid levels were determined in muscles of the rats killed with drowning than the other group. In the control and electric shock rats, ATP decreased in different levels in the three different muscle types mentioned above in control group, being much decline in masseter and then in quadriceps. This may be caused by lower mass and less glycogen storage of masseter. No different ATP levels were measured in drowning group with respect to the muscle type possibly because of the severe activity of triceps and quadriceps and because of smaller mass of masseter. One can conclude that the occurrence of rigor mortis is closely related to the mode of death.

Objective: To observe changes of thromboxane B2 (TXB2), 6-keto-prostaglandin F1 alpha(6-Keto-PGF1 alpha) and TXB2/6-Keto-PGF1 alpha (T/P) in lungs of rats drowned in hypothermic sea water and to assess their influence on the blood-gas., Methods: Rats of different groups were drowned nearly to death in hypothermic sea water and then taken out of the water rapidly, observed at room temperature, after that the following steps were taken in 5, 15, 30, 60, 240 min and 360 min groups, that were 1 ml arterial blood taken from left heart for blood-gas analysis including pH, PaO2 and PaCO2, rectal temperature observed; at last, the ratio of left dry lungs with left wet lungs was assessed, TXB2 and 6-Keto-PGF1 alpha in right lungs were examined in all above groups and dead group(14 rats dead, only 4 examined)., Results: The rectal temperature[(20.13 +/- 0.48) degree C], pH(6.68 +/- 0.03), PaO2[(45.00 +/- 6.30) mm Hg)], TXB2[(97.46 +/- 17.46) ng/L] and 6-Keto-PGF1 alpha[(25.59 +/- 8.12) ng/L] dropped to the lowest point in the 5 minutes group(P < 0.01), while PaCO2[(89.18 +/- 5.10) mm Hg] reached the highest point(P < 0.01), all above items from 5 minutes group then showed a recovering tendency, but only the pH in 240 minutes and 360 minutes groups as well as TXB2 in 360 minutes group and dead group reached near the level of normal control groups (P > 0.05); T/P had a rising tendency and reached the highest point in the 360 minutes group., Conclusions: The production and secretion of TXB2 and 6-Keto-PGF1 alpha were influenced by hypothermia, hypoxemia and acidosis, the imbalance of T/P could be one of factors influencing the improvement of blood gas index.

To evaluate the significance of immunohistochemical staining of ubiquitin (heat shock protein) in the midbrain for the medico-legal diagnosis of fatal asphyxiation and drowning, we investigated forensic autopsy cases of fatal mechanical asphyxia (n = 18), manual/ligature strangulation (n = 9), hanging (n = 4), aspiration/choking (n = 5) and drowning (n = 16). These were compared to control groups (n = 30) consisting of fatalities from brainstem injury (n = 12) and acute myocardial infarction (n = 18). Ubiquitin was clearly demonstrated in the nuclei of pigmented substantia nigra neurons, showing two intranuclear staining patterns: a type of inclusion (possibly Marinesco bodies) and a diffuse staining. The diffuse staining was significantly more frequently observed in cases of drowning. The percentage of total ubiquitin positive neurons was frequently higher in strangulation (5.1-28.4%, mean 17.0%), aspiration/choking (5.3-32.0%, mean 17.6%) and drowning (7.0-34.1%, mean 19.8%), but relatively low in hanging (5.1-12.7%, mean 8.6%), brainstem injury (0-10.4%, mean 5.0%) and acute myocardial infarction (1.5-16.9%, mean 8.3%). These observations suggest that intranuclear ubiquitin immunoreactivity of the pigmented substantia nigra neurons in the midbrain was induced by a fatal severe stress on the central nervous system in asphyxiation and drowning.

The brain stem hypoglossal nucleus (HN) is the center of nerves innervating the upper respiratory tract and is related to control of mastication, deglutition, speech and respiration. To elucidate the relationship between asphyxia and the HN, we investigated the change of hypoglossal neurons in cases of hanging, strangulation, smothering, choking, drowning and respiratory failure. Using immunohistochemical techniques, we observed the brain stem HN with antibodies against microtubule-associated protein 2 (MAP2), muscarinic acetylcholine receptor (mAChR), c-fos gene product (c-Fos) and 72 kD heat-shock protein (HSP70). MAP2, a cytoskeletal protein of the neuron, is a marker of neuronal damage. Muscarinic AChR was used as a marker of neuronal membrane and ACh signaling. We employed both HSP70 and c-Fos as markers of stress- or damage-related events. We measured the percentage of immunopositive neurons in total neurons of HN. Drowning produced higher expression of HSP70 and c-Fos than other causes of asphyxia, suggesting that drowning induces more severe damage in HN neurons. Furthermore, it was suspected that neuronal changes in drowning might relate to functions of the HN. These observations indicate that immunohistochemical examination of the brain stem HN could provide useful information for determining the cause of asphyxia.

Clinical, laboratory and X-ray findings in 34 victims of submersion are presented. Five people died and 29 survived (age range 12-60 years). Severe hypoxia was found in all patients (mean PO2 of 58 mmHg with some oxygen support). Arterial blood gas analysis showed significant metabolic acidosis in 19 patients and significant respiratory acidosis in 15 patients. Pulmonary oedema was the most common X-ray finding. Fourteen patients were put on mechanical ventilation on the basis of their clinical picture and blood gases analysis. Clinical and laboratory data are very similar to those reported in international studies.

Adipocere formation is well known as a later postmortem change. We collected adipocere from five male victims which had been submerged under the sea or fresh water for 1 mon to 4 yr. Fresh subcutaneous fat of a male victim who died from a cerebral contusion was used as the control. The samples were homogenized, and the lipids were extracted with chloroform and methanol followed by injection into a gas chromatograph and a gas chromatograph-mass spectrometer. We detected hydroxy fatty acids (10-hydroxyoctadecanoic acid and 10-hydroxyhexadecanoic acid) as well as 10-ketooctadecanoic acid in adipocere, but not in the control. In addition, we found for the first time a cholesterol-related peak with a molecular ion of 388 in adipocere and identified it as epicoprostanol, suggesting not only oxidation but also reduction had occurred during the formation of adipocere. In addition, we showed the time-course of epicoprostanol accumulation. The relationship between the time of adipocere formation and the characteristic lipid composition is discussed.

We examined changes in plasma during drowning and hanging to determine the mechanism of acute asphyxia. Rabbits were used for the acute drowning and hanging experiments (using artificial sea water or fresh water for the former and complete or incomplete hanging for the latter). The plasma was examined for changes in haptoglobin and total protein contents. For acute drowning, the haptoglobin level reduced dramatically after one minute of inhalation of artificial sea and fresh water by about 46.0% and about 61.0%, respectively. Total protein content showed the same pattern of reduction. Afterward, haptoglobin remained generally unchanged but the total protein content returned to the original level 3 minutes after inhalation. For the handing, the haptoglobin level reduced within 1 minute. When respiratory arrest was near, both haptoglobin and total protein contents rose gradually. The changes in the haptoglobin level caused by hanging were similar to those that occur in acute asphxia when the air passage is completely obstructed and in subacute asphxia when the air passage is strangulated. Totally different and unique patterns were seen in association with drowning in comparison with obstructive asphyxia. These results suggested that the pathological physiology of acute drowning is different from that of fatal neck compression or other types of acute asphyxia.

In an attempt to detect signs of dilution of blood 38 autopsy cases of drowning in fresh water and 35 cases with other causes of death were investigated. Osmolarity and sodium and potassium concentrations were determined in serum from the right and left sides of the heart, cerebrospinal fluiid and vitreous humor...