RGC32, a novel p53-inducible gene, is located on centrosomes during mitosis and results in G2/M arrest.

To identify target genes for the hemizygous deletions of chromosome 13 that are recurrently observed in malignant gliomas, we performed genome-wide DNA copy-number analysis using array-based comparative genomic hybridization and gene expression analysis using an oligonucleotide-array. The response gene to complement 32 (RGC32) at 13q14.11 was identified as a deletion target, and its expression was frequently silenced in glioma cell lines compared with normal brain. Levels of RGC32 mRNA tended to decrease toward higher grades of primary astrocytomas, especially in tumors with mutations of p53. Expression of RGC32 mRNA was dramatically increased by exogenous p53 in a p53-mutant glioma cell line, and also by endogenous p53 in response to DNA damage in p53+/+ colon-cancer cells, but not in isogenic p53-/- cells. Chromatin immunoprecipitation and reporter assays demonstrated binding of endogenous p53 protein to the promoter region of the RGC32 gene, implying p53-dependent transcriptional activity. Transiently and stably overexpressed RGC32 suppressed the growth of glioma cells, probably owing to induction of G2/M arrest. Immunocytochemical analysis revealed a concentration of RGC32 protein at the centrosome during mitosis. RGC32 formed a protein complex with polo-like kinase 1 and was phosphorylated in vitro. These observations implied a novel mechanism by which p53 might negatively regulate cell-cycle progression by way of this newly identified transcriptional target. Our results provide the first evidence that RGC32 might be a possible tumor-suppressor for glioma, that it is directly induced by p53, and that it mediates the arrest of mitotic progression.

Comparison of the likelihood ratio and identity-by-state scoring methods for analyzing sib-pair test cases: a study using computer simulation.

To assess the power and significance of the likelihood ratio (LR) and the identity-by-state scoring (IBS) methods for a pair of siblings, we performed computer simulations by use of 10 DNA markers (HLA-DQalpha, D1S80, and 8 short tandem repeat loci) that were frequently analyzed in paternity tests in Japan. The combined power of discrimination of these 10 markers in the Japanese population is 0.999 999 999 98. Pedigrees each consisting of 10,000 pair of full-siblings, half-siblings and unrelated individuals were generated and typed on all markers as random samples. Both the summation of log10 LR and IBS of each group had approximate standard normal distribution with significant differences between the means. Statistical studies showed that the LR method has 91% power to detect unrelated individuals and 38% power to detect half-siblings as not full-siblings with a 5% false-positive rate, whereas the IBS method does 87% and 42% powers, respectively. In 62% of full-siblings, in contrast with only 0.2% of unrelated individuals, the values of LR exceeded 100 which was equivalent to 0.99 of probability of full-sibship at 50% prior probability. The advantage of the LR method over IBS method was convincing especially for the detection of unrelated individuals as not half-siblings, however, the latter would be also informative for sib-pair tests if sufficient number of polymorphic markers are available.

Forced oral opening for cadavers with rigor mortis: two approaches for the myotomy on the temporal muscles.

Forensic dentistry plays an essential role in personal identification procedures. An adequate interincisal space of cadavers with rigor mortis is required to obtain detailed dental findings. We have developed intraoral and two directional approaches, for myotomy of the temporal muscles. The intraoral approach, in which the temporalis was dissected with scissors inserted via an intraoral incision, was adopted for elderly cadavers, females and emaciated or exhausted bodies, and had a merit of no incision on the face. The two directional approach, in which myotomy was performed with thread-wire saw from behind and with scissors via the intraoral incision, was designed for male muscular youths. Both approaches were effective to obtain a desired degree of an interincisal opening without facial damage.

A histological study on the mechanism of epidermal nuclear elongation in electrical and burn injuries.

Epidermal nuclear elongation is one of the most important signs for the diagnosis of electrical injury. In this study, we investigated the mechanism responsible for this phenomenon by comparing the findings from burn injuries and those from contusions. Electrical and burn injuries were made in the dorsal skin of rats using energy ranging from 100 to 790 joules for electrical injury, and 170-690 joules for burn injury. Contusions were also made by compressing the skin with a vice. In electrical and burn injuries, the dermis under the epidermal elongated nuclei was homogeneous and without empty spaces between collagen bundles and the number of dermal fibroblasts per 0.01 mm2 below the damaged epidermis decreased significantly (P < 0.05). The incidence of this change correlated with the depth of denatured dermal collagen fibres and in both types of injuries, dermal cells had no nuclear antigenicity for ubiquitin. The width of the injured epidermis with nuclear elongation decreased significantly (P < 0.05) and the elongated nuclei were parallel to the basal membrane. In electrical injury however, nuclear elongation occurred more frequently near the external root sheath. Nuclear elongation of fibroblasts and external root sheath cells was also found, but those of sebaceous gland cells were not detected. Epidermal elongated nuclei were also found in contusions. The evidence strongly suggests that epidermal nuclear elongation in electrical and burn injuries is due to dermal expansion by heat.

Hypoplasia of the left lobe of the liver.

In a forensic autopsy, hypoplasia of the left lobe (lateral segment) of the liver was found. Although the cause was unknown, the morphologic anomaly observed in this case was regarded as congenital because there was no history of surgery. Moreover, the main vessels of the left lobe were present despite complete absence of the parenchyma.

An application of PCR-single strand conformation polymorphism to MN genotyping.

A PCR-based genotyping of MN blood group system was investigated for DNA samples taken from a population of 409 northern Japanese. DNA fragment (257bp) including exon 2 of glycophorin A (GPA) gene, in which encodes the determinants of MN antigens, was specifically amplified. On the analysis of PCR-single-strand conformation polymorphism (PCR-SSCP) for M alleles, band patterns of M(G) and M(T) were easily discriminated each other. For N alleles, three band patterns were observed, and we tentatively named these alleles as N(1), N(2) and N(V). The N(1) allele appeared predominantly and N(2) had two base substitutions at 1st (C-->A) and 56th (C-->T) in exon 2 of N(1). The other N(V), which was detected from a pair of a mother and her child, possessed a single base substitution at 23rd (A-->G) in intron 2. The allele frequencies of M(G), M(T), N(1) and N(2) were 0.4450, 0.0978, 0.4303 and 0.0269, respectively. The polymorphism information content and the probability of paternity exclusion by this MN genotyping were estimated to be 0.5252 and 0.3219, respectively.

Sudden infant death with anomalous origin of the left coronary artery from the pulmonary artery.

Autopsy of a 2-month-old boy revealed an anomalous origin of the left coronary artery from the pulmonary trunk. The endomyocardial fibrosis associated with myocardial ischemia was noted in the left ventricular wall of the enlarged heart. Death was attributed to silent heart failure due to the anomalous left coronary artery. Significant clinical symptoms are generally observed in patients with such anomalies, however, they might cause sudden death in childhood.

Simple methods of forced oral opening for cadavers oriented to causes of fixed jaw.

Forensic dentistry plays an essential role in personal identification procedures. An adequate interincisal space of relatively fresh cadavers would be required to obtain detailed dental findings. We developed novel and simple procedures of oral opening which are oriented to the causes of the fixed jaw and conscious of facial damage. Preliminary inspection disclosed three causes of the difficulty of oral opening, namely exceptional drying or parchmenting, rigor mortis and loss of extensibility of the temporalis muscles. Tenotomy on the temporalis muscles for cases with the last two factors, and/or mandibular periosteal stripping for those with the first one were employed to approach the oral cavity of cadavers, and a desired degree of interincisal opening was attained. Single or combinational operation of these methods is definitely effective to obtain odontological data without altering facial configuration.