Akiko Tsuji

Estimating age of humans based on telomere shortening.

To estimate age using DNA based on telomere shortening, we determined the terminal restriction fragment (TRF) length, as telomere length, using Southern blot analysis of peripheral human blood and blood stains. All blood stains had been stored at room temperature for 5 months. The average TRF length clearly showed a tendency to shortening with aging. The formula for age estimation was based on a correlation between average TRF length and age of the subjects. The estimated age calculated from TRF length widely depends on environmental and genetic factors. However, as long as the DNA is well preserved, use of our method is feasible regardless of age of the subject and can give a rough estimation of age of subjects in forensic samples that carry no morphological information.

Age estimation in dental pulp DNA based on human telomere shortening

Age estimation based on evidence found in teeth has received considerable attention within the field of forensic science. We determined the terminal restriction fragment (TRF) length, as telomere length, to estimate age. Using dental pulp DNA we found the average TRF length showed a tendency to shortening with aging. Our findings show that telomere shortening, based on dental pulp DNA is a new and useful approach to estimate age of the subject at the time of death.

Pattern of poisoning in Japan: selection of drugs and poisons for systematic toxicological analysis

Patterns of poisoning are known to be different in different countries, because of the local environmental, cultural, and religious situations. Therefore, in Japan, it is important to know the pattern of poisoning in our own country and to prepare for every poisoning case by establishing an efficient systematic toxicological analysis system in forensic practice. We conducted a retrospective study of the kinds of compounds causing poisonings and the frequency of their use based on two series of reports dealing with poisoning cases in Japan prepared by the National Research Institute of Police Science and the Japanese Society of Legal Medicine for 2003 to 2006. From these reports, 459 and 177 compounds, respectively, were extracted as poisonous compounds over the study period. After data analysis, we selected 314 drugs and poisons as important target compounds for systematic drug analysis in Japan; they included 36 volatile compounds, 14 abused drugs, 170 medical drugs, 60 pesticides, 13 natural toxins, and 21 others. This is the first study to show the toxic drugs and poisons to be analyzed in Japan based on frequency of use, and as such the list will be useful in establishing the most efficient screening system in forensic practice.

Construction of calibration-locking databases for rapid and reliable drug screening by gas chromatography-mass spectrometry

Unique calibration-locking databases were constructed for rapid and semiquantitative drug screening by gas chromatography-mass spectrometry (GCMS). In addition to the free-drug database of 127 drugs, a drug database with acetylating reagents was constructed to increase the number of detectable compounds in the analysis by GC-MS; 156 drugs, including 30 drugs of abuse, 42 hypnotics and their metabolites, 18 antipsychotic drugs, 15 antidepressants, and 12 antipyretic analgesic agents, were registered with parameters, such as the mass spectrum, retention time, qualifier ion/target ion percentage, and calibration curve using the novel GC-MS software NAGINATA. Diazepam-d5 was used as internal standard for construction of each calibration curve in the range of 0.01–5.0 μg/ml for most drugs. We examined the applicability of the constructed database to analyzing whole blood samples spiked with 40 drugs most commonly encountered in toxicological cases in Japan. The drugs in blood were extracted using enhanced polymer columns (Focus), subjected to GC-MS after incubation with acetylating reagents, and screened by the drug database. Among the 40 drugs examined, 38 and 30 drugs were successfully identifi ed at the level of 1 and 0.1 μg/ml, respectively, without using standard compounds. The time required for data analysis was less than 1 min, and semiquantitative data were also obtained simultaneously. Because new drugs and metabolites can easily be added to the databases, we can recommend them as useful tools in clinical and forensic toxicological screening.

Genetic polymorphisms of methylenetetrahydrofolate reductase and aldehyde dehydrogenase 2, alcohol use and risk of colorectal adenomas: Self-Defense Forces Health Study

Methylenetetrahydrofolate reductase is a key enzyme in folate metabolism, which affects DNA synthesis and methylation and is possibly linked to colorectal carcinogenesis. Alcohol and acetaldehyde have an adverse effect on folate metabolism. This study investigated the relationship of functional MTHFR C677T and ALDH2 polymorphisms to colorectal adenomas with reference to alcohol consumption in a case‐control study of male officials in the Self‐Defense Forces (SDF) who received a preretirement health examination at two SDF hospitals. The study subjects were 452 cases of colorectal adenoma and 1050 controls with no polyp who underwent total colonoscopy. Genotypes were determined by the PCR‐RFLP method using genomic DNA extracted from the buffy coat. Statistical adjustment was made for age, hospital, rank in the SDF, body mass index, cigarette‐years and alcohol intake. Neither MTHFR C677T nor ALDH2 showed a measurable association with colorectal adenoma. While high alcohol consumption was associated with a moderately increased risk of colorectal adenoma, neither of the two polymorphisms showed a significant effect on the association between alcohol and colorectal adenoma. Individuals with the variant alleles ALDH2*2 and MTHFR 677T had a decreased risk of colorectal adenomas, showing adjusted odds ratios of 0.70 (95% confidence interval 0.49–1.00) for all adenomas and 0.57 (0.34–0.95) for large adenomas (≥ 5 mm), as compared to individuals with ALDH2*1/1 and MTHFR 677CC genotypes combined. The findings may be interpreted as suggesting that folate inhibits the growth of colorectal adenomas, but further confirmation is needed. (Cancer Sci 2005; 96: 513 –518)

Electrolyte analysis of pleural effusion as an indicator of drowning in seawater and freshwater

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.

Unusual death of a baby: a dog attack and confirmation using human and canine STRs

We performed an autopsy on a 3-month-old baby boy who had only one area severe and extensive wound to his head and face. Three unrelated miniature dachshunds were in the house. After our investigation, we were able to confirm that the wound had in fact been caused by a dog attack, and we were able to identify the offending dog among the three dogs using both human and canine short tandem repeat obtained from samples taken from the suspected dog and from the scene of the attack.

Forensic significance of conduction system abnormalities as a precise cause of accidental death

Abstract. We examined the pathology findings of the cardiac conduction system in 42 human autopsies who were considered to have died an accidental death. Abnormalities in serial sections of the conduction system were found in seven and of these three had an accessory pathway in the conduction system. In one of these there was mild Ebsteins anomaly (ME), two had fibromuscular dysplasia of the atrioventricular node artery with mitral valve prolapse, and two others had abnormal routes showing fragmentation of the bundle of His with ME. A full investigation of the cardiac conduction system can therefore be useful for determining the precise cause in cases of accidental death.

Sudden unexpected death following stellate ganglion block

A 29-year-old woman, who had been suffering from left facial palsy, died 3.5 hours after undergoing stellate ganglion block (SGB). Autopsy revealed subcutaneous emphysema of the body, bilateral pneumothorax and a huge post-tracheal hematoma. The lower half of the trachea was markedly flattened by pressure from this hematoma. The cause of death was certified as airway obstruction due to a post-tracheal hematoma following SGB. The doctor cut both the anterior and the posterior wall of the trachea, because the trachea was flattened by the hematoma. He then intubated and sent air into the post-tracheal space. We consider that the patient could have recovered if she had received the correct tracheotomy. We therefore conclude that there was professional error on the part of the doctor in the form of medical malpractice.

Pulmonary fat embolism induced by exposure to high ambient temperature in rats with a fatty liver

The aim of this study was to investigate whether a fatty liver contributes to pulmonary embolism under a high ambient temperature. As an experimental model, we exposed fatty liver rats to a high temperature (45°C) and then looked for fat emboli in the alveolar capillaries using the fat-staining method. Fat emboli were detected in the alveolar capillaries of the fatty liver rats, but not in those of the normal liver rats. Moreover, the degree of pulmonary fat embolism tended to become more severe in proportion to the severity of the fatty liver. In addition, fat emboli did not appear at a core body temperature of 40°C, but were detected at a core body temperature of 44°C. From these results, we conclude that a fatty liver may contribute to the formation of pulmonary fat embolism and that high temperatures act as a trigger for the onset of pulmonary embolism. Moreover, it is possible that fatty liver affects the development of heat stroke induced by exposure to a high ambient temperature and that pulmonary fat embolism is a significant finding which helps to enable a diagnosis of heat stroke in autopsy cases.