DNA degradation within mouse brain and dental pulp cells 72 hours postmortem

In this study, we sought to elucidate the process of DNA degradation in brain and dental pulp cells of mice, within postmortem 0-72 hours, by using the single cell gel electrophoresis assay and professional comet image analysis and processing techniques. The frequency of comet-like cells, the percentage of tail DNA, tail length, tail moment, Olive moment and tail area increased in tandem with increasing postmortem interval. In contrast, the head radius, the percentage of head DNA and head area showed a decreasing trend. Linear regression analysis revealed a high correlation between these parameters and the postmortem interval. The findings suggest that the single cell gel electrophoresis assay is a quick and sensitive method to detect DNA degradation in brain and dental pulp cells, providing an objective and accurate new way to estimate postmortem interval.

Spectrophotometric Determination of Trimethylamine-nitrogen in Cadaver Tissues for the Estimation of Late Postmortem Interval:A Pilot Study

To study the relationship between the late postmortem interval （PMI） and trimethyl-amine-nitrogen （TMA-N） in postmortem tissues of cadaver, TMA-N in muscles, livers and kidneys of rats was measured at different postmortem intervals （PMI） by using a modified spectrophotometric method. The results indicated that the detection sensitivity of TMA-N was 1 mg/L, and there was a good linear correlation between the value of absorbance （A value） and TMA-N at the concentration of 1-10 mg/L （R2=0.9991）. Although TMA variation in muscles was different from that in inner organs during the time since death, TMA-N changes in cadaver tissues was positively correlated with PMI. During 2 to 7d since death, the best correlation between PMI and TMA-N concentration was found in muscles. With PMI as an independent variable, the cubic polynomial regression equation was y=-0.457x3 ＋6.519x2-24.574x＋27.207 （R2=0.969）. During 3 to 8 days since death, PMI was best correlated with TMA-N concentration in inner organs. With PMI as the independent variable, the cubic polynomial regression equation was y=0.509x3-9.153x2＋55.727x-95.819 （R2=0.953）. It was concluded that TMA-N in tissues could be used as a new estimator for late PMI. The method used in this study offered advantages such as accuracy, sensitivity, little samples required and wide PMI estimation.

Bioluminescent Assay of Microbial ATP in Postmortem Tissues for the Estimation of Postmortem Interval

To study the relationship between changes of microbial ATP in four kinds of murine tis-sues and the postmortem interval （PMI）, healthy SD rats were sacrificed and their muscles, livers, spleens and kidneys were sampled at different postmortem intervals. The concentration of microbial ATP was detected using bioluminescent assay and the data was statistically analyzed. The concentra-tion of microbial ATP in muscle increased with PMI time. The peak appeared at the 7th day after death, and at the 10th day, microbial ATP in muscle tissue increased again. In internal organs, the peaks of microbial ATP were observed at the 8th day after death and the level decreased during 8–10 d. The differences in microbial ATP concentration in liver, spleen and kidney were not statistically significant. During day 0 to day 9 after death, the correlation was best between PMI and microbial ATP in muscle. With PMI as the independent variable, the cubic polynomial regression equation was Y=0.02X3–0.166X2–0.666X＋13.412 （R2=0.989, P〈0.01）. In internal organs, the best correlation was found between PMI and microbial ATP during day 0 to day 10. With PMI as the independent variable, the cubic polynomial regression equation was Y=0.016X3–0.127X2–0.809X＋13.324 （R2=0.986, P〈0.01）. There existed high correlations between PMI and microbial ATP concentration in rat tissues. Since only a small amount of tissue was needed for the detection and the sample was not affected by self-decomposition, the method may extend the time range of PMI estimation.

Changes in ATP Levels in Rabbit Blood and Its Application for Estimation of the Postmortem Interval

Summary： Relationship between ATP changes of rabbit blood and postmortem interval （PMI） was studied. Twenty-four healthy rabbits were sacrificed and randomly divided into 3 groups with 8 rab- bits of each group. The bodies of three groups were placed in calorstat at temperature of 15℃, 25℃ and 35℃, respectively. The blood from the right ventricle was sampled through indwelling needle each 4 h until 72 h after death. ATP levels in the blood samples were measured by using ATP fluo- rescence rapid detection technique at different PMIs. Blood ATP levels slightly increased in the early stage after death and then constantly declined at all temperatures （15℃, 25℃, and 35℃）. Cubic polynomial regression equations with log[ATP] as dependent variable （y） and PMI as independent variable （x） at different temperatures and the optimal time period were established as followed： Under 15℃ and during 16-64 h after death, y=-3.027×10^-5x^3＋0.003x^2-0.096x-10.625 （Ra^2=0.992, P〈0.001）; under 25℃ and during 8-56 h after death, y=-2.921×10^-5x^3＋0.002x^2- 0.059x-11.186 （Ra^2=0.989, P〈0.001）; under 35℃ and during 4-36 h after death, y=-9.769×10^-5x^3＋ 0.005x^2 -0.117x-11.166 （Ra^2=0.991, P〈0.001）. The changes in ATP levels in blood collected from right ven- tricle of rabbit cadavers showed relatively stable and regular degradation within 72 h after death at different temperatures.

Image Analysis on Corneal Opacity:A Novel Method to Estimate Postmortem Interval in Rabbits

Corneal opacity is one of the most commonly used parameters for estimating postmortem interval （PMI）. This paper proposes a new method to study the relationship between changes of corneal opacity and PMI by processing and analyzing cornea images. Corneal regions were extracted from images of rabbits＇ eyes and described by color-based and texture-based features, which could represent the changes of cornea at different PMI. A KNN classifier was used to reveal the association of image features and PMI. The result of the classification showed that the new method was reliable and effective.

Oleic Acid (OA), A Potential Dual Contrast Agent for Postmortem MR Angiography (PMMRA): A Pilot Study

Summary:Choosing proper perfusates as contrast agents is an important aspect for postmortem magnetic resonance angiography(PMMRA).However,in this emerging field,the number of suitable kinds of liquid is still very limited.The objective of this research is to compare MR images of oleic acid(OA)with paraffin oil(PO)in vitro and in ex situ animal hearts,in order to evaluate the feasibility to use OA as a novel contrast agent for PMMRA.In vitro,OA,PO and water(control)were introduced into three tubes separately and T,weighted-spin echo(Tw SE)and T2w-SE images were acquired on a 1.5T MR scanner.In the second experiment,0A and PO were injected into left coronary artery(LCA)and left ventricle(LV)of ex situ bovine hearts and their Tw-SE,Tzw-SE,Tw-multipoint Dixon(Tjw-mDixon)and 3DT2w-mDixon images were acquired.The overall results indicate that OA may have a potential to be used as a dual(T and T2 based)contrast agent for PMMRA when proper sequence parameters are utilized.However,as the pilot study was based on limited number of animal hearts,more researches using OA in cadavers are needed to validate our findings.

POSTMORTEM REDISTRIBUTION KINETICS OF ACONITINE IN RABBITS

The postmortem redistribution of aconitine(AC) and its influencing factors by orally ingested Aconitum brachypodum Diels (AbD) in rabbits were studied. The results showed that postmortem AC redistribution did exist, and the diffusion along a concentration gradient was the major influencing factor on it. Change of temperature and incomplete distribution in life also influenced it.Besides those mentioned above, there were other influencing factors. These may be related to postmortem blood movement and toxin released from cells occurring as part of the processes of autolysis and putrefaction.

Formation of advanced glycation end products in raw and subsequently boiled broiler muscle:biological variation and effects of postmortem ageing and storage

The study aimed to investigate and compare the contents of carboxymethyllysine(CML)in two kinds of broilers during postmortem ageing and storage.The contents of CML in raw and boiled(100℃,30 min)broiler briskets and legs which were from white feather broilers(n=8)and yellow feather broilers(n=8)with ageing and storage at 4℃for 0-168 h were determined.Postmortem ageing and storage had a significant(P<0.05)effect on the color and A;in both boiled broilers meat.In addition,with the ageing and storage time increasing,CML content in raw white feather broiler brisket meat had no significant(P>0.05)change,while that in boiled brisket meat significantly(P<0.05)increased during 0-6 h,then decreased during 6-24 h,finally increased again.CML content in leg meat increased significantly(P<0.05)with the ageing and storage time prolonging.But postmortem ageing and storage had no significant(P>0.05)effect on the CML contents in raw/boiled yellow feather broilers.Meanwhile,CML contents in white feather broilers were much higher than that in yellow feather broilers.Thus,white feather broilers can be selected as the research object to study the mechanism of ageing and storage on CML content in the postmortem broiler in the future.

Detection of human papillomavirus L1 gene DNA fragments in postmortem blood and spleen after Gardasil^(■) vaccination—A case report

A same-nested PCR was used to re-amplify the amplicon of a hypervariable region of the HPV-16 L1 gene DNA in the postmortem blood and splenic tissue obtained at autopsy of a formerly healthy teenage girl who suffered a sudden unexpected death in sleep 6 months after 3 intramuscular injections of a quadrivalent HPV vaccine, Gardasil?. A full autopsy analysis revealed no cause of death. The HPV-16 gene DNA detected in the postmortem materials was similar to the HPV-16 gene DNA fragments in Gardasil? in that both were in non-B-conformation, requiring nondegenerate GP6 and MY11 primers to re-amplify the PCR amplicon for detection and to generate a template useful for direct DNA sequencing. A sequence excised from the base-calling DNA sequencing electropherogram was analyzed by Basic Local Alignment Search Tool (BLAST) alignment and a 45 - 60 base sequence fully matched with a standard hypervariable region of the HPV-16 L1 gene retrieved from the National Center for Biotechnology Information database validated the correct genotyping for HPV- 16 L1 gene DNA. These naked non-proliferating HPV- 16 L1 gene DNA fragments appeared to be in the macrophages of the postmortem blood and spleen, and were protected from degradation by binding firmly to the particulate aluminum adjuvant used in vaccine formulation. The significance of these HPV DNA fragments of a vaccine origin found in post-mortem materials is not clear and warrants further investigation.

DNA methylation of the Monoamine Oxidases A and B genes in postmortem brains of subjects with schizophrenia

Aims: We focused on DNA methylation of the promoter regions of the Monoamine Oxidase (MAO) A and B genes from postmortem brains of subjects with schizophrenia. Methods: We determined levels of DNA methylation using genomic DNA samples purified from four brain areas: prefrontal cortex (PFC), hippocampus, occipital cortex and nucleus accumbens (NAc), by a bisulfite sequencing method from seven normal subjects and six subjects with schizophrenia. Results: Although very few methylated CpGs of the MAOA and MAOB genes were detected in male samples, various DNA methylation patterns were present in female samples, and some differences were found in such patterns between normal subjects and subjects with schizophrenia. In the PFC, the average level of methylation of both genes was significantly higher in subjects with schizophrenia than in normal subjects. The content of highly methylated alleles of the MAOA gene in the NAc was significantly associated with schizophrenia, with similar results obtained for the MAOB gene in both the NAc and PFC. Some CpG sites showed higher levels of methylation in schizophrenia than in normal subjects. Conclusions: Levels of methylation were quite high in NAc and PFC in female subjects with schizophrenia compared with those in female normal subjects.

Some worlds about postmortem blood atropine concentrations

Atropine is an anticholinergic drug, used in treatment of spasm and pain. Postmortem blood atropine concentrations tend to be regionally dependent. We reported in this work the analytical findings of atropine in the peripheral and heart blood from a case of suspected death. Atropine was?determined in both peripheral and heart blood by liquid chromatography with tandem mass spectrometry. Towards the reference ranges, the concentration of atropine in the peripheral blood is therapeutic, and in the heart blood is lethal. The high concentration of atropine in the heart blood reflects postmortem redistribution rather than cardiotoxicity. The findings have great implications for forensic toxicology.

Protein S-nitrosylation regulates the energy metabolism of early postmortem pork using the in vitro model

This research aimed to investigate the regulation of energy metabolism by protein S-nitrosylation utilizing the in vitro muscle glycolysis model.Longissimus thoracis(LT)muscles homogenates were treated with nitric oxide donor NOR-3((±)-(E)-4-Ethyl-2-(E)-hydroxyimino-5-nitro-3-hexenamide)and control(0.1 M K_(2)HPO_(4))under different pH conditions(6.5,6.0 and 5.5)in vitro buffer system for 24 h,respectively.Results indicated that the NOR-3 treatment group had a significantly higher pH decline rate than the control group and resulted in a higher lactate accumulation and glycogen degradation at 24 h compared with the control group(p<0.05).Moreover,NOR-3 treatment significantly increased the activities along with S-nitrosylation levels of lactate dehydrogenase and glycogen phosphorylase at pH 6.5 in a concentration-dependent manner(p<0.05).In addition,low pH could weaken the NOR-3 treatment effect and inhibit glycolysis rate.Thus,protein S-nitrosylation could play a role in regulating postmortem glycolysis in vitro model even at low pH conditions.

Zimbabwe Election Postmortem

The people of Zimbabwe cast their votes in a general election on July 31,2013 in the hope of ending the shaky coalition government comprising the ZANU-PF led by President Robert Mugabe,the MDC-T led by Prime Minister Morgan Tsvangirai and the MDC led by Industry and Commerce Minister Welshman Ncube.According to the presidential election results.

The Impact of Gender, Age and Tissues in vitro on Estimat- ing Postmortem Interval by FTIR Spectroscopy

To investigate the influence of such individual factors as gender,age and tissues in vitro to the postmortem interval(PMI) by the Fourier transform infrared(FTIR) spectrometer in animal experiments.SD rats were classified into male and female groups,different age groups(21-day,42-day and 63-day group),and tissues in vitro and in vivo groups.The rats were sacrificed by cervical dislocation,whose bodies were kept in a controlled environmental chamber set at(20±2) ℃ and 50% humidity.The liver,kidney,spleen,myocardium,brain,lung and skeletal muscle tissues were collected for measurement from time zero to 48 h postmortem.With the change of PMI,no obvious changes were found in the main FTIR absorbance peaks and their ratios at different time points.All the experimental groups showed no significant changes when compared with the controls.The gender,age and tissues in vitro were not found to be contributing factors in the estimation of PMI via FTIR spectroscopy.

Postmortem regional distribution of morphine in dependent rats

Objective: Morphine concentration measured in postmortem tissues may or may not reflect antemortem concentration. We measured levels of morphine in autopsied tissues to determine whether morphine distribution in morphine-dependent rats is altered after death. Methods: Solid-phase extraction was used to extract morphine from the samples, and morphine levels were measured at 0-96 h postmortem using gas chromatography. Results: The study of the morphine dependent rats showed a significant (P<0.05) increase of morphine concentration in postmortem cardiac blood, liver tissues and kidneys tissues. A significant increase was also observed at 72 h and 96 h postmortem in the brain, while morphine levels in cardiac tissues only increased at 24 h and 96 h postmortem. These changes were associated with an observed pH rapid decrease: pH of cardiac blood dropped from 7.36±0.15 to 6.86±0.09 (P<0.01), pH of liver tissues from 6.98±0.04 to 6.34±0.03 (P<0.05). Conclusion: The postmortem regional distribution of morphine occurs in dependent rats, but different from the change that occurs in acute poisoning rats. The morphine concentration in cardiac blood and tissues tends to increase during the period of 0-96 h postmortem in dependent rats. Morphine concentration increases with pH rapid decrease. The antemortem internal amount of morphine affects its postmortem regional distribution. It appears that several mechanisms are accountable for postmortem morphine distribution. The understanding of the mechanisms and patterns may eventually lead to better choices of samples which may better represent antemortem drug levels.

Time-Dependent FTIR Spectral Changes in Rats of Massive Hemorrhage Death during the Later Postmortem Period

The aim of the current study was to investigate the spectra in the different organs of the rats which died of massive hemorrhage;to explore their spectral changes 15 days postmortem and the best mathematical model with different band absorption ratio changes to postmortem interval(PMI);and to compare the spectral changes of different temperature.Thirty male Sprague-Dawley rats were sacrificed by cutting abdominal aorta,and the cadavers were divided equally and kept at 4 ℃,20 ℃ and 30 ℃ in the control chamber.From the same rat,seven different organs were sampled at intervals of 1-15 days postmortem,and then measured by Fourier transfom infrared(FTIR) spectrometer.Six mathematical model functions were explored.The absorbance of bands and band absorbance ratios of absorption peak in each organ showed a time-dependent increase or decrease,most band absorbance ratios remaining stable for 7-15 days postmortem.Cubic model functions of the various bands absorbance ratios against PMI showed a stronger related coefficient.The absorbance bands with obvious changes at 20 ℃ showed stabilized tendencies at 4 ℃ and significant changes at 30 ℃ within 15 days postmortem.In addition,FTIR spectroscopy revealed a time-dependent metabolic process,with potential of being used to estimate PMI during 7 days postmortem,which merits further investigation.

Inferring the Postmortem Interval of Rat Cadaver after Boiling Water Treatment Based on Microbial Community Succession

Background:In forensic investigations,accurate estimation of the postmortem interval(PMI)is an important task,but also an ongoing challenge.Especially in cases where the cadaver has been specially treated,for example,by boiling,the determination of PMI becomes extremely difficult.Previous studies have shown that the succession of the microbial community after decomposition of the cadaver can be used to infer PMI.However,the feasibility of determining the PMI of boiled cadavers has not yet been demonstrated.Aims and Objectives:The main objective of this study was to test whether we can infer PMI of boiled cadavers based on the succession of microbial communities.Materials and Methods:SD rats were killed by cervical dislocation.Subsequently,the rat cadavers were divided into the case(boiled cadavers)and control(unboiled cadavers)groups.Rectal samples were collected from the rats for 45 days and at nine time points.High-throughput sequencing of the 16S rRNA gene was performed to characterize the microbial community in the rectum.Results:The results showed that the composition and relative abundance of bacterial communities at the phylum level were significantly different between the case and control groups.The alpha diversity of the microbial community showed a decreasing trend with the decomposition process.Principal coordinate analysis showed that the case and control groups had obvious patterns along the succession of microbial communities.The rectal microbial communities showed a significant linear trend in the time course of decomposition.A random forest model was used to infer PMI.The goodness-of-fit(R2)of the model was 68.00%and 84.00%,and the mean absolute errors were 2.05 and 1.48 days within 45 days of decomposition for the case and control groups,respectively.Conclusions:Our results suggest that microbial community succession could be a potential method to infer PMI of boiled cadavers.

Application and Prospect of Postmortem Imaging Technology in Forensic Cardiac Pathology:A Systemic Review

Postmortem imaging(PMI)technology known as virtual autopsy or virtopsy is regarded as a useful method of noninvasive or minimally invasive autopsy in forensic practice.Postmortem computed tomography is applicable to traumatic investigation,acute hemorrhage,pulmonary parenchyma disease,calcification(calculus,atherosclerosis),and gas accumulation.Postmortem magnetic resonance(PMMR)has been proven to have advantages in soft tissue identification.Cardiac death is one of the keys and difficult points in forensic practice.With the introduction and development of PMCT angiography and PMMR,it was proved to be a very promising tool in the investigation of cardiac death,including vascular cavities and ischemic myocardium.This article reviewed the applications of the latest PMI and its related technologies in forensic cardiac pathology,including advantages,limitations,and development prospects.

Postmortem imaging as a complementary tool for the investigation of cardiac death

In the past 2 decades,modern radiological methods,such as multiple detector computed tomography (MDCT),MDCT-angiography,and cardiac magnetic resonance imaging (MRI)were introduced into postmortem practice for investigation of sudden death (SD),including cases of sudden cardiac death (SCD).In forensic cases,the underlying cause of SD is most frequently cardiovascular with coronary atherosclerotic disease as the leading cause.There are many controversies about the role of postmortem imaging in establishing the cause of death and especially the value of minimally invasive autopsy techniques.This paper discusses the state of the art for postmortem radiological evaluation of the heart compared to classical postmortem examination,especially in cases of SCD.In SCD cases,postmortem CT is helpful to estimate the heart size and to visualize haemopericardium and calcified plaques and valves,as well as to identify and locate cardiovascular devices.Angiographic methods are useful to provide a detailed view of the coronary arteries and to analyse them,especially regarding the extent and location of stenosis and obstruction.In postsurgical cases,it allows verification and documentation of the patency of stents and bypass grafts before opening the body.Postmortem MRI is used to investigate soft tissues such as the myocardium,but images are susceptible to postmortem changes and further work is necessary to increase the understanding of these radiological aspects,especially of the ischemic myocardium.In postsurgery cases,the value of postmortem imaging of the heart is reportedly for the diagnostic and documentation purposes.The implementation of new imaging methods into routine postmortem practice is challenging,as it requires not only an investment in equipment but,more importantly,investment in the expertise of interpreting the images.Once those requirements are implemented,however,they bring great advantages in investigating cases of SCD,as they allow documentation of the body,orientation of sampling for further analyses and gathering of other information that cannot be obtained by conventional autopsy such as a complete visualization of the vascular system using postmortem angiography.