Research Progress of Exosomes and Their Forensic Significance.

Exosomes are membranous tiny vesicles secreted by cells, which are widely found in the extracellular matrix and various body fluids and carry a variety of biologically functional molecules such as proteins, lipids, messenger RNA (mRNA) and microRNA (miRNA). Exosomes not only play important biological roles in the field of immunology and oncology, but also have potential application value in the field of forensic medicine. This article reviews the discovery, production and degeneration mechanism, biological functions, isolation and identification methods of exosomes, summarizes the research on exosomes and their significance in the field of forensic science, and discusses their applications in body fluid identification, individual identification, postmortem interval estimation to provide ideas for the application of exosomes in forensic work.

Three new benzolactones from Lavandula angustifolia and their bioactivities.

Three new benzolactones (1-3), together with four known ones (4-7), were isolated from the whole herb of Lavandula angustifolia. Their structures were established on the basis of detailed spectroscopic analysis (1D- and 2D-NMR, HRESIMS, UV, and IR) and comparison with data reported in the literature. New compounds were evaluated for their anti-tobacco mosaic virus (TMV) activities and cytotoxic activities. The results revealed that compounds 1-3 showed obvious anti-TMV activities with inhibition rates of 26.9, 30.2, and 28.4%, which were at the same grade as positive control. Compounds 1-3 also showed weak inhibitory activities against some tested human tumor cell lines with IC50 values in the range of 32.1-7.6 µM.

Bacterial Succession on Rat Carcasses and Applications for PMI Estimation.

UNLABELLED: Abstract: OBJECTIVE: To investigate the bacterial succession on rat carcasses and to evaluate the use of bacterial succession for postmortem interval (PMI) estimation. METHODS: Adult female SD rat remains were placed in carton boxes. The bacterial colonization of circumocular skin, mouth and vagina was collected to be identified using culture-dependent biochemical methods. The changes in community composition were regularly documented. RESULTS: The bacterial succession in three habitats showed that Staphylococcus and Neisseria were predominated in early PMI, especially Staphylococcus aureus and Neisseria lactamica in 6 hours after death. Lactobacillus casei developed on the 3-4 days regularly, and kept stable at a certain level in late PMI. CONCLUSION: The involvement of normal and putrefactive bacteria in three body habitats of rat remains can be used for PMI estimation.

Sesquiterpenoids from Tussilago farfara and their inhibitory effects on nitric oxide production.

Tussilagone (TSL) and its allied sesquiterpenoids were considered as the main active principles of the flower buds of Tussilago farfara, which has been widely used in China as an antitussive herbal medicine. Six new bisabolane-type sesquiterpenoids, tussfararins A-F (1-6), along with 12 known sesquiterpenoids, were isolated from the flower buds of T. farfara. Structures of the new compounds were elucidated by extensive spectroscopic analysis. The biological analysis showed that compounds 1, 3, 6, and 7 inhibited nitric oxide (NO) production in lipopolysaccharide (LPS)-stimulated RAW 264.7 cells with IC50 values of 13.6-24.4 µM.

Genetic polymorphism of nine non-CODIS STR loci in Hunan Province-based Chinese Han population.

OBJECTIVE: To determine the allelic frequency distribution and genetic parameters of nine non-CODIS DNA index systems of the short tandem repeat (STR) loci (D2S1772, D6S1043, D7S3048, D8S1132, D11S2368, D12S391, D13S325, D18S1364, and GATA198B05). METHODS: A total of 353 blood samples were collected, extracted, amplified, and analyzed from unrelated healthy individuals of Han nationality in Hunan Province, China. RESULTS: One hundred and fourteen alleles were observed in the population with corresponding allelic frequencies ranged from 0.001 0 to 0.323 0. For all the nine non-CODIS STR loci, the observed genotypic data showed no significant deviations from the Hardy-Weinberg equilibrium. The Ho, He, PIC, DP, and PE of the studied non-CODIS STR loci ranged from 0.1080 to 0.1950, 0.8050 to 0.8920, 0.7700 to 0.8600, 0.9250 to 0.9660 and 0.6070 to 0.7800, respectively. CONCLUSION: Nine non-CODIS STR loci have high degrees of polymorphisms, which may be useful in individual forensic identification and parentage testing in forensic practice.

[Determination of chlorogenic acid, rutin, scopoletin and total polyphenol in tobacco by Fourier transform near infrared spectroscopy].

The objective of the present study was to investigate the feasibility of predicting chlorogenic acid, rutin, scopoletin and total polyphenol in tobacco by Fourier transform near-infrared (FT-NIR) spectroscopy. The partial least squares(PLS) regression method, second derivative and Norris derivative filter were applied in the NIR spectroscopy prediction of chlorogenic acid, rutin, scopoletin and total polyphenol in the range of 7 500 to 4 000 cm(-1). For chlorogenic acid, rutin, scopoletin and total polyphenol, the determination coefficients were 0.976 6, 0.941 9, 0.957 1 and 0.966 6, respectively. The SEP/SEC values for them were < 1.2, and the SD/SEP values for them were > 2. The root mean square error of cross validation (RMSECV) of the four calibration models were 1.938 9, 1.046 2, 0.047 9 and 2.745 2, respectively. NIR spectroscopy was compared with the conventional methods. The results show that the two methods showed no significant difference at the significant level of 0.05. NIR spectroscopy technology can accurately analyze chlorogenic acid, rutin, scopoletin and total polyphenol in tobacco.

[Reflection of estimating postmortem interval in forensic entomology and the Daubert standard].

Estimating postmortem interval (PMI) is always the emphasis and difficulty in forensic practice. Forensic entomology plays a significant indispensable role. Recently, the theories and technologies of forensic entomology are increasingly rich. But many problems remain in the research and practice. With proposing the Daubert standard, the reliability and accuracy of estimation PMI by forensic entomology need more demands. This review summarizes the application of the Daubert standard in several aspects of ecology, quantitative genetics, population genetics, molecular biology, and microbiology in the practice of forensic entomology. It builds a bridge for basic research and forensic practice to provide higher accuracy for estimating postmortem interval by forensic entomology.

[Identification of sarcosaphagous Calliphorid flies by analyzing the sequence of 16S rDNA].

OBJECTIVE: To explore the application of a 289bp fragment of the 16S rDNA gene to identify various species of sarcosaphagous Calliphorid flies. METHODS: Twenty-six Calliphorid flies were collected from 14 Chinese provinces. All specimens were properly assigned into three genera and six species. The DNA of the pectoralis was extracted using CTAB method. Then PCR amplification was done for the 289 bp fragment of the 16S rDNA gene. The PCR products were then purified and sequenced, and the obtained sequences were uploaded to GenBank. The phylogenetic tree was built by the neighbor-joining method and intraspecific and interspecific divergences were calculated by sequence analysis. RESULTS: The above 26 sarcosaphagous flies could be well clustered according to different genera and species. The evolutional intraspecific values were all zero, the evolutional interspecific variations varied from 0.3% to 6.5%. CONCLUSION: The 289 bp fragment of the 16S rDNA of sarcosaphagous flies can be effectively used to identify most of the flies at species level. This method appears to be fast and low dissipative, which might be used to estimate postmortem interval by sarcosaphagous flies.

Case Report: Identification of the First Synonymous Variant of Myosin Binding Protein C3 (c.24A>C, p.P8P) Altering RNA Splicing in a Cardiomyopathy and Sudden Cardiac Death Case.

Background: Sudden cardiac death (SCD), based on sudden cardiac ejection cessation, is an unexpected death. Primary cardiomyopathies, including dilated cardiomyopathy (DCM), are one of main causes of SCD. The DCM is characterized by a cardiac dilatation and a reduced systolic function with a prevalence of 1/250 in adults. The DCM has been reported with more than 60 disease-causing genes, and MYBPC3 variants are one of the most common and well-known causes of DCM. Methods: We identified a 29-year-old female who died of SCD. We performed a whole-exome sequencing (WES) to detect her genetic etiology and used minigene modeling and immunohistochemistry staining to verify the pathogenicity. Results: We determined that the woman died of SCD caused by DCM due to an identified novel synonymous variant of MYBPC3 (NM_000256.3: c.24A>C, p.P8P) in the deceased. The variant can result in abnormal splicing, which was confirmed by minigene models and immunohistochemistry staining. Conclusion: We may have identified the first deleterious synonymous variant of MYBPC3 in an SCD case and verified its significant impact on RNA splicing. Our description enriched the spectrum of MYBPC3 variants and emphasized the significance of synonymous variants that are always disregarded in genetic screening.

[Comparison of mtDNA extracting methods for common sarcosaphagous insects].

OBJECTIVE: To compare effects of three different methods for mtDNA extraction from common sarcosaphagous insects including cetyl trimethyl ammonium bromide (CTAB) method, sodium dodecyl sulfate-potassium acetate (SDS-KAc) method and sodium dodecyl sulfate-proteinase K (SDS-PK) method. METHODS: Seventy-two insects from four species [Chrysomya megacephala (Fabricius, 1784), Eusilpha bicolor (Fairmaire, 1896), Paraeutrichopus pecoudi (Mateu, 1954), Vespa velutina (Lepeletier, 1836)] were collected from the corpses of the rabbits in Changsha district. The total DNA of above samples was extracted by CTAB, SDS-Kac and SDS-PK methods. The purity and concentration of DNA were examined by protein-nucleic acid spectrophotometry, and mtDNA were amplified by specific primers and PCR products were detected by agarose gel electrophoresis. Then PCR products were sequenced and subsequently up-loaded to GenBank. RESULTS: mtDNA was successfully extracted with three methods from most of the samples. The SDS-PK method was better in DNA purity compared to other methods and the CTAB method was superior in extracting DNA from old samples, while SDS-KAc method showed no significant difference for extraction effects of different samples. CONCLUSION: The most appropriate method should be chosen depending on different situations. SDS-PK method is expected to obtain high-quality DNA, while CTAB method is preferred in extracting obsolete samples. SDS-KAc method is low cost and can be used in various kinds of preliminary experiments.

Mesenchymal stem cells desensitize castration-resistant prostate cancer to docetaxel chemotherapy via inducing TGF-ß1-mediated cell autophagy.

BACKGROUND: Mesenchymal stem cells (MSCs) have been proved to drive castration resistant prostate cancer (CRPC). In this study, we aim to investigate the contribution of MSCs to the development of docetaxel resistance in CRPC cells and its potential mechanisms. METHODS: The effect of MSCs on CRPC cells resistance to docetaxel was determined using in vivo and in vitro approaches. CCK8 and PI/Annexin V-FITC assay were used to examined the cell viability and apoptosis. The concentration of transforming growth factor-ß1 was measured by enzyme-linked immunosorbent assay and small interfering RNA was used for functional analyses. RESULTS: MSCs significantly reduced the sensitivity of CRPC cells to docetaxel-induced proliferation inhibition and apoptosis promotion in vivo and in vitro. CRPC cells cocultured with MSCs under docetaxel administration have an increased autophagy activation, while autophagy inhibitor could effectively reversed MSCs-induced resistance to docetaxel. Additionally, MSCs-induced CRPC cell autophagy increase under docetaxel administration depends on MSCs secreting TGF-ß1 and inhibition of TGF-ß1 secretion in MSCs could consequently increase the sensitivity of CRPC cells to docetaxel. CONCLUSIONS: These results suggest that docetaxel administrated CRPC cells may elicit MSCs secreting TGF-ß1 increase, which desensitizes CRPC to docetaxel chemotherapy accelerating chemoresistance occurrence via inducing cell autophagy.

[Forensic analysis of 74 tumor related medical malpractice cases].

OBJECTIVE: To analyze the causes of medical malpractice in patients with tumor, to determine the medical responsibility, and to recommend the related preventions. METHODS: Seventy four medical malpractice cases, which were involved in tumor and collected from 2000 to 2009 in medicolegal expertise center of west China, were analyzed retrospectively. RESULTS: The medical malpractice cases in the patients with tumor showed an increasing tendency in recent years. The main causes are missed diagnosis, misdiagnosis, improper chemotherapy and neglect of complications. The causes of medical malpractice were different in the different levels of medical services. The occurrence of medical malpractice in surgery and OB-GYN showed more frequent than the others. CONCLUSION: Forensic pathology autopsy is important to resolve medical malpractice of tumor patients by finding out the cause of death and clarifying the medical responsibility. The occurrence of medical malpractice could be reduced by the clinical doctors through improving serve consciousness, obtaining the patients' trust, improving the medical treatment, following related laws and rules, fulfiling duty of medical careness.

[Comparison of mtDNA extraction from different parts of sarcosaphagous insects].

OBJECTIVE: To explore mitochondrial DNA (mtDNA) extraction effects of different parts from sarcosaphagous insects using improved cetyltriethylammnonium bromide (CTAB) method. METHODS: Thirteen Lucilia sericata (Meigen) and 13 Nicrophorus fossor (Erichson) were collected from the corpses of rabbits placed on the outdoor lawn in Huhehot district. Four parts (head, chest muscle, legs and wings) of insect were collected, and the mtDNA of all samples were extracted using CTAB method. The purity and concentration were tested using protein and nucleic acid spectrophotometry. The integrity of the extracted mtDNA and PCR products were checked by agarose gel electrophoresis. The PCR products were sequenced and the obtained sequences were imputed into GenBank for comparison. RESULTS: mtDNA were successfully extracted from 10 head samples, 6 legs samples, 4 wing samples and 13 chest muscle samples of the Lucilia sericata (Meigen). Also, mtDNA were successfully extracted from 5 head samples, 8 legs samples, 3 wing samples and 13 chest muscle samples of the Nicrophorus fossor (Erichson). CONCLUSION: mtDNA can be obtained from chest muscle and other parts of sarcosaphagous insects using the improved CTAB method.

[The statin dosage for achieving goal of cholesterol-lowering based on risk stratification in patients with ischemic cerebrovascular diseases].

OBJECTIVE: To explore statin dosages for targeting goal of LDL-C lowering on the basis of stroke risk stratification and the dosage-effective relation of statin and LDL-C lowering in Chinese patients with ischemic stroke and transient ischemic attack (TIA). METHODS: This is a prospective and open clinical trial patients with ischemic stroke/TIA within 6 months were enrolled and the dosages of atorvastatin were calculated based on risk stratification according to "Chinese Consensus for Prevention of Ischemic Stroke/TIA with Statin" (Chinese Consensus). A dose of 10 mg of atorvastatin daily to target LDL-C goal was taken as the standard dosage targeting goal (SDTG). Patients taking this dosage of atorvastatin constituted a SDTG group. Those who needed a daily dose of 20 mg or more of atorvastatin were randomized into an intensive dosage targeting goal (IDTG) group (atorvastatin 20 - 80 mg/d) and a standard dosage non-targeting goal (SDNTG) group (atorvastatin 10 mg/d without targeting goal). All patients took atorvastatin for 12 weeks. The primary outcome was the rate of targeting goal for LDL-C lowering at 2, 4 and 12 weeks, respectively and the secondary outcome was the occurence of recurrent stroke and other vascular events within 12 weeks. The main safety endpoint was serial adverse events including symptomatic intracranial hemorrhage. RESULTS: Altogether 102 cases were enrolled and 99 cases were followed up for 12 weeks. According to the Chinese Consensus, the rate of high risk, very high risk-I and very high risk-II was 44%, 28% and 28%, respectively. Targeting rate for LDL-C lowering was 77% - 85% at each time point in the SDTG and IDTG groups, being significantly higher than those in the SDNTG group (12% - 16%, P < 0.01). No significant difference was found concerning the occurrence of recurrent stroke, other vascular events and safety endpoints among the three groups. The amplitude of LDL-C lowering was 32% - 35%, 46% - 49%, 51% - 52% and 60% - 65% with corresponding to daily dosage of 10 mg, 20 mg, 40 mg and 80 mg atorvastatin. CONCLUSIONS: At least more than half of the patients after ischemic stroke/TIA need intensive statin therapy to target the LDL-C lowering goal. The dosage-effective relation of atorvastatin and LDL-C lowering in Chinese is similar to the reported data in other races.

[Study on the distribution and location of neuroglobin in human tissues and cells].

OBJECTIVE: To explore the distribution and location of neuroglobin (NGB) and its function in human tissues and cells. METHODS: The distribution and localization of NGB in human tissues and cells were examined by immunohistochemical method. RESULTS: NGB-positive cells were mainly distributed in neurons of central nervous system and peripheral nervous system, and some in endocrine tissues and genital system. NGB-immunoreactive product located in the cytoplasm of these cells. CONCLUSION: The expression of NGB in human nervous tissues, some endocrine tissues and genital system suggested that NGB might play an important role in the utilizations of oxygen and physiological functions.

Diagnostic value of STAF score in combination with D-dimer in cardioembolism.

The aim of this study was to evaluate the diagnostic value of the Score for the Targeting of Atrial Fibrillation (STAF) in combination with the serum D-dimer (DD) levels in cardioembolism(CE).This study was a retrospective case-onlystudy, consecutively including patients with acute ischemic stroke. All patients were evaluated following the STAF scoring criteria and were classified according to the Trial of ORG 10172 in Acute Stroke Treatment (TOAST) etiology classification criteria. A total of 317 patients were enrolled, including 37 CE cases (11.67%). STAF ≥5 showed a sensitivity of 89% and a specificity of 91% for the diagnosis of CE, whereas DD >791.30 ng/mL had a sensitivity of 58% and a specificity of 78%. When the STAF was used in combination with the DD level, the sensitivity was 95%, and the specificity was 100%.STAF score is an excellent tool for the diagnosis of CE when combined with DD, and can facilitate the etiological classification of acute ischemic stroke.

Discovering biomarkers in bladder cancer by metabolomics.

It has become increasingly clear that the development of cancer, a multifactorial disease, cannot be explained by a single molecule or gene mutation. As a new discipline, metabolomics focuses on the body's metabolite changes, and attempts to find differences to explain the development of cancer; it has proven to be effective and credible. Metabolic studies of bladder cancer (BCa) lag behind those of other tumors. This review systematically outlines the specific process of metabolomics and the use of metabolomics in BCa studies in recent years. We have reviewed the in vitro cell line, bladder tumor tissue and biofluid (urine, plasma and serum) studies used in metabolomics analyses of BCa. The advantages and drawbacks of the use of different samples were compared. Based on the available studies, we have further described the aberrant metabolic pathways of BCa and have suggested some metabolites that may be potential biomarkers for BCa detection.

An investigation and pathological analysis of two fatal cases of cadmium poisoning.

Pollution associated with population growth, and with industrial and urban development has led to a serious decline in the water quality of Chinese rivers. Cadmium (Cd) is recognized as one of the most toxic metals and is strongly accumulated by organisms. Humans are exposed to cadmium originating from the environment and from industrial pollution. In spite of thousands of published studies on Cd, there is little information on its pathological features seen in human autopsy. The gross and pathological findings of forensic autopsies of two case of cadmium poisoning are presented and related to an epidemiological investigation. In both cases, multiple organ damage was observed, involving brain, lung, liver, kidney, red blood cells, and platelets, which is consistent with reports in the literature. In particular, in both cases, transmission electron microscopy revealed a large number of dense lysosomal and phagocytic particles in the cytoplasm near the nucleus, indicating the need for a genotoxic study of cadmium. Our observations provide new clues for the future recognition and prevention of Cd poisoning.

[Determination of caffeine in teas by supercritical fluid chromatography].

A rapid supercritical fluid chromatographic method for the determination of caffeine in tea is described. It is shown that a mobile phase of carbon dioxide, modified with 5% methanol achieved excellent results. The method is simple in the sample preparation. Other components contained in the sample do not interfere the measurement. It can be used to determine caffeine in tea rapidly.