Sequencing mitochondrial DNA from a tooth and application to forensic odontology.

Genetic identification can be complicated by long intervals between the time of death and examination of tissues, and sometimes only bone and teeth may be available for analysis. Several investigators have described the isolation of nuclear DNA from these materials, but all have indicated that the DNA is significantly degraded. Recently, the polymerase chain reaction (PCR) and direct DNA sequencing have enabled rapid and reliable characterization of specific highly polymorphic DNA sequences from different individuals. Above all, mitochondrial DNA sequences offer several unique advantages for the identification of human remains. The isolation of mtDNA from a tooth and the symmetrical PCR amplification and direct DNA sequencing of its most polymorphic regions are reported.

Possibility of paternity testing using RFLP analysis on a very small amount of material.

Paternity testing by DNA analysis was carried out using dental pulpal and chorionic villous tissue from two children respectively, and fresh blood samples obtained from the alleged parents. DNA was extracted spectroscopically from the pulp of an upper wisdom tooth (16 micrograms) and the chorionic villi (53 micrograms). The RFLP method was used for DNA analysis of the parent-child relationships because both of the DNAs extracted had a high molecular weight. Distinct bands were detected with 32P-labelled multi-locus (Myo) and single locus (pYNH24) DNA probes. In the case of the dental specimen all of the bands of the child's DNA were found to be derived from either of the alleged parents, demonstrating a consistent parent-child relationship (the probability of established paternity was 99.86%) whilst in the case of the villous specimen the father-child relationship was denied. This procedure can provide much information using very little material for analysis but where the samples are in a good condition.

The type A behavior pattern and immune reactivity to brief stress: change of volume of secretory immunoglobulin A in saliva.

This article presents findings of a laboratory experiment on the association of the Type A behavior pattern with reactivity of secretory immune functioning to brief stress. 38 female undergraduate students classified as Type A (n = 19) or as Type B (n = 19) on the basis of their scores on the Kwansei Gakuin Type A scale performed a continuous arithmetic task in a situation in which they were exposed to aversive loud noise. Secretory immunoglobulin A (s-IgA) in saliva and autonomic measures (heart rate and frequency of eyeblink) were evaluated before and after the manipulation of stress. The volume of s-IgA at baseline was significantly higher for the Type A group than for the Type B group, suggesting that the former relative to the latter might be chronically higher in mucosal immune functioning. Also, the volume of s-IgA significantly increased after exposure to a brief stress for the Type B group but did not change for the Type A group, a finding which might indicate that the Type A group may have less immune reactivity to a brief stress.

[Two cases of personal identification from dental information].

We describe two cases in which unknown bodies were positively identified from dental information and biochemical examination using tooth materials. In one case, a charred body was positively identified with little effort by comparison of antemortem dental records (dental chart and dental X-ray film) with postmortem data. In the other case, although the unknown individual had dental treatment, the police were unable to obtain the antemortem dental records of the victim. We then conducted biochemical analysis of teeth, facilitating personal identification using DNA analysis and age estimation based on aspartic acid racemization. The mutation obtained from the sequence of mtDNA and the genotypes of HLADQ alpha, HPRTB and ABO blood groups including the data for estimated age supported the kinship between the unknown individual and his mother. The data for maternally inherited mtDNA were of great importance in this case, since it was possible to obtain DNA from the mother. Dental identification in one of the most accurate methods of personal identification if suitable antemortem records are available. In the absence of such records, biochemical analysis of teeth also makes it possible to increase the probability of correct personal identification.

Advantages of dental mitochondrial DNA for detection and classification of the sequence variation using restriction fragment length polymorphisms.

After amplification by polymerase chain reaction (PCR), the nucleotide sequences of a 452-bp section of the D-loop region of human mitochondrial DNA (mtDNA) were determined in 40 teeth extracted from patients living in Kanagawa prefecture, Japan. Dental DNA was extracted separately from the dental pulp and dentin (i.e., the attached pulp cells from the most superficial layer of the pulp cavity wall) of the same tooth. Comparison of the nucleotide sequences of the 452-bp region of the D-loop demonstrated that nucleotide substitutions and insertion/deletion events were identical in material from both sources. Thus, dentin produces equivalent results when the dental pulp of a tooth is unsuitable for mtDNA analysis. To establish the reliability of the screening procedure for the sequence analysis, we identified restriction sites for the enzymes KpnI and MnlI in the 452-bp region of the D-loop. Thirteen of 14 patterns of four polymorphisms analyzed using the mtDNA from the 40 tooth samples were identifiable by an initial screening procedure involving restriction fragment length polymorphism (RFLP) analysis. Combined use of sequence analysis and RFLP analysis proved extremely efficient in analyzing mtDNA polymorphisms, allowing identification of individuals.

Adherence of cyanoacrylate which leaked from gastric varices to the left renal vein during endoscopic injection sclerotherapy: a histopathologic study.

We report a case involving leakage of cyanoacrylate (CA) to the inferior vena cava (IVC) through a gastrorenal shunt and left renal vein. A 72-year-old man with liver cirrhosis was admitted to our hospital to undergo emergency treatment for massive hemorrhage of gastric varices. Endoscopic injection sclerotherapy (EIS) using CA was performed on the varices. Radiographic fluoroscopy revealed that most of the injected CA had adhered firmly to the gastric varices, but a certain portion of the CA had flowed to the IVC through the gastrorenal shunt and left renal vein. At that point, the patient did not complain of any symptoms. However, 6 months later, he died of hepatic failure and an autopsy was performed. Histopathologic examination of the wall of the IVC and renal vein, to which CA had adhered, revealed that the CA was covered with endothelial cells of the vessel and no nearby thrombus was present. Long-term anticoagulant therapy may not be indicated in cases of leakage of CA from the gastric varices to other veins, since the leaked CA may be readily covered with endothelium without thrombus formation as in our patient. It is possible for CA to flow to the IVC and have a fatal impact. Our patient was fortunate, and for safe EIS it is important that these complications are prevented.